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Sökning: WFRF:(Henning Petra 1974)

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1.
  • Magnusson, Maria K, 1972, et al. (författare)
  • Adenovirus 5 vector genetically re-targeted by an Affibody molecule with specificity for tumor antigen HER2/neu.
  • 2007
  • Ingår i: Cancer gene therapy. - : Springer Science and Business Media LLC. - 0929-1903 .- 1476-5500. ; 14:5, s. 468-79
  • Tidskriftsartikel (refereegranskat)abstract
    • In order to use adenovirus (Ad) type 5 (Ad5) for cancer gene therapy, Ad needs to be de-targeted from its native receptors and re-targeted to a tumor antigen. A limiting factor for this has been to find a ligand that (i) binds a relevant target, (ii) is able to fold correctly in the reducing environment of the cytoplasm and (iii) when incorporated at an optimal position on the virion results in a virus with a low physical particle to plaque-forming units ratio to diminish the viral load to be administered to a future patient. Here, we present a solution to these problems by producing a genetically re-targeted Ad with a tandem repeat of the HER2/neu reactive Affibody molecule (ZH) in the HI-loop of a Coxsackie B virus and Ad receptor (CAR) binding ablated fiber genetically modified to contain sequences for flexible linkers between the ZH and the knob sequences. ZH is an Affibody molecule specific for the extracellular domain of human epidermal growth factor receptor 2 (HER2/neu) that is overexpressed in inter alia breast and ovarian carcinomas. The virus presented here exhibits near wild-type growth characteristics, infects cells via HER2/neu instead of CAR and represents an important step toward the development of genetically re-targeted adenoviruses with clinical relevance.
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2.
  • Myhre, Susanna, 1974, et al. (författare)
  • Decreased immune reactivity towards a knobless, affibody-targeted adenovirus type 5 vector
  • 2007
  • Ingår i: Gene Therapy. - : Springer Science and Business Media LLC. - 0969-7128 .- 1476-5462. ; 14:4, s. 376-381
  • Tidskriftsartikel (refereegranskat)abstract
    • In this study, a prototype Adenovirus type 5 (Ad5) vector deleted of the fiber knob domain and carrying an Affibody molecule as the targeting ligand showed decreased susceptibility to human pre-existing antibodies. This vector, Ad5/R7-Z(taq)Z(taq), has short fibers carrying seven shaft repeats, a non-native trimerization signal and an affibody molecule (Z(taq)) reactive to Taq polymerase. Ad5/R7-Z(taq)Z(taq) could be specifically targeted to 293 cells stably expressing membrane-bound anti-Z(taq) idiotypic affibody called Z(ztaq) (293Z(ztaq)). Sera from 50 blood donors were analyzed for neutralization activity (NA) against the parental Ad5/Fiwt vector and knobless Ad5/R7-Z(taq)Z(taq) on 293Z(ztaq) cells. Twenty-three sera had NA titers (>= 1:64) against Ad5/Fiwt (46%) and only two against Ad5/R7-Z(taq)Z(taq) (4%). Characterization of sera with NA titers showed that the knob domain is one of the targets of the antibodies. Neutralization assays using sera pre-adsorbed on knob and hexon proteins showed that the NA of the sera was carried mainly by anti-knob and anti-hexon antibodies, but in certain sera the anti-hexon antibodies represent the major population of the neutralizing antibodies (NAbs). Our results suggested that a combination of knob deletion and hexon switching could be an effective strategy for Ad vectors to better evade the anti-Ad NAbs.
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3.
  • Myhre, Susanna, 1974, et al. (författare)
  • Re-targeted adenovirus vectors with dual specificity; binding specificities conferred by two different Affibody molecules in the fiber.
  • 2009
  • Ingår i: Gene therapy. - : Springer Science and Business Media LLC. - 1476-5462 .- 0969-7128. ; 16:2, s. 252-61
  • Tidskriftsartikel (refereegranskat)abstract
    • Vectors based on Adenovirus type 5 (Ad5) are among the most common vectors in cancer gene therapy trials to date. However, for increased efficiency and safety, Ad5 should be de-targeted from its native receptors and re-targeted to a tumor antigen. We have described earlier an Ad5 vector genetically re-targeted to the tumor antigen HER2/neu by a dimeric version of the Affibody molecule ZH inserted in the HI-loop of the fiber knob of a coxsackie and adenovirus receptor-binding ablated fiber. This virus showed almost wild-type growth characteristics and infected cells through HER2/neu. Here we generate vectors with double specificity by incorporating two different Affibody molecules, ZH (HER2/neu-binding) and ZT (Taq polymerase-binding), at different positions relative to one another in the HI-loop. Receptor-binding studies together with viral production and gene transfer assays showed that the recombinant fiber with ZT in the first position and ZH in the second position (ZTZH) bound to both its targets, whereas surprisingly, the fiber with ZHZT was devoid of binding to HER2/neu. Hence, it is possible to construct a recombinant adenovirus with dual specificity after evaluating the best position for each ligand in the fiber knob.
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4.
  • Stattin, Eva-Lena, et al. (författare)
  • SNX10 gene mutation leading to osteopetrosis with dysfunctional osteoclasts
  • 2017
  • Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 7:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Autosomal recessive osteopetrosis (ARO) is a heterogeneous disorder, characterized by defective osteoclastic resorption of bone that results in increased bone density. We have studied nine individuals with an intermediate form of ARO, from the county of Vasterbotten in Northern Sweden. All afflicted individuals had an onset in early infancy with optic atrophy, and in four patients anemia was present at diagnosis. Tonsillar herniation, foramen magnum stenosis, and severe osteomyelitis of the jaw were common clinical features. Whole exome sequencing, verified by Sanger sequencing, identified a splice site mutation c.212 + 1 G > T in the SNX10 gene encoding sorting nexin 10. Sequence analysis of the SNX10 transcript in patients revealed activation of a cryptic splice site in intron 4 resulting in a frame shift and a premature stop (p.S66Nfs * 15). Haplotype analysis showed that all cases originated from a single mutational event, and the age of the mutation was estimated to be approximately 950 years. Functional analysis of osteoclast progenitors isolated from peripheral blood of patients revealed that stimulation with receptor activator of nuclear factor kappa-B ligand (RANKL) resulted in a robust formation of large, multinucleated osteoclasts which generated sealing zones; however these osteoclasts exhibited defective ruffled borders and were unable to resorb bone in vitro.
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5.
  • Bergmann, Berglind, et al. (författare)
  • Antibiotics with Interleukin-15 inhibition reduces joint inflammation and bone erosions but not cartilage destruction in Staphylococcus aureus-induced arthritis.
  • 2018
  • Ingår i: Infection and immunity. - 1098-5522. ; 86:5
  • Tidskriftsartikel (refereegranskat)abstract
    • Background:Staphylococcus aureus-induced arthritis causes rapid joint destruction, often leading to disabling joint damage despite antibiotics. We have previously shown that IL-15 inhibition without antibiotics is beneficial in S. aureus-induced arthritis. We therefore hypothesized that inhibition of IL-15, in combination with antibiotics, might represent a useful therapy that would both reduce inflammation and joint destruction, but preserve the host's ability to clear the infection.Methods: Female wildtype C57BL/6 mice were intravenously inoculated with the TSST-1-producing LS-1 strain of S. aureus with 0.8x108S. aureus LS-1/mouse. Three days later the treatment was started consisting of cloxacillin followed by flucloxacillin, together with either anti-IL-15 antibodies (aIL-15ab) or control antibodies. Outcomes included survival, weight change, bacterial clearance, and joint damage.Results: The addition of aIL-15ab to antibiotics in S. aureus-induced arthritis reduced synovitis and bone erosions compared to controls. The number of bone-resorbing osteoclasts in the joints was reduced, whereas cartilage destruction was not significantly altered. Importantly, the combination therapy did not adversely affect the clinical outcome of S. aureus-induced arthritis, such as survival, weight change or compromise the host's ability to clear the infection.Conclusions: As the clinical outcome of S. aureus-induced arthritis was not affected, the addition of aIL-15ab to antibiotics ought to be safe. Taken together, the combination of aIL-15ab and antibiotics is a beneficial, but not optimal, treatment of S. aureus-induced arthritis as it reduces synovitis and bone erosions but has a limited effect on cartilage destruction.
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6.
  • Brommage, Robert, et al. (författare)
  • NOTUM inhibition increases endocortical bone formation and bone strength
  • 2019
  • Ingår i: Bone Research. - : Springer Science and Business Media LLC. - 2095-4700 .- 2095-6231. ; 7:2
  • Tidskriftsartikel (refereegranskat)abstract
    • The disability, mortality and costs caused by non-vertebral osteoporotic fractures are enormous. Existing osteoporosis therapies are highly effective at reducing vertebral but not non-vertebral fractures. Cortical bone is a major determinant of non-vertebral bone strength. To identify novel osteoporosis drug targets, we phenotyped cortical bone of 3 366 viable mouse strains with global knockouts of druggable genes. Cortical bone thickness was substantially elevated in Notum(-/-) mice. NOTUM is a secreted WNT lipase and we observed high NOTUM expression in cortical bone and osteoblasts but not osteoclasts. Three orally active small molecules and a neutralizing antibody inhibiting NOTUM lipase activity were developed. They increased cortical bone thickness and strength at multiple skeletal sites in both gonadal intact and ovariectomized rodents by stimulating endocortical bone formation. Thus, inhibition of NOTUM activity is a potential novel anabolic therapy for strengthening cortical bone and preventing non-vertebral fractures.
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7.
  • Conaway, H. H., et al. (författare)
  • Activation of dimeric glucocorticoid receptors in osteoclast progenitors potentiates RANKL induced mature osteoclast bone resorbing activity
  • 2016
  • Ingår i: Bone. - : Elsevier BV. - 8756-3282 .- 1873-2763. ; 93, s. 43-54
  • Tidskriftsartikel (refereegranskat)abstract
    • Glucocorticoid (GC) therapy is the greatest risk factor for secondary osteoporosis. Pathogenic mechanisms involve an initial increase in bone resorption followed by decreased bone formation. To gain a better understanding of the resorptive activity of GCs, we have used mouse bone marrow macrophages (BMM) to determine if GCs can directly modulate RANKL stimulated osteoclast formation and/or activity. In agreement with previous studies, experiments performed in plastic wells showed that GCs (dexamethasone, hydrocortisone, and prednisolone) inhibited osteoclast number and size during the initial phases of RANKL stimulated osteoclastogenesis; however, in prolonged cultures, decreased apoptosis was observed and escape from GC induced inhibition occurred with an enhanced number of osteoclasts formed, many with an increased area. When BMM cells were seeded on bone slices, GCs robustly enhanced RANKL stimulated formation of resorption pits and release of CTX without affecting the number or size of osteoclasts formed and with no effect on apoptosis. Stimulation of pit formation was not associated with increased life span of osteoclasts or an effect on mRNA expression of several osteoclastic or osteoclastogenic genes. The potentiation of RANKL induced CTX release by dexamethasone was significantly less in BMM cells from mice with conditional knockout of the osteoclastic glucocorticoid receptor and completely absent in cells from GRdim mice, which carry a point mutation in one dimerizing interface of the GC receptor. These data suggest that: 1. Plastic is a poor medium to use for studying direct effects of GCs on osteoclasts 2. GCs can enhance bone resorption without decreasing apoptosis, and 3. A direct enhancement of RANKL mediated resorption is stimulated by the dimeric GC-receptor. © 2016 Elsevier Inc.
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8.
  • Conaway, H. Herschel, et al. (författare)
  • Glucocorticoids employ the monomeric glucocorticoid receptor to potentiate vitamin D3 and parathyroid hormone–induced osteoclastogenesis
  • 2019
  • Ingår i: The FASEB Journal. - : Federation Amer Soc Exp Biol. - 0892-6638 .- 1530-6860. ; 33:12, s. 14394-14409
  • Tidskriftsartikel (refereegranskat)abstract
    • Glucocorticoid (GC) therapy decreases bone mass and increases the risk of fractures. We investigated interactions between the GC dexamethasone (DEX) and the bone resorptive agents 1,25(OH)2-vitamin D3 (D3) and parathyroid hormone (PTH) on osteoclastogenesis. We observed a synergistic potentiation of osteoclast progenitor cell differentiation and formation of osteoclasts when DEX was added to either D3- or PTH-treated mouse bone marrow cell (BMC) cultures. Cotreatment of DEX with D3 or PTH increased gene encoding calcitonin receptor (Calcr), acid phosphatase 5, tartrate resistant (Acp5), cathepsin K (Ctsk), and TNF superfamily member 11 (Tnfsf11) mRNA, receptor activator of NF-κB ligand protein (RANKL), numbers of osteoclasts on plastic, and pit formation and release of C-terminal fragment of type I collagen from cells cultured on bone slices. Enhanced RANKL protein expression caused by D3 and DEX was absent in BMC from mice in which the GC receptor (GR) was deleted in stromal cells/osteoblasts. Synergistic interactions between DEX and D3 on RANKL and osteoclast formation were present in BMC from mice with attenuated GR dimerization. These data demonstrate that the GR cooperates with D3 and PTH signaling, causing massive osteoclastogenesis, which may explain the rapid bone loss observed with high dosages of GC treatment.
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9.
  • Conaway, H Herschel, et al. (författare)
  • Retinoids stimulate periosteal bone resorption by enhancing the protein RANKL, a response inhibited by monomeric glucocorticoid receptor.
  • 2011
  • Ingår i: The Journal of biological chemistry. - 1083-351X .- 0021-9258. ; 286:36, s. 31425-36
  • Tidskriftsartikel (refereegranskat)abstract
    • Increased vitamin A (retinol) intake has been suggested to increase bone fragility. In the present study, we investigated effects of retinoids on bone resorption in cultured neonatal mouse calvarial bones and their interaction with glucocorticoids (GC). All-trans-retinoic acid (ATRA), retinol, retinalaldehyde, and 9-cis-retinoic acid stimulated release of (45)Ca from calvarial bones. The resorptive effect of ATRA was characterized by mRNA expression of genes associated with osteoclast differentiation, enhanced osteoclast number, and bone matrix degradation. In addition, the RANKL/OPG ratio was increased by ATRA, release of (45)Ca stimulated by ATRA was blocked by exogenous OPG, and mRNA expression of genes associated with bone formation was decreased by ATRA. All retinoid acid receptors (RARα/β/γ) were expressed in calvarial bones. Agonists with affinity to all receptor subtypes or specifically to RARα enhanced the release of (45)Ca and mRNA expression of Rankl, whereas agonists with affinity to RARβ/γ or RARγ had no effects. Stimulation of Rankl mRNA by ATRA was competitively inhibited by the RARα antagonist GR110. Exposure of calvarial bones to GC inhibited the stimulatory effects of ATRA on (45)Ca release and Rankl mRNA and protein expression. This inhibitory effect was reversed by the glucocorticoid receptor (GR) antagonist RU 486. Increased Rankl mRNA stimulated by ATRA was also blocked by GC in calvarial bones from mice with a GR mutation that blocks dimerization (GR(dim) mice). The data suggest that ATRA enhances periosteal bone resorption by increasing the RANKL/OPG ratio via RARα receptors, a response that can be inhibited by monomeric GR.
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10.
  • Conaway, H Herschel, et al. (författare)
  • Vitamin a metabolism, action, and role in skeletal homeostasis.
  • 2013
  • Ingår i: Endocrine reviews. - : The Endocrine Society. - 1945-7189 .- 0163-769X. ; 34:6, s. 766-97
  • Tidskriftsartikel (refereegranskat)abstract
    • Vitamin A (retinol) is ingested as either retinyl esters or carotenoids and metabolized to active compounds such as 11-cis-retinal, which is important for vision, and all-trans-retinoic acid, which is the primary mediator of biological actions of vitamin A. All-trans-retinoic acid binds to retinoic acid receptors (RARs), which heterodimerize with retinoid X receptors. RAR-retinoid X receptor heterodimers function as transcription factors, binding RAR-responsive elements in promoters of different genes. Numerous cellular functions, including bone cell functions, are mediated by vitamin A; however, it has long been recognized that increased levels of vitamin A can have deleterious effects on bone, resulting in increased skeletal fragility. Bone mass is dependent on the balance between bone resorption and bone formation. A decrease in bone mass may be caused by either an excess of resorption or decreased bone formation. Early studies indicated that the primary skeletal effect of vitamin A was to increase bone resorption, but later studies have shown that vitamin A can not only stimulate the formation of bone-resorbing osteoclasts but also inhibit their formation. Effects of vitamin A on bone formation have not been studied in as great a detail and are not as well characterized as effects on bone resorption. Several epidemiological studies have shown an association between vitamin A, decreased bone mass, and osteoporotic fractures, but the data are not conclusive because other studies have found no associations, and some studies have suggested that vitamin A primarily promotes skeletal health. In this presentation, we have summarized how vitamin A is absorbed and metabolized and how it functions intracellularly. Vitamin A deficiency and excess are introduced, and detailed descriptions of clinical and preclinical studies of the effects of vitamin A on the skeleton are presented.
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11.
  • Corjon, Stéphanie, et al. (författare)
  • Cell entry and trafficking of human adenovirus bound to blood factor X is determined by the fiber serotype and not hexon:heparan sulfate interaction
  • 2011
  • Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:5
  • Tidskriftsartikel (refereegranskat)abstract
    • Human adenovirus serotype 5 (HAdV5)-based vectors administered intravenously accumulate in the liver as the result of their direct binding to blood coagulation factor X (FX) and subsequent interaction of the FX-HAdV5 complex with heparan sulfate proteoglycan (HSPG) at the surface of liver cells. Intriguingly, the serotype 35 fiber-pseudotyped vector HAdV5F35 has liver transduction efficiencies 4-logs lower than HAdV5, even though both vectors carry the same hexon capsomeres. In order to reconcile this apparent paradox, we investigated the possible role of other viral capsid proteins on the FX/HSPG-mediated cellular uptake of HAdV5-based vectors. Using CAR- and CD46-negative CHO cells varying in HSPG expression, we confirmed that FX bound to serotype 5 hexon protein and to HAdV5 and HAdV5F35 virions via its Gla-domain, and enhanced the binding of both vectors to surface-immobilized hypersulfated heparin and cellular HSPG. Using penton mutants, we found that the positive effect of FX on HAdV5 binding to HSPG and cell transduction did not depend on the penton base RGD and fiber shaft KKTK motifs. However, we found that FX had no enhancing effect on the HAdV5F35-mediated cell transduction, but a negative effect which did not involve the cell attachment or endocytic step, but the intracellular trafficking and nuclear import of the FX-HAdV5F35 complex. By cellular imaging, HAdV5F35 particles were observed to accumulate in the late endosomal compartment, and were released in significant amounts into the extracellular medium via exocytosis. We showed that the stability of serotype 5 hexon:FX interaction was higher at low pH compared to neutral pH, which could account for the retention of FX-HAdV5F35 complexes in the late endosomes. Our results suggested that, despite the high affinity interaction of hexon capsomeres to FX and cell surface HSPG, the adenoviral fiber acted as the dominant determinant of the internalization and trafficking pathway of HAdV5-based vectors. © 2011 Corjon et al.
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12.
  • de Souza, P. P. C., et al. (författare)
  • Stimulation of Osteoclast Formation by Oncostatin M and the Role of WNT16 as a Negative Feedback Regulator
  • 2022
  • Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1422-0067. ; 23:6
  • Tidskriftsartikel (refereegranskat)abstract
    • Oncostatin M (OSM), which belongs to the IL-6 family of cytokines, is the most potent and effective stimulator of osteoclast formation in this family, as assessed by different in vitro assays. Osteoclastogenesis induced by the IL-6 type of cytokines is mediated by the induction and paracrine stimulation of the osteoclastogenic cytokine receptor activator of nuclear factor kappa-B ligand (RANKL), expressed on osteoblast cell membranes and targeting the receptor activator of nuclear factor kappa-B (RANK) on osteoclast progenitor cells. The potent effect of OSM on osteoclastogenesis is due to an unusually robust induction of RANKL in osteoblasts through the OSM receptor (OSMR), mediated by a JAK-STAT/MAPK signaling pathway and by unique recruitment of the adapter protein Shc1 to the OSMR. Gene deletion of Osmr in mice results in decreased numbers of osteoclasts and enhanced trabecular bone caused by increased trabecular thickness, indicating that OSM may play a role in physiological regulation of bone remodeling. However, increased amounts of OSM, either through administration of recombinant protein or of adenoviral vectors expressing Osm, results in enhanced bone mass due to increased bone formation without any clear sign of increased osteoclast numbers, a finding which can be reconciled by cell culture experiments demonstrating that OSM can induce osteoblast differentiation and stimulate mineralization of bone nodules in such cultures. Thus, in vitro studies and gene deletion experiments show that OSM is a stimulator of osteoclast formation, whereas administration of OSM to mice shows that OSM is not a strong stimulator of osteoclastogenesis in vivo when administered to adult animals. These observations could be explained by our recent finding showing that OSM is a potent stimulator of the osteoclastogenesis inhibitor WNT16, acting in a negative feedback loop to reduce OSM-induced osteoclast formation.
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13.
  • Farman, Helen H., 1983, et al. (författare)
  • Membrane estrogen receptor alpha is essential for estrogen signaling in the male skeleton
  • 2018
  • Ingår i: Journal of Endocrinology. - : Bioscientifica. - 0022-0795 .- 1479-6805. ; 239:3, s. 303-312
  • Tidskriftsartikel (refereegranskat)abstract
    • The importance of estrogen receptor alpha (ER alpha) for the regulation of bone mass in males is well established. ERa mediates estrogenic effects both via nuclear and membraneinitiated ER alpha (mER alpha) signaling. The role of mERa signaling for the effects of estrogen on bone in male mice is unknown. To investigate the role of mERa signaling, we have used mice (Nuclear-Only-ER; NOER) with a point mutation (C451A), which results in inhibited trafficking of ER alpha to the plasma membrane. Gonadal-intact male NOER mice had a significantly decreased total body areal bone mineral density (aBMD) compared to WT littermates at 3, 6 and 9 months of age as measured by dual-energy X-ray absorptiometry (DEXA). High-resolution microcomputed tomography (mu CT) analysis of tibia in 3-month-old males demonstrated a decrease in cortical and trabecular thickness in NOER mice compared to WT littermates. As expected, estradiol (E2) treatment of orchidectomized (ORX) WT mice increased total body aBMD, trabecular BV/TV and cortical thickness in tibia compared to placebo treatment. E2 treatment increased these skeletal parameters also in ORX NOER mice. However, the estrogenic responses were significantly decreased in ORX NOER mice compared with ORX WT mice. In conclusion, mER alpha is essential for normal estrogen signaling in both trabecular and cortical bone in male mice. Increased knowledge of estrogen signaling mechanisms in the regulation of the male skeleton may aid in the development of new treatment options for male osteoporosis.
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14.
  • Franqueville, Laure, et al. (författare)
  • Protein crystals in Adenovirus type 5-infected cells: requirements for intranuclear crystallogenesis, structural and functional analysis.
  • 2008
  • Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 3:8
  • Tidskriftsartikel (refereegranskat)abstract
    • Intranuclear crystalline inclusions have been observed in the nucleus of epithelial cells infected with Adenovirus serotype 5 (Ad5) at late steps of the virus life cycle. Using immuno-electron microscopy and confocal microscopy of cells infected with various Ad5 recombinants modified in their penton base or fiber domains, we found that these inclusions represented crystals of penton capsomers, the heteromeric capsid protein formed of penton base and fiber subunits. The occurrence of protein crystals within the nucleus of infected cells required the integrity of the fiber knob and part of the shaft domain. In the knob domain, the region overlapping residues 489-492 in the FG loop was found to be essential for crystal formation. In the shaft, a large deletion of repeats 4 to 16 had no detrimental effect on crystal inclusions, whereas deletion of repeats 8 to 21 abolished crystal formation without altering the level of fiber protein expression. This suggested a crucial role of the five penultimate repeats in the crystallisation process. Chimeric pentons made of Ad5 penton base and fiber domains from different serotypes were analyzed with respect to crystal formation. No crystal was found when fiber consisted of shaft (S) from Ad5 and knob (K) from Ad3 (heterotypic S5-K3 fiber), but occurred with homotypic S3K3 fiber. However, less regular crystals were observed with homotypic S35-K35 fiber. TB5, a monoclonal antibody directed against the Ad5 fiber knob was found by immunofluorescence microscopy to react with high efficiency with the intranuclear protein crystals in situ. Data obtained with Ad fiber mutants indicated that the absence of crystalline inclusions correlated with a lower infectivity and/or lower yields of virus progeny, suggesting that the protein crystals might be involved in virion assembly. Thus, we propose that TB5 staining of Ad-infected 293 cells can be used as a prognostic assay for the viability and productivity of fiber-modified Ad5 vectors.
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15.
  • Funck-Brentano, Thomas, et al. (författare)
  • Porcupine inhibitors impair trabecular and cortical bone mass and strength in mice
  • 2018
  • Ingår i: Journal of Endocrinology. - : Bioscientifica. - 0022-0795 .- 1479-6805. ; 238:1, s. 13-23
  • Tidskriftsartikel (refereegranskat)abstract
    • WNT signaling is involved in the tumorigenesis of various cancers and regulates bone homeostasis. Palmitoleoylation of WNTs by Porcupine is required for WNT activity. Porcupine inhibitors are under development for cancer therapy. As the possible side effects of Porcupine inhibitors on bone health are unknown, we determined their effects on bone mass and strength. Twelve-week-old C57BL/6N female mice were treated by the Porcupine inhibitors LGK974 (low dose = 3 mg/kg/day; high dose = 6 mg/kg/day) or Wnt-C59 (10 mg/kg/day) or vehicle for 3 weeks. Bone parameters were assessed by serum biomarkers, dual-energy X-ray absorptiometry, mu CT and histomorphometry. Bone strength was measured by the 3-point bending test. The Porcupine inhibitors were well tolerated demonstrated by normal body weight. Both doses of LGK974 and Wnt-C59 reduced total body bone mineral density compared with vehicle treatment (P < 0.001). Cortical thickness of the femur shaft (P < 0.001) and trabecular bone volume fraction in the vertebral body (P < 0.001) were reduced by treatment with LGK974 or Wnt-C59. Porcupine inhibition reduced bone strength in the tibia (P < 0.05). The cortical bone loss was the result of impaired periosteal bone formation and increased endocortical bone resorption and the trabecular bone loss was caused by reduced trabecular bone formation and increased bone resorption. Porcupine inhibitors exert deleterious effects on bone mass and strength caused by a combination of reduced bone formation and increased bone resorption. We suggest that cancer targeted therapies using Porcupine inhibitors may increase the risk of fractures.
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16.
  • Grahnemo, Louise, et al. (författare)
  • Increased bone mass in a mouse model with low fat mass.
  • 2018
  • Ingår i: American journal of physiology. Endocrinology and metabolism. - : American Physiological Society. - 1522-1555 .- 0193-1849. ; 315:6
  • Tidskriftsartikel (refereegranskat)abstract
    • Mice with impaired acute inflammatory responses within adipose tissue display reduced diet-induced fat mass gain associated with glucose intolerance and systemic inflammation. Therefore, acute adipose tissue inflammation is needed for a healthy expansion of adipose tissue. Because inflammatory disorders are associated with bone loss, we hypothesized that impaired acute adipose tissue inflammation leading to increased systemic inflammation results in a lower bone mass. To test this hypothesis, we used mice overexpressing an adenoviral protein complex - the receptor internalization and degradation (RID) complex that inhibits pro-inflammatory signaling - under the control of the aP2-promotor (RID tg mice), resulting in suppressed inflammatory signaling in adipocytes. As expected, RID tg mice had a lower high-fat diet-induced weight and fat mass gain and higher systemic inflammation than their littermate wild type controls. Contrary to our hypothesis, the RID tg mice had increased bone mass in long bones and vertebrae, affecting trabecular and cortical parameters, as well as improved humeral biomechanical properties. We did not find any differences in bone formation or resorption parameters as determined by histology or enzyme immunoassay. However, bone marrow adiposity, often negatively associated with bone mass, was decreased in male RID tg mice as determined by histological analysis of tibia. In conclusion, mice with reduced fat mass, due to impaired adipose tissue inflammation, have increased bone mass.
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17.
  • Granholm, Susanne, et al. (författare)
  • Comparisons between the effects of calcitonin receptor-stimulating peptide and intermedin and other peptides in the calcitonin family on bone resorption and osteoclastogenesis
  • 2011
  • Ingår i: Journal of Cellular Biochemistry. - : Wiley. - 0730-2312 .- 1097-4644. ; 112:11, s. 3300-3312
  • Tidskriftsartikel (refereegranskat)abstract
    • Calcitonin receptor-stimulating peptide (CRSP) and intermedin (IMD) are two recently discovered peptides in the calcitonin (CT) family of peptides. CRSP and IMD, similar to CT, calcitonin gene-related peptide (CGRP) and amylin (AMY), but in contrast to adrenomedullin (ADM), inhibited bone resorption in mouse calvarial bones. CRSP and IMD, similar to CT, CGRP, AMY, but in contrast to ADM, decreased formation of osteoclasts and number of pits in bone marrow macrophage cultures stimulated by M-CSF and RANKL, with no effect on the expression of a number of genes associated with osteoclast progenitor cell differentiation. CRSP and IMD inhibited osteoclastogenesis at a late stage but had no effect on DC-STAMP mRNA. IMD, similar to CGRP, AMY and ADM stimulated cyclic AMP formation in M-CSF expanded osteoclast progenitor cells lacking CT receptors. RANKL induced CT receptors and a cyclic AMP response also to CT and CRSP, and increased the cyclic AMP response to CGRP, AMY and IMD but decreased the response to ADM. Our data demonstrate that CRSP and IMD share several functional properties of peptides in the CT family of peptides, including inhibition of bone resorption and osteoclast formation. The data also show that the reason why ADM does not inhibit osteoclast activity or formation is related to the fact that RANKL decreases ADM receptor signalling through the adenylate cyclase-cyclic AMP pathway. Finally, the findings indicate that activation by CGRP, AMY and IMD may include activation of both CT and CT receptor-like receptors. J. Cell. Biochem. © 2011 Wiley-Liss, Inc.
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18.
  • Granholm, Susanne, et al. (författare)
  • Osteoclast progenitor cells present in significant amounts in mouse calvarial osteoblast isolations and osteoclastogenesis increased by BMP-2
  • 2013
  • Ingår i: Bone. - : Elsevier BV. - 8756-3282 .- 1873-2763. ; 52:1, s. 83-92
  • Tidskriftsartikel (refereegranskat)abstract
    • Enzymatically released cells from neonatal mouse calvarial bones are frequently used as primary mouse osteoblasts for in vitro studies. We, here, report that although these cells lack mRNA expression of the osteoclastic genes Calcr, Acp5 and Mmp-9 at early time points after their isolation, these transcripts are gradually upregulated when the calvarial osteoblast cultures are differentiated to more mature osteoblasts in long term cultures. Similarly, Calcr, Acp5, Mmp-9, as well as Rank and Nfatc1 mRNA expressions are robustly increased when the osteoblast cultures were induced to differentiate by treatment with bone morphogenetic protein (BMP-2). The increased Calcr mRNA resulted in functionally active calcitonin receptors. Enhanced osteoblastic differentiation was associated with increased Rankl mRNA expression and decreased Opg and Cfs1 mRNA expression. Treatment of the osteoblastic cells with BMP-2 or RANKL, either on plastic dishes or bone slices, resulted in the formation of multinucleated tartrate-resistant acid phosphatase positive cells, which were able to excavate resorption pits and release CTX from the bones. In contrast, increased osteoblastic differentiation induced by BMP-2 in the mouse calvarial osteoblastic cell line MC3T3-E1 was not associated with increased mRNA expression of Calcr, Acp5, Rank, c-Fms or Oscar. Interestingly, Ctsk mRNA was increased during osteoblastic differentiation in both mouse calvarial osteoblast cultures and in MC3T3-E1 cultures. Also osteoblasts isolated from mouse long bones by outgrowth from explant cultures were contaminated with osteoclast progenitors able to differentiate into bone resorbing osteoclasts. These data indicate that mouse calvarial osteoblast cultures contain osteoclast progenitor cells, which will be differentiated along the osteoclastic lineage during osteoblastic differentiation. Moreover, the data show that BMP-2 not only stimulates osteoblastic differentiation but can also induce osteoclastogenesis through increased RANKL.
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19.
  • Granio, Ophélia, et al. (författare)
  • Adenovirus 5-fiber 35 chimeric vector mediates efficient apical correction of the CFTR defect in cystic fibrosis primary airway epithelia.
  • 2010
  • Ingår i: Human gene therapy. - : Mary Ann Liebert Inc. - 1557-7422 .- 1043-0342. ; 21:3, s. 251-269
  • Tidskriftsartikel (refereegranskat)abstract
    • In vivo gene transfer to the human respiratory tract using Adenovirus serotype 5 (Ad5) vectors has revealed their limitations related to inefficient gene transfer, host antiviral response and innate adenoviral toxicity. In the present work, we compared the cytotoxicity and efficiency of Ad5 and a chimeric Ad5F35 vector with respect to CFTR gene transfer to cystic fibrosis (CF) and non-CF human airway epithelial cells. We found that high doses of Ad5 vector had an adverse effect on the function of exogenous and endogenous CFTR. Results obtained with Ad5 capsid mutants suggested that the RGD motifs on the penton base capsomers were responsible for the negative effect on the CFTR function. This negative interference was not a result of a lower level of biosynthesis and/or altered cellular trafficking of the CFTR protein, but rather from an indirect mechanism of functional blockage of CFTR, related to the RGD-integrin-mediated endocytic pathway of Ad5. No negative interference with CFTR was observed for Ad5F35, an Ad5-based vector pseudotyped with fibers from Ad35, a serotype which uses another cell entry pathway. In vitro, Ad5F35 vector expressing the GFP-tagged CFTR (Ad5F35-GFP-CFTR) showed a 30-fold higher efficiency of transduction and chloride channel correction in CFTR-deficient cells, compared to Ad5GFP-CFTR. Ex vivo, Ad5F35-GFP-CFTR had the capacity to transduce efficiently reconstituted airway epithelia from CF patients (CF-HAE) via the apical surface, restored the chloride channel function at relatively low vector doses, and showed a relatively stable expression of GFP-CFTR for several weeks.
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20.
  • Granio, Ophélia, et al. (författare)
  • Improved adenovirus type 5 vector-mediated transduction of resistant cells by piggybacking on coxsackie B-adenovirus receptor-pseudotyped baculovirus.
  • 2009
  • Ingår i: Journal of virology. - 1098-5514. ; 83:12, s. 6048-66
  • Tidskriftsartikel (refereegranskat)abstract
    • Taking advantage of the wide tropism of baculoviruses (BVs), we constructed a recombinant BV (BV(CAR)) pseudotyped with human coxsackie B-adenovirus receptor (CAR), the high-affinity attachment receptor for adenovirus type 5 (Ad5), and used the strategy of piggybacking Ad5-green fluorescent protein (Ad5GFP) vector on BV(CAR) to transduce various cells refractory to Ad5 infection. We found that transduction of all cells tested, including human primary cells and cancer cell lines, was significantly improved using the BV(CAR)-Ad5GFP biviral complex compared to that obtained with Ad5GFP or BV(CAR)GFP alone. We determined the optimal conditions for the formation of the complex and found that a high level of BV(CAR)-Ad5GFP-mediated transduction occurred at relatively low adenovirus vector doses, compared with transduction by Ad5GFP alone. The increase in transduction was dependent on the direct coupling of BV(CAR) to Ad5GFP via CAR-fiber knob interaction, and the cell attachment of the BV(CAR)-Ad5GFP complex was mediated by the baculoviral envelope glycoprotein gp64. Analysis of the virus-cell binding reaction indicated that the presence of BV(CAR) in the complex provided kinetic benefits to Ad5GFP compared to the effects with Ad5GFP alone. The endocytic pathway of BV(CAR)-Ad5GFP did not require Ad5 penton base RGD-integrin interaction. Biodistribution of BV(CAR)-Ad5Luc complex in vivo was studied by intravenous administration to nude BALB/c mice and compared to Ad5Luc injected alone. No significant difference in viscerotropism was found between the two inocula, and the liver remained the preferred localization. In vitro, coagulation factor X drastically increased the Ad5GFP-mediated transduction of CAR-negative cells but had no effect on the efficiency of transduction by the BV(CAR)-Ad5GFP complex. Various situations in vitro or ex vivo in which our BV(CAR)-Ad5 duo could be advantageously used as gene transfer biviral vector are discussed.
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21.
  • Gupta, Priti, et al. (författare)
  • Bazedoxifene does not share estrogens effects on IgG sialylation
  • 2023
  • Ingår i: Plos One. - 1932-6203. ; 18:5
  • Tidskriftsartikel (refereegranskat)abstract
    • The incidence of rheumatoid arthritis (RA) increases at the same time as menopause when estrogen level decreases. Estrogen treatment is known to reduce the IgG pathogenicity by increasing the sialylation grade on the terminal glycan chain of the Fc domain, inhibiting the binding ability to the Fc gamma receptor. Therefore, treatment with estrogen may be beneficial in pre-RA patients who have autoantibodies and are prone to get an autoimmune disease. However, estrogen treatment is associated with negative side effects, therefore selective estrogen receptor modulators (SERMs) have been developed that have estrogenic protective effects with minimal side effects. In the present study, we investigated the impact of the SERM bazedoxifene on IgG sialylation as well as on total serum protein sialylation. C57BL6 mice were ovariectomized to simulate postmenopausal status, followed by ovalbumin immunization, and then treated with estrogen (estradiol), bazedoxifene, or vehicle. We found that estrogen treatment enhanced IgG levels and had a limited effect on IgG sialylation. Treatment with bazedoxifene increased the sialic acids in plasma cells in a similar manner to E2 but did not reach statistical significance. However, we did not detect any alteration in IgG-sialylation with bazedoxifene treatment. Neither estrogen nor bazedoxifene showed any significant alteration in serum protein sialylation but had a minor effect on mRNA expression of glycosyltransferase in the bone marrow, gonadal fat, and liver.
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22.
  • Gupta, Priti, et al. (författare)
  • Impact of estrogen on IgG glycosylation and serum protein glycosylation in a murine model of healthy postmenopause.
  • 2023
  • Ingår i: Frontiers in endocrinology. - 1664-2392. ; 14
  • Tidskriftsartikel (refereegranskat)abstract
    • The glycosylation of immunoglobulin (Ig) G regulates IgG interaction capability with Fc gamma receptors found in all immune cells. In pathogenic conditions, estrogen can impact IgG levels and glycosylation. Following menopause, when estrogen levels decline affecting the immune system and potentially leading to a heightened susceptibility of immune activation.In this study, we aim to determine if estrogen levels can regulate IgG glycosylation in postmenopausal healthy situations.Mice were ovariectomized to simulate an estrogen-deficient postmenopausal status and then treated with 17-beta-estradiol (E2) at different doses and different administration strategies.Using a highly sensitive liquid chromatography-tandem mass spectrometry (MS/MS) glycoproteomic method, we demonstrated that E2 treatment increased the degree of glycosylation on IgG-Fc with both galactosylation and sialylation in the position required for interaction with Fc gamma receptors. We also observed that only long-term estrogen deficiency reduces IgG levels and that estrogen status had no impact on total IgG sialylation on both Fab and Fc domains or general glycoprotein sialylation evaluated by ELISA. Furthermore, E2 status did not affect the total sialic acid content of total cells in lymphoid organs and neither B cells nor plasma cells.The study concluded that E2 treatment does not affect total serum glycoprotein sialylation but alters IgG glycosylation, including IgG sialylation, implying that estrogen functions as an intrinsic modulator of IgG sialylation and could thereby be one pathway by which estrogen modulates immunity.
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23.
  • Gustafsson, Karin L., 1987, et al. (författare)
  • A tissue-specific role of membrane-initiated ERα signaling for the effects of SERMs
  • 2022
  • Ingår i: Journal of Endocrinology. - 0022-0795. ; 253:2, s. 75-84
  • Tidskriftsartikel (refereegranskat)abstract
    • Selective estrogen receptor modulators (SERMs) act as estrogen receptor (ER) agonists or antagonists in a tissue-specific manner. ERs exert effects via nuclear actions but can also utilize membrane-initiated signaling pathways. To dete rmine if membrane-initiated ERα (mERα) signaling affects SERM action in a tissue-specific manner, C451 A mice, lacking mERα signaling due to a mutation at palmitoylation site C451, were treated with Lasofoxifene (Las), Bazedoxifene (Bza), or estradi ol (E2), and various tissues were evaluated. Las and Bza treatment increased uterine weight to a similar extent in C451A and control mice, demonstrating mERα-independent uterine SERM effects, while the E2 effect on the uterus was predominantly mER α-dependent. Las and Bza treatment increased both trabecular and cortical bone mass in controls to a similar degree as E2, while both SERM and E2 treatment effects were abse nt in C451A mice. This demonstrates that SERM effects, similar to E2 effects, in th e skeleton are mERα- dependent. Both Las and E2 treatment decreased thymus weight in controls, while neither treatment affected the thymus in C451A mice, demonstrati ng mERα-dependent SERM and E2 effects in this tissue. Interestingly, both SERM and E2 treatments decreased the total body fat percent in C451A mice, demonstrating the ability of these treatments to affect fat tissue in the absence of functional mER α signaling. In conclusion, mERα signaling can modulate SERM responses in a tissue-specific manne r. This novel knowledge increases the understanding of the mechanisms behind SERM effects and may thereby facilitate the development of new improved SERMs.
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24.
  • Gustafsson, Karin L., 1987, et al. (författare)
  • Arginine site 264 in murine estrogen receptor alpha is dispensable for the regulation of the skeleton.
  • 2021
  • Ingår i: American journal of physiology. Endocrinology and metabolism. - 1522-1555. ; 320:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Estrogen protects against bone loss, but is not a suitable treatment due to adverse effects in other tissues. Increased knowledge regarding estrogen signaling in estrogen-responsive tissues is therefore warranted to aid the development of bone-specific estrogen treatments. Estrogen receptor alpha (ERα), the main mediator of estrogenic effects in bone, is widely subjected to posttranslational modifications (PTMs). In vitro studies have shown that methylation at site R260 in the human ERα affects receptor localization and intracellular signaling. The corresponding amino acid R264 in murine ERα has been shown to have a functional role in endothelium in vivo; albeit the methylation of R264 in the murine gene is yet to be empirically demonstrated. The aim of this study was to investigate if R264 in ERα is involved in the regulation of the skeleton in vivo. DXA analysis at three, six, nine, and twelve months of age showed no differences in total body areal BMD between R264A and WT in either female or male mice. Furthermore, analyses using CT demonstrated that trabecular bone mass in tibia and vertebra, and cortical thickness in tibia, were similar between R264A and WT mice. In addition, R264A females displayed a normal estrogen treatment response in trabecular bone mass, as well as in cortical thickness. Furthermore, uterus, thymus, and adipose tissue responded similarly in R264A and WT female mice after estrogen treatment. In conclusion, our novel finding that mutation of R264 in ERα does not affect the regulation of the skeleton, together with the known role of R264 for ERα-mediated endothelial effects, supports the concept that R264 determines tissue specificity of ERα.
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25.
  • Gustafsson, Karin L., 1987, et al. (författare)
  • ER alpha expression in T lymphocytes is dispensable for estrogenic effects in bone
  • 2018
  • Ingår i: Journal of Endocrinology. - : Bioscientifica. - 0022-0795 .- 1479-6805. ; 238:2, s. 129-136
  • Tidskriftsartikel (refereegranskat)abstract
    • Estrogen treatment has positive effects on the skeleton, and we have shown that estrogen receptor alpha (ERa) expression in cells of hematopoietic origin contributes to a normal estrogen treatment response in bone tissue. T lymphocytes are implicated in the estrogenic regulation of bone mass, but it is not known whether T lymphocytes are direct estrogen target cells. Therefore, the aim of this study was to determine the importance of ERa expression in T lymphocytes for the estrogenic regulation of the skeleton using female mice lacking ERa expression specifically in T lymphocytes (Lck-ERa-/-) and ERaflox/flox littermate (control) mice. Deletion of ERa expression in T lymphocytes did not affect bone mineral density (BMD) in sham-operated Lck-ERa-/compared to control mice, and ovariectomy (ovx) resulted in a similar decrease in BMD in control and Lck-ERa-/- mice compared to sham-operated mice. Furthermore, estrogen treatment of ovx Lck-ERa-/- led to an increased BMD that was indistinguishable from the increase seen after estrogen treatment of ovx control mice. Detailed analysis of both the appendicular (femur) and axial (vertebrae) skeleton showed that both trabecular and cortical bone parameters responded to a similar extent regardless of the presence of ERa in T lymphocytes. In conclusion, ERa expression in T lymphocytes is dispensable for normal estrogenic regulation of bone mass in female mice.
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26.
  • Gustafsson, Karin L., 1987, et al. (författare)
  • The role of membrane ER alpha signaling in bone and other major estrogen responsive tissues
  • 2016
  • Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
  • Tidskriftsartikel (refereegranskat)abstract
    • Estrogen receptor a (ER alpha) signaling leads to cellular responses in several tissues and in addition to nuclear ER alpha-mediated effects, membrane ER alpha (mER alpha) signaling may be of importance. To elucidate the significance, in vivo, of mER alpha signaling in multiple estrogen-responsive tissues, we have used female mice lacking the ability to localize ER alpha to the membrane due to a point mutation in the palmitoylation site (C451A), so called Nuclear-Only-ER (NOER) mice. Interestingly, the role of mER alpha signaling for the estrogen response was highly tissue-dependent, with trabecular bone in the axial skeleton being strongly dependent (>80% reduction in estrogen response in NOER mice), cortical and trabecular bone in long bones, as well as uterus and thymus being partly dependent (40-70% reduction in estrogen response in NOER mice) and effects on liver weight and total body fat mass being essentially independent of mER alpha (<35% reduction in estrogen response in NOER mice). In conclusion, mER alpha signaling is important for the estrogenic response in female mice in a tissue-dependent manner. Increased knowledge regarding membrane initiated ER alpha actions may provide means to develop new selective estrogen receptor modulators with improved profiles.
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27.
  • Henning, Petra, 1974, et al. (författare)
  • Adenovirus type 5 fiber knob domain has a critical role in fiber protein synthesis and encapsidation
  • 2006
  • Ingår i: JOURNAL OF GENERAL VIROLOGY. - : Microbiology Society. - 0022-1317 .- 1465-2099. ; 87, s. 3151-3160
  • Tidskriftsartikel (refereegranskat)abstract
    • Adenovirus serotype 5 (Ad5) vectors carrying knobless fibers designed to remove their natural tropism were found to have a lower fiber content than recombinant Ad5 with wild-type (WT) capsid, implying a role for the knob-coding sequence or/and the knob domain in fiber encapsidation. Experimental data using a variety of fiber gene constructs showed that the defect did not occur at the fiber mRNA level, but at the protein level. Knobless fiber proteins were found to be synthesized at a significant slower rate compared with knob-carrying fibers, and the trimerization process of knobless fibers paralleled their slow rate of synthesis. A recombinant Ad5 diploid for the fiber gene (referred to as Ad5/R7-ZZwt/E1 : WT-fiber) was constructed to analyse the possible rescue of the knobless low-fiber-content phenotype by co-expression of WT fiber. Ad5/R7-ZZwt/E1 : WT-fiber contained a knobless fiber gene in its natural location (L5) in the viral genome and an additional WT fiber gene in an ectopic position in E1. Knobless fiber was still synthesized at low levels compared with the co-expressed E1 : WT fiber and the recovery of the two fiber species in virus progeny reflected their respective amounts in the infected cells. Our results suggested that deletion of the fiber knob domain had a negative effect on the translation of the fiber mRNA and on the intracellular concentration of fiber protein. They also suggested that the knob control of fiber protein synthesis and encapsidation occurred as aciseffect, which was not modified by WT fiber protein providedin transby the same Ad5 genome.
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28.
  • Henning, Petra, 1974, et al. (författare)
  • Genetic modification of adenovirus 5 tropism by a novel class of ligands based on a three-helix bundle scaffold derived from staphylococcal protein A.
  • 2002
  • Ingår i: Human gene therapy. - : Mary Ann Liebert Inc. - 1043-0342 .- 1557-7422. ; 13:12, s. 1427-39
  • Tidskriftsartikel (refereegranskat)abstract
    • The use of adenovirus (Ad) as an efficient and versatile vector for in vivo tumor therapy requires the modulation of its cellular tropism. We previously developed a method to genetically alter the tropism of Ad5 fibers by replacing the fiber knob domain by an extrinsic trimerization motif and a new cellular ligand. However, fibers carrying complex ligands such as single-chain antibody fragments did not assemble into functional pentons in vitro in the presence of penton base, and failed to be rescued into infectious virions because of their inability to fold correctly within the cytoplasm of Ad-infected cells. Here we show that the coding sequence for a disulfide bond-independent three-helix bundle scaffold Z, derived from domain B of Staphylococcal protein A and capable of binding to the Fc portion of immunoglobulin (Ig) G1, could be incorporated into modified knobless Ad fiber gene constructs with seven shaft repeats. These fiber gene constructs could be rescued into viable virions that were demonstrated to enter 293 cells engineered for IgG Fc surface expression but not unmodified 293 cells, via a mechanism that could be specifically blocked with soluble Fc target protein. However, the tropism modified viruses showed a slightly impaired cellular entry and a lower infectivity than wildtype (WT) virus. In addition, we generated recombinant fibers containing an IgA binding Affibody ligand, derived from combinatorial specificity-engineering of the Z domain scaffold. Such fiber constructs also showed the expected target specific binding, indicating that the affibody protein class is ideally suited for genetic engineering of Ad tropism.
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29.
  • Henning, Petra, 1974 (författare)
  • Retargeting of adenovirus vectors. Viral ligands and cellular receptors
  • 2004
  • Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
    • Gene therapy is one of the newest technologies attempted to treat human cancer. Progress in the field has however been hampered by the lack of efficient and safe vectors. Ideal vectors should specifically transduce desired cells in vivo. This thesis is devoted to the development and evaluation of retargeted adenovirus type 5 (Ad5) vectors. Different types of complex cell binding ligands have been incorporated into various Ad5 capsid proteins to study ligands and anchoring proteins suitable for Ad5 retargeting. Retargeted Ad5 has been evaluated in terms of e.g. specificity, infectivity and capsid protein incorporation. In addition, immunoglobulin binding Ad5 has been generated and used to evaluate potential tumor receptors. Complex ligands containing disulfide bonds can generally not be used to retarget Ad since they are insoluble and do not bind to their corresponding receptor when expressed in the reducing cell cytosol where the virus proteins are synthesized. Affibody moleculesTM and hyperstable single chain antibodies (scFv) could however be incorporated into Ad capsids with retained binding specificity and can predictably be used as scaffolds for generation of tumor specific Ad5 vectors. The incorporation of new viral ligands into truncated knobless fibers resulted in retargeted Ad5 but the infectivity, growth and fiber content of these viruses were decreased. The decreased fiber content was, at least partially, caused by impaired translation of the fiber protein. Several efforts were made to increase the fiber content of retargeted Ad with truncated, affibody liganded fibers without success. Incorporation of a hyperstable scFv into pIX resulted in normal pIX content on virus capsids with retained binding specificity of the scFv and caused less decrease in virus infectivity than incorporation into de-knobbed fibers. It was found that Ad can use EGF-R, CA242, Her2/neu and possibly PSMA as new cellular receptors. However, no gene transfer via CA242 or EGF-R could be detected in two colorectal tumor cell lines even though the receptors are expressed and the virus was shown to bind to the cell surface. In addition, gene transfer mediated by all the tested cellular receptors was shown to be dependent on an intact integrin-binding RGD motif in the penton base.In conclusion, cytoplasmic solubility and ligand reactivity of fiber-ligand fusions seems to be the best prediction criterions for recovery of genetically retargeted Ad. Affibody molecules and hyperstable single chain antibodies can be used for genetic retargeting of Ad. The fiber content, growth and infectivity is impaired in viruses with truncated knobless fibers. The minor capsid protein pIX is an interesting alternative to fibers for incorporation of new viral ligands since a hyperstable scFv can be genetically added to the protein with less decrease in infectivity of the virus than when the ligand is put on truncated deknobbed fibers. Ad5 can use alternative receptors for virus cellular entry but it is possible that blocks to Ad5 transduction exist in tumor cells downstream of virus binding to cellular receptors. One of these factors is the integrin-binding RGD motif of the penton base.
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30.
  • Henning, Petra, 1974, et al. (författare)
  • Retinoid receptors in bone and their role in bone remodeling
  • 2015
  • Ingår i: Frontiers in Endocrinology. - : Frontiers Media SA. - 1664-2392. ; 6
  • Tidskriftsartikel (refereegranskat)abstract
    • Vitamin A (retinol) is a necessary and important constituent of the body which is provided by food intake of retinyl esters and carotenoids. Vitamin A is known best for being important for vision, but in addition to the eye, vitamin A is necessary in numerous other organs in the body, including the skeleton. Vitamin A is converted to an active compound, all-trans-retinoic acid (ATRA), which is responsible for most of its biological actions. ATRA binds to intracellular nuclear receptors called retinoic acid receptors (RARα, RARβ, RARγ). RARs and closely related retinoid X receptors (RXRα, RXRβ, RXRγ) form heterodimers which bind to DNA and function as ligand-activated transcription factors. It has been known for many years that hypervitaminosis A promotes skeleton fragility by increasing osteoclast formation and decreasing cortical bone mass. Some epidemiological studies have suggested that increased intake of vitamin A and increased serum levels of retinoids may decrease bone mineral density and increase fracture rate, but the literature on this is not conclusive. The current review summarizes how vitamin A is taken up by the intestine, metabolized, stored in the liver, and processed to ATRA. ATRA's effects on formation and activity of osteoclasts and osteoblasts are outlined, and a summary of clinical data pertaining to vitamin A and bone is presented. © 2015 Henning, Conaway and Lerner.
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31.
  • Henning, Petra, 1974, et al. (författare)
  • Stimulation of osteoclast formation and bone resorption by glucocorticoids: Synergistic interactions with the calcium regulating hormones parathyroid hormone and 1,25(OH)2-vitamin D3
  • 2022
  • Ingår i: Vitamins and Hormones Vol 120 Parathyroid Hormone. - : Elsevier. - 0083-6729. ; , s. 231-270
  • Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
    • Osteoporosis is a significant health problem, with skeletal fractures increasing morbidity and mortality. Excess glucocorticoids (GC) represents the leading cause of secondary osteoporosis. The first phase of glucocorticoid-induced osteoporosis is increased bone resorption. In this Chapter, in vitro studies of the direct glucocorticoid receptor (GR) mediated cellular effects of GC on osteoclasts to affect bone resorption and indirect effects on osteoblast lineage cells to increase the RANKL/OPG ratio and stimulate osteoclastogenesis and bone resorption are reviewed in detail, together with detailed descriptions of in vivo effects of GC in different portions of the skeleton in research animals and humans. Brief sections are devoted to contrasting functions of GC in osteonecrosis, vitamin D formation, in vitro and in vivo bone resorptive actions dependent on vitamin D receptor and vitamin D toxicity, as well as the molecular basis of GR action. Included are also more detailed assessments of the interactions of GC with the major calcium regulating hormones, 1,25(OH)2-vitamin D3 and parathyroid hormone, describing the in vitro increases in RANKL/OPG ratios, osteoclastogenesis and synergistic bone resorption that occurs when GC is combined with either 1,25(OH)2-vitamin D3 or parathyroid hormone. Additionally, a molecular basic for the synergistic interaction of GC with 1,25(OH)2-vitamin D3 is provided along with a suggested molecular basic for the interaction between GC and parathyroid hormone.
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32.
  • Henning, Petra, 1974, et al. (författare)
  • The effect of estrogen on bone requires ER alpha in nonhematopoietic cells but is enhanced by ER alpha in hematopoietic cells
  • 2014
  • Ingår i: American Journal of Physiology-Endocrinology and Metabolism. - : American Physiological Society. - 0193-1849 .- 1522-1555. ; 307:7
  • Tidskriftsartikel (refereegranskat)abstract
    • The effects of estrogen on bone are mediated mainly via estrogen receptor (ER)alpha. ER alpha in osteoclasts (hematopoietic origin) is involved in the trabecular bone-sparing effects of estrogen, but conflicting data are reported on the role of ER alpha in osteoblast lineage cells (nonhematopoietic origin) for bone metabolism. Because Cre-mediated cell-specific gene inactivation used in previous studies might be confounded by nonspecific and/or incomplete cell-specific ER alpha deletion, we herein used an alternative approach to determine the relative importance of ER alpha in hematopoietic (HC) and nonhematopoietic cells (NHC) for bone mass. Chimeric mice with selective inactivation of ER alpha in HC or NHC were created by bone marrow transplantations of wild-type (WT) and ER alpha-knockout (ER alpha(-/-)) mice. Estradiol treatment increased both trabecular and cortical bone mass in ovariectomized WT/WT (defined as recipient/donor) and WT/ER alpha(-/-) mice but not in ER alpha(-/-)/WT or ER alpha(-/-)/ER alpha(-/-) mice. However, estradiol effects on both bone compartments were reduced (similar to 50%) in WT/ER alpha(-/-) mice compared with WT/WT mice. The effects of estradiol on fat mass and B lymphopoiesis required ER alpha specifically in NHC and HC, respectively. In conclusion, ER alpha in NHC is required for the effects of estrogen on both trabecular and cortical bone, but these effects are enhanced by ER alpha in HC.
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33.
  • Henning, Petra, 1974, et al. (författare)
  • The novel cytotoxic polybisphosphonate osteodex decreases bone resorption by enhancing cell death of mature osteoclasts without affecting osteoclastogenesis of RANKL-stimulated mouse bone marrow macrophages
  • 2024
  • Ingår i: INVESTIGATIONAL NEW DRUGS. - : Springer. - 0167-6997 .- 1573-0646.
  • Tidskriftsartikel (refereegranskat)abstract
    • It has previously been demonstrated that the polybisphosphonate osteodex (ODX) inhibits bone resorption in organ-cultured mouse calvarial bone. In this study, we further investigate the effects by ODX on osteoclast differentiation, formation, and function in several different bone organ and cell cultures. Zoledronic acid (ZOL) was used for comparison. In retinoid-stimulated mouse calvarial organ cultures, ODX and ZOL significantly reduced the numbers of periosteal osteoclasts without affecting Tnfsf11 or Tnfrsf11b mRNA expression. ODX and ZOL also drastically reduced the numbers of osteoclasts in cell cultures isolated from the calvarial bone and in vitamin D3-stimulated mouse crude bone marrow cell cultures. These data suggest that ODX can inhibit osteoclast formation by inhibiting the differentiation of osteoclast progenitor cells or by directly targeting mature osteoclasts. We therefore assessed if osteoclast formation in purified bone marrow macrophage cultures stimulated by RANKL was inhibited by ODX and ZOL and found that the initial formation of mature osteoclasts was not affected, but that the bisphosphonates enhanced cell death of mature osteoclasts. In agreement with these findings, ODX and ZOL did not affect the mRNA expression of the osteoclastic genes Acp5 and Ctsk and the osteoclastogenic transcription factor Nfatc1. When bone marrow macrophages were incubated on bone slices, ODX and ZOL inhibited RANKL-stimulated bone resorption. In conclusion, ODX does not inhibit osteoclast formation but inhibits osteoclastic bone resorption by decreasing osteoclast numbers through enhanced cell death of mature osteoclasts.
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34.
  • Henning, Petra, 1974, et al. (författare)
  • The subcellular location of antigen expressed by adenoviral vectors modifies adaptive immunity but not dependency on cross-presenting dendritic cells.
  • 2011
  • Ingår i: European journal of immunology. - : Wiley. - 1521-4141 .- 0014-2980. ; 41:8, s. 2185-96
  • Tidskriftsartikel (refereegranskat)abstract
    • Adenoviral (Ad) vaccine vectors can generate protective immunity to various pathogens in animal studies. However, recent failures in clinical vaccine trials have underscored the need for a better understanding of how mucosal immune responses to Ad-encoded vaccine Ags are generated in vivo. In this study, we addressed whether directing Ad-encoded ovalbumin (OVA) to different subcellular compartments influences the generation of OVA-specific acquired immunity and the APCs required following i.n. immunization of mice. We show that both secreted and membrane-anchored OVA activate CD4(+) T cells, induce cytotoxic CD8(+) T lymphocytes (CTLs) and generate serum IgG. Additionally, vaginal IgG is induced when OVA is expressed at these subcellular locations, but only the secreted form generates a significant IgA response in the lungs. On the contrary, intracellular expression of OVA efficiently expands CD8(+) T cells but fails to activate CD4(+) T cells, results in poor CTL activity, and does not generate Abs. Finally, we show that regardless of the subcellular localization of OVA, conventional DCs (cDCs) are required for the activation of T cells. However, the direct transduction of conventional DCs is not essential. These findings have important implications for the improvement of Ad vector design and vaccine-induced mucosal immunity.
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35.
  • Henning, Petra, 1974, et al. (författare)
  • Toll-like receptor-2 induced inflammation causes local bone formation and activates canonical Wnt signaling.
  • 2024
  • Ingår i: Frontiers in immunology. - : Frontiers Media S.A.. - 1664-3224. ; 15:5
  • Tidskriftsartikel (refereegranskat)abstract
    • It is well established that inflammatory processes in the vicinity of bone often induce osteoclast formation and bone resorption. Effects of inflammatory processes on bone formation are less studied. Therefore, we investigated the effect of locally induced inflammation on bone formation. Toll-like receptor (TLR) 2 agonists LPS from Porphyromonas gingivalis and PAM2 were injected once subcutaneously above mouse calvarial bones. After five days, both agonists induced bone formation mainly at endocranial surfaces. The injection resulted in progressively increased calvarial thickness during 21 days. Excessive new bone formation was mainly observed separated from bone resorption cavities. Anti-RANKL did not affect the increase of bone formation. Inflammation caused increased bone formation rate due to increased mineralizing surfaces as assessed by dynamic histomorphometry. In areas close to new bone formation, an abundance of proliferating cells was observed as well as cells robustly stained for Runx2 and alkaline phosphatase. PAM2 increased the mRNA expression of Lrp5, Lrp6 and Wnt7b, and decreased the expression of Sost and Dkk1. In situ hybridization demonstrated decreased Sost mRNA expression in osteocytes present in old bone. An abundance of cells expressed Wnt7b in Runx2-positive osteoblasts and ß-catenin in areas with new bone formation. These data demonstrate that inflammation, not only induces osteoclastogenesis, but also locally activates canonical WNT signaling and stimulates new bone formation independent on bone resorption.
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36.
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37.
  • Henning, Petra, 1974, et al. (författare)
  • WNT16 is Robustly Increased by Oncostatin M in Mouse Calvarial Osteoblasts and Acts as a Negative Feedback Regulator of Osteoclast Formation Induced by Oncostatin M
  • 2021
  • Ingår i: Journal of Inflammation Research. ; 14, s. 4723-4741
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Bone loss is often observed adjacent to inflammatory processes. The WNT signaling pathways have been implicated as novel regulators of both immune responses and bone metabolism. WNT16 is important for cortical bone mass by inhibiting osteoclast differentiation, and we have here investigated the regulation of WNT16 by several members of the pro-inflammatory gp130 cytokine family. Methods: The expression and regulation of Wnt16 in primary murine cells were studied by qPCR, scRNAseq and in situ hybridization. Signaling pathways were studied by siRNA silencing. The importance of oncostatin M (OSM)-induced WNT16 expression for osteoclastogenesis was studied in cells from Wnt16-deficient and wild-type mice. Results: We found that IL-6/sIL-6R and OSM induce the expression of Wnt16 in primary mouse calvarial osteoblasts, with OSM being the most robust stimulator. The induction of Wnt16 by OSM was dependent on gp130 and OSM receptor (OSMR), and downstream signaling by the SHC1/STAT3 pathway, but independent of ERK. Stimulation of the calvarial cells with OSM resulted in enhanced numbers of mature, oversized osteoclasts when cells were isolated from Wnt16 deficient mice compared to cells from wild-type mice. OSM did not affect Wnt16 mRNA expression in bone marrow cell cultures, explained by the finding that Wnt16 and Osmr are expressed in distinctly different cells in bone marrow, nor was osteoclast differentiation different in OSM-stimulated bone marrow cell cultures isolated from Wnt16-/- or wild-type mice. Furthermore, we found that Wnt16 expression is substantially lower in cells from bone marrow compared to calvarial osteoblasts. Conclusion: These findings demonstrate that OSM is a robust stimulator of Wnt16 mRNA in calvarial osteoblasts and that WNT16 acts as a negative feedback regulator of OSMinduced osteoclast formation in the calvarial bone cells, but not in the bone marrow.
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38.
  • Hong, Saw See, et al. (författare)
  • Adenovirus stripping: a versatile method to generate adenovirus vectors with new cell target specificity.
  • 2003
  • Ingår i: Molecular therapy : the journal of the American Society of Gene Therapy. - 1525-0016. ; 7:5 Pt 1, s. 692-9
  • Tidskriftsartikel (refereegranskat)abstract
    • We developed a new type of adenovirus type 5 (Ad5)-derived vector with genetically modified fiber proteins whose knob domains could be stripped off due to the insertion of a single Factor Xa cleavage site in the fiber shaft, between a cellular ligand and the knob domain. This Ad vector did not require a specific cell line for propagation and could be grown in HEK-293 cells. Stripping off the knob domains removed the endogenous cell-binding moiety of Ad but retained the new cell ligand for retargeting purposes. As experimental models for cell ligands, we used two peptides with different sequence complexities: (i) the integrin-binding tripeptide RGD and (ii) a 58-residue oligopeptide termed affibody (Zwt). Zwt binds specifically to the human IgG1 Fc domain or to its Fc3(1) homolog. The modified fibers were efficiently encapsidated into virions, and the Factor Xa sites were fully accessible to proteolysis. In vitro binding assays using recombinant Fc3(1) protein and Ad5-mediated gene transduction of Fc3(1)-expressing cells demonstrated that the proteolytically deknobbed Ad5-Zwt vector was functional and specific for receptor targeting.
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39.
  • Horkeby, Karin L, et al. (författare)
  • Phosphorylation of S122 in ERα is important for the skeletal response to estrogen treatment in male mice
  • 2022
  • Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 12:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Estrogen receptor alpha (ERα) signaling has beneficial skeletal effects in males. ERα signaling also affects other tissues, and to find bone-specific treatments, more knowledge regarding tissue-specific ERα signaling is needed. ERα is subjected to posttranslational modifications, including phosphorylation, which can influence ERα function in a tissue-specific manner. To determine the importance of phosphorylation site S122 (corresponding to human ERα site S118) for the skeleton and other tissues, male mice with a S122A mutation were used. Total areal bone mineral density was similar between gonadal intact S122A and WT littermates followed up to 12months of age, and weights of estrogen-responsive organs normalized for body weight were unchanged between S122A and WT males at both 3 and 12months of age. Interestingly, 12-month-old S122A males had decreased body weight compared to WT. To investigate if site S122 affects the estrogen response in bone and other tissues, 12-week-old S122A and WT males were orchidectomized (orx) and treated with estradiol (E2) or placebo pellets for four weeks. E2 increased cortical thickness in tibia in both orx WT (+ 60%, p < 0.001) and S122A (+ 45%, p < 0.001) males. However, the E2 effect on cortical thickness was significantly decreased in orx S122A compared to WT mice (−24%, p < 0.05). In contrast, E2 affected trabecular bone and organ weights similarly in orx S122A and WT males. Thus, ERα phosphorylation site S122 is required for a normal E2 response specifically in cortical bone in male mice, a finding that may have implications for development of future treatments against male osteoporosis.
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40.
  • Jiang, Yiwen, et al. (författare)
  • Membrane estrogen receptor alpha signaling modulates the sensitivity to estradiol treatment in a dose- and tissue- dependent manner
  • 2023
  • Ingår i: Scientific Reports. - 2045-2322. ; 13:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Estradiol (E2) affects both reproductive and non-reproductive tissues, and the sensitivity to different doses of E2 varies between tissues. Membrane estrogen receptor alpha (mER alpha)-initiated signaling plays a tissue-specific role in mediating E2 effects, however, it is unclear if mER alpha signaling modulates E2 sensitivity. To determine this, we treated ovariectomized C451A females, lacking mER alpha signaling, and wildtype (WT) littermates with physiological (0.05 mu g/mouse/day (low); 0.6 mu g/mouse/day (medium)) or supraphysiological (6 mu g/mouse/day (high)) doses of E2 (17 beta-estradiol-3-benzoate) for three weeks. Low-dose treatment increased uterus weight in WT, but not C451A mice, while non-reproductive tissues (gonadal fat, thymus, trabecular and cortical bone) were unaffected in both genotypes. Medium-dose treatment increased uterus weight and bone mass and decreased thymus and gonadal fat weights in WT mice. Uterus weight was also increased in C451A mice, but the response was significantly attenuated (- 85%) compared to WT mice, and no effects were triggered in non-reproductive tissues. High-dose treatment effects in thymus and trabecular bone were significantly blunted (- 34% and - 64%, respectively) in C451A compared to WT mice, and responses in cortical bone and gonadal fat were similar between genotypes. Interestingly, the high dose effect in uterus was enhanced (+ 26%) in C451A compared to WT mice. In conclusion, loss of mER alpha signaling reduces the sensitivity to physiological E2 treatment in both non-reproductive tissues and uterus. Furthermore, the E2 effect after high-dose treatment in uterus is enhanced in the absence of mER alpha, suggesting a protective effect of mER alpha signaling in this tissue against supraphysiological E2 levels.
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41.
  • Kassem, Ali, et al. (författare)
  • Porphyromonas gingivalis Stimulates Bone Resorption by Enhancing RANKL (Receptor Activator of NF-kappa B Ligand) through Activation of Toll-like Receptor 2 in Osteoblasts
  • 2015
  • Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 290:33, s. 20147-20158
  • Tidskriftsartikel (refereegranskat)abstract
    • Periodontitis has been associated with rheumatoid arthritis. In experimental arthritis, concomitant periodontitis caused by oral infection with Porphyromonas gingivalis enhances articular bone loss. The aim of this study was to investigate how lipopolysaccharide (LPS) from P. gingivalis stimulates bone resorption. The effects by LPS P. gingivalis and four other TLR2 ligands on bone resorption, osteoclast formation, and gene expression in wild type and Tlr2-deficient mice were assessed in ex vivo cultures of mouse parietal bones and in an in vivo model in which TLR2 agonists were injected subcutaneously over the skull bones. LPS P. gingivalis stimulated mineral release and matrix degradation in the parietal bone organ cultures by increasing differentiation and formation of mature osteoclasts, a response dependent on increased RANKL (receptor activator of NF-kappa B ligand). LPS P. gingivalis stimulated RANKL in parietal osteoblasts dependent on the presence of TLR2 and through a MyD88 and NF-kappa B-mediated mechanism. Similarly, the TLR2 agonists HKLM, FSL1, Pam2, and Pam3 stimulated RANKL in osteoblasts and parietal bone resorption. LPS P. gingivalis and Pam2 robustly enhanced osteoclast formation in periosteal/endosteal cell cultures by increasing RANKL. LPS P. gingivalis and Pam2 also up-regulated RANKL and osteoclastic genes in vivo, resulting in an increased number of periosteal osteoclasts and immense bone loss in wild type mice but not in Tlr2-deficient mice. These data demonstrate that LPS P. gingivalis stimulates periosteal osteoclast formation and bone resorption by stimulating RANKL in osteoblasts via TLR2. This effect might be important for periodontal bone loss and for the enhanced bone loss seen in rheumatoid arthritis patients with concomitant periodontal disease.
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42.
  • Kassem, Ali, et al. (författare)
  • TLR5, a novel mediator of innate immunity-induced osteoclastogenesis and bone loss
  • 2015
  • Ingår i: Faseb Journal. - : Wiley. - 0892-6638 .- 1530-6860. ; 29:11, s. 4449-4460
  • Tidskriftsartikel (refereegranskat)abstract
    • Accumulating evidence points to the importance of the innate immune system in inflammation-induced bone loss in infectious and autoimmune diseases. TLRs are well known for being activated by ligands expressed by bacteria, viruses, and fungi. Recent findings indicate that also endogenous ligands in inflammatory processes are important, one being a TLR5 agonist present in synovial fluid from patients with rheumatoid arthritis (RA). We found that activation of TLR5 by its specific ligand, flagellin, caused robust osteoclast formation and bone loss in cultured mouse neonatal parietal bones dependent on increased receptor activator of NF-kappa B ligand (RANKL): osteoprotegerin ratio, with half-maximal stimulation at 0.01 mu g/ml. Flagellin enhanced Rankl mRNA in isolated osteoblasts by a myeloid differentiation primary response gene 88 and NF-kappa B-dependent mechanism. Injection of flagellin locally over skull bones in 5-wk-oldmice resulted in increased mRNA expression of Rankl and osteoclastic genes, robust osteoclast formation, and bone loss. The effects in vitro and in vivo were absent in Tlr5(-/-) mice. These data show that TLR5 is a novel activator of RANKL and osteoclast formation and, therefore, a potential key factor in inflammation-induced bone erosions in diseases like RA, reactive arthritis, and periodontitis. TLR5 might be a promising novel treatment target for prevention of inflammatory bone loss.
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43.
  • Lagerquist, Marie K, et al. (författare)
  • Acute fat loss does not affect bone mass
  • 2021
  • Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 11:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Obesity has previously been thought to protect bone since high body weight and body mass index are associated with high bone mass. However, some more recent studies suggest that increased adiposity negatively impacts bone mass. Here, we aimed to test whether acute loss of adipose tissue, via adipocyte apoptosis, alters bone mass in age-related obese mice. Adipocyte apoptosis was induced in obese male FAT-ATTAC mice through AP20187 dimerizer-mediated activation of caspase 8 selectively in adipocytes. In a short-term experiment, dimerizer was administered to 5.5 month-old mice that were terminated 2 weeks later. At termination, the total fat mass weighed 58% less in dimerizer-treated mice compared with vehicle-treated controls, but bone mass did not differ. To allow for the detection of long-term effects, we used 9-month-old mice that were terminated six weeks after dimerizer administration. In this experiment, the total fat mass weighed less (- 68%) in the dimerizer-treated mice than in the controls, yet neither bone mass nor biomechanical properties differed between groups. Our findings show that adipose tissue loss, despite the reduced mechanical loading, does not affect bone in age-related obese mice. Future studies are needed to test whether adipose tissue loss is beneficial during more severe obesity.
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44.
  • Lagerquist, Marie K, et al. (författare)
  • Reduction of Mature B Cells and Immunoglobulins Results in Increased Trabecular Bone
  • 2022
  • Ingår i: Jbmr Plus. - : Wiley. - 2473-4039. ; 6:9
  • Tidskriftsartikel (refereegranskat)abstract
    • Inflammation has a significant effect on bone remodeling and can result in bone loss via increased stimulation of osteoclasts. Activated immunoglobulins, especially autoantibodies, can increase osteoclastogenesis and are associated with pathological bone loss. Whether immunoglobulins and mature B lymphocytes are important for general bone architecture has not been completely determined. Here we demonstrate, using a transgenic mouse model, that reduction of mature B cells and immunoglobulins leads to increased trabecular bone mass compared to wild-type (WT) littermate controls. This bone effect is associated with a decrease in the number of osteoclasts and reduced bone resorption, despite decreased expression of osteoprotegerin. We also demonstrate that the reduction of mature B cells and immunoglobulins do not prevent bone loss caused by estrogen deficiency or arthritis compared to WT littermate controls. In conclusion, the reduction of mature B cells and immunoglobulins results in disturbed regulation of trabecular bone turnover in healthy conditions but is dispensable for pathological bone loss. (c) 2022 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.
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45.
  • Lawenius, Lina, et al. (författare)
  • Pasteurized Akkermansia muciniphila protects from fat mass gain but not from bone loss
  • 2020
  • Ingår i: American Journal of Physiology-Endocrinology and Metabolism. - : American Physiological Society. - 0193-1849 .- 1522-1555. ; 318:4
  • Tidskriftsartikel (refereegranskat)abstract
    • Probiotic bacteria can protect from ovariectomy (ovx)-induced bone loss in mice. Akkermansia muciniphila is considered to have probiotic potential due to its beneficial effect on obesity and insulin resistance. The purpose of the present study was to determine if treatment with pasteurized Akkermansia muciniphila (pAkk) could prevent ovx-induced bone loss. Mice were treated with vehicle or pAkk for 4 wk, starting 3 days before ovx or sham surgery. Treatment with pAkk reduced fat mass accumulation confirming earlier findings. However, treatment with pAkk decreased trabecular and cortical bone mass in femur and vertebra of gonadal intact mice and did not protect from ovx-induced bone loss. Treatment with pAkk increased serum parathyroid hormone (PTH) levels and increased expression of the calcium transporter Trpv5 in kidney suggesting increased reabsorption of calcium in the kidneys. Serum amyloid A 3 (SAA3) can suppress bone formation and mediate the effects of PTH on bone resorption and bone loss in mice and treatment with pAkk increased serum levels of SAA3 and gene expression of Saa3 in colon. Moreover, regulatory T cells can be protective of bone and pAkk-treated mice had decreased number of regulatory T cells in mesenteric lymph nodes and bone marrow. In conclusion, treatment with pAkk protected from ovx-induced fat mass gain but not from bone loss and reduced bone mass in gonadal intact mice. Our findings with pAkk differ from some probiotics that have been shown to protect bone mass, demonstrating that not all prebiotic and probiotic factors have the same effect on bone.
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46.
  • Lima Teixeira, Jorge F., et al. (författare)
  • Osteoprotective effect by interleukin-4 (IL-4) on lipoprotein-induced periodontitis
  • 2023
  • Ingår i: Cytokine. - 1043-4666 .- 1096-0023. ; 172
  • Tidskriftsartikel (refereegranskat)abstract
    • Lipoproteins are immunostimulatory bacterial components suggested to participate in inflammation-induced bone loss in periodontal disease through stimulation of osteoclast differentiation. Toll-like receptor 2 activation by Pam2CSK4 (PAM2), known to mimic bacterial lipoproteins, was previously shown to enhance periodontal bone resorption in mice. The anti-inflammatory cytokine interleukin-4 (IL-4) is a known inhibitor of RANKL-induced bone resorption in vitro. Here, we have investigated whether IL-4 could decrease PAM2-induced periodontal bone loss and osteoclastogenesis in vivo. In a model of periodontitis induced by gingival injections of PAM2 in mice, concomitant injections of IL-4 reduced bone loss. Histologically, IL-4 reduced the recruitment of inflammatory cells and the formation of TRAP+ osteoclasts stimulated by PAM2. Mouse bone marrow macrophages (BMMs) and neonatal calvarial osteoblasts were used to assess the effect of IL-4 on PAM2-induced osteoclastogenesis in vitro. In RANKL-primed BMMs stimulated by PAM2 Nfatc1, Ctsk, and Acp5 gene expression was up-regulated and resulted in robust formation of TRAP+ multinucleated osteoclasts, effects which were impaired by IL-4. These effects were mediated by impairment in PAM2-induced c-fos expression. In primary calvarial osteoblast cultures, IL-4 decreased PAM2-induced Tnfsf11 (encoding RANKL) mRNA and enhanced Tnfrsf11b (encoding OPG) expression. Our data demonstrate that the osteoprotective effect by IL-4 on lipoprotein-induced periodontal disease occurs through the inhibition of osteoclastogenesis by three mechanisms, one by acting directly on osteoclast progenitors, another by acting indirectly through decreasing the expression of osteoclast-regulating cytokines in osteoblasts and a third by decreasing inflammation.
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47.
  • Lindholm, Leif, 1940, et al. (författare)
  • Novel strategies in tailoring human adenovirus into therapeutic cencer gene therapy vectors
  • 2008
  • Ingår i: Future Virology. - : Future Medicine Ltd. - 1746-0794 .- 1746-0808. ; 3:1, s. 45-59
  • Forskningsöversikt (refereegranskat)abstract
    • Gene therapy is a novel approach for the treatment of cancer that has so far not been realized. The scope of this review is to try to define the remaining barriers to the successful use of adenovirus vectors for gene and viral therapy of human tumors and to suggest solutions whereby these barriers can be bypassed. It is the conviction of the authors that too many studies have been performed in animal models that are not sufficiently comprehensive to allow conclusions to be drawn for application in humans. For example, in the case of the murine experimental model, in which most studies have been performed, mice are devoid of circulating antibodies to adenovirus type 5 and adenovirus cannot replicate in mouse cells. While the problems are real enough, as witnessed by the quite limited success in human trials, some of the solutions that will be suggested here are hypothetical and have not as yet been tried, even in animals. The review has no ambition to be exhaustive but is intended as a contribution in order to forward the field of gene therapy vectors for systemic clinical application
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48.
  • Lionikaite, Vikte, et al. (författare)
  • Clinically relevant doses of Vitamin A decrease cortical bone mass in mice
  • 2018
  • Ingår i: Journal of Endocrinology. - 0022-0795 .- 1479-6805. ; 239:3, s. 389-402
  • Tidskriftsartikel (refereegranskat)abstract
    • Excess vitamin A has been associated with decreased cortical bone thickness and increased fracture risk. While most studies in rodents have employed high dosages of vitamin A for short periods of time, we investigated the bone phenotype in mice after longer exposure to more clinically relevant doses. For 1, 4 and 10 weeks, mice were fed a control diet (4.5µg retinyl acetate/g chow), a diet modeled from the human upper tolerable limit (UTL; 20µg retinyl acetate/g chow) and a diet three times UTL (supplemented; 60µg retinyl acetate/g chow). Time-dependent decreases in periosteal circumference and bone mineral content were noted with the supplemented dose. These reductions in cortical bone resulted in a significant time-dependent decrease of predicted strength and a non-significant trend toward reduced bone strength as analyzed by three-point bending. Trabecular bone in tibiae and vertebrae remained unaffected when vitamin A was increased in the diet. Dynamic histomorphometry demonstrated that bone formation was substantially decreased after 1 week of treatment at the periosteal site with the supplemental dose. Increasing amount of vitamin A decreased endocortical circumference, resulting in decreased marrow area, a response associated with enhanced endocortical bone formation. In the presence of bisphosphonate, vitamin A had no effect on cortical bone, suggesting that osteoclasts are important, even if effects on bone resorption were not detected by osteoclast counting, genes in cortical bone or analysis of serum TRAP5b and CTX. In conclusion, our results indicate that even clinically relevant doses of vitamin A have a negative impact on the amount of cortical bone. © 2018 The authors Published by Bioscientifica Ltd.
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49.
  • Lionikaite, Vikte, et al. (författare)
  • Effects of retinoids on physiologic and inflammatory osteoclastogenesis in vitro
  • 2018
  • Ingår i: Journal of leukocyte biology. - 1938-3673. ; 104:6, s. 1133-1145
  • Tidskriftsartikel (refereegranskat)abstract
    • Increased intake of vitamin A (retinoids) is associated with decreased bone mass and increased fracture risk in humans. Mechanistic studies in rodents have shown that hypervitaminosis A results in decreased bone mass caused by an increase in periosteal osteoclasts while simultaneously decreasing endocortic osteoclasts. In vivo and ex vivo bone organ cultures have demonstrated that excess retinoids increase osteoclast formation due to increased receptor activator of nuclear factor kappa B-ligand (RANKL) expression. In vitro, studies using murine bone marrow macrophages (BMM) have shown that retinoids inhibit osteoclast formation induced by recombinant RANKL. These opposing in vivo/ex vivo versus in vitro effects may elucidate why excess retinoids affect periosteal and endocortic osteoclast formation differently. In addition, it has been reported that retinoids can inhibit osteoclast formation under inflammatory conditions such as experimentally induced arthritis in mice. In the present study, we have compared the effect of all-trans-retinoic acid (ATRA) on physiologically and inflammatory induced osteoclastogenesis. ATRA inhibited physiologically induced (RANKL) osteoclast formation of human peripheral blood monocytes and mouse BMM as well as human monocytes stimulated with the pro-inflammatory compounds, TNF-α and LPS. The inhibition was due to impeded differentiation, rather than fusion, of mononucleated progenitor cells. ATRA disrupted differentiation by interfering with osteoclastogenic intracellular signaling. In line with this view, overexpression of Tnfrsf11a (encodes for RANK) in BMM could not overcome the inhibition of osteoclastogenesis by ATRA. The data suggest that ATRA inhibits both physiologic and inflammatory osteoclast differentiation of progenitors from the bone marrow and peripheral blood. ©2018 Society for Leukocyte Biology.
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50.
  • Lionikaite, Vikte, et al. (författare)
  • Vitamin A decreases the anabolic bone response to mechanical loading by suppressing bone formation
  • 2019
  • Ingår i: FASEB journal : official publication of the Federation of American Societies for Experimental Biology. - : Federation of American Societies for Experimental Biology. - 1530-6860 .- 0892-6638. ; 33:4, s. 5237-5247
  • Tidskriftsartikel (refereegranskat)abstract
    • Increased vitamin A consumption is associated with decreased cortical bone mass and increased fracture risk in humans. Rodent studies have demonstrated that hypervitaminosis A increases cortical bone resorption, whereas the importance of the effects on bone formation is less well defined. We used an experimental model of increased bone formation by loading of the tibiae to investigate the effect of vitamin A on bone formation. Control [retinol activity equivalents (RAE) 4.5 µg/g chow] or vitamin A (RAE 60 µg/g chow) diets were given to female C57BL/6N mice for 4 wk, after which the tibiae were subjected to axial loading on alternate days for 2 wk, while the diets were continued. Vitamin A inhibited the loading-induced increase in trabecular and cortical bone volume. This was attributed to inhibition of loading-induced increase in osteoblast number and activity, and expression of osteoblastic genes Sp7, Alpl, and Col1a1 in cortical bone. Vitamin A, loading, and combination thereof also resulted in site-specific effects on bone composition measured by Raman spectroscopy. In summary, a clinically relevant dose of vitamin A suppresses the loading-induced gain of bone mass by decreasing bone formation. These observations may have implications for regulation of bone mass caused by physical activity and the risk of osteoporosis in humans.-Lionikaite, V., Henning, P., Drevinge, C., Shah, F. A., Palmquist, A., Wikström, P., Windahl, S. H., Lerner, U. H. Vitamin A decreases the anabolic bone response to mechanical loading by suppressing bone formation.
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