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| 2. |
- Mikkola, H, et al.
(författare)
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Lentivirus gene transfer in murine hematopoietic progenitor cells is compromised by a delay in proviral integration and results in transduction mosaicism and heterogeneous gene expression in progeny cells
- 2000
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Ingår i: Journal of Virology. - : American Society for Microbiology. - 0022-538X .- 1098-5514. ; 74:24, s. 8-11911
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Tidskriftsartikel (refereegranskat)abstract
- Human immunodeficiency virus type 1-based lentivirus vectors containing the green fluorescent protein (GFP) gene were used to transduce murine Lin(-) c-kit(+) Sca1(+) primitive hematopoietic progenitor cells. Following transduction, the cells were plated into hematopoietic progenitor cell assays in methylcellulose and the colonies were scored for GFP positivity. After incubation for 20 h, lentivirus vectors transduced 27.3% +/- 6.7% of the colonies derived from unstimulated target cells, but transduction was more efficient when the cells were supported with stem cell factor (SCF) alone (42. 0% +/- 5.5%) or SCF, interleukin-3 (IL-3), and IL-6 (53.3 +/- 1.8%) during transduction. The, vesicular stomatitis virus glycoprotein-pseudotyped MGIN oncoretrovirus control vector required IL-3, IL-6, and SCF for significant transduction (39.3 +/- 9.4%). Interestingly, only a portion of the progeny cells within the lentivirus-transduced methylcellulose colonies expressed GFP, in contrast to the homogeneous expression in oncoretrovirus-transduced colonies. Secondary plating of the primary GFP(+) lentivirus vector-transduced colonies revealed vector PCR(+) GFP(+) (42%), vector PCR(-) GFP(-) (46%), and vector PCR(+) GFP(-) (13%) secondary colonies, indicating true genetic mosaicism with respect to the viral genome in the progeny cells. The degree of vector mosaicism in individual colonies could be reduced by extending the culture time after transduction and before plating into the clonal progenitor cell assay, indicating a delay in the lentiviral integration process. Furthermore, supplementation with exogenous deoxynucleoside triphosphates during transduction decreased mosaicism within the colonies. Although cytokine stimulation during transduction correlates with higher transduction efficiency, rapid cell division after transduction may result in loss of the viral genome in the progeny cells. Therefore, optimal transduction may require activation without promoting intense cell proliferation prior to vector integration.
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| 3. |
- Persson-Sjögren, S, et al.
(författare)
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Mouse islets cultured with vasoactive intestinal polypeptide : effects on insulin release and immunoreactivity for tyrosine hydroxylase.
- 2001
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Ingår i: Pancreas. - 0885-3177 .- 1536-4828. ; 22:1, s. 84-90
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Tidskriftsartikel (refereegranskat)abstract
- Mouse islets cultured for 1 or 4 days with or without 10 nM vasoactive intestinal polypeptide (VIP) were stained for tyrosine hydroxylase (TH) and examined for insulin secretion during culture and in a postculture perifusion system. Exposure to exogenous VIP for 4 days increased the frequency of islet cells expressing TH-like immunoreactivity. Regardless of the culturing conditions, the islets exhibited significant insulin secretory responses to 16.7 mM glucose, the effect being potentiated by 10 nM VIP in the perifusion medium. The insulin-releasing action of glucose and the potentiating effect of VIP were less pronounced in islets cultured for 1 day with VIP than in islets cultured without this neuropeptide. The following conclusions are suggested: (a) VIP stimulates the expression of TH in mouse islet cells; (b) the latency of the VIP-induced TH is a postreceptor phenomenon; (c) islet cultures exposed to VIP represent a new instance of the association between increased functional demands on beta cells and enhanced expression of TH and a new instance of VIP having trophic effects.
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| 4. |
- Persson-Sjögren, S, et al.
(författare)
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Tyrosine hydroxylase in mouse pancreatic islet cells, in situ and after syngeneic transplantation to kidney.
- 2002
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Ingår i: Histology and Histopathology. - 0213-3911 .- 1699-5848. ; 17:1, s. 113-21
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Tidskriftsartikel (refereegranskat)abstract
- Tyrosine hydroxylase (TH) is co-expressed with islet hormones in the fetal mouse pancreas. In the adult animal, the enzyme has been considered as a marker of ageing beta-cells. By immunohistochemical staining, we analyzed the expression of TH-like immunoreactivity (TH-LI), insulin-LI (INS-LI) and somatostatin-LI (SOM-LI) in adult mouse islets, in situ and after isolation and transplantation to kidney. In pancreas in situ, most TH-LI cells expressed INS-LI while less than 5% expressed SOM-LI. The total number of TH-LI cells/mm2 was significantly increased directly after isolation and in 0-day, 12-week and 52-week old grafts, but not in 3-day grafts. The proportion of TH-LI cells expressing SOM-LI increased after transplantation, amounting to about one-third by 52 weeks. As expressed per unit islet area, the frequencies of both TH/INS and TH/SOM cells increased significantly in the transplants. The results demonstrate that TH occurs in both beta-cells and D-cells of adult islets. In both cell types the enzyme appears to be responsive to the microenvironmental changes inherent in transplantation. This cellular phenotype plasticity might contribute to the altered insulin secretory dynamics in islet grafts.
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| 5. |
- Regland, Björn, 1947, et al.
(författare)
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Treatment of Alzheimer's disease with clioquinol.
- 2001
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Ingår i: Dementia and geriatric cognitive disorders. - 1420-8008. ; 12:6, s. 408-14
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Tidskriftsartikel (refereegranskat)abstract
- As heavy metal ions may be implicated in the formation of senile plaques in Alzheimer-afflicted brains, treatment with clioquinol was tested in 20 patients with Alzheimer's disease. Clioquinol is a chelator that crosses the blood-brain barrier and has greater affinity for zinc and copper ions than for calcium and magnesium ions. Treatment was given for 21 days at doses of 20 mg/day to 10 patients and 80 mg/day to another 10 patients. The study was blind to the dosages but included no controls. Cerebrospinal fluid (CSF) investigations revealed a significant increase at day 7 and a decrease at day 21 in Tau protein and growth-associated protein (GAP43). These proteins are increased in Alzheimer's disease and considered as rather stable markers. The initial increase may indicate a temporary cytotoxicity to the brain and/or an increased release into the CSF from stores in the tissue, possibly from senile plaques where the proteins are accumulated. The levels of CSF-Tau protein correlated positively and significantly with the serum levels of copper and also with the serum copper/zinc ratio. Clinical ratings showed slight improvement after 3 weeks treatment with clioquinol in this open study.
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| 8. |
- Tallheden, Tommi, 1972, et al.
(författare)
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Phenotypic plasticity of human articular chondrocytes.
- 2003
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Ingår i: The Journal of bone and joint surgery. American volume. - 0021-9355. ; 85-A Suppl 2, s. 93-100
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Tidskriftsartikel (refereegranskat)abstract
- Progenitor cells in mesenchymal tissues are important in the maintenance of tissue homeostasis and regeneration capacity. Articular cartilage is a tissue with a very low capacity for repair. One explanation could be the lack of chondrogenic progenitor cells within the adult tissue. As a test of chondrogenic differentiation potential, we examined the ability of isolated chondrocytes to take on several phenotypic identities within the mesenchymal lineage by applying culture techniques and markers used in the study of the phenotypic plasticity of marrow-derived mesenchymal stem cells (MSCs).
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| 9. |
- Wilmot, Carrie M., et al.
(författare)
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Defining redox state of X-ray crystal structures by single-crystal ultraviolet visible microspectrophotometry
- 2002
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Ingår i: Methods in Enzymology. - 0076-6879 .- 1557-7988. ; 353, s. 301-318
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Tidskriftsartikel (refereegranskat)abstract
- Exciting results have been emerging from the field of single-crystal X-ray crystallography, giving unprecedented detail of freeze-trapped reaction intermediates from important classes of macromolecules that contain chromophores. These structures have been coupled with single-crystal UV-visible microspectrophotometry. This has defined the distinct catalytic intermediates present in the crystal structures, allowing the correlation of electronic transitions with the observed structural transitions. Of particular note is that many of these structures have been generated “on the fly” during kinetic turnover in the crystal. Most enzymatic reactions proceed through distinct catalytic intermediates that, under favorable conditions, may accumulate transiently in the crystal during turnover. In some cases, the physical constraints of the contacts within crystals may also lead to a significant slowing of the reaction at certain points along the pathway where conformational changes are required. This can lead to a transient build-up of spectrally distinct intermediates in the crystal that can be trapped by flash freezing in liquid nitrogen, allowing a complete single-crystal data set to be collected to the highest possible resolution at a later time. Similar build-up of intermediates may be achieved by altering the pH, temperature, or the solvent environment around the protein in the crystal, or by producing engineered variants that build up an intermediate of interest. The chapter focuses on the technical considerations required to carry out UV-visible microspectroscopy of single crystals.
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