| 1. |
- Padma, Arvind M., et al.
(författare)
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Decellularization protocol-dependent damage-associated molecular patterns in rat uterus scaffolds differentially affect the immune response after transplantation.
- 2021
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Ingår i: Journal of tissue engineering and regenerative medicine. - : Hindawi Limited. - 1932-7005 .- 1932-6254. ; 15:7, s. 674-685
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Tidskriftsartikel (refereegranskat)abstract
- Scaffolds derived from decellularized tissue possess many advantages for bioengineering applications, including for novel infertility treatments. However, the decellularization process results in allogenic-independent damage-associated molecular patterns (DAMPs). This field is poorly studied, in particular for uterus bioengineering applications. An increased knowledge concerning the immune system activation after transplantation of decellularized tissue will enable safer construct development and thereby accelerate translation from research to clinic. We therefore transplanted rat uterus scaffolds produced by three different decellularization protocols based on Triton X-100 (P1 and P2) or sodium deoxycholate (P3) in a syngeneic animal model and assessed the immune response towards DAMPs exposed by the decellularization process. Biopsies were retrieved on day 5, 15, and 30 post transplantation and immunohistochemistry-stained CD45+ (leucocytes), CD4+ (T-cells), CD8a+ (cytotoxic T-cells), CD22+ (B-cells), NCR1+ (NK-cells), CD68+ (pan-macrophages), and CD163+ (M2 macrophages) cells within the grafts were quantified. The gene expression for interferon γ, interleukin (IL)-1β, IL-2, IL-6, and tumor necrosis factor (TNF) eotaxin-2, RANTES, MCP-1, MIP-1α, MIP-3α, IL-8 were also measured. Scaffolds from P1 induced a rapid cell infiltration after transplantation, presumably induced by DNA-based DAMPs. However, this response was only transient. Protocol 3 derived scaffolds induced an early pro-inflammatory cytokine response at the transcript level which remained high throughout the study. This response may be caused by the stronger decellularization detergent that could expose more extracellular matrix-related DAMPs. However, earlier proteomics analysis also identified significantly more abundant heat shook proteins-related DAMPs in this scaffold type. Protocol 2 caused the least immunogenic scaffolds and should thus be the future focus for in vivo uterus bioengineering applications.
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| 2. |
- Padma, Arvind M., et al.
(författare)
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Immune response after allogeneic transplantation of decellularized uterine scaffolds in the rat.
- 2021
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Ingår i: Biomedical Materials. - 1748-605X. ; 16:4
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Tidskriftsartikel (refereegranskat)abstract
- Data on how the immune system reacts to decellularized scaffolds after implantation is scarce and difficult to interpret due to many heterogeneous parameters such as tissue-type match, decellularization method and treatment application. The engraftment of these scaffolds must prove safe and that they remain inert to the recipient's immune system to enable successful translational approaches and potential future clinical evaluation. Herein, we investigated the immune response after the engraftment of three decellularized scaffold types that previously showed potential to repair a uterine injury in the rat. Protocol (P) 1 and P2 were based on Triton-X100 and generated scaffolds containing 820 ng mg-1and 33 ng mg-1donor DNA per scaffold weight, respectively. Scaffolds obtained with a sodium deoxycholate-based protocol (P3) contained 160 ng donor DNA per mg tissue. The total number of infiltrating cells, and the population of CD45+leukocytes, CD4+T-cells, CD8a+cytotoxic T-cells, CD22+B-cells, NCR1+NK-cells, CD68+and CD163+macrophages were quantified on days 5, 15 and 30 after a subcutaneous allogenic (Lewis to Sprague Dawley) transplantation. Gene expression for the pro-inflammatory cytokines INF-γ, IL-1β, IL-2, IL-6 and TNF were also examined. P1 scaffolds triggered an early immune response that may had been negative for tissue regeneration but it was stabilized after 30 d. Conversely, P3 initiated a delayed immune response that appeared negative for scaffold survival. P2 scaffolds were the least immunogenic and remained similar to autologous tissue implants. Hence, an effective decellularization protocol based on a mild detergent was advantageous from an immunological perspective and appears the most promising for futurein vivouterus bioengineering applications.
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| 3. |
- Padma, Arvind M., et al.
(författare)
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The development of an extended normothermic ex vivo reperfusion model of the sheep uterus to evaluate organ quality after cold ischemia in relation to uterus transplantation
- 2019
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Ingår i: Acta Obstetricia et Gynecologica Scandinavica. - : Wiley. - 0001-6349 .- 1600-0412. ; 98:9, s. 1127-1138
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Uterus transplantation has recently proved that infertility in women with uterine factor infertility can be cured. It is still an experimental procedure with numerous critical details remaining to be established, including tolerance to warm and cold ischemic insults. In preparation for human uterus transplantation trials, most teams use the sheep as a model system for research and team training, since the vasculature and the uterus is of similar size as in the human. We, therefore, aimed to develop an ex vivo sheep uterus reperfusion platform that mimics the reperfusion situation so that initial assessments and comparisons can be performed without the need for costly and labor-intensive in vivo transplantation experiments. Material and methods: Isolated sheep uteri were perfused with the preservation solution IGL-1 and were then exposed to cold ischemia for either 4(n=6) or 48hours (n=7). Uteri were then reperfused for 48hours under normothermic conditions with an oxygenated recirculating perfusate containing growth factors and synthetic oxygen carriers. Histological and biochemical analysis of the perfusate was conducted to assess reperfusion injury. Results: Quantification of cell density indicated no significant edema in the myometrium or in the endometrium of uteri exposed to 4hours cold ischemia and then a normothermic ex vivo reperfusion for 48hours. Only the outer serosa layer and the inner columnar luminal epithelial cells were affected by the reperfusion. However, a much faster and severe reperfusion damage of all uterine layers were evident during the reperfusion experiment following 48hours of cold ischemia. This was indicated by major accumulation of extracellular fluid, presence of apoptotic-labeled glandular epithelial layer and vascular endothelium. A significant accumulation of lactate was measured in the perfusate with a subsequent decrease in pH. Conclusions: We developed a novel ex vivo sheep uterus model for prolonged perfusion. This model proved to be able to distinguish reperfusion injury-related differences associated to organ preservation. The experimental setup is a platform that can be used to conduct further studies on uterine ischemia- and reperfusion injury that may lead to improved human uterus transplantation protocols. © 2019 Nordic Federation of Societies of Obstetrics and Gynecology
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| 4. |
- Padma, Arvind M., et al.
(författare)
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Towards a bioengineered uterus: bioactive sheep uterus scaffolds are effectively recellularized by enzymatic preconditioning
- 2021
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Ingår i: Npj Regenerative Medicine. - : Springer Science and Business Media LLC. - 2057-3995. ; 6:1
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Tidskriftsartikel (refereegranskat)abstract
- Uterine factor infertility was considered incurable until recently when we reported the first successful live birth after uterus transplantation. However, risky donor surgery and immunosuppressive therapy are factors that may be avoided with bioengineering. For example, transplanted recellularized constructs derived from decellularized tissue restored fertility in rodent models and mandate translational studies. In this study, we decellularized whole sheep uterus with three different protocols using 0.5% sodium dodecyl sulfate, 2% sodium deoxycholate (SDC) or 2% SDC, and 1% Triton X-100. Scaffolds were then assessed for bioactivity using the dorsal root ganglion and chorioallantoic membrane assays, and we found that all the uterus scaffolds exhibited growth factor activity that promoted neurogenesis and angiogenesis. Extensive recellularization optimization was conducted using multipotent sheep fetal stem cells and we report results from the following three in vitro conditions; (a) standard cell culturing conditions, (b) constructs cultured in transwells, and (c) scaffolds preconditioned with matrix metalloproteinase 2 and 9. The recellularization efficiency was improved short-term when transwells were used compared with standard culturing conditions. However, the recellularization efficiency in scaffolds preconditioned with matrix metalloproteinases was 200-300% better than the other strategies evaluated herein, independent of decellularization protocol. Hence, a major recellularization hurdle has been overcome with the improved recellularization strategies and in vitro platforms described herein. These results are an important milestone and should facilitate the production of large bioengineered grafts suitable for future in vivo applications in the sheep, which is an essential step before considering these principles in a clinical setting.
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| 5. |
- Sehic, Edina, et al.
(författare)
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Mesenchymal stem cells establish a pro-regenerative immune milieu after decellularized rat uterus tissue transplantation
- 2022
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Ingår i: Journal of Tissue Engineering. - : SAGE Publications. - 2041-7314. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Decellularized tissue is generally considered immune privileged after transplantation and is an attractive scaffold type for tissue regeneration, including applications for infertility treatment. However, the immune response following transplantation of decellularized tissue is insufficiently studied, in particular after they have been recellularized with mesenchymal stem cells (MSCs). Therefore, we replaced a large uterus segment with a bioengineered graft developed from decellularized uterus tissue and analyzed the immune response during the first 4 months in acellular or MSCs-recellularized scaffolds in the rat. Immunohistochemistry-stained infiltrating immune cells and plasma levels for 16 cytokines and chemokines were quantified. Results revealed that MSCs created an advantageous microenvironment by increasing anti-inflammatory interleukin 10 levels, and increasing the population of FOXP3(+) T-Regs and CD163(+) M2 macrophages, and by reducing the CD8(+) cytotoxic T cell population. Hence, MSCs should be considered an immunotherapeutic cell source with the ability to dictate regeneration success after decellularized tissue transplantation.
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| 6. |
- Tiemann, Tom T, et al.
(författare)
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Towards uterus tissue engineering: a comparative study of sheep uterus decellularisation
- 2020
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Ingår i: Molecular human reproduction. - : Oxford University Press (OUP). - 1460-2407 .- 1360-9947. ; 26:3, s. 167-178
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Tidskriftsartikel (refereegranskat)abstract
- © The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. Uterus tissue engineering may dismantle limitations in current uterus transplantation protocols. A uterine biomaterial populated with patient-derived cells could potentially serve as a graft to circumvent complicated surgery of live donors, immunosuppressive medication and rejection episodes. Repeated uterine bioengineering studies on rodents have shown promising results using decellularised scaffolds to restore fertility in a partially impaired uterus and now mandate experiments on larger and more human-like animal models. The aim of the presented studies was therefore to establish adequate protocols for scaffold generation and prepare for future in vivo sheep uterus bioengineering experiments. Three decellularisation protocols were developed using vascular perfusion through the uterine artery of whole sheep uteri obtained from slaughterhouse material. Decellularisation solutions used were based on 0.5% sodium dodecyl sulphate (Protocol 1) or 2% sodium deoxycholate (Protocol 2) or with a sequential perfusion of 2% sodium deoxycholate and 1% Triton X-100 (Protocol 3). The scaffolds were examined by histology, extracellular matrix quantification, evaluation of mechanical properties and the ability to support foetal sheep stem cells after recellularisation. We showed that a sheep uterus can successfully be decellularised while maintaining a high integrity of the extracellular components. Uteri perfused with sodium deoxycholate (Protocol 2) were the most favourable treatment in our study based on quantifications. However, all scaffolds supported stem cells for 2weeks in vitro and showed no cytotoxicity signs. Cells continued to express markers for proliferation and maintained their undifferentiated phenotype. Hence, this study reports three valuable decellularisation protocols for future in vivo sheep uterus bioengineering experiments.
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| 7. |
- Alshaikh, Ahmed Baker, et al.
(författare)
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Decellularization and recellularization of the ovary for bioengineering applications; studies in the mouse.
- 2020
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Ingår i: Reproductive biology and endocrinology : RB&E. - : Springer Science and Business Media LLC. - 1477-7827. ; 18:1
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Tidskriftsartikel (refereegranskat)abstract
- Fertility preservation is particularly challenging in young women diagnosed with hematopoietic cancers, as transplantation of cryopreserved ovarian cortex in these women carries the risk for re-introducing cancer cells. Therefore, the construction of a bioengineered ovary that can accommodate isolated small follicles was proposed as an alternative to minimize the risk of malignancy transmission. Various options for viable bioengineered scaffolds have been reported in the literature. Previously, we reported three protocols for producing mouse ovarian scaffolds with the decellularization technique. The present study examined these scaffolds further, specifically with regards to their extracellular composition, biocompatibility and ability to support recellularization with mesenchymal stem cells.Three decellularization protocols based on 0.5% sodium dodecyl sulfate (Protocol 1; P1), or 2% sodium deoxycholate (P2), or a combination of the two detergents (P3) were applied to produce three types of scaffolds. The levels of collagen, elastin and sulfated glycosaminoglycans (sGAGs) were quantified in the remaining extracellular matrix. Detailed immunofluorescence and scanning electron microscopy imaging were conducted to assess the morphology and recellularization efficiency of the constructs after 14days in vitro utilizing red fluorescent protein-labelled mesenchymal stem cells.All protocols efficiently removed the DNA while the elastin content was not significantly reduced during the procedures. The SDS-protocol (P1) reduced the sGAG and the collagen content more than the SDC-protocol (P2). All scaffolds were biocompatible and recellularization was successful, particularly in several P2-derived scaffolds. The cells were extensively distributed throughout the constructs, with a denser distribution observed towards the ovarian cortex. The cell density was not significantly different (400 to 550 cells/mm2) between scaffold types. However, there was a tendency towards a higher cell density in the SDC-derived constructs. Scanning electron microscope images showed fibrous scaffolds with a dense repopulated surface structure.While there were differences in the key structural macromolecules between protocols, all scaffolds were biocompatible and showed effective recellularization. The results indicate that our SDC-protocol might be better than our SDS-protocol. However, additional studies are necessary to determine their suitability for attachment of small follicles and folliculogenesis.
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| 8. |
- Alshaikh, Ahmed Baker, et al.
(författare)
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Decellularization of the mouse ovary: comparison of different scaffold generation protocols for future ovarian bioengineering.
- 2019
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Ingår i: Journal of ovarian research. - : Springer Science and Business Media LLC. - 1757-2215. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- In order to preserve fertility in young women with disseminated cancer, e.g. leukemia, an approach that has been suggested is to retransplant isolated small follicles within an ovarian matrix free from malignant cells and with no risk for contamination. The present study evaluates the first step to create a bioengineered ovarian construct that can act as growth-supporting tissue for isolated small follicles that are dependent on a stroma for normal follicular maturation. The present study used the intact mouse ovary to develop a mouse ovarian scaffold through various protocols of decellularization.Potential Immunogenic DNA and intracellular components were removed from whole mouse ovaries by agitation in a 0.5% sodium dodecyl sulfate solution (Protocol 1; P1), or in a 2% sodium deoxycholate solution (P2) or by a combination of the two (P3). The remaining decelluralized ovarian extracellular matrix structure was then assessed based on the DNA- and protein content, and was further evaluated histologically by haematoxylin and eosin-, Verhoeff's van gieson- (for elastin), Masson's trichrome- (for collagens) and Alcian blue (for glycosaminoglycans) staining. We also evaluated the decellularization efficiency using the mild detergent Triton-X100 (1%).Sodium dodecyl sulfate efficiently removed DNA and intracellular components from the ovarian tissue but also significantly reduced the integrity of the remaining ovarian extracellular matrix. Sodium deoxycholate, a considerably milder detergent compared to sodium dodecyl sulfate, preserved the ovarian extracellular matrix better, evident by a more distinct staining for glycosaminoglycan, collagen and elastic fibres. Triton-X100 was found ineffective as a decellularization reagent for mouse ovaries in our settings.The sodium dodecyl sulfate generated ovarian scaffolds contained minute amounts of DNA that may be an advantage to evade a detrimental immune response following engraftment. The sodium deoxycholate generated ovarian scaffolds had higher donor DNA content, yet, retained the extracellular composition better and may therefore have improved recellularization and other downstream bioengineering applications. These two novel types of mouse ovarian scaffolds serve as promising scaffold-candidates for future ovarian bioengineering experiments.
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| 9. |
- Bagge, Jasmine, et al.
(författare)
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Mucosal Recovery after Intestinal Transplantation in the Rat: A Sequential Histological and Molecular Assessment
- 2023
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Ingår i: European Surgical Research. - : S. Karger AG. - 0014-312X .- 1421-9921. ; 64:2, s. 201-210
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Tidskriftsartikel (refereegranskat)abstract
- Introduction Intestinal cold ischemia and subsequent reperfusion during transplantation result in various degrees of mucosal injury ranging from mild edema to extensive mucosal loss. Mucosal barrier impairment favours bacterial translocation and fluid loss and raises nutritional challenges. The injured intestine also releases proinflammatory mediators and upregulates various epitopes towards an inflammatory phenotype. We studied the process of mucosal injury and repair during the early period after intestinal transplantation from a histological and molecular standpoint.Materials and Methods Adult Sprague Dawley rats were used as donors and recipients. Donor intestines were perfused and stored in saline for 3 hours, then transplanted heterotopically using microvascular anastomoses. Intestinal graft segments were obtained after 20 minutes, 6 hours, 12 hours, and 24 hours after reperfusion. Histology studies (goblet cell count, morphometry), immunofluorescence and western blot for several tight junction proteins, apoptosis and inflammation related proteins were performed.Results Cold storage led to extensive epithelial detachment, whereas reperfusion resulted in extensive villus loss (about 60 % of the initial length) and goblet cell numbers were drastically reduced. Over the first 24 hours, gradual morphologic and molecular recovery was noted, although several molecular alterations persisted (increased apoptosis and inflammation, altered expression of several tight junctions).Conclusions The current data suggest that a near-complete morphologic recovery from a moderate mucosal injury occurs within the first 24 hours after intestinal transplantation. However, several molecular alterations persist and need to be considered when designing intestinal transplant experiments and choosing sampling and endpoints.
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| 10. |
- Casselbrant, Anna, 1970, et al.
(författare)
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Luminal Polyethylene Glycol Alleviates Intestinal Preservation Injury Irrespective of Molecular Size
- 2018
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Ingår i: Journal of Pharmacology and Experimental Therapeutics. - : American Society for Pharmacology & Experimental Therapeutics (ASPET). - 0022-3565 .- 1521-0103. ; 366:1, s. 29-36
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Tidskriftsartikel (refereegranskat)abstract
- Intestinal preservation injury (IPI) and the resulting mucosa injury raise several serious challenges early after intestinal transplantation. The current clinical approach using only vascular perfusion allows the shortest preservation period among the abdominal organs. The experimental addition of luminal polyethylene glycol (PEG) solutions has been repeatedly suggested to alleviate preservation injury, improve graft quality, and prolong the preservation time. We investigated whether the molecular mass of PEG in solution influences the development of intestinal preservation injury. Small intestines of Sprague-Dawley rats were perfused with University of Wisconsin solution. Group 1 underwent vascular perfusion only (clinical control), group 2 received additional luminal PEG3350 Da, group 3 received luminal PEG10000 Da, and group 4 received luminal PEG20000 Da (n = 8/group). Tissue samples were obtained after 4, 8, and 14 hours. We studied the tissue damage (Chiu/Park score, Goblet cells, apoptosis, tight junctions), activation of c-Jun NH2-terminal kinase (JNK), and p38-mitogen-activated protein kinase (MAPK), and we performed Ussing chamber assessments. Mucosal morphologic and electrophysiologic parameters were significantly improved in the groups receiving luminal PEG. There was significantly less apoptotic activity in groups 2, 3, and 4. Both MAPKs revealed an activation peak after 4 hours with group 3 showing lesser p38-MAPK activation. PEG 20 kDa interfered with protein immunodetection. The results indicate that luminal solutions of PEG of medium and large molecular mass significantly delay the onset and development of IPI, providing further evidence that luminal interventions may allow for longer cold storage intervals of intestinal grafts.
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| 11. |
- Frances-Herrero, E., et al.
(författare)
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Bioengineering trends in female reproduction: a systematic review
- 2022
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Ingår i: Human Reproduction Update. - : Oxford University Press (OUP). - 1355-4786 .- 1460-2369. ; 28:6, s. 798-837
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND To provide the optimal milieu for implantation and fetal development, the female reproductive system must orchestrate uterine dynamics with the appropriate hormones produced by the ovaries. Mature oocytes may be fertilized in the fallopian tubes, and the resulting zygote is transported toward the uterus, where it can implant and continue developing. The cervix acts as a physical barrier to protect the fetus throughout pregnancy, and the vagina acts as a birth canal (involving uterine and cervix mechanisms) and facilitates copulation. Fertility can be compromised by pathologies that affect any of these organs or processes, and therefore, being able to accurately model them or restore their function is of paramount importance in applied and translational research. However, innate differences in human and animal model reproductive tracts, and the static nature of 2D cell/tissue culture techniques, necessitate continued research and development of dynamic and more complex in vitro platforms, ex vivo approaches and in vivo therapies to study and support reproductive biology. To meet this need, bioengineering is propelling the research on female reproduction into a new dimension through a wide range of potential applications and preclinical models, and the burgeoning number and variety of studies makes for a rapidly changing state of the field. OBJECTIVE AND RATIONALE This review aims to summarize the mounting evidence on bioengineering strategies, platforms and therapies currently available and under development in the context of female reproductive medicine, in order to further understand female reproductive biology and provide new options for fertility restoration. Specifically, techniques used in, or for, the uterus (endometrium and myometrium), ovary, fallopian tubes, cervix and vagina will be discussed. SEARCH METHODS A systematic search of full-text articles available in PubMed and Embase databases was conducted to identify relevant studies published between January 2000 and September 2021. The search terms included: bioengineering, reproduction, artificial, biomaterial, microfluidic, bioprinting, organoid, hydrogel, scaffold, uterus, endometrium, ovary, fallopian tubes, oviduct, cervix, vagina, endometriosis, adenomyosis, uterine fibroids, chlamydia, Asherman's syndrome, intrauterine adhesions, uterine polyps, polycystic ovary syndrome and primary ovarian insufficiency. Additional studies were identified by manually searching the references of the selected articles and of complementary reviews. Eligibility criteria included original, rigorous and accessible peer-reviewed work, published in English, on female reproductive bioengineering techniques in preclinical (in vitro/in vivo/ex vivo) and/or clinical testing phases. OUTCOMES Out of the 10 390 records identified, 312 studies were included for systematic review. Owing to inconsistencies in the study measurements and designs, the findings were assessed qualitatively rather than by meta-analysis. Hydrogels and scaffolds were commonly applied in various bioengineering-related studies of the female reproductive tract. Emerging technologies, such as organoids and bioprinting, offered personalized diagnoses and alternative treatment options, respectively. Promising microfluidic systems combining various bioengineering approaches have also shown translational value. WIDER IMPLICATIONS The complexity of the molecular, endocrine and tissue-level interactions regulating female reproduction present challenges for bioengineering approaches to replace female reproductive organs. However, interdisciplinary work is providing valuable insight into the physicochemical properties necessary for reproductive biological processes to occur. Defining the landscape of reproductive bioengineering technologies currently available and under development for women can provide alternative models for toxicology/drug testing, ex vivo fertility options, clinical therapies and a basis for future organ regeneration studies.
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| 12. |
- Hellström, Mats, 1976, et al.
(författare)
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Bioengineered uterine tissue supports pregnancy in a rat model
- 2016
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Ingår i: Fertility and Sterility. - : Elsevier BV. - 0015-0282. ; 106:2
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Tidskriftsartikel (refereegranskat)abstract
- Objective: To create a bioengineered uterine patch for uterine repair of a partially defect uterus. Design: Three different decellularized uterine scaffolds were recellularized in vitro with primary uterine cells and green fluorescent protein-(GPF-) labeled bone marrow-derived mesenchymal stem cells (GFP-MSCs). The patches were transplanted in vivo to investigate their tissue adaptation and supporting capacity during pregnancy. Animal(s): Female Lewis rats (n = 9) as donors to generate whole-uterus scaffolds using three different protocols (n = 3 per protocol); Sprague Dawley rats (n = 40) for primary uterus cell isolation procedures (n = 10) and for transplantation/pregnancy studies (n = 30); and male Sprague Dawley rats (n = 12) for mating. Intervention(s): Decellularization was achieved by whole-uterus perfusion with buffered or nonbuffered Triton-X100 and dimethyl sulfoxide (DMSO; group P1/P2) or with sodium deoxycholate (group P3). Primary uterine cells and GFP-MSCs were used to develop uterine tissue constructs, which were grafted to uteri with partial tissue defects. Main Outcome Measure(s): Recellularization efficiency and graft quality were analyzed morphologically, immunohistochemically, and by real-time quantitative polymerase chain reaction (PCR). The location and number of fetuses were documented during pregnancy days 16-20. Result(s): Pregnancy and fetal development were normal in groups P1 and P2, with fetal development over patched areas. Group P3 showed significant reduction of fetal numbers, and embryos were not seen in the grafted area. Quantitative PCR and immunohistochemistry revealed uterus-like tissue in the patches, which had been further reconstructed by infiltrating host cells after transplantation. Conclusion(s): Primary uterine cells and MSCs can be used to reconstruct decellularized uterine tissue. The bioengineered patches made from triton-X100+DMSO-generated scaffolds were supportive during pregnancy. These protocols should be explored further to develop suitable grafting material to repair partially defect uteri and possibly to create a complete bioengineered uterus. ((C) 2016 by American Society for Reproductive Medicine.)
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| 13. |
- Hellström, Mats, 1976, et al.
(författare)
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The Bioengineered Uterus – a possible future
- 2019
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Ingår i: Uterus Transplantation. Brännsträm, M. (red.). - Cham : Springer. - 9783319941615 ; , s. 219-230
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Bokkapitel (refereegranskat)abstract
- © Springer Nature Switzerland AG 2020. This book discusses all aspects of the uterus transplantation procedure in women who are childless because of a dysfunctional or missing uterus, from initial pre-operative investigations to follow-up after the uterine graft has been removed. All the international experts who are engaged in clinical activities and/or research in this field have contributed one or more of the book’s chapters. Since the first live birth following uterus transplantation was reported in Sweden in September 2014, the field has been rapidly evolving around the globe: the first cases in USA, Germany, China, Brazil, and Czech Republic were performed in 2016 and several other centres are preparing for clinical trials of uterus transplantation. Uterus transplantation is a subject that attracts the attention of many bioethicists, and like all other major procedures in transplantation surgery and assisted reproduction techniques, it is a complex process that requires a multidisciplinary team if it is to be successful. As such the book is a valuable resource not only for clinicians from many disciplines, but also for nurses, ethicists, psychologists and researchers.
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| 14. |
- Hellström, Mats, 1976, et al.
(författare)
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Uterine Tissue Engineering and the Future of Uterus Transplantation.
- 2017
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Ingår i: Annals of biomedical engineering. - : Springer Science and Business Media LLC. - 1573-9686 .- 0090-6964. ; 45:7, s. 1718-1730
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Tidskriftsartikel (refereegranskat)abstract
- The recent successful births following live donor uterus transplantation are proof-of-concept that absolute uterine factor infertility is a treatable condition which affects several hundred thousand infertile women world-wide due to a dysfunctional uterus. This strategy also provides an alternative to gestational surrogate motherhood which is not practiced in most countries due to ethical, religious or legal reasons. The live donor surgery involved in uterus transplantation takes more than 10h and is then followed by years of immunosuppressive medication to prevent uterine rejection. Immunosuppression is associated with significant adverse side effects, including nephrotoxicity, increased risk of serious infections, and diabetes. Thus, the development of alternative approaches to treat absolute uterine factor infertility would be desirable. This review discusses tissue engineering principles in general, but also details strategies on how to create a bioengineered uterus that could be used for transplantation, without risky donor surgery and any need for immunosuppression. We discuss scaffolds derived from decellularized organs/tissues which may be recellularized using various types of autologous somatic/stem cells, in particular for uterine tissue engineering. It further highlights the hurdles that lay ahead in developing an alternative to an allogeneic source for uterus transplantation.
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| 15. |
- Oltean, Mihai, 1976, et al.
(författare)
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A sequential assessment of the preservation injury in porcine intestines
- 2017
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Ingår i: Journal of Surgical Research. - : Elsevier BV. - 0022-4804. ; 216, s. 149-157
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Tidskriftsartikel (refereegranskat)abstract
- Background: Clinical and experimental evidence strongly suggest that ischemia-reperfusion injury after intestinal transplantation has deleterious short-and long-term effects and finding means to reduce ischemia-reperfusion injury is a major research area. The anatomical and physiological similarities between the human and porcine digestive tract favor its use as a preclinical model for translational research. Intriguingly, no systematic appraisal of the development of the intestinal preservation injury in pigs is available. Materials and methods: Intestinal procurement was performed in nine pigs using histidine-tryptophan-ketoglutarate solution as preservation fluid. Ileal biopsies were obtained after 8, 14, and 24 h of static cold storage (SCS), and the preservation injury was assessed morphologically (Chiu score) as well as on the molecular level. Tight junction (zonula occludens, claudin-3 and 4, tricellulin, and occludin) and adherens junctions (E-cadherin) proteins were studied using immunofluorescence and Western blot. Results: Eight hours of SCS induced minimal mucosal changes (Chiu grade 1) that advanced to significant subepithelial edema (Chiu grade 3) after 24 h; progressive Goblet cell depletion was also noted. Apoptosis (studied by cleaved caspase-3 staining significantly increased after 24 h of SCS. Significant molecular changes with decreasing expression of zonula occludens, tricellulin, and occludin were evident already after 8 h of SCS and continuously worsened. Claudin-3 and Claudin-4 and E-cadherin expression remained relatively unaltered during SCS. Conclusions: Important molecular alterations precede histologic changes during SCS of the porcine intestine and may be used as more sensitive injury markers than histologic changes in intestinal ischemia and transplantation. (C) 2017 Elsevier Inc. All rights reserved.
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| 16. |
- Oltean, Mihai, 1976, et al.
(författare)
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Luminal solutions protect mucosal barrier during extended preservation.
- 2015
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Ingår i: The Journal of surgical research. - : Elsevier BV. - 1095-8673 .- 0022-4804. ; 194:1, s. 289-96
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Tidskriftsartikel (refereegranskat)abstract
- Mucosal barrier injury during intestinal preservation (IP) and transplantation favors life-threatening infections. Luminal delivery of solutions containing amino acids or polyethylene glycols (PEGs) may improve preservation results and reduce this injury. We tested if solutions containing glutamine and PEG influence the mucosal injury.Rat intestines were perfused and stored in Viaspan-University of Wisconsin solution. Before IP, a PEG 3350 solution was introduced intraluminally alone (group 1) or supplemented with 40 mmol/L L-glutamine (group 2). Controls underwent vascular flush alone (group 3). Preservation injury was evaluated after 8, 14, and 24 h by histology and goblet cell count. Tight-junction proteins zonula occludens-1, claudin-3, claudin-4, and caveolin-1 were studied by immunofluorescence. Maltase and caspase-3 activity were also analyzed.Group 1 showed mild edema at 8 h and mucosal disruption by 24 h; these features were greatly improved in group 2 where continuous mucosa was found after 24 h of IP. Intestines in group 3 did worse at all time points with subepithelial edema (Park/Chiu grade 3) and marked goblet cell depletion; caspase-3 activity was lowest in group 2. Tight-junction proteins varied continuously during IP; zonula occludens-1 expression and colocalization with claudins decreased significantly in group 3 but not in other groups. Claudin-3 was distinctly localized in the membrane, but stained diffuse, cytoplasmic at later time-points. Claudin-4 changed to a cytoplasmic granular pattern. No caveolin-1 colocalization was observed.Luminal PEG and glutamine delay epithelial breakdown and preserve several important mucosal features during extended IP.
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| 17. |
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| 18. |
- Oltean, Mihai, 1976, et al.
(författare)
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Optimal Solution Volume for Luminal Preservation: A Preclinical Study in Porcine Intestinal Preservation
- 2016
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Ingår i: Transplantation Proceedings. - : Elsevier BV. - 0041-1345. ; 48:2, s. 532-535
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Tidskriftsartikel (refereegranskat)abstract
- Background. Rodent studies suggest that luminal solutions alleviate the mucosal injury and prolong intestinal preservation but concerns exist that excessive volumes of luminal fluid may promote tissue edema. Differences in size, structure, and metabolism between rats and humans require studies in large animals before clinical use. Methods. Intestinal procurement was performed in 7 pigs. After perfusion with histidine-tryptophan-ketoglutarate (HTK), 40-cm-long segments were cut and filled with 13.5% polyethylene glycol (PEG) 3350 solution as follows: VO (controls, none), V1 (0.5 mL/cm), V2 (1 mL/cm), V3 (1.5 mL/cm), and V4 (2 mL/cm). Tissue and luminal solutions were sampled after 8, 14, and 24 hours of cold storage (CS). Preservation injury (Chiu score), the apical membrane (Z0-1, brush-border maltase activity), and the electrolyte content in the luminal solution were studied. Results. In control intestines, 8-hour CS in HTK solution resulted in minimal mucosal changes (grade 1) that progressed to significant subepithelial edema (grade 3) by 24 hours. During this time, a gradual loss in ZO-1 was recorded, whereas maltase activity remained unaltered. Moreover, variable degrees of submucosal edema were observed. Luminal introduction of high volumes (2 mUmL) of PEG solution accelerated the development of the subepithelial edema and submucosal edema, leading to worse histology. However, ZO-1 was preserved better over time than in control intestines (no luminal solution). Maltase activity was reduced in intestines receiving luminal preservation. Luminal sodium content decreased in time and did not differ between groups. Conclusions. This PEG solution protects the apical membrane and the tight-junction proteins but may favor water absorption and tissue (submucosal) edema, and luminal volumes >2 mL/cm may result in worse intestinal morphology.
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| 19. |
- Oltean, Mihai, 1976, et al.
(författare)
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The microsurgical training programme in Gothenburg, Sweden: early experiences
- 2017
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Ingår i: Journal of Plastic Surgery and Hand Surgery. - : Medical Journals Sweden AB. - 2000-656X .- 2000-6764. ; 51:3, s. 193-198
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Microsurgical techniques are increasingly used in routine surgical practice as well as in biomedical research. The training opportunities at standardised training courses are limited, and no microsurgical training facility or programme existed in Scandinavia before 2013. Methods: A microsurgery laboratory was set up and two different courses were started, aiming separately at biomedical researchers and surgeons. The course for biomedical researchers teaches basic microsurgical skills such as vessel isolation, cannulation, and arterial microvascular suture under magnification. The more advanced course for surgeons focuses on various techniques of microvascular and nerve anastomosis. Both courses use a combination of theory and practice, with emphasis on the practical part, the course for surgeons also includes clinically relevant information. Results: Twelve 5-day courses using both non-living models and exercises on laboratory animals have been conducted and attended by 49 researchers and 44 surgeons. The organisation and the programme of the training courses as well as 'The 4E concept' behind the course (educational curriculum, equipment, ergonomy, and evaluation) are further detailed. Conclusions: We have successfully established the first training laboratory and series of microsurgical training courses in Scandinavia at two different levels. The experience from the first 12 courses shows the need for this type of structured training, and confirms that the microsurgical education curriculums needs to be adapted to participants' prerequisites and expectations, and various difficulty levels should be considered.
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| 20. |
- Oltean, Mihai, 1976, et al.
(författare)
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The proteomic signature of intestinal acute rejection in the mouse
- 2022
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Ingår i: Metabolites. - : MDPI AG. - 2218-1989. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Intestinal acute rejection (AR) lacks a reliable non-invasive biomarker and AR surveillance is conducted through frequent endoscopic biopsies. Although citrulline and calprotectin have been suggested as AR biomarkers, these have limited clinical value. Using a mouse model of intestinal transplantation (ITx), we performed a proteome-wide analysis and investigated rejection-related proteome changes that may eventually be used as biomarkers. ITx was performed in allogenic (Balb/C to C57Bl) and syngeneic (C57Bl) combinations. Graft samples were obtained three and six days after transplantation (n = 4/time point) and quantitative proteomic analysis with iTRAQ-labeling and mass spectrometry of whole tissue homogenates was performed. Histology showed moderate AR in all allografts post-transplantation at day six. Nine hundred and thirty-eight proteins with at least three unique peptides were identified in the intestinal grafts. Eighty-six proteins varying by >20% between time points and/or groups had an alteration pattern unique to the rejecting allografts: thirty-seven proteins and enzymes (including S100-A8 and IDO-1) were significantly upregulated whereas forty-nine (among other chromogranin, ornithine aminotransferase, and arginase) were downregulated. Numerous proteins showed altered expression during intestinal AR, several of which were previously identified to be involved in acute rejection, although our results also identified previously unreported proteome changes. The metabolites and downstream metabolic pathways of some of these proteins and enzymes may become potential biomarkers for intestinal AR.
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| 21. |
- Padma, Arvind M., et al.
(författare)
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Uterus bioengineering as a future alternative to uterus transplantation
- 2022
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Ingår i: Clinical and Experimental Obstetrics & Gynecology. - : IMR Press. - 0390-6663. ; 49:3
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Tidskriftsartikel (refereegranskat)abstract
- Objective: To review the current knowledge on uterus bioengineering and discuss potential future directives. Uterus bioengineering may solve two major hurdles in organ transplantation of a uterus, organ shortage and control of rejection by immunosuppression. Mechanism: Literature search using PubMed. Findings in brief: Sixty-seven references were summarized that describe the scientific progress made on uterus bioengineering, including other studies related to the topic. Most articles describe work on rat models, including proof of-concept that uterus bioengineering can be used to restore fertility after a partial uterine injury. These promising results are currently being translated to larger and more clinically relevant animal models. In particular, uterus-specific scaffolds produced by a process called "decellularization" that were developed for the mouse, rat, rabbit, pig, goat, and sheep. These scaffolds stimulated angiogenesis and regeneration in vitro and in vivo, and successfully harbored various types of cells for an extended time in vitro. Additionally, applications for endometrial extracellular matrix-specific hydrogels derived from decellularized uterus tissue is discussed. Current challenges for uterus bioengineering are also addressed, e.g., the cellular reconstruction phase, and how they might be improved. Conclusions: Significant progress was made during the last decade with convincing evidence from multiple independent groups in experiments with small animal models. Initial steps towards large animal uterus bioengineering were made. The future continuation of such studies will provide important data required to translate these ideas to an experimental phase in the human. Partial uterus reconstruction through a bioengineered tissue transplantation is closer to a clinical reality compared to whole uterus bioengineering principles aimed to replace a donor in a UTx setting.
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| 22. |
- Söfteland, John M., 1977, et al.
(författare)
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Intestinal Preservation Injury: A Comparison Between Rat, Porcine and Human Intestines.
- 2019
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Ingår i: International journal of molecular sciences. - : MDPI AG. - 1422-0067. ; 20:13
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Tidskriftsartikel (refereegranskat)abstract
- Advanced preservation injury (PI) after intestinal transplantation has deleterious short- and long-term effects and constitutes a major research topic. Logistics and costs favor rodent studies, whereas clinical translation mandates studies in larger animals or using human material. Despite diverging reports, no direct comparison between the development of intestinal PI in rats, pigs, and humans is available. We compared the development of PI in rat, porcine, and human intestines. Intestinal procurement and cold storage (CS) using histidine-tryptophan-ketoglutarate solution was performed in rats, pigs, and humans. Tissue samples were obtained after 8, 14, and 24 h of CS), and PI was assessed morphologically and at the molecular level (cleaved caspase-3, zonula occludens, claudin-3 and 4, tricellulin, occludin, cytokeratin-8) using immunohistochemistry and Western blot. Intestinal PI developed slower in pigs compared to rats and humans. Tissue injury and apoptosis were significantly higher in rats. Tight junction proteins showed quantitative and qualitative changes differing between species. Significant interspecies differences exist between rats, pigs, and humans regarding intestinal PI progression at tissue and molecular levels. These differences should be taken into account both with regards to study design and the interpretation of findings when relating them to the clinical setting.
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| 23. |
- Söfteland, John M., 1977, et al.
(författare)
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Luminal polyethylene-glycol solution delays the onset of preservation injury in the human intestine.
- 2021
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Ingår i: American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons. - : Elsevier BV. - 1600-6143. ; 21:6, s. 2220-2230
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Tidskriftsartikel (refereegranskat)abstract
- The organ damage incurred during the cold storage (CS) of intestinal grafts has short and long-term consequences. Animal studies suggest that additional luminal preservation (LP) with polyethylene-glycol (PEG) may alleviate this damage. This study aims to validate these findings using human intestines. Ileal segments, perfused intravascularly with IGL-1 solution, were procured from 32 multiorgan donors and divided into two parts: one containing a PEG 3350-based solution introduced luminally (LP group) and another one without luminal treatment (control). Sampling was performed after 4h, 8h, 14h, and 24h of CS. Histology was assessed using the Chiu/Park score. Tight junctions (TJ), several inflammatory markers, and transcription factors were examined by immunofluorescence, ddPCR, and Western blot. Tissue water content (edema) was also measured. Apoptotic activity was assessed with caspase 2,3 and 9 assays. LP significantly lowered mucosal injury at all time points. Redistribution of TJ proteins occurred earlier and more severely in the control group. After 24h of CS, LP intestines showed an emerging unfolding protein response. Increased caspase-3 and -9 activity were found in the control group. The current results indicate that luminal PEG is safe and effective in reducing damage to the intestinal epithelium during CS.
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| 24. |
- Söfteland, John M., 1977, et al.
(författare)
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The impact of age and luminal preservation on the development of intestinal preservation injury in rats
- 2020
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Ingår i: Transplantation. - 1534-6080. ; 104:1
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Organs from older donors are believed to withstand ischemia worse than those from younger donors. The effect of age on the development of intestinal preservation injury (IPI) is unclear. METHODS: We compared the development of IPI in intestines from young (3 mo), adult (14 mo), and old (20 mo) rat donors and assessed if luminal preservation (LP) is effective in delaying IPI. Small intestines were perfused with, and stored in, preservation solution (Custodiol) with or without LP solution (polyethylene glycol 3350). IPI was studied using histology (Chiu score, Alcian blue staining), Western blot, and electrophysiological assessment (Ussing chamber) at 4, 8, and 14 hours. RESULTS: Intestines of old rats did not show major histological alterations, whereas their aortas and kidneys revealed typical age-related changes (arteriosclerosis and glomerulosclerosis). Intestines from old rats fared similarly to their younger counterparts at all time points regarding preservation injury and goblet cells count. Intestines undergoing LP showed fewer histological signs of damage and higher goblet cells count when compared with samples without LP, regardless of donor age. Ussing chamber experiments indicated a time-dependent deterioration of all parameters studied, which was delayed by the use of LP. CONCLUSIONS: Older intestines did not convincingly demonstrate a faster IPI compared with intestines from adult and young donors. The small differences between the age groups were nullified by the use of LP. LP significantly delayed the IPI in all age groups and may allow for longer preservation periods without an increased risk of mucosal damage.
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| 25. |
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| 26. |
- Ericson, Jenny, 1976-
(författare)
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Breastfeeding in mothers of preterm infants : Prevalence and effects of support
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The overall aim of this thesis was to describe the prevalence of breastfeeding in preterm infants and to evaluate the effectiveness and mother’s experiences of proactive person-centred telephone support after discharge. Furthermore, to describe the duration of breastfeeding and risks of ceasing breastfeeding up to 12 months. The first study, a register study with data from the Swedish Neonatal Quality register (SNQ), included breastfeeding data at discharge from 29 445 preterm infants born from 2004-2013. The results demonstrated that the prevalence of exclusive breastfeeding among preterm infants in Sweden decreased during the study period, especially among extremely preterm infants (<28 weeks). We also performed a multicentre randomised controlled trial (RCT) of 493 breastfeeding mothers of preterm infants discharged from six neonatal units in Sweden. The intervention consisted of a proactive breastfeeding telephone support system in which a breastfeeding support team called the mothers once everyday up to 14 days after discharge. The control group received reactive support; the mothers were invited to call the breastfeeding support team if they wanted to talk or ask any questions (i.e., usual care).The RCT demonstrated that the intervention did not affect exclusive breastfeeding at eight weeks after discharge (primary outcome) or up to 12 months. The proactive support did not affect maternal breastfeeding satisfaction, attachment, quality of life or method of feeding (secondary outcomes). However, parental stress was significantly reduced in mothers in the intervention group. Mothers in the intervention group were significantly more satisfied and involved in the support and felt empowered compared with mothers in the control group, who experienced reactive support as dual. Further findings showed that a lower maternal educational level, partial breastfeeding at discharge and longer stay in the neonatal unit increased the risk of ceasing breastfeeding during the first 12 months of postnatal age. In conclusion, the trend for exclusive breastfeeding at discharge in preterm infants is declining, which necessitates concern. The evaluated intervention of telephone support did not affect breastfeeding, in the short-or long-term. However, maternal stress was reduced and mothers were significantly more satisfied with the proactive support and felt empowered by the support.
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| 27. |
- Hellström, Mats, 1976
(författare)
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Creation of bioengineered reproductive organs: The end of organ transplantation?
- 2018
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Ingår i: Encyclopedia of Reproduction. ; , s. 395-400
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Bokkapitel (refereegranskat)abstract
- © 2018 Elsevier Inc. Bioengineering for reproductive aid may become a clinical reality. For instance, healthy mouse pups were born with oocytes generated from matured follicles grown in bioengineered ovaries; sperm and oocytes generated from reprogramed adult- or embryonic stem cells have successfully been used for preclinical in vitro fertilization studies. Additionally, meshes coated with endometrial stem cells may reduce negative side-effects in prolapse surgery, and uterine bioengineering protocols may someday lead to the generation of a patient-specific donor organ in a uterus transplantation setting. Additional applications are also briefly discussed, together with the current progress and challenges that face reproductive bioengineering research.
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| 28. |
- Hwang, C. J., et al.
(författare)
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Parkin deficiency exacerbate ethanol-induced dopaminergic neurodegeneration by P38 pathway dependent inhibition of autophagy and mitochondrial function
- 2017
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Ingår i: Redox Biology. - : Elsevier BV. - 2213-2317. ; 11, s. 456-468
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Tidskriftsartikel (refereegranskat)abstract
- Parkinson's disease (PD) is a neurodegenerative disease characterized by selective degeneration of dopaminergic neurons in the substantia nigra. Parkin (which encoded by Park2), an E3 ubiquitin ligase, is the most frequently mutated gene that has casually been linked to autosomal recessive early onset familial PD. We tested the effect of Park2 on ethanol-induced dopaminergic neurodegeneration in Park2 knockout (KO) transgenic mice after chronic ethanol feeding. Male Park2 wild type (WT) and KO mice (8 weeks old) were fed on a Lieber-DeCarli diet containing 6.6% ethanol for 2 weeks, and compared their responses. We found that knockout of Park2 exacerbates ethanol-induced behavioral impairment as well as dopamine depletion. In the mechanism study, we found that knockout of Park2 increased reactive oxygen species (ROS) production, mitophagy formation, mitochondrial dysfunction, and expression of pro-apoptotic proteins, but decreased expression of pro-autophagic proteins. Knockout of Park2 also increased ethanol-induced activation of p38 mitogen-activated protein kinase. In addition, ROS production, mitophagy formation, mitochondrial dysfunction, and expression of pro-apoptotic proteins were increased, but expression of pro-autophagic proteins were decreased by a treatment of ethanol (100 mu M) in Park2 siRNA-transfacted PC12 cells (5 mu M). Moreover, the exacerbating effects of Park2 deletion on ethanol-induced ROS generation, mitophagy, mitochondrial dysfunction as well as cell death were reduced by p38 specific inhibitor (SB203580) in in vitro (10 mu M) and in vivo 10 mg/kg). Park2 deficiency exacerbates ethanol-induced dopaminergic neuron damage through p38 kinase dependent inhibition of autophagy and mitochondrial function.
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| 29. |
- Kim, Y. E., et al.
(författare)
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Inhibitory effect of punicalagin on lipopolysaccharide-induced neuroinflammation, oxidative stress and memory impairment via inhibition of nuclear factor-kappaB
- 2017
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Ingår i: Neuropharmacology. - : Elsevier BV. - 0028-3908. ; 117, s. 21-32
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Tidskriftsartikel (refereegranskat)abstract
- Neuroinflammation is significant in the pathogenesis and development of Alzheimer's disease (AD). Previously, we showed lipopolysaccharide (LPS)-induced neuroinflammation caused memory impairment. We investigated the possible preventive effects of punicalagin (PUN), a component of pomegranate, on memory deficiency caused by LPS, along with the fundamental mechanisms. LPS-treated cultured astrocytes and microglial BV -2 cells were investigated for anti-neuroinflammatory effects of PUN. PUN (1.5 mu g/kg) ameliorates LPS (250 mu g/kg daily 7 times) -induced memory impairment as well as prevents the LPS-induced expression of inflammatory proteins. In in vitro study, we also found that PUN (1 mu g/ml) inhibited the LPS-(10, 20 and 50 mu M) induced expression of iNOS and Cox -2 as well as the production of ROS, NO, TNF-alpha and IL-1 beta(PUN also suppress activation of NF-kappa B via inhibition of I kappa B degradation as well as p50 and p65 translocation into the nucleus in LPS treated mouse brain and cultured astrocytes and microglial BV -2 cells. Consistent with the inhibitory effect on neuro inflammation, PUN inhibited LPS-induced A beta(1-42) generation through down-regulation of APP and BACE1 expression in in vivo and in vitro study. Moreover, PUN directly binds to NF-kappa B subunit p50 evidenced by a docking model and pull down assay. These results suggest that PUN inhibits LPS-induced memory impairment via anti-inflammatory and anti-amylogenic mechanisms through inhibition of NF-kappa B activation. (C) 2017 Elsevier Ltd. All rights reserved.
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| 30. |
- LeVaillant, C. J., et al.
(författare)
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Significant changes in endogenous retinal gene expression assessed 1 year after a single intraocular injection of AAV-CNTF or AAV-BDNF
- 2016
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Ingår i: Molecular Therapy - Methods and Clinical Development. - : Elsevier BV. - 2329-0501. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Use of viral vectors to deliver therapeutic genes to the central nervous system holds promise for the treatment of neurodegenerative diseases and neurotrauma. Adeno-associated viral (AAV) vectors encoding brain-derived neurotrophic factor (BDNF) or ciliary derived neurotrophic factor (CNTF) promote the viability and regeneration of injured adult rat retinal ganglion cells. However, these growth-inducing transgenes are driven by a constitutively active promoter, thus we examined whether long-term AAV-mediated secretion of BDNF or CNTF affected endogenous retinal gene expression. One year after the intravitreal injection of AAV-green fluorescent protein (GFP), bi-cistronic AAV-BDNF-GFP or AAV-CNTF-GFP, mRNA was extracted and analyzed using custom 96 well polymerase chain reaction arrays. Of 93 test genes, 56% showed significantly altered expression in AAV-BDNF-GFP and/or AAV-CNTF-GFP retinas compared with AAV-GFP controls. Of these genes, 73% showed differential expression in AAV-BDNF versus AAV-CNTF injected eyes. To focus on retinal ganglion cell changes, quantitative polymerase chain reaction was undertaken on mRNA (16 genes) obtained from fixed retinal sections in which the ganglion cell layer was enriched. The sign and extent of fold changes in ganglion cell layer gene expression differed markedly from whole retinal samples. Sustained and global alteration in endogenous mRNA expression after gene therapy should be factored into any interpretation of experimental/clinical outcomes, particularly when introducing factors into the central nervous system that require secretion to evoke functionality. © 2016 Official journal of the American Society of Gene & Cell Therapy
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| 31. |
- Naeimi Kararoudi, Meisam, et al.
(författare)
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Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 Gene Editing Technique in Xenotransplantation.
- 2018
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Ingår i: Frontiers in immunology. - : Frontiers Media SA. - 1664-3224. ; 9
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Forskningsöversikt (refereegranskat)abstract
- Genetically modified pigs have been considered favorable resources in xenotransplantation. Microinjection of randomly integrating transgenes into zygotes, somatic cell nuclear transfer, homologous recombination, zinc finger nucleases, transcription activator-like effector nucleases, and most recently, clustered regularly interspaced short palindromic repeats-cas9 (CRISPR/Cas9) are the techniques that have been used to generate these animals. Here, we provide an overview of the CRISPR approaches that have been used to modify genes which are vital in improving xenograft survival rate, including cytidine monophosphate-N-acetylneuraminic acid hydroxylase, B1,4N-acetylgalactosaminyltransferase, isoglobotrihexosylceramide synthase, class I MHC, von Willebrand factor, C3, and porcine endogenous retroviruses. In addition, we will mention the importance of potential candidate genes which could be targeted using CRISPR/Cas9.
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| 32. |
- Nyman, Ulf, et al.
(författare)
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Revised Swedish guidelines on intravenous iodine contrast medium-induced acute kidney injury 2022: A summary
- 2023
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Ingår i: Acta Radiologica. - : SAGE Publications. - 0284-1851 .- 1600-0455. ; 64:5
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Forskningsöversikt (refereegranskat)abstract
- The Swedish Society of Uroradiology has revised their computed tomography (CT) guidelines regarding iodine contrast media-induced acute kidney injury (CI-AKI). They are more cautious compared to the European Society of Urogenital Radiology and the American College of Radiology since the actual risk of CI-AKI remains uncertain in patients with moderate to severe kidney damage due to a lack of prospective controlled studies and mainly based on retrospective propensity score-matched studies with low-grade evidence. Another source of uncertainty is the imprecision of glomerular filtration rate (GFR) estimating equations. However, randomized hydration studies indictae an upper limit risk of CI-AKI of about 5% for outpatients with a GFR in the range of 30-44 or 45-59 mL/min/1.73m(2) combined with multiple risk factors. Apart from GFR limits, the guideline also includes limits for systemic contrast medium exposure expressed in gram-iodine/GFR ratio.
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| 33. |
- Padma, Arvind M., et al.
(författare)
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Protocols for Rat Uterus Isolation and Decellularization: Applications for Uterus Tissue Engineering and 3D Cell Culturing.
- 2018
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Ingår i: Decellularized Scaffolds and Organogenesis. Kursad Turksen (red.). - New York, NY : Springer. - 1940-6029. - 9781493976553 ; , s. 161-175
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Bokkapitel (refereegranskat)abstract
- Sophisticated culturing conditions are required to grow cells in a three-dimensional (3D) environment. Cells then require a type of scaffold rich in proteins, growth factors, and signaling molecules that simulates their natural environment. Tissues from all species of animals have an organ-specific extracellular matrix (ECM) structure that plays a key role in cell proliferation and migration. Hence, the scaffold composition plays a significant role for any successful 3D cell culturing system. We developed a whole rat uterus ECM scaffold by the perfusion of detergents and ionic solutions through the vascular system of an isolated normal rat uterus in a process termed "decellularization." The generated rat uterus scaffolds consist of a cell-free ECM structure similar to that of the normal rat uterus, and are thus excellent platforms on to which new cells can be added. Rat uterus 3D cell culturing systems based on these scaffolds could become valuable to decidual differentiation- and embryo implantation studies, or for investigating invasion mechanisms of endometrial cancer cells. They could also be used for the creation of tissue engineered uterine tissue, for partial or whole organogenesis developed for transplantation applications to treat absolute uterine infertility. This is a condition affecting about 1 in 500 women, and is only treatable by a uterus transplantation. This article provides valuable troubleshooting notes and describes in detail how to generate rat uterus scaffolds, including the delicate surgery required to isolate the uterus with an intact vascular tree which facilitates vascular perfusion and re-transplantation.
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| 34. |
- Park, Ju Ho, et al.
(författare)
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Anti-Inflammatory Effect of Titrated Extract of Centella asiatica in Phthalic Anhydride-Induced Allergic Dermatitis Animal Model.
- 2017
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Ingår i: International journal of molecular sciences. - : MDPI AG. - 1422-0067. ; 18:4
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Tidskriftsartikel (refereegranskat)abstract
- Centella asiatica has potent antioxidant and anti-inflammatory properties. However, its anti-dermatitic effect has not yet been reported. In this study, we investigated the anti-dermatitic effects of titrated extract of Centella asiatica (TECA) in a phthalic anhydride (PA)-induced atopic dermatitis (AD) animal model as well as in vitro model. An AD-like lesion was induced by the topical application of five percent PA to the dorsal skin or ear of Hos:HR-1 mouse. After AD induction, 100 μL of 0.2% and 0.4% of TECA (40 μg or 80 μg/cm²) was spread on the dorsum of the ear or back skin three times a week for four weeks. We evaluated dermatitis severity, histopathological changes and changes in protein expression by Western blotting for inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and NF-κB activity, which were determined by electromobility shift assay (EMSA). We also measured TNF-α, IL-1β, IL-6, and IgE concentration in the blood of AD mice by enzyme-linked immunosorbent assay (ELISA). TECA treatment attenuated the development of PA-induced atopic dermatitis. Histological analysis showed that TECA inhibited hyperkeratosis, mast cells and infiltration of inflammatory cells. TECA treatment inhibited expression of iNOS and COX-2, and NF-κB activity as well as the release of TNF-α, IL-1β, IL-6, and IgE. In addition, TECA (1, 2, 5 μg/mL) potently inhibited Lipopolysaccharide (LPS) (1 μg/mL)-induced NO production, expression of iNOS and COX-2, and NF-κB DNA binding activities in RAW264.7 macrophage cells. Our data demonstrated that TECA could be a promising agent for AD by inhibition of NF-κB signaling.
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| 35. |
- Simsa, Robin, et al.
(författare)
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Systematic in vitro comparison of decellularization protocols for blood vessels.
- 2018
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Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 13:12
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Tidskriftsartikel (refereegranskat)abstract
- Decellularization of native blood vessels is a promising technology to generate 3D biological scaffolds for vascular grafting. Blood vessel decellularization has been performed in previous studies under various experimental conditions, that complicates comparison and optimization of suitable protocols. The goal of this work was to systematically compare the decellularization and recellularization efficacy of 5 different protocols utilizing the detergents sodium dodecyl sulfate (SDS), sodium deoxycholate (SDC), CHAPS and TritonX-100 together with DNA-removing enzymes on porcine vena cava in a perfusion bioreactor setup. Additionally, we tested the effect of DNase on the extracellular matrix (ECM) properties. We found that all protocols could efficiently decellularize blood vessels. Mechanical strength, collagen preservation and ECM integrity were similar among all tested detergents, yet TritonX protocols required long-term DNase application for complete decellularization. However, TritonX-based protocols showed the greatest recellularization efficacy with HUVECs in vitro. Furthermore, we developed a novel protocol for TritonX which improved recellularization and reduced total process time and ECM stiffness compared to previous protocols. SDS, SDC and CHAPS based protocols had a lower recellularization potential. In conclusion, decellularization of blood vessels can be achieved with all tested reagents, but TritonX treated ECM can be most efficiently recellularized with endothelial cells.
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| 36. |
- Tsiartas, Panos, et al.
(författare)
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P–459 Ex vivo perfusion of whole ewe ovaries with follicular maturation for up to seven days: towards the development of an alternative fertility preservation method
- 2021
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Ingår i: Human Reproduction Vol 36 Issue Supplement 1. - : Oxford University Press (OUP). - 0268-1161.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Abstract Study question To develop an alternative fertility preservation method for young female cancer patients based on an ex vivo perfusion of whole ovaries serving as a platform for future ovarian stimulation studies. Summary answer It is possible to maintain viable follicles and to retrieve oocytes after ex vivo perfusion of ewe ovaries for up to 7 days. What is known already Some progress has been made in terms of follicular growth and the isolation of mature oocytes in vitro. However, full development, from early follicular stages to a viable offspring, has only been described in rodent models. The complex events controlling follicular expansion and the long time required for folliculogenesis and oocyte maturity in large mammalian species creating challenges and limitations for in vitro studies. Ex vivo perfusion of a whole ovary could potentially be a solution by exploiting the intact ovarian architecture to support folliculogenesis and oocyte maturation. Study design, size, duration Thirty-one ewe ovaries were divided into 4 groups and ex vivo perfused in a bioreactor. Group 1 (n=14) perfusion for 48hours with no hormone supplementation; Group 2 (n=4) perfusion 96–101hours with follicle stimulating hormone (FSH); Group 3 (n=3) perfusion 120–168hours with human menopausal gonadotropin (hMG); Group 4 (n=10) perfusion 72–144hours with hMG. Participants/materials, setting, methods Ewe ovaries from sexually mature ewes were ex vivo perfused in a bioreactor under normothermic conditions for up to 7 days (max total 168hours). Histomorphological, immunohistochemical, hormonal and biochemical analyses were performed to assess ovarian structure and viability after cold ischemia and after perfusion which was subsequently compared to control ovaries. Main results and the role of chance The perfused ovaries in group 2 and 3 showed no significant differences in follicular density, viability and oocyte quality after ischemia and perfusion compared to control ovaries. Estradiol and progesterone levels did not increase during the perfusion. The perfused ovaries in group 1 and 4 showed a significant decrease in the ovarian reserve and oocyte quality. In total, 16 GV-MI oocytes were retrieved from groups 3 and 4. Limitations, reasons for caution 1. Ovaries were retrieved from ewes of unknown cycle and reproductive history. 2. The perfusion medium was changed after 24hours from perfusion start to remove detrimental metabolites and this could affect the measured concentrations of hormones and metabolites in the perfusion medium. Wider implications of the findings: These results pave the way to propose ex vivo perfusion as a good platform for fertility preservation studies on whole mammalian and human ovaries to retrieve fully mature oocytes. Trial registration number Not applicable
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| 37. |
- Tsiartas, P., et al.
(författare)
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Seven days ex vivo perfusion of whole ewe ovaries with follicular maturation and oocyte retrieval: towards the development of an alternative fertility preservation method
- 2022
-
Ingår i: Reproduction Fertility and Development. - : CSIRO Publishing. - 1031-3613 .- 1448-5990. ; 34:3, s. 331-342
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Tidskriftsartikel (refereegranskat)abstract
- Fertility preservation methods for prepubertal women about to undergo gonadotoxic chemo and/or radiation therapy are limited. Therefore, the aim of this study was to investigate the feasibility to develop an alternative fertility preservation method based on an ex vivo perfusion platform for whole ewe ovaries. Thirteen ewe ovaries were divided into two groups (group 1 and 2) that were perfused in a bioreactor for up to 7 days. Group 1 (n = 3) were stimulated with human menopausal gonadotropin (hMG) administered in single daily dose, while group 2 (n = 10) were stimulated continuously for 24 h. The perfused ovaries in group 1 showed no significant differences in follicular density, sub-follicular morphology and oocyte quality after ischaemia and after ex vivo perfusion compared with non-perfused control ovaries. The perfused ovaries in group 2 showed a significant decrease in the follicular reserve and oocyte quality compared with the control group. In total, 16 GV-MI oocytes were retrieved from both groups. This study describes for the first time the ex vivo maintenance of viable follicles of ewe ovaries with oocyte integrity and the retrieval of oocytes after ex vivo hormonal perfusion with two different protocols for up to 7 days.
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| 38. |
- You, S. W., et al.
(författare)
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Large-scale reconstitution of a retina-to-brain pathway in adult rats using gene therapy and bridging grafts: An anatomical and behavioral analysis
- 2016
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Ingår i: Experimental Neurology. - : Elsevier BV. - 0014-4886. ; 279, s. 197-211
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Tidskriftsartikel (refereegranskat)abstract
- Peripheral nerve (PN) grafts can be used to bridge tissue defects in the CNS. Using a PN-to-optic nerve (ON) graft model, we combined gene therapy with pharmacotherapy to promote the long-distance regeneration of injured adult retinal ganglion cells (RGCs). Autologous sciatic nerve was sutured onto the transected ON and the distal end immediately inserted into contralateral superior colliculus (SC). Control rats received intraocular injections of saline or adeno-associated virus (AAV) encoding GFP. In experimental groups, three bi-cistronic AAV vectors encoding ciliary neurotrophic factor (CNTF) were injected into different regions of the grafted eye. Each vector encoded a different fluorescent reporter to assess retinotopic order in the regenerate projection. To encourage sprouting/synaptogenesis, after 6 weeks some AAV-CNTF injected rats received an intravitreal injection of recombinant brain-derived neurotrophic factor (rBDNF) or AAV-BDNF. Four months after surgery, cholera toxin B was used to visualize regenerate RGC axons. RGC viability and axonal regrowth into SC were significantly greater in AAV-CNTF groups. In some cases, near the insertion site, regenerate axonal density resembled retinal terminal densities seen in normal SC. Complex arbors were seen in superficial but not deep SC layers and many terminals were immunopositive for presynaptic proteins vGlut2 and SV2. There was improvement in visual function via the grafted eye with significantly greater pupillary constriction in both AAV-CNTF + BDNF groups. In both control and AAV-CNTF + rBDNF groups the extent of light avoidance correlated with the maximal distance of axonal penetration into superficial SC. Despite the robust regrowth of RGC axons back into the SC, axons originating from different parts of the retina were intermixed at the PN graft/host SC interface, indicating that there remained a lack of order in this extensive regenerate projection. (C) 2016 Elsevier Inc. All rights reserved.
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