Dataset Information

Decellularized bovine ovarian niche restored the function of cumulus and endothelial cells.

ABSTRACT:

Objective

Recently, the decellularization technique is introduced as one of the tissue engineering procedures for the treatment of various deficiencies. Here, we aimed to assess the dynamic activity of CCs and HUVECs within decellularized bovine ovarian tissue transplanted subcutaneously in rats. Ovarian tissue was decellularized using a cocktail consisting of different chemicals, and the efficiency of decellularization was assessed using hematoxylin-eosin and DAPI staining. The cell survival was evaluated using an LDH leakage assay. Thereafter, decellularized samples were recellularized using HUVECs and CCs, encapsulated inside alginate (1.2%)-gelatin, (1%) hydrogel, and transplanted subcutaneously to rats. The existence of CD31- and estrogen-positive cells was assessed using immunohistochemistry staining.

Results

Bright-field imaging and DAPI staining revealed the lack of nuclei with naive matrix structure in ovarian tissue subjected to decellularization protocol. SEM imaging revealed a normal matrix in decellularized ovaries. LDH assay showed a lack of cytotoxicity for CCs after 7-days compared to the control group. Immunohistochemistry staining showed both CD31- and estrogen-positive cells in CCs + HUVECs compared to the CCs group. CD31 cells appeared with flattened morphology aligned with matrix fibers. The existence of estrogen and CD31 positive cells showed the efficiency of decellularized ovarian tissue to restore cellular function and activity.

SUBMITTER: Amjadi F

PROVIDER: S-EPMC9644448 | biostudies-literature | 2022 Nov

REPOSITORIES: biostudies-literature

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Decellularized bovine ovarian niche restored the function of cumulus and endothelial cells.

Amjadi Farhad F Beheshti Rahim R Nasimi Fatemeh Sokouti FS Hassani Ayla A Shirazi Reza R Tamadon Amin A Rahbarghazi Reza R Mahdipour Mahdi M

BMC research notes 20221108 1

<h4>Objective</h4>Recently, the decellularization technique is introduced as one of the tissue engineering procedures for the treatment of various deficiencies. Here, we aimed to assess the dynamic activity of CCs and HUVECs within decellularized bovine ovarian tissue transplanted subcutaneously in rats. Ovarian tissue was decellularized using a cocktail consisting of different chemicals, and the efficiency of decellularization was assessed using hematoxylin-eosin and DAPI staining. The cell sur ...[more]

PMID: 36348463

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Project description:BackgroundMany patients suffering from peripheral arterial disease (PAD) are dependent on bypass surgery. However, in some patients no suitable replacements (i.e. autologous or prosthetic bypass grafts) are available. Advances have been made to develop autologous tissue engineered vascular grafts (TEVG) using endothelial colony forming cells (ECFC) obtained by peripheral blood draw in large animal trials. Clinical translation of this technique, however, still requires additional data for usability of isolated ECFC from high cardiovascular risk patients. Bovine carotid arteries (BCA) were decellularized using a combined SDS (sodium dodecyl sulfate) -free mechanical-osmotic-enzymatic-detergent approach to show the feasibility of xenogenous vessel decellularization. Decellularized BCA chips were seeded with human ECFC, isolated from a high cardiovascular risk patient group, suffering from diabetes, hypertension and/or chronic renal failure. ECFC were cultured alone or in coculture with rat or human mesenchymal stromal cells (rMSC/hMSC). Decellularized BCA chips were evaluated for biochemical, histological and mechanical properties. Successful isolation of ECFC and recellularization capabilities were analyzed by histology.ResultsDecellularized BCA showed retained extracellular matrix (ECM) composition and mechanical properties upon cell removal. Isolation of ECFC from the intended target group was successfully performed (80% isolation efficiency). Isolated cells showed a typical ECFC-phenotype. Upon recellularization, co-seeding of patient-isolated ECFC with rMSC/hMSC and further incubation was successful for 14 (n = 9) and 23 (n = 5) days. Reendothelialization (rMSC) and partial reendothelialization (hMSC) was achieved. Seeded cells were CD31 and vWF positive, however, human cells were detectable for up to 14 days in xenogenic cell-culture only. Seeding of ECFC without rMSC was not successful.ConclusionUsing our refined decellularization process we generated easily obtainable TEVG with retained ECM- and mechanical quality, serving as a platform to develop small-diameter (< 6 mm) TEVG. ECFC isolation from the cardiovascular risk target group is possible and sufficient. Survival of diabetic ECFC appears to be highly dependent on perivascular support by rMSC/hMSC under static conditions. ECFC survival was limited to 14 days post seeding.

Project description:BackgroundOocyte maturation and preimplantation embryo development are controlled by array of genes that are post-transcriptionally regulated by microRNAs. With respect to this, previously, we identified altered expression of microRNA-130b (miR-130b) during oocyte maturation. Here, we aimed to investigate the role of miR-130b in bovine granulosa and cumulus cell function, oocyte maturation and preimplantation embryo development using gain- and loss-of- function approach.MethodsFor this study, the granulosa cells, cumulus cells and the oocytes were collected from ovaries obtained from slaughterhouse. The genes targeted by miR-130b were identified using dual-luciferase reporter assay. The role of miR-130b in granulosa and cumulus cell function was investigated by increasing and inhibiting its expression in in vitro cultured cells using miR-130b precursor and inhibitor, respectively while the role of miR-130b on oocyte development, immature oocytes were microinjected with miR-130b precursor and inhibitor and the polar body extrusion, the proportion of oocytes reaching to metaphase II stage and the mitochondrial were determined in each oocyte group 22 h after microinjection. Moreover, to investigate the role of miR-130b during preimplantation embryo development, zygote stage embryos were microinjected with miR-130b precursor or inhibitor and the cleavage rate, morula and blastocyst formation was analyzed in embryos derived from each zygote group after in vitro culture.ResultsThe luciferase assay showed that SMAD5 and MSK1 genes were identified as the direct targets of miR-130b. Overexpression of miR-130b increased the granulosa and cumulus cell proliferation, while inhibition showed the opposite phenotype. Apart from these, modulation of miR-130b altered the lactate production and cholesterol biosynthesis in cumulus cells. Furthermore, inhibition of miR-130b expression during oocyte in vitro maturation reduced the first polar body extrusion, the proportion of oocytes reaching to metaphase II stage and the mitochondrial activity, while inhibition of miR-130b during preimplantation embryo development significantly reduced morula and blastocyst formation.ConclusionThis study demonstrated that in vitro functional modulation of miR-130b affected granulosa and cumulus cell proliferation and survival, oocyte maturation, morula and blastocyst formation suggesting that miR-130b is involved in bovine oocyte maturation and preimplantation embryo development.

Dataset Information

Decellularized bovine ovarian niche restored the function of cumulus and endothelial cells.

Objective

Results

Publications

Decellularized bovine ovarian niche restored the function of cumulus and endothelial cells.

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