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    Effects of vitrification solution supplemented with platelet-rich plasma in rat ovarian tissue cryopreservation
    (Tubitak Scientific & Technological Research Council Turkey, 2023) Gokhan, Aylin; Cavusoglu, Tuerker; Kilic, Kubilay Dogan; Sirin, Cansin; Tomruk, Canberk; Yigitturk, Guerkan; Erbas, Oytun
    Background/aim: The subject of this study was to investigate the utility of platelet-rich plasma (PRP) in the cryopreservation process to reduce cryodamage and increase tissue viability.Materials and methods: Twenty-one female Wistar rats were randomly allocated to three groups. In Group 1 (G1), rats were not subjected to vitrification (n = 7). Group 2 (G2) was the vitrification group in which PRP was added to the basic vitrification solution (n = 7). Group 3 (G3) was the vitrification group in which fetal bovine serum was added to the basic vitrification solution (n = 7). Warmed tissues were evaluated with histochemical (HC) and immunohistochemical (IHC) staining, the TUNEL method, immunofluorescence (IF) staining, and biochemical analyses.Results: The percentages of IHC staining, TUNEL method positivity, and IF staining were significantly higher in G2 compared to both G1 and G3 (P < 0.05). G2 ovaries exhibited a significant increase in both malondialdehyde and catalase values in comparison to G1 (P < 0.05). In HC staining, degenerations in primary and secondary follicles and in ovarian tissue were more common in the PRP-supplemented group. The calcium used in PRP activation was suspected to have increased the degeneration and prevented the possible positive effects of PRP.Conclusion: To the best of our knowledge, PRP-supplemented vitrification solution was used for the first time in the literature in this study in whole rat ovarian tissue vitrification. If PRP is to be used as a component in vitrification solution for rat ovarian tissue, the use of lower amounts of calcium or different methods in PRP activation, or the use of nonactivated PRP, should be considered from the beginning.
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    Investıgatıon of ursodeoxycholıc acıd effects on sırolımus treated adıpose tıssue-derıved mesenchymal stem cells
    (2022) Uyanıkgil, Yiğit; Gündüz, Cumhur; Yiğittürk, Gürkan; Çavuşoğlu, Türker; Gökçe, Burak; None, Senem; Tomruk, Canberk
    Objective The usage of mesenchymal stem cells (MSC) with immunosuppressive drugs after organ transplantation is becoming remarkable in clinical applications. However, the drugs negatively affect MSCs. Ursodeoxycholic acid (UDCA), which is an antioxidant molecule, may reverse these effects. The study aims that to determine the effects of sirolimus and UDCA on human adipose tissue-derived MSCs (ADMSCs) individually and in combination. Material and Method The cytotoxicity of the agents was evaluated by WST-1 test in time and dose-dependent manner. The combinational effects were determined using isobologram analysis. Muse cell analyzer was used for the evaluation of apoptosis and cell cycle. Oxidative stress markers were measured by biochemical methods. Results IC50 dose of sirolimus was determined as 18.58?M in the 48th hour. Because no cytotoxic effect was observed at the studied doses of UDCA, the apoptosis, cell cycle, and oxidative stress indicator analyses were continued with a safe dose of 100 ?M. Sirolimus promoted apoptosis and inhibited cell proliferation. It was determined that UDCA reduced the apoptotic and anti-proliferative effects of sirolimus on ADMSCs with its anti-oxidant property. Conclusion The UDCA treatment in combination with immunosuppressive therapy after organ and tissue transplantation may have positive effects on ADMSCs.
  • Küçük Resim
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    Platelet-Rich Plasma in Vitrification; is it Helpful or Harmful?
    (Univ Agriculture, Fac Veterinary Science, 2023) Cavusoglu, Turker; Gokhan, Aylin; Tomruk, Canberk; Sirin, Cansin; Kilic, Kubilay Dogan; Yigitturk, Guerkan
    Human and animal studies on cryoprotectants and freezing solutions are still needed to establish a simple yet reliable protocol and increase the success of cryopreservation. The main aim of this study was to evaluate the short- and long-term effects of platelet-rich plasma, a well-known antioxidant substance due to its contents including bioactive molecules and growth factors, on whole ovarian tissue cryopreservation. Fresh tissues (control group, G1) were subjected to histological tissue processing without any treatment. Ovaries treated with platelet-rich plasma (PRP)-supplemented vitrification solution were subjected to tissue processing without cryostorage group 2 (G2) or following six months of cryostorage group 3 (G3). Steps in G2 and G3 were also performed for group 4 (G4) and group 5 (G5), respectively, except that the vitrification solution was supplemented with fetal bovine serum. PRP was activated with calcium chloride (CaCl2) after double centrifugation. Ethylene glycol, dimethyl sulfoxide, and sucrose were used as cryoprotective agents in all groups. Histomorphological changes were evaluated with the semi-quantitative histochemical-scoring algorithm. Apoptotic and antiapoptotic effects and intercellular connections were evaluated with immunohistochemical staining of Bax, Bcl-2, Caspase-3 (C3), Connexin-43 (Cx-43), and TUNEL analysis. Cryopreservation with PRP-supplementation (G3) significantly increased tissue degeneration (p<0.05). There was an increase in the number of degenerated both primary and secondary follicles (p<0.05), and an increase in the immune expression of Bax, C3 and Cx-43 and TUNEL assay in G3 was observed compared to other groups (p<0.05). Since the morphology of primordial follicles was more preserved than other follicles in all groups, primordial follicles were not included in the follicle count. Our study suggested that cryopreservation with PRP-supplemented vitrification solution caused excessive damage to rat ovaries. We assumed that CaCl2 might have further provoked this cellular damage.