Recovering Spermatogenesis By Protected Cryopreservation Using Metformin and Transplanting Spermatogonial Stem Cells Into Testis in an Azoospermia Mouse Model.

IF 1.6 4区生物学

Biopreservation and Biobanking Pub Date : 2024-02-01 Epub Date: 2023-08-15 DOI:10.1089/bio.2022.0178

Alieh Bashghareh, Tayebeh Rastegar, Peyman Modarresi, Shokoofeh Kazemzadeh, Maryam Salem, Azim Hedayatpour

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引用次数: 0

Abstract

Cryopreservation and transplantation of spermatogonial stem cells (SSCs) may serve as a new method to restore male fertility in patients undergoing chemotherapy or radiotherapy. However, SSCs may be damaged during cryopreservation due to the production of reactive oxygen species (ROS). Therefore, different antioxidants have been used as protective agents. Studies have shown that metformin (MET) has antioxidant activity. The aim of this study was to assess the antioxidant and antiapoptotic effects of MET in frozen-thawed SSCs. In addition, the effect of MET on the proliferation and differentiation of SSCs was evaluated. To this end, SSCs were isolated from mouse pups aged 3-6 days old, cultured, identified by flow cytometry (ID4, INTEGRIN β1⁺), and finally evaluated for survival and ROS rate. SSCs were transplanted after busulfan and cadmium treatment. Cryopreserved SSCs with and without MET were transplanted after 1 month of cryopreservation. Eight weeks after transplantation, the recipient testes were evaluated for the expression of apoptosis (BAX, BCL2), proliferation (PLZF), and differentiation (SCP3, TP1, TP2, PRM1) markers using immunohistochemistry, Western blot, and quantitative real-time polymerase chain reaction. The findings revealed that the survival rate of SSCs was higher in the 500 μm/mL MET group compared to the other groups (50 and 5000 μm/mL). MET significantly decreased the intracellular ROS production. Transplantation of SSCs increased the expression level of proliferation (PLZF) and differentiation (SCP3, TP1, TP2, PRM1) markers compared to azoospermia group, and their levels were significantly higher in the MET group compared to the cryopreservation group containing basic freezing medium (p < 0.05). MET increased the survival rate of SSCs, proliferation, and differentiation and decreased the ROS production and the apoptosis rate. Cryopreservation by MET seems to be effective in treating infertility.

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在无精子症小鼠模型中使用二甲双胍进行保护性冷冻保存并将精原干细胞移植到睾丸，从而恢复生精功能

冷冻保存和移植精原干细胞（SSCs）可作为恢复化疗或放疗患者男性生育能力的新方法。然而，由于活性氧（ROS）的产生，精原干细胞可能会在冷冻保存过程中受损。因此，不同的抗氧化剂被用作保护剂。研究表明，二甲双胍（MET）具有抗氧化活性。本研究旨在评估二甲双胍在冷冻解冻的造血干细胞中的抗氧化和抗凋亡作用。此外，还评估了 MET 对造血干细胞增殖和分化的影响。为此，研究人员从 3-6 天大的小鼠幼崽中分离出造血干细胞，对其进行培养、流式细胞术鉴定（ID4、INTEGRIN β1+），最后评估其存活率和 ROS 率。硫丹和镉处理后移植 SSCs。有MET和无MET的冷冻造血干细胞在冷冻1个月后移植。移植八周后，使用免疫组化、Western 印迹和实时定量聚合酶链反应评估受体睾丸的凋亡（BAX、BCL2）、增殖（PLZF）和分化（SCP3、TP1、TP2、PRM1）标志物的表达。研究结果显示，与其他组（50 和 5000 μm/mL MET）相比，500 μm/mL MET 组的 SSCs 存活率更高。MET 能明显减少细胞内 ROS 的产生。与无精子症组相比，移植 SSCs 增加了增殖（PLZF）和分化（SCP3、TP1、TP2、PRM1）标记物的表达水平，与含有基本冷冻培养基的冷冻组相比，MET 组的这些标记物表达水平明显更高（p

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来源期刊

Biopreservation and Biobanking Biochemistry, Genetics and Molecular Biology-General Biochemistry,Genetics and Molecular Biology

自引率

12.50%

发文量

114

期刊介绍： Biopreservation and Biobanking is the first journal to provide a unifying forum for the peer-reviewed communication of recent advances in the emerging and evolving field of biospecimen procurement, processing, preservation and banking, distribution, and use. The Journal publishes a range of original articles focusing on current challenges and problems in biopreservation, and advances in methods to address these issues related to the processing of macromolecules, cells, and tissues for research. In a new section dedicated to Emerging Markets and Technologies, the Journal highlights the emergence of new markets and technologies that are either adopting or disrupting the biobank framework as they imprint on society. The solutions presented here are anticipated to help drive innovation within the biobank community. Biopreservation and Biobanking also explores the ethical, legal, and societal considerations surrounding biobanking and biorepository operation. Ideas and practical solutions relevant to improved quality, efficiency, and sustainability of repositories, and relating to their management, operation and oversight are discussed as well.