Hassani Fatemeh, Ebrahimi Bita, Moini Ashraf, Ghiaseddin Ali, Bazrafkan Mahshid, Hassanzadeh G Holamreza, Valojerdi Mojtaba Rezazadeh
Department of Anatomy, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Department of Embryology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran.
Cell J. 2020 Jan;21(4):479-493. doi: 10.22074/cellj.2020.6393. Epub 2019 Jul 29.
Testing novel biomaterials for the three dimensional (3D) culture of ovarian follicles may ultimately lead to a culture model which can support the integrity of follicles during culture (IVC). The present study reports the first application of a chitosan (CS) hydrogel in culturing mouse preantral follicles.
In this interventional experiment study, CS hydrogels with the concentrations of 0.5, 1, and 1.5% were first tested for fourier transform infrared spectroscopy (FT-IR), Compressive Strength, viscosity, degradation, swelling ratio, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity and live/dead assay. Thereafter, mouse ovarian follicles were encapsulated in optimum concentration of CS (1%) and compared with those in alginate hydrogel. The follicular morphology, quality of matured oocyte and steroid secretion in both CS and alginate were assessed by enzyme-linked immunosorbent assay (ELISA). The expression of folliculogenesis, endocrine, and apoptotic related genes was also evaluated by quantitative real-time polymerase chain reaction (qRT-PCR) and compared with day that in 0.
The rates of survival, and diameter of the follicles, secretion of estradiol, normal appearance of meiotic spindle and chromosome alignment were all higher in CS group compared with those in alginate group (P≤0.05). The expression of and in CS group was significantly higher than that of the alginate group (P≤0.05).
The results showed that CS is a permissive hydrogel and has a beneficial effect on encapsulation of ovarian follicle and its further development during 3D culture.
测试用于卵巢卵泡三维（3D）培养的新型生物材料，最终可能会得到一种能够在培养过程中（体外成熟培养）维持卵泡完整性的培养模型。本研究报道了壳聚糖（CS）水凝胶在培养小鼠腔前卵泡中的首次应用。
在这项干预性实验研究中，首先对浓度为0.5%、1%和1.5%的CS水凝胶进行傅里叶变换红外光谱（FT-IR）、抗压强度、粘度、降解、溶胀率、3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）细胞毒性和活/死检测。此后，将小鼠卵巢卵泡包裹在最佳浓度的CS（1%）中，并与藻酸盐水凝胶中的卵泡进行比较。通过酶联免疫吸附测定（ELISA）评估CS和藻酸盐中卵泡的形态、成熟卵母细胞的质量和类固醇分泌。还通过定量实时聚合酶链反应（qRT-PCR）评估卵泡发生、内分泌和凋亡相关基因的表达，并与第0天的表达进行比较。
与藻酸盐组相比，CS组的卵泡存活率、直径、雌二醇分泌、减数分裂纺锤体的正常外观和染色体排列均更高（P≤0.05）。CS组中[基因名称未给出]和[基因名称未给出]的表达明显高于藻酸盐组（P≤0.05）。
结果表明，CS是一种适宜的水凝胶，对卵巢卵泡的包裹及其在三维培养中的进一步发育具有有益作用。
