Reproductive Medical Department of West China Second University Hospital, Key Laboratory of Birth Defects and Related Diseases of Women and Children, Ministry of Education, Sichuan University, Chengdu 610041, China.
Cryo Letters. 2024 May-Jun;45(3):177-184.
Ovarian tissue cryopreservation for fertility preservation carries a risk of malignant cell re-seeding. Artificial ovary is a promising method to solve such a problem. However, ovary decellularization protocols are limited. Hence, further studies are necessary to get better ovarian decellularization techniques for the construction of artificial ovary scaffolds.
To establish an innovative decellularization technique for whole porcine ovaries by integrating liquid nitrogen with chemical agents to reduce the contact time between the scaffolds and chemical reagents.
Porcine ovaries were randomly assigned to three groups: novel decellularized group, conventional decellularized group and fresh group. The ovaries in the novel decellularized group underwent three cycles of freezing by liquid nitrogen and thawing at temperatures around 37 degree C before decellularization. The efficiency of the decellularization procedure was assessed through histological staining and DNA content analysis. The maintenance of ovarian decellularized extracellular matrix(ODECM) constituents was determined by analyzing the content of matrix proteins. Additionally, we evaluated the biocompatibility of the decellularized extracellular matrix(dECM) by observing the growth of granulosa cells on the ODECM scaffold in vitro.
Hematoxylin and eosin staining, DAPI staining and DNA quantification techniques collectively confirm the success of the novel decellularization methods in removing cellular and nuclear components from ovarian tissue. Moreover, quantitative assessments of ODECM contents revealed that the novel decellularization technique preserved more collagen and glycosaminoglycan compared to the conventional decellularized group (P<0.05). Additionally, the novel decellularized scaffold exhibited a significantly higher number of granulosa cells than the conventional scaffold during in vitro co-culture (P<0.05).
The novel decellularized method demonstrated high efficacy in eliminating DNA and cellular structures while effectively preserving the extracellular matrix. As a result, the novel decellularized method holds significant promise as a viable technique for ovarian decellularization in forthcoming studies. Doi.org/10.54680/fr24310110212.
卵巢组织冷冻保存用于保留生育能力存在恶性细胞再种植的风险。人工卵巢是解决这个问题的一种很有前途的方法。然而，卵巢去细胞化方案有限。因此，需要进一步研究以获得更好的卵巢去细胞化技术，用于构建人工卵巢支架。
通过将液氮与化学试剂结合，减少支架与化学试剂的接触时间，建立一种新颖的猪整个卵巢去细胞化技术。
猪卵巢被随机分为三组：新型去细胞组、传统去细胞组和新鲜组。新型去细胞组的卵巢在去细胞化前经历了三次液氮冷冻和 37°C 左右解冻循环。通过组织学染色和 DNA 含量分析评估去细胞化程序的效率。通过分析基质蛋白含量来确定卵巢去细胞外基质（ODECM）成分的保留情况。此外，我们通过观察体外在 ODECM 支架上生长的颗粒细胞来评估去细胞外基质（dECM）的生物相容性。
苏木精-伊红染色、DAPI 染色和 DNA 定量技术共同证实了新型去细胞化方法成功地从卵巢组织中去除了细胞和核成分。此外，ODECM 含量的定量评估显示，与传统去细胞组相比，新型去细胞化技术保留了更多的胶原蛋白和糖胺聚糖（P<0.05）。此外，在体外共培养期间，新型去细胞化支架上的颗粒细胞数量明显多于传统支架（P<0.05）。
新型去细胞化方法在有效去除 DNA 和细胞结构的同时，表现出高效性，同时有效保留了细胞外基质。因此，新型去细胞化方法作为卵巢去细胞化的可行技术在未来的研究中具有重要意义。Doi.org/10.54680/fr24310110212。
