BACKGROUND Spermatogonial stem cell (SSC) transplantation technology as a promising option for male fertility preservation has received increasing attention, along with efficient SSC purification technology as a necessary technical support; however, the safety of such application in patients with tumors remains controversial. METHODS In this study, we used a green fluorescent protein mouse xenograft model of B cell acute lymphocytic leukemia. We isolated and purified SSCs from the testicular tissue of model mice using density gradient centrifugation, immune cell magnetic bead separation, and flow cytometry. The purified SSCs were transplanted into convoluted seminiferous tubules of the nude mice and C57BL/6 male mice subjected to busulfan. The development and proliferation of SSCs in the recipient testis were periodically tested, along with whether B cell acute lymphocytic leukemia was induced following SSC implantation. The genetic characteristics of the offspring obtained from natural mating were also observed. RESULTS In testicular leukemia model mice, a large number of BALL cells infiltrated into the seminiferous tubule, spermatogenic cells, and sperm cells in the testis tissue decreased. After spermatogonial stem cell transplantation, the transplanted SSCs purified by immunomagnetic beads and flow cytometry methods colonized and proliferated extensively in the basement of the seminiferous tubules of mice; a large number of spermatogenic cells and sperm were found in recipient testicular tissue after 12 weeks of SSC transplantation. In leukemia detection in nude mice after transplantation in the three SSC purification groups, a large number of BALL cells could be detected in the blood of recipient mice 2-3 weeks after transplantation in the density gradient centrifugation group, but not in the blood of the flow cytometry sorting group and the immunomagnetic bead group after 16 weeks of observation. CONCLUSIONS In this study, we confirmed that immunomagnetic beads and flow cytometry methods of purifying SSCs from the testicular tissue of the testicular leukemia mouse model could be safely applied to the SSC transplantation technology without concomitant tumor implantation. The results thus provide a theoretical basis for the application of tumor SSC cryopreservation for fertility preservation in patients with tumors.

中文翻译：

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各种生精干细胞纯化方法应用于生精干细胞移植的相对安全性。


背景技术精原干细胞（SSC）移植技术作为男性生育力保存的一种有前景的选择越来越受到关注，高效的SSC纯化技术作为必要的技术支撑。然而，这种应用在肿瘤患者中的安全性仍存在争议。方法在本研究中，我们使用了B细胞急性淋巴细胞白血病的绿色荧光蛋白小鼠异种移植模型。我们使用密度梯度离心、免疫细胞磁珠分离和流式细胞术从模型小鼠的睾丸组织中分离和纯化SSC。将纯化的SSC移植到接受白消安处理的裸鼠和C57BL/6雄性小鼠的曲细精管中。定期检测受者睾丸中SSC的发育和增殖，以及SSC植入后是否诱发B细胞急性淋巴细胞白血病。还观察了自然交配获得的后代的遗传特征。结果睾丸白血病模型小鼠生精小管内大量BALL细胞浸润，睾丸组织中生精细胞、精子细胞减少。精原干细胞移植后，通过免疫磁珠和流式细胞术方法纯化的移植SSC在小鼠生精小管基底广泛定植和增殖； SSC移植12周后，在受体睾丸组织中发现大量生精细胞和精子。 3个SSC纯化组移植后裸鼠白血病检测中，密度梯度离心组移植后2-3周受体小鼠血液中可检测到大量BALL细胞，而移植组小鼠血液中未检测到大量BALL细胞。流式细胞术分选组和免疫磁珠组经过16周的观察。结论在本研究中，我们证实从睾丸白血病小鼠模型睾丸组织中纯化SSC的免疫磁珠和流式细胞术方法可以安全地应用于SSC移植技术，而无需伴随肿瘤植入。该结果为肿瘤SSC冷冻保存在肿瘤患者生育力保存中的应用提供了理论依据。