Bostani Aliasghar, Hoveizi Elham, Naddaf Hadi, Razeghi Jafar
Department of Plant, Cell and Molecular Biology, Faculty of Natural Sciences, University of Tabriz, Tabriz, Iran.
Department of Biology, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, Iran.
Mol Neurobiol. 2025 Feb;62(2):1493-1510. doi: 10.1007/s12035-024-04344-9. Epub 2024 Jul 13.
Nervous injuries are common in humans. One of the most advanced treatment methods is neural tissue engineering. This research aims to utilize nerve-like cells (NLCs) derived from endometrial mesenchymal stem cells (EnMSCs) on a polyacrylonitrile/chitosan (PAN/CS) scaffold, along with berberine, for the reconstruction of a rat sciatic nerve injury model. In this experimental study, EnMSCs were obtained through enzymatic digestion and identified using flow cytometry and their differentiation into adipocyte and osteoblast. PAN nanofiber scaffolds were produced through electrospinning, and EnMSCs were neurally differentiated on these scaffolds for grafting into an animal model. The expression of Nestin, Map-2, Tuj-1, and NF genes in NLCs was confirmed through RT-PCR and immunocytochemistry. Twenty-five adult male rats were used in this study, divided into 5 groups: (1) Scaffold/Cells/Berberine, (2) Scaffold/Cells, (3) Scaffold, (4) Berberine, and (5) Control. The animals were maintained for 8 weeks, and their sciatic nerve function (SFI) was assessed. Additionally, histological examinations were performed using hematoxylin/eosin, luxol fast blue staining, and immunohistochemistry. According to the results, extraction, identification, and differentiation of EnMSCs and fabrication of PAN conduit and its transplantation were successfully performed. The best behavioral performance and histology were observed in the Scaffold/Cells/Berberine group. The SFI test results were -24.08 for the Scaffold/Cells/Berberine group and -39.27 for the control group. The nerve diameter in these two groups was 591 µm and 80 µm, respectively, and the percentage of new nerve formation was 18.5% in the Scaffold/Cells/Berberine group and 0.2% in the control group. The immunohistochemistry results demonstrated that the intensity of the green color was higher in the groups with cells compared to the groups without cells. Furthermore, in the luxol staining results, all groups showed a significant improvement compared to the control group. In the Scaffold/Cells/Berberine group, fibers, and axons appeared denser, more organized, and displayed a higher intensity of blue staining. According to the results of this study, EnMSCs demonstrated efficient differentiation into NLCs. With the assistance of PAN/CS scaffolds and simultaneous administration of berberine, EnMSCs have the potential for nerve regeneration and recovery from sciatic nerve injury in the rat animal model.
神经损伤在人类中很常见。最先进的治疗方法之一是神经组织工程。本研究旨在将源自子宫内膜间充质干细胞（EnMSCs）的神经样细胞（NLCs）与小檗碱一起用于聚丙烯腈/壳聚糖（PAN/CS）支架上，以重建大鼠坐骨神经损伤模型。在本实验研究中，通过酶消化获得EnMSCs，并使用流式细胞术对其进行鉴定以及检测其向脂肪细胞和成骨细胞的分化情况。通过静电纺丝制备PAN纳米纤维支架，并将EnMSCs在这些支架上进行神经分化以用于移植到动物模型中。通过RT-PCR和免疫细胞化学证实了NLCs中巢蛋白、微管相关蛋白2（Map-2）、βIII微管蛋白（Tuj-1）和神经丝蛋白（NF）基因的表达。本研究使用了25只成年雄性大鼠，分为5组：（1）支架/细胞/小檗碱组，（2）支架/细胞组，（3）支架组，（4）小檗碱组，（5）对照组。将动物饲养8周，并评估其坐骨神经功能（SFI）。此外，使用苏木精/伊红染色、髓鞘染色和免疫组织化学进行组织学检查。根据结果，成功进行了EnMSCs的提取、鉴定和分化以及PAN导管的制备及其移植。在支架/细胞/小檗碱组中观察到最佳的行为表现和组织学结果。支架/细胞/小檗碱组的SFI测试结果为-24.08，对照组为-39.27。这两组的神经直径分别为591 µm和80 µm，支架/细胞/小檗碱组的新神经形成百分比为18.5%，对照组为0.2%。免疫组织化学结果表明，与无细胞组相比，有细胞组的绿色荧光强度更高。此外，在髓鞘染色结果中，所有组与对照组相比均有显著改善。在支架/细胞/小檗碱组中，纤维和轴突显得更密集、更有组织，并且蓝色染色强度更高。根据本研究结果，EnMSCs表现出向NLCs的高效分化。在PAN/CS支架的辅助下并同时给予小檗碱，EnMSCs在大鼠动物模型中具有促进坐骨神经损伤后神经再生和恢复的潜力。
