Institute of Veterinary Anatomy, Histology and Embryology, Justus-Liebig-University of Giessen, Giessen, Germany.
Clinic of Small Animals, c/o Institute of Veterinary Anatomy, Histology and Embryology, Justus-Liebig-University of Giessen, Giessen, Germany.
J Anat. 2021 Aug;239(2):336-350. doi: 10.1111/joa.13420. Epub 2021 Feb 27.
Muscle stem cells (MSCs) are a promising tool for cell-based therapy and tissue regeneration in veterinary medicine. Evaluation of MSCs from muscles of different origins improves our understanding of their regenerative potential. The present study compared the stemness, cell proliferation, migration potential, myogenic differentiation (MD), and multipotency of MSCs for four developmentally different muscles of ovine origin. MSCs were isolated from the hind limb (HL), diaphragm (DI), extraocular (EO), and masseter (MS) muscles. Cell proliferation, migration, and stemness were examined using sulforhodamine B, and colony formation assays. Evaluation of multipotency was examined using histological and morphometric analyses, alkaline phosphatase (ALP) activity, and the expression of myogenic, adipogenic, and osteogenic markers using RT-qPCR. Data were statistically analysed using analysis of variance. The results revealed that all experimental groups expressed stem cell markers paired box transcription factor Pax7, α7-integrin, CD90, and platelet-derived growth factor receptor alpha. DI and HL muscle cells displayed higher proliferation, migration, and colony formation capacities compared to the EO and MS muscle cells. HL and DI muscle cells showed increased MD, as indicated by myotube formation and relative expression of MyoD at day 7 and Myogenin at day 14. Although MS and EO muscle cells displayed impaired MD, these cells were more prone to adipogenic differentiation, as indicated by Oil Red O staining and upregulated fatty acid-binding protein 4 and peroxisome proliferator-activated receptor gamma expression. DI muscle cells demonstrated a higher osteogenic differentiation capability, as shown by the upregulation of osteopontin expression and an elevated ALP activity. Our data indicate that ovine HL and DI MSCs have a higher regenerative and multipotent potential than the EO and MS muscle cells. These results could be valuable for regional muscle biopsies and cell-based therapies.
肌肉干细胞（MSCs）是兽医细胞治疗和组织再生的有前途的工具。评估来自不同来源的肌肉中的 MSCs 可以提高我们对其再生潜力的理解。本研究比较了源自绵羊不同起源的四肢（HL）、膈肌（DI）、眼外肌（EO）和咬肌（MS）的 MSCs 的干性、细胞增殖、迁移潜力、成肌分化（MD）和多能性。从后肢（HL）、膈肌（DI）、眼外肌（EO）和咬肌（MS）中分离出 MSCs。使用磺酰罗丹明 B 和集落形成测定法检查细胞增殖、迁移和干性。使用组织学和形态计量学分析、碱性磷酸酶（ALP）活性以及使用 RT-qPCR 评估成肌、成脂和成骨标志物的表达来评估多能性。使用方差分析对数据进行统计学分析。结果表明，所有实验组均表达了干细胞标志物配对盒转录因子 Pax7、α7-整合素、CD90 和血小板衍生生长因子受体 alpha。与 EO 和 MS 肌肉细胞相比，DI 和 HL 肌肉细胞显示出更高的增殖、迁移和集落形成能力。HL 和 DI 肌肉细胞表现出更高的 MD，这表现为在第 7 天形成肌管和相对表达 MyoD，在第 14 天表达 Myogenin。尽管 MS 和 EO 肌肉细胞显示出受损的 MD，但这些细胞更容易向成脂分化，这表现为油红 O 染色和脂肪酸结合蛋白 4 和过氧化物酶体增殖物激活受体 gamma 表达上调。DI 肌肉细胞表现出更高的成骨分化能力，表现为骨桥蛋白表达上调和 ALP 活性升高。我们的数据表明，绵羊 HL 和 DI MSCs 比 EO 和 MS 肌肉细胞具有更高的再生和多能性潜力。这些结果对于区域性肌肉活检和基于细胞的治疗可能具有重要价值。
