Cell Biology-Inspired Tissue Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, 6229 ER Maastricht, The Netherlands.
Musculoskeletal Gene Therapy Laboratory, Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, MN 55905, USA.
Int J Mol Sci. 2023 May 10;24(10):8556. doi: 10.3390/ijms24108556.
MicroRNAs (miRNAs) are short non-coding RNA sequences with the ability to inhibit the expression of a target mRNA at the post-transcriptional level, acting as modulators of both the degenerative and regenerative processes. Therefore, these molecules constitute a potential source of novel therapeutic tools. In this study, we investigated the miRNA expression profile that presented in enthesis tissue upon injury. For this, a rodent enthesis injury model was developed by creating a defect at a rat's patellar enthesis. Following injury, explants were collected on days 1 ( = 10) and 10 ( = 10). Contra lateral samples ( = 10) were harvested to be used for normalization. The expression of miRNAs was investigated using a "Fibrosis" pathway-focused miScript qPCR array. Later, target prediction for the aberrantly expressed miRNAs was performed by means of the Ingenuity Pathway Analysis, and the expression of mRNA targets relevant for enthesis healing was confirmed using qPCRs. Additionally, the protein expression levels of collagens I, II, III, and X were investigated using Western blotting. The mRNA expression pattern of , , , , and in the injured samples indicated their possible regulation by their respective targeting miRNA, which included miR-16, -17, -100, -124, -133a, -155 and -182. Furthermore, the protein levels of collagens I and II were reduced directly after the injury (i.e., day 1) and increased 10 days post-injury, while collagens III and X showed the opposite pattern of expression.
微小 RNA（miRNAs）是具有在转录后水平抑制靶 mRNA 表达能力的短非编码 RNA 序列，作为退行性和再生过程的调节剂。因此，这些分子构成了新型治疗工具的潜在来源。在本研究中，我们研究了损伤时附着组织中呈现的 miRNA 表达谱。为此，通过在大鼠髌腱附着处创建一个缺陷来开发了一种啮齿动物附着损伤模型。损伤后，在第 1 天（= 10）和第 10 天（= 10）收集外植体。收集对侧样本（= 10）用于归一化。使用“纤维化”途径聚焦的 miScript qPCR 阵列研究 miRNA 的表达。然后，通过 Ingenuity 通路分析对异常表达的 miRNA 进行靶预测，并使用 qPCR 确认与附着愈合相关的 mRNA 靶的表达。此外，使用 Western blot 研究胶原 I、II、III 和 X 的蛋白表达水平。受伤样本中 、 、 、 和 的 mRNA 表达模式表明它们可能受到各自靶向 miRNA 的调节，包括 miR-16、-17、-100、-124、-133a、-155 和 -182。此外，胶原 I 和 II 的蛋白水平在损伤后直接降低（即第 1 天），并在损伤后 10 天增加，而胶原 III 和 X 表现出相反的表达模式。
