The myotendinous junction (MTJ) is a specialised interface for transmitting high forces between muscle and tendon and yet the MTJ is a common site of strain injury with a high recurrence rate. The aim of this study was to identify previously unknown MTJ components in mature animals and humans. Samples were obtained from the superficial digital flexor (SDF) muscle-tendon interface of 20 horses and the tissue was separated through a sequential cryo-sectioning approach into muscle, MTJ (muscle tissue enriched in myofiber tips attached to the tendon), and tendon fractions. RT-PCR was performed for genes known to be expressed in the three tissue fractions and t-SNE plots were used to select the muscle, MTJ and tendon samples from 5 horses for RNA-sequencing. The expression of previously known and unknown genes identified through RNA-sequencing was studied by immunofluorescence on human hamstring MTJ tissue. The main finding was that RNA-sequencing identified expression of a panel of 61 genes enriched at the MTJ. 48 of these genes were novel for the MTJ, and 13 genes had been reported to be associated with the MTJ in earlier studies. The expression of known (COL22A1, NCAM, POSTN, NES, OSTN) and previously undescribed (MNS1 and LCT) MTJ genes was confirmed at the protein level by immunofluorescence on tissue sections of human MTJ. In conclusion, in muscle-tendon interface tissue enriched with myofiber tips, we identified expression of previously unknown MTJ genes representing diverse biological processes, which may be important in the maintenance of the specialized MTJ.

中文翻译：

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成熟马和人类肌腱连接处的 RNA 测序和免疫荧光

肌腱连接处 (MTJ) 是肌肉和肌腱之间传递高力的专门接口，但 MTJ 是常见的劳损部位，复发率高。本研究的目的是确定成熟动物和人类中以前未知的 MTJ 成分。从 20 匹马的浅指屈肌 (SDF) 肌肉-肌腱界面获取样本，并通过连续冷冻切片方法将组织分离为肌肉、MTJ（富含肌纤维尖端的肌肉组织，附着在肌腱上）和肌腱部分. 对已知在三个组织部分中表达的基因进行 RT-PCR，并使用 t-SNE 图从 5 匹马中选择肌肉、MTJ 和肌腱样本进行 RNA 测序。通过对人腘绳肌 MTJ 组织的免疫荧光研究了通过 RNA 测序鉴定的先前已知和未知基因的表达。主要发现是 RNA 测序鉴定了一组 61 个在 MTJ 富集的基因的表达。这些基因中有 48 个是 MTJ 的新基因，并且在早期研究中报告了 13 个基因与 MTJ 相关。已知（COL22A1、NCAM、POSTN、NES、OSTN）和以前未描述的（MNS1 和 LCT）MTJ 基因的表达通过人 MTJ 组织切片上的免疫荧光在蛋白质水平得到证实。总之，在富含肌纤维尖端的肌肉 - 肌腱界面组织中，我们确定了代表不同生物过程的先前未知的 MTJ 基因的表达，这可能对维持专门的 MTJ 很重要。