Our preliminary experiments found that miR-21-5p was significantly up-regulated in spinal cord injured (SCI) rat by next-generation sequencing. Nevertheless, to our knowledge, no studies have investigated the role of miR-21-5p in bladder of SCI rats.

Female Wistar rats underwent spinal cord transection at T9-10 and were randomly divided into negative control (NC) and experimental groups (n=10 for each group). Tail intravenous injection of miR-NC or antagomiR-21-5p were performed every three days after spinal cord transection, which were nine times in total respectively. Four weeks after transection, cystometric analyses were conducted and bladder tissues were collected for Masson staining, qRT-PCR and western-blot analyses. The normal rats were used as blank control group.

Compared with NC group, the level of miR-21-5p was significantly decreased in experimental group; while it was much higher in NC than blank group. In addition, Masson staining showed that bladder fibrosis was significantly lessened in experimental group. Decreased non-voiding contraction number and increased intercontraction interval were also found in experimental group. qRT-PCR and western-blot analyses showed that inhibiting miR-21-5p downregulated the levels of Smad7 mRNA and protein, which subsequently increased the levels of p-Smad3 mRNA and protein.

All of these days, miR-21-5p was reported to act as an oncogene through inhibiting cellular apoptosis by targeting tumor suppressor genes.(1) However, it should be noted that the over-expression of miR-21-5p abnormally activates transforming growth factor-β1 (TGF-β1) and Hedgehog signaling pathways, promoting tumor invasion by the induction of EMT. As we all know, TGF-β1 signaling pathway plays a pivotal role in EMT and fibrogenesis. Recently, the up-regulated expression of miR-21-5p was reported to be involved in the renal, myocardial, pulmonary and peritendinous fibrosis and may serve as an alternative target to directly inhibit these fibrosis.(2) Further research indicates that miR-21-5p overexpression could enhance TGF-β1 induced EMT by inhibiting smad7.(3) Moreover, proliferation, migration, and pro-fibrotic activities of fibroblasts were promoted by miR-21-5p through reducing smad7 expression. Specifically speaking, the increase of intracellular miR-21-5p induced fibroblasts differentiation into myofibroblasts and overexpression of extracellular matrix (ECM) and fibrogenic markers. What’s more, tissue inhibitor of metalloproteinases (TIMPs) which was implicated in collagen synthesis and accumulation during fibrosis were also targeted by miR-21-5p.
Importantly, previous study suggests that miR-21-5p is upregulated by TGF-β1 via activation of Smad3 rather than Smad2. In normal state, Smad-3 activation can induce expression of smad7, which forms a negative feedback mechanism. Nevertheless, in pathological situations, the expression of smad7 was found to be suppressed and the negative feedback was damaged, which may due to the upregulated expression of miR-21-5p. In contrast, conditional knockout of Smad2 could enhance miR-21-5p expression. Further research is needed to explore the mechanisms underlying the interactions between miR-21-5p and TGF-β1 signaling pathway. In addition, bladder fibrosis after spinal cord injury may bear responsibility for the high intravesical pressures, low bladder compliance, bladder wall stiffer and vesicoureteral reflux. Up to now, however, there is no effective method for preventing bladder fibrosis. Therefore, it is also of great significance to investigate the functional role of miR-21-5p in bladder fibrosis after spinal cord injury.

Inhibiting miR-21-5p alleviates bladder fibrosis and detrusor overactivity in SCI rats, which may potentially serve as a new molecular target for neurogenic bladder.

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Nonaka CKV, Macedo CT, Cavalcante BRR, et al. Circulating miRNAs as Potential Biomarkers Associated with Cardiac Remodeling and Fibrosis in Chagas Disease Cardiomyopathy. Int J Mol Sci 2019;20.
Wang JY, Gao YB, Zhang N, et al. miR-21 overexpression enhances TGF-beta1-induced epithelial-to-mesenchymal transition by target smad7 and aggravates renal damage in diabetic nephropathy. Mol Cell Endocrinol 2014;392:163-72.