LncRNA SNHG14/miR⁃148a⁃3p/PODXL轴在瘢痕疙瘩形成过程中的分子机制

Molecular mechanism of LncRNA SNHG14/miR⁃148a⁃3p/PODXL axis in process of keloid formation

目的探讨LncRNA SNHG14/miR⁃148a⁃3p/PODXL轴在瘢痕疙瘩形成过程中的分子机制。方法qRT⁃PCR法与Western blot法分别检测正常皮肤组织、瘢痕疙瘩组织中SNHG14、miR⁃148a⁃3p、PODXL的表达量;Pearson法分析瘢痕疙瘩组织中SNHG14与miR⁃148a⁃3p的相关性,以及miR⁃148a⁃3p与PODXL的相关性;体外培养人瘢痕疙瘩成纤维细胞,si⁃NC、si⁃SNHG14分别转染入瘢痕疙瘩成纤维细胞;MTT实验、平板克隆形成实验、Transwell小室实验分别检测细胞增殖、克隆形成、迁移及侵袭;双荧光素酶报告基因实验检测SNHG14与miR⁃148a⁃3p的靶向关系,以及miR⁃148a⁃3p与PODXL的靶向关系;Western blot法检测ProcollagenⅠ、α⁃SMA、Col1蛋白表达量。结果与正常皮肤组织比较,瘢痕疙瘩组织中SNHG14与PODXL的表达量升高(P<0.05),miR⁃148a⁃3p的表达量降低(P<0.05);SNHG14与miR⁃148a⁃3p呈负相关(r=-0.8031,P<0.001),miR⁃148a⁃3p与PODXL呈负相关(r=-0.8559,P<0.001),SNHG14与PODXL呈正相关(r=0.7241,P<0.001);转染si⁃SNHG14可明显降低PODXL的表达量,促进miR⁃148a⁃3p的表达,并可降低细胞活力、克隆形成、迁移及侵袭能力,ProcollagenⅠ、α⁃SMA、Col1蛋白水平降低,差异有统计学意义(P<0.05);SNHG14可靶向调控miR⁃148a⁃3p,miR⁃148a⁃3p可靶向调控PODXL。结论干扰SNHG14表达可通过调控miR⁃148a⁃3p/PODXL轴而抑制瘢痕疙瘩成纤维细胞增殖、克隆形成、迁移、侵袭及胶原蛋白合成。

Objective To explore the molecular mechanism of LncRNA SNHG14/miR⁃148a⁃3p/PODXL axis in the process of keloid formation.Methods qRT⁃PCR and Western blot methods were used to detect the expression of SNHG14,miR⁃148a⁃3p and PODXL in normal skin tissue and keloid tissue.Pearson method was used to detect the correlation between SNHG14 and miR⁃148a⁃3p in keloid tissue,and the correlation between miR⁃148a⁃3p and PODXL.Human keloid fibroblasts were cultured in vitro,and si⁃NC and si⁃SNHG14 were respectively transfected into keloid fibroblasts.MTT experiment,plate clone formation experiment,and transwell chamber experiment were used to detect cell proliferation,clone formation,migration and invasion respectively.The dual luciferase reporter gene experiment was used to detect the targeting relationship between SNHG14 and miR⁃148a⁃3p,and the targeting relationship between miR⁃148a⁃3p and PODXL.Western blot method was used to detect the protein expression of ProcollagenⅠ,α⁃SMA and Col1.Results Compared with that in normal skin tissue,the expression of SNHG14 and PODXL in keloid tissue was increased(P<0.05),while that of miR⁃148a⁃3p was decreased(P<0.05).SNHG14 was negatively correlated with miR⁃148a⁃3p(r=-0.8031,P<0.001);miR⁃148a⁃3p was negatively with PODXL(r=-0.8559,P<0.001),but SNHG14 was positively with PODXL(r=0.7241,P<0.001).Transfection of si⁃SNHG14 could significantly reduce the expression of PODXL,promote the expres⁃sion of miR⁃148a⁃3p,and could reduce cell viability,clone formation,migration and invasion,while the protein levels of ProcollagenⅠ,α⁃SMA,and Col1 were decreased;the difference was statistically significant(P<0.05).SNHG14 could target and regulate miR⁃148a⁃3p,and miR⁃148a⁃3p could target and regulate PODXL.Conclusion Interfering with the expression of SNHG14 could inhibit the proliferation,clone formation,migration,invasion and collagen synthesis of keloid fibroblasts by regulating the miR⁃148a⁃3p/PODXL axis.