Zhong Hua, Guo Lebin, Guo Wei, Liu Fusheng, Zheng Bowen, Zhang Shiquan, Wang Pengyu, Tian Chenjun, Xu Zhun, Zou Ming-Xiang
Yiyang Central Hospital Affiliated to Hunan University of Traditional Chinese Medicine, Yiyang , China.
Department of Spine Surgery, The Second Xiangya Hospital, Central South University, Changsha , China.
Neurosurgery. 2025 Mar 25;97(3):752-761. doi: 10.1227/neu.0000000000003406.
Currently, the role of long noncoding RNA (lncRNA) DiGeorge syndrome critical region gene 5 ( DGCR5 ) in intervertebral disk degeneration (IDD) remains unclear. This study explored the molecular mechanisms of how lncRNA DGCR5 promoted human nucleus pulposus cells (NPCs) degeneration and pyroptosis during IDD.
NPCs were identified by microscopic observation, flow cytometry, and immunofluorescence. An in vitro NPC degeneration model was induced using lipopolysaccharides treatment. SP600125 and SB203580 were used to inhibit the c-Jun N-terminal kinase (JNK) and p38 MAPK signaling, respectively. Quantitative real-time polymerase chain reaction or Western blot was performed to detect lncRNA DGCR5 , JNK, phospho-JNK, p38 MAPK, and phospho-p38 MAPK expressions in nucleus pulposus tissues and NPCs. Cell Counting Kit-8 assay was conducted to detect NPC activity. Western blot was performed to detect the expression levels of extracellular matrix-associated proteins (including collagen II, aggrecan, and matrix metalloproteinase 3 [MMP3]) and pyroptosis-associated proteins (including nucleotide oligomerization domain-like receptors family pyrin domain containing 3), cleaved caspase-1, lactate dehydrogenase, and N-terminal fragment of Gasdermin D (GSDMD) in NPCs. Enzyme-linked immunosorbent assay was conducted to detect the expressions of interleukin (IL)-1beta (IL-1β) and IL-18 in NPC supernatants.
DGCR5 was upregulated, and the JNK/p38 MAPK signaling was activated both in nucleus pulposus tissues of IDD patients and lipopolysaccharides-induced NPCs. Inhibition of the JNK/p38 MAPK signaling enhanced NPC proliferation, promoted collagen II and aggrecan expression, while inhibited MMP 3 expression. Silencing of DGCR5 suppressed JNK/p38 MAPK signaling activity, inhibited NPC proliferation, and reduced collagen II and aggrecan expression but promoted MMP 3 expression. Similar findings were also observed for the expressions of nucleotide oligomerization domain-like receptors family pyrin domain containing 3, cleaved caspase-1, N-terminal fragment of GSDMD, IL-1β, and IL-18 as well as the released lactate dehydrogenase level. However, overexpression of DGCR5 yielded opposite results.
These data suggest that lncRNA DGCR5 regulates NPC degeneration and pyroptosis to promote IDD through the JNK/p38 MAPK signaling pathway.
目前，长链非编码RNA（lncRNA）狄乔治综合征关键区域基因5（DGCR5）在椎间盘退变（IDD）中的作用尚不清楚。本研究探讨了lncRNA DGCR5在IDD过程中促进人髓核细胞（NPC）退变和焦亡的分子机制。
通过显微镜观察、流式细胞术和免疫荧光鉴定NPC。使用脂多糖处理诱导建立体外NPC退变模型。分别用SP600125和SB203580抑制c-Jun氨基末端激酶（JNK）和p38丝裂原活化蛋白激酶（MAPK）信号通路。采用定量实时聚合酶链反应或蛋白质印迹法检测髓核组织和NPC中lncRNA DGCR5、JNK、磷酸化JNK、p38 MAPK和磷酸化p38 MAPK的表达。进行细胞计数试剂盒-8检测以检测NPC活性。采用蛋白质印迹法检测NPC中细胞外基质相关蛋白（包括Ⅱ型胶原、聚集蛋白聚糖和基质金属蛋白酶3 [MMP3]）和焦亡相关蛋白（包括含核苷酸寡聚化结构域样受体家族pyrin结构域3）、裂解的半胱天冬酶-1、乳酸脱氢酶和Gasdermin D（GSDMD）N端片段的表达水平。采用酶联免疫吸附测定法检测NPC上清液中白细胞介素（IL）-1β（IL-1β）和IL-18的表达。
在IDD患者的髓核组织和脂多糖诱导的NPC中，DGCR5表达上调，JNK/p38 MAPK信号通路被激活。抑制JNK/p38 MAPK信号通路可增强NPC增殖，促进Ⅱ型胶原和聚集蛋白聚糖表达，同时抑制MMP 3表达。沉默DGCR5可抑制JNK/p38 MAPK信号通路活性，抑制NPC增殖，降低Ⅱ型胶原和聚集蛋白聚糖表达，但促进MMP 3表达。在含核苷酸寡聚化结构域样受体家族pyrin结构域3、裂解的半胱天冬酶-1、GSDMD N端片段、IL-1β和IL-18的表达以及释放的乳酸脱氢酶水平方面也观察到类似结果。然而，过表达DGCR5则产生相反的结果。
这些数据表明，lncRNA DGCR5通过JNK/p38 MAPK信号通路调节NPC退变和焦亡，从而促进IDD。
