Inhibitory effect on the apoptosis of disc nucleus pulposus cells in rats by silencing COL9A3 gene to mediate MAPK signaling pathway: a study on the function and mechanism.

OBJECTIVE: To explore the effect of collagen IX alpha 3 chain (COL9A3) gene silencing on apoptosis of nucleus pulposus cells in rats with intervertebral disc degeneration (IVDD) and its regulatory mechanism, so as to provide potential reference for the treatment of IVDD. MATERIALS AND METHODS: The model of IVDD in rats were constructed to isolate, culture and identify nucleus pulposus cells for subsequent experiment. With the construction of lentivirus vectors, cells were divided into Blank group, negative control (NC) group, COL9A3 shRNA group, COL9A3 overexpression group, mitogen-activated protein kinase (MAPK) pathway inhibitor (Theaflavin 3,3'-digallate, TF3) group and COL9A3 shRNA+TF3 group according to different transfection treatments. After cell transfection, the expression of COL9A3, extracellular regulated protein kinase 1/2 (ERK1/2) and phosphorylated-ERK1/2 (p-ERK1/2), MEK1/2 and p-MEK1/2, as well apoptosis related indexes were detected by using quantitative real-time PCR (qRT-PCR) and Western Blot. Furthermore, 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect the proliferation of transfected cells, and flow cytometry to detect changes of cell cycle and apoptosis. RESULTS: The model of IVDD in rats was established successfully, and rat nucleus pulposus cells were cultured subsequently. After transfection, compared with NC group, there was significant downregulation in COL9A3 expression, significant upregulation in ERK1/ERK2 and MEK1/MEK2 expressions, evident increase in Bax and Caspase3 expressions, and a decrease Bcl-2 expression in transfected cells of COL9A3 shRNA group (all p<0.05). The opposite trends were detected in the above indexes in cells of COL9A3 overexpression group (all p<0.05). Furthermore, in TF3 group, there was significant decrease in ERK1/ERK2 and MEK1/MEK2 expressions, reduction in Bax and Caspase3 expressions, increase in Bcl-2 expression (all p<0.05), yet without evident change in the expression of COL9A3. Meanwhile, COL9A3 expression was decreased in COL9A3 shRNA+TF3 group (p<0.05); however, there was no significant statistical difference in the remaining indexes when compared with those in the NC group (all p>0.05). Subsequently, compared with NC group, COL9A3 overexpression group and TF3 group revealed evident increased cell proliferation and significant decreased cell apoptosis (all p<0.05); whereas it was remarkably the opposite in COL9A3 shRNA group (all p<0.05). However, no evident difference was detected in the comparison of cell proliferation and apoptosis among Blank group, NC group and COL9A3 shRNA+TF3 group (all p>0.05). CONCLUSIONS: Overexpression of COL9A3 gene and inhibition of MAPK signaling pathway can induce proliferation and inhibit apoptosis of nucleus pulposus cells. Significantly, silence of COL9A3 gene expression can activate MAPK signaling pathway and affect the expression of apoptosis related factors, so as to inhibit the proliferation of nucleus pulposus cells and promote cell apoptosis in rats with IVDD.

Renal manifestations of concurrent systemic lupus erythematosus and HIV infection.

Autoimmune phenomena are common in human immunodeficiency virus (HIV) infection, yet systemic lupus erythematosus (SLE) and HIV infection rarely are seen concurrently in the same patient. Many of the cases of combined HIV infection and SLE reported in the literature are patients with SLE before HIV infection and who did not undergo renal biopsy at a time when both processes were present. We report the clinical manifestations and renal biopsy findings in four subjects with concurrent HIV infection and SLE and compare them with the seven previously reported cases in the literature. Taken together, most patients were black (91%) and male (73%), and approximately half (55%) were children with perinatal HIV infection. These demographics differ markedly from those of idiopathic SLE, a disease that predominantly affects female adults. Renal presentations included proteinuria and hypocomplementemia, frequently with hematuria and renal insufficiency. Renal biopsy findings in 10 cases included all classes of lupus nephritis (class IIb in two cases, class III in one case, class IV in three cases, class V in three cases, class III and V in one case), two of which also displayed overlapping features of HIV-associated nephropathy (HIVAN). One case had isolated findings of HIVAN. This cohort provides a unique population in which to study interacting pathomechanisms between HIV infection and SLE.