| Background and Objective The annual incidence of oral cancer is about 350,000-400,000.The World Health Organization(WHO)reported in 1985 that the estimated age-standardized incidence of oral and pharyngeal cancer in China was 8.7/100,000 for men and 60,000/100,000 for women.The number of actual cases of oral cancer in China is among the highest in the world.Therefore,prevention and treatment of oral cancer is an arduous task before us.The occurrence of oral cancer is related to smoking,drinking,viral infection,malnutrition,eating habits and local irritation.Bit-l is a new gene discovered in 2004.It is located in human 17q23.2,its m RNA length is about 1kb,encoding 179 aa,and the molecular weight of Bit-1 protein is about 27 k Da.Its clear physiological function is still unclear.Research result.Upregulation of Bcl-2 in Bcl-2 in adherent cells is achieved by activating the FAK-P13K-AFK-NFk B signaling pathway,thereby promoting cell survival,and down-regulating endogenous Bit-1 will ultimately enhance adhesion.Apoptosis of cells.The Bit-l gene is a new gene discovered by researchers in 2004 when studying the regulation of Bcl-2 expression by integrin.Inhibitor of transcription 1).Later genomic studies showed that the evolution of bacteria from human to human Bit-l is highly conserved.In human tissues,Bit-l is highly expressed in the testis,prostate,skeletal muscle and liver,in the heart,spleen and placenta.And the colon is also expressed,but its expression is almost undetectable in human thymus and peripheral blood leukocytes.The higher the concentration of Bit-l in the cytoplasm,the more obvious the promotion of apoptosis.Bit-l protein is a new molecule that is very close to apoptosis.Therefore,studying this mitochondrial protein that determines apoptosis has important clinical treatment for exploring the occurrence and regulation of cancer cell apoptosis.significance.Studies have shown that in the esophageal squamous cell carcinoma,after silencing of Bit-1,microarray analysis confirmed that IL-6 levels were significantly reduced.IL-6 is a known multifunctional molecule that was originally thought to be involved in immune and inflammatory responses.More and more studies have shown that IL-6 is highly expressed in patients including oral squamous cell carcinoma and is associated with poor prognosis.In addition,IL-6 also promotes tumor cell growth,invasion and migration,angiogenesis,and metastasis through IL-6R-mediated signaling pathways;STAT3 signaling is involved in tumor progression through IL-6R-mediated activation.Based on this,we hypothesized that "Bit-1 may regulate the progression of oral squamous cell carcinoma by inducing IL-6/IL-6R-STAT3-mediated angiogenesis,growth,and metastasis." The aim of this study was to investigate the role of Bit1 in inhibiting the growth inhibition and reducing migration and invasion of oral squamous cell carcinoma by inhibiting the IL-6/IL-6R-STAT3 pathway,and to explore the mechanism of its action in order to serve as an oral scale.The treatment of squamous cell carcinoma offers some new ideas.Methods 1.Clinical sample screening 60 clinical samples(60 cases of oral squamous cell carcinoma and adjacent tissues)were collected,and the expression of Bit-1 and IL-6 was detected by PCR and WB.2.Cell line screening Human CAL-27,SCC-25,KB cells,PCR and WB were used to detect the expression of Bit-1 and IL-6,and a cell line with high expression of Bit-1(high expression of IL-6)was selected.3.Co-IP verifies the relationship between bit-1 and IL-6 The experimental results show that there is a positive correlation between bit-1 and IL-6.4.Interference with bit-1 on cell migration,invasion and apoptosis 1)Design and synthesize 3 bit-1 sh RNAs,PCR and WB screening.2)Cell grouping(6 groups): blank cells,NC control,sh RNA interference,IL-6/STAT3 inhibitor(Corylifol A,10 u M,40 min),interference + IL-6(added IL-6 100 ng/ml after 24 h of transfection),24h),interference + IL-6/STAT3 inhibitor(Corylifol A,10 u M,40 min after 24 h transfection)Detection indicators:(1)Transwell detection of changes in cell migration ability;(2)Transwell detects changes in cell invasion ability;(3)Flow detection of apoptosis.3)PCR and WB detection of BIT-1,IL-6,IL-6R,STAT3.Results 1.Western Blot method to determine the expression of Bit-1 and IL-6 in adjacent tissues and tumor tissues of patients,and concluded that Bit-1 and IL-6 are weakly expressed in normal tissues and highly expressed in OSCC tissues.The results of real-time PCR showed that Bit-1 and IL-6 are weakly expressed in normal tissues and highly expressed in OSCC tissues.2.Cell screening experiments,we found that all OSCC cells used in this study have high levels of both Bit1 and IL-6 protein expression.However,the expression levels of Bit1 and IL-6 are different in these OSCC cells,and the expression shows a relatively high level in SCC-25,and the expression levels of Bit1 and IL-6 are lower in CAL-27 and KB cells.3.Co-IP verified the role of Bit-1 and IL-6: IL-6 was detected in Bit1 immunopreci pitation,and Bit-1 was detected in IL-6 immunoprecipitation.There is sufficient evidence that the positive correlation between Bit-1 and IL-6 was confirmed by WB results.4.The knockout of Bit-1 can effectively reduce the migration and invasion ability of SCC-25 cells and promote the apoptosis of SCC-25 cells.Moreover,the expression level of Bit-1 is positively correlated with the expression levels of IL-6 and its downstream signaling molecules IL-6R and STAT3.Conclusions 1.Bit-1 and IL-6 are highly expressed in human oral squamous cell carcinoma,and Bit-1 is positively correlated with the expression of IL-6 in OSCC.2.Bit-1 regulates OSCC migration,invasion and apoptosis by activating IL-6 / IL-6 R-STAT3 signaling pathway. |
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