Effect of troponin-related proteins on proliferation and migration of lung adenocarcinoma cells and its relationship with prognosis of patients
Objective An Investigation into the Expression of Troponin-Related Protein(TROAP)in lung adenocarcinoma and its effects on the biological function of lung adenocarcinoma cells and infiltration of immune cells.Methods This study involved 68 samples of lung adenocarcinoma tissues obtained from patients who received surgical treatment and were pathologically diagnosed at Hebei Chest Hospital between January 1,2014,and June 30,2016.Additionally,20 samples of adjacent normal tissues(located more than 5cm away from the tumour edge)were included as the subjects of this re-search.An analysis was conducted on the expression of TROAP mRNA in lung adenocarcinoma using the Cancer Genome Atlas(TCGA)database and the Gene Expression Omnibus(GEO)database.The study also examined the link between TROAP mRNA expression and clinicopathological characteristics as well as prognosis.TROAP expression in lung cancer tissues and cell lines was assessed using real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR),Western blotting,and immunohistochemistry techniques.The TROAP interference sequence was created and introduced into A549 cells by transfection.The cells were subsequently categorised into four groups:a control group with no treat-ment(Blank group),a control group with a non-targeting siRNA(si-NC),and two groups treated with siRNA that spe-cifically targeted TROAP(si-TROAP-1 group and si-TROAP-2 group).The cell's ability to multiply,move,and invade other cells was measured using cell counting kit 8(CCK8),colony formation,transwell,and wound healing tests.An a-nalysis was conducted to examine the biological roles of TROAP in lung cancer using GO function and KEGG pathway en-richment.In addition,we investigated the correlation between TROAP expression levels and the extent of immune cell in-filtration in tumours using single-sample gene set enrichment analysis(ssGSEA).Results In the TCGA,GSE19804,and GSE43458 datasets,the levels of TROAP expression were found to be higher in lung adenocarcinoma compared to normal lung tissues.The t-values for these comparisons were 12.260,5.547 and 6.848,respectively,all with a signifi-cance level of P＜0.001.The expression of TROAP showed a strong correlation with TNM staging(x2=11.708,P＜0.001),tumour size(x2=5.968,P=0.015),and lymph node metastasis(x2=18.634,P＜0.001),indicating substan-tial differences between the groups.The analysis of patients with lung adenocarcinoma showed that the expression of TROAP was linked to both overall survival and progression-free survival.The x2 values for overall survival and progres-sion-free survival were 13.310 and 10.040,respectively,with corresponding P-values of＜0.001 and 0.002.Moreover,the study found that a high level of TROAP expression(HR=4.724,95％CI:1.307-17.071,P=0.018)and the pres-ence of distant metastasis in the tumour(HR=4.906,95％CI:1.234-19.508,P=0.024)were identified as separate prognostic indicators for lung adenocarcinoma.The CCK8 proliferation assay results indicated that the cell growth rates at 72 hours were 0.897±0.060,0.653±0.021,and 0.513±0.045 in the si-NC group,si-TROAP-1 group,and si-TROAP-2 group,respectively.A substantial statistical difference was seen between the groups(F=55.510,P＜0.001).The clone formation experiment yielded the following results:the si-NC group had 59.330±4.040 cell clones,the si-TROAP-1 group had 17.330±2.517 cell clones,and the si-TROAP-2 group had 21.000±5.292 cell clones.There was a statistically significant difference between the groups(F=96.130,P＜0.001).The cell apoptosis data demonstra-ted that the apoptosis rates in the si-NC group,si-TROAP-1 group,and si-TROAP-2 group were(12.390±1.072)％,(37.850±6.914)％and(33.520±2.275)％,respectively.There was a statistically significant difference in apoptosis rates across the groups(F=30.860,P＜0.001).The wound healing assay results indicated that the cell healing rates at 24 hours were(61.790±1.524)％in the si-NC group,(23.320±0.386)％in the si-TROAP-1 group,and(23.520±1.781)％in the si-TROAP-2 group.A statistically significant difference was observed between the groups,with a large effect size(F=782.500,P＜0.001).The findings of the Transwell invasion experiment showed that the number of cells that entered the chamber in the si-NC group,si-TROAP-1 group,and si-TROAP-2 group were 100.000±4.847,43.850±3.026,and 39.540±6.482,respectively.Furthermore,there was a notable and meaningful distinction observed be-tween the groups,as indicated by the statistical analysis(F=137.200,P＜0.001).TROAP was potentially implicated in cellular processes such as the cell cycle and DNA replication pathways,with a false discovery rate(FDR)below 0.05 and a P value below 0.05.Conclusions Patients with lung adenocarcinoma who have a high expression of TROAP are more likely to have a bad prognosis.Suppressing the expression of TROAP can impede the growth,infiltration,and movement of A549 lung cancer cells.Furthermore,the presence of TROAP in tumor-infiltrating immune cells indicates a potential correlation,implying that TROAP could be utilised as a prognostic indicator for patients with lung adenocarcinoma.
NETL
NSTL
万方数据
