Objective To study the effect of neurotrophic tyrosine kinase receptor B(TrKB) on in vitro invasiveness of malignant transformed hFOB1.19 cells and the role of TrKB in invasion and metastasis of osteosarcoma.Methods Expression of TrKB in malignant transformed hFOB1.19 cells and SaOS-2 osteosarcoma cells was detected by RT-PCR and immunofluorescence.Function of TrKB of malignant transformed hFOB1.19 cells was further studied.Malignant transformed hFOB1.19 cells were treated with specific tyrosine kinase inhibitor K252a for 24 h as a treatment group,and untreated malignant transformed hFOB1.19 cells into which DMSO was added served as a control group.Morphology of cells was observed under an inverted phase contrast microscope.Cell invasiveness was detected by Transwell assays.Microfilaments of cells were detected by actin immunofluorescence.Results The expression of TrKB was significantly higher in malignant transformed hFOB1.19 cells than in SaOS-2 osteosarcoma cells(P

Objective To investigate the effect and underlying mechanism of roxadustat on apoptosis and senescence of renal tubular epithelial cell line HK-2 induced by heat stress.Methods After HK-2 cells were treated with different concentrations of roxadustat(10,20,30,40 and 50 μmol/L)for 24 h,CCK-8 assay was used to determine the optimal intervention concentration of roxadustat.HK-2 cells were divided into 4 groups(n=3):control group,roxadustat group(30 μmol/L,24 h),heat-stress group(43 ℃,2 h),and heat-stress+roxadustat group(30 μmol/L roxadustat treatmnet for 24 h followed by heat-stress 2 h).Cell viability was detected by CCK-8 assay.Expression of hypoxia-inducible factor-1α(HIF-1α),Cleaved Caspase-3,p16 and p21 at protein level was detected by Western blotting.Immunofluorescence assay was employed to observe the distribution of HIF-1α.β-galactosidase staining kit was utilized to detect SA-β-Gal activity.TUNEL staining was used to measure cell apoptosis.Results The highest cell viability was observed in the cells after 30 μmol/L roxadustat treatment.Heat stress resulted in a significant decrease in cell viability(P＜0.05),elevated protein levels of HIF-1α,Cleaved Caspase-3,p16 and p21(P＜0.05),enhanced SA-β-Gal activity(P＜0.05)and increased percentage of TUNEL-positive cells(P＜0.05)when compared with the cells in the control group.In comparison with the heat-stress group,the heat-stress+roxadustat group showed significant decrease in the protein levels of Cleaved Caspase-3,p16 and p21(P＜0.05),reduced activity of SA-β-Gal[(65.44±5.00)％vs(77.15±2.61)％,P＜0.05]and decreased percentage of TUNEL-positive cells[(16.73±2.20)％vs(46.40±13.87)％,P＜0.05],but increase in cell viability[(86.33±4.51)％vs(66.33±8.50)％,P＜0.05]as well as HIF-1α protein expression(P＜0.05).Furthermore,immunofluorescence assay showed that HIF-1α was mainly distributed in the nucleus and perinucleus.Conclusion Roxadustat attenuates heat stress-induced apoptosis and senescence of renal tubular epithelial cells by upregulating HIF-1α.

The heterogeneity of exhausted T cells (Tex) is a critical determinant of immune checkpoint blockade therapy efficacy. However, few studies have explored exhausted T cell subpopulations in human cancers. In the present study, we examined samples from two cohorts of 175 patients with head and neck squamous cell cancer (HNSCC) by multiplex immunohistochemistry (mIHC) to investigate two subsets of Tex, CD8⁺PD1⁺TCF1⁺ progenitor exhausted T cells (TCF1⁺Tex^prog) and CD8⁺PD1⁺TCF1^- terminally exhausted T cells (TCF1^-Tex^term). Moreover, fresh tumor samples from 34 patients with HNSCC were examined by flow cytometry and immunohistochemistry to further investigate their properties and cytotoxic capabilities and their correlation with regulatory T cells (Tregs) in the tumor immune microenvironment (TIME). mIHC and flow cytometry analysis showed that TCF1^-Tex^term represented a greater proportion of CD8⁺PD1⁺Tex than TCF1⁺Tex^prog in most patients. TCF1⁺Tex^prog produced abundant TNFα, while TCF1^-Tex^term expressed higher levels of CD103, TIM-3, CTLA-4, and TIGIT. TCF1^-Tex^term exhibited a polyfunctional TNFα⁺GZMB⁺IFNγ⁺ phenotype; and were associated with better overall survival and recurrence-free survival. The results also indicated that larger proportions of TCF1^-Tex^term were accompanied by an increase in the proportion of Tregs. Therefore, it was concluded that TCF1^-Tex^term was the major CD8⁺PD1⁺Tex subset in the HNSCC TIME and that these cells favor patient survival. A high proportion of TCF1^-Tex^term was associated with greater Treg abundance.
CD8-Positive T-Lymphocytes ; Head and Neck Neoplasms/therapy* ; Humans ; Immunotherapy/methods* ; Prognosis ; Programmed Cell Death 1 Receptor ; Squamous Cell Carcinoma of Head and Neck/therapy* ; Tumor Microenvironment ; Tumor Necrosis Factor-alpha