Objective To establish a flow cytometry method for detecting C1s protein on platelet surface and preliminarily explore its potential application value in the auxiliary diagnosis of primary immune thrombocytopenia(ITP).Methods C1s-conjugated 2 μm car-boxylated magnetic beads(C1s beads)were prepared and used as quality control particles.Fluorescein isothiocyanate(FITC)-labeled anti-C1s antibody was employed as the detection antibody to develop a flow cytometric assay for detecting C1s protein expression on platelets.The intra-assay and inter-assay precision,as well as the dilution linearity of the method,were evaluated.Subsequently,the expression levels of C1 s protein on the surface of platelets were compared among the ITP group,the non-ITP thrombocytopenia group,and the healthy control group.Results Light microscopy showed that both unconjugated carboxylated magnetic beads(blank beads)and C1s-conjugated beads were uniformly dispersed without aggregation.Under fluorescence microscopy,C1s beads exhibited strong yellow-green fluorescence,whereas the blank beads showed no fluorescence signal.The established flow cytometry assay exhibited ac-ceptable precision,with intra-assay coefficient of variation(CV)values of 7.02％,7.12％,and 3.91％for low,medium,and high con-centrations of C1s beads,respectively,and inter-assay CV values of 13.49％,6.15％,and 0.78％,respectively.The dilution linearity was satisfactory,coefficient of determination(R2)=0.998 8.Clinical sample testing revealed that the proportion of C1s-positive plate-lets in ITP group(2.56±0.79)％was significantly higher than that in healthy control group(0.23±0.18)％and the non-ITP thrombo-cytopenia control group(0.22±0.10)％,with statistically significant differences(both P＜0.05).Conclusion This study successfully established a stable and reliable flow-cytometry method for quantifying C1s expression on platelet surface and preliminarily demonstrated that C1s expression is significantly elevated on platelets of ITP patients,suggesting that C1s could serve as a potential auxiliary diag-nostic marker for ITP.

Objective To establish a flow cytometry method for detecting C1s protein on platelet surface and preliminarily explore its potential application value in the auxiliary diagnosis of primary immune thrombocytopenia(ITP).Methods C1s-conjugated 2 μm car-boxylated magnetic beads(C1s beads)were prepared and used as quality control particles.Fluorescein isothiocyanate(FITC)-labeled anti-C1s antibody was employed as the detection antibody to develop a flow cytometric assay for detecting C1s protein expression on platelets.The intra-assay and inter-assay precision,as well as the dilution linearity of the method,were evaluated.Subsequently,the expression levels of C1 s protein on the surface of platelets were compared among the ITP group,the non-ITP thrombocytopenia group,and the healthy control group.Results Light microscopy showed that both unconjugated carboxylated magnetic beads(blank beads)and C1s-conjugated beads were uniformly dispersed without aggregation.Under fluorescence microscopy,C1s beads exhibited strong yellow-green fluorescence,whereas the blank beads showed no fluorescence signal.The established flow cytometry assay exhibited ac-ceptable precision,with intra-assay coefficient of variation(CV)values of 7.02％,7.12％,and 3.91％for low,medium,and high con-centrations of C1s beads,respectively,and inter-assay CV values of 13.49％,6.15％,and 0.78％,respectively.The dilution linearity was satisfactory,coefficient of determination(R2)=0.998 8.Clinical sample testing revealed that the proportion of C1s-positive plate-lets in ITP group(2.56±0.79)％was significantly higher than that in healthy control group(0.23±0.18)％and the non-ITP thrombo-cytopenia control group(0.22±0.10)％,with statistically significant differences(both P＜0.05).Conclusion This study successfully established a stable and reliable flow-cytometry method for quantifying C1s expression on platelet surface and preliminarily demonstrated that C1s expression is significantly elevated on platelets of ITP patients,suggesting that C1s could serve as a potential auxiliary diag-nostic marker for ITP.

To explore the clinical effect of saxagliptin on release of peripheral blood neutrophil-to-lymphocyte ratio(NLR)in elder patients with type 2 diabetes mellitus. A total of 110 newly-diagnosed elder patients with type 2 diabetes were assigned into 2 groups(both n=55). Trial group was treated with exercise-diet therapy and saxagliptin at 5 mg/d(SG group),while control group with exercise-diet therapy and repaglinide(RG group). NLR were evaluated in both groups before treatment without significant difference(P>0.05). After 12 weeks'treatment, NLR were both lowered. Decrease of NLR in SG group was more significant than that of RG group(P<0.05).

Aim To observe the effects of heat shock protein 70 ( Hsp70 ) activator SW02 on lipopolysaccha-ride( LPS)-induced expression of inducible nitric oxide synthase ( iNOS ) and LPS-induced production of nitric oxide ( NO ) in macrophages. Methods RAW264. 7 cells were stimulated by LPS, and were divided into DMSO,DMSO+LPS(1 mg·L-1),SW02,and SW02+LPS ( 1 mg · L-1 ) groups. The protein expression was detected by Western blot. NO concentration was measured by Griess kit. The iNOS mRNA was detected by real-time PCR. The NF-κB binding to iNOS promot-ers was measured by chromatin immunoprecipitation ( ChIP ) assays. Results SW02 significantly blocked the protein and mRNA expression of iNOS as well as the production of NO in LPS-stimulated RAW264 . 7 cells(P<0. 01 or P<0. 05,SW02+LPS group vs DM-SO+LPS group) . SW02 did not affect the LPS-induced degradation of IκB-α( P>0. 05 , SW02 +LPS group vs DMSO+LPS group ) and nuclear translocation of NF-κB ( P > 0. 05 , SW02 + LPS group vs DMSO + LPS group) . However,SW02 reduced the NF-κB binding to iNOS promoters inside the cell( P<0. 05,SW02+LPS group vs DMSO+LPS group) . Conclusion These re-sults show that SW02 prevents iNOS expression and NO induction likely through attenuation of the NF-κB bind-ing to iNOS promoters in macrophages.