OBJECTIVETo investigate the effect of lonidamine on apoptosis of human breast carcinoma cells MCF-7 and the possible mechanisms.

METHODSMTT assay and colony-forming assay were used to evaluate the growth inhibition induced by lonidamine in breast cancer MCF-7 cells. PI/Annexin-V staining was used to detect the apoptotic cells. The ATP levels in the cells were detected using an ATP assay kit. The expression of glucose regulated protein 78 (GRP78), inhibitor of apoptosis protein (cIAP1) and caspase-8 were analyzed with Western blotting.

RESULTSMTT assay and colony-forming assay showed that 50-250 mmol/L lonidamine caused a time- and concentration-dependent inhibition of MCF-7 cell proliferation. Exposure to increased concentrations of lonidamine resulted in significantly increased apoptosis rate in MCF-7 cells. In MCF-7 cells treated with 50, 150 and 250 mmol/L lonidamine for 5 h, the intracellular ATP levels were lowered to 80.67%, 62.78% and 30.73% of the control level, respectively. Western blotting showed that lonidamine up-regulated the expression of GRP78, down-regulated the expression of cIAP1 and promoted caspase-8 activation as the treatment time extended.

CONCLUSIONLonidamine can inhibit the proliferation and induce apoptosis in MCF-7 cells, and these effects are probably mediated by reducing ATP level, inducing endoplasmic reticulum stress response, down-regulating cIAP1, and promoting caspase-8 activation in the cells.

Apoptosis ; drug effects ; Breast Neoplasms ; metabolism ; pathology ; Caspase 8 ; metabolism ; Cell Proliferation ; Down-Regulation ; Endoplasmic Reticulum Stress ; Heat-Shock Proteins ; metabolism ; Humans ; Indazoles ; pharmacology ; Inhibitor of Apoptosis Proteins ; metabolism ; MCF-7 Cells ; drug effects ; Ubiquitin-Protein Ligases ; metabolism ; Up-Regulation

Objective To investigate the effect of lonidamine on apoptosis of human breast carcinoma cells MCF-7 and the possible mechanisms. Methods MTT assay and colony-forming assay were used to evaluate the growth inhibition induced by lonidamine in breast cancer MCF-7 cells. PI/Annexin-V staining was used to detect the apoptotic cells. The ATP levels in the cells were detected using an ATP assay kit. The expression of glucose regulated protein 78 (GRP78), inhibitor of apoptosis protein (cIAP1) and caspase-8 were analyzed with Western blotting. Results MTT assay and colony-forming assay showed that 50-250 mmol/L lonidamine caused a time- and concentration-dependent inhibition of MCF-7 cell proliferation. Exposure to increased concentrations of lonidamine resulted in significantly increased apoptosis rate in MCF-7 cells. In MCF-7 cells treated with 50, 150 and 250 mmol/L lonidamine for 5 h, the intracellular ATP levels were lowered to 80.67%, 62.78%and 30.73%of the control level, respectively. Western blotting showed that lonidamine up-regulated the expression of GRP78, down-regulated the expression of cIAP1 and promoted caspase-8 activation as the treatment time extended. Conclusion Lonidamine can inhibit the proliferation and induce apoptosis in MCF-7 cells, and these effects are probably mediated by reducing ATP level, inducing endoplasmic reticulum stress response, down-regulating cIAP1, and promoting caspase-8 activation in the cells.

Objective To investigate the effect of lonidamine on apoptosis of human breast carcinoma cells MCF-7 and the possible mechanisms. Methods MTT assay and colony-forming assay were used to evaluate the growth inhibition induced by lonidamine in breast cancer MCF-7 cells. PI/Annexin-V staining was used to detect the apoptotic cells. The ATP levels in the cells were detected using an ATP assay kit. The expression of glucose regulated protein 78 (GRP78), inhibitor of apoptosis protein (cIAP1) and caspase-8 were analyzed with Western blotting. Results MTT assay and colony-forming assay showed that 50-250 mmol/L lonidamine caused a time- and concentration-dependent inhibition of MCF-7 cell proliferation. Exposure to increased concentrations of lonidamine resulted in significantly increased apoptosis rate in MCF-7 cells. In MCF-7 cells treated with 50, 150 and 250 mmol/L lonidamine for 5 h, the intracellular ATP levels were lowered to 80.67%, 62.78%and 30.73%of the control level, respectively. Western blotting showed that lonidamine up-regulated the expression of GRP78, down-regulated the expression of cIAP1 and promoted caspase-8 activation as the treatment time extended. Conclusion Lonidamine can inhibit the proliferation and induce apoptosis in MCF-7 cells, and these effects are probably mediated by reducing ATP level, inducing endoplasmic reticulum stress response, down-regulating cIAP1, and promoting caspase-8 activation in the cells.

convalescents, and to screen for broad-spectrum neutralizing antibodies against the SARS-CoV-2 RBD. Methods Using biotinylated RBD as a molecular probe, flow cytometry was employed to perform single-cell sorting of B cells from peripheral blood mononuclear cells (PBMCs) of convalescents. The obtained B cells were lysed and subjected to reverse transcription, followed by nested PCR amplification of the heavy and light chains of antibodies was conducted using random primers. The amplified products were cloned into corresponding expression vectors, and the respective matched heavy-light chain plasmids were co-transfected into 293F cells for expression. Monoclonal antibodies were then purified using Protein A column chromatography. Neutralization experiments were conducted with the wild-type (WT) pseudovirus, and antibodies with IC50<0.1 μg/mL were selected for further testing of neutralizing breadth and potency against the wild-type (WT), Beta variant (B.1.351), Delta variant (B.1.617.2), and currently prevalent pseudovirus strains (XBB, BA.5, BF.7). Results A total of 21 RBD-specific monoclonal B cells were obtained from two recovered patients, resulting in the isolation of 13 pairs of antibody light/heavy chains. Nine antibodies were successfully expressed, with P1-A1, P1-B6, and P1-B9 exhibiting IC50 values below 0.1 μg/mL against the pseudovirus of the wild-type strain (WT). Specifically, P1-B6 effectively neutralized the wild-type strain (WT), Beta variant (B.1.351), and Delta variant (B.1.617.2), with IC50 values reaching 0.01 μg/mL. P1-B9 demonstrated effective neutralization against the wild-type strain (WT), Beta variant (B.1.351), Delta variant (B.1.617.2), and Gamma variant (P.1) pseudoviruses, with IC50 values of 0.42 μg/mL, 0.63 μg/mL, 0.28 μg/mL, and 2.50 μg/mL, respectively. Additionally, P1-B6 exhibited good neutralization against BA.5 and BF.7 pseudoviruses, with IC50 values of 0.06 μg/mL and 0.09 μg/mL, respectively. Conclusions Infection with the SARS-CoV-2 WT strain can induce the generation of neutralizing antibodies with broad-spectrum activity. Generating these broadly neutralizing antibodies does not require an excessively high somatic hypermutation. The obtained antibodies can be used as candidates for SARS-CoV-2 diagnosis and prevention.

OBJECTIVE: To study the clinical effect and mechanism of total glucosides of paeony (TGP) in the adjuvant therapy for children with Henoch-Schönlein purpura nephritis (HSPN). METHODS: Sixty-four HSPN children with moderate proteinuria were divided into a TGP treatment group ( RESULTS: Compared with the healthy children before treatment, the children with HSPN had higher proportion of Tfh cells and expression levels of IL-21 and IL-4 ( CONCLUSIONS TGP has a marked clinical effect in the treatment of HSPN and can reduce the inflammatory response of the kidney and exert a protective effect on the kidney by inhibiting the proliferation of Tfh cells and downregulating the expression of IL-21 and IL-4 in plasma.
Child ; Glucosides/therapeutic use* ; Humans ; Nephritis ; Paeonia ; Prospective Studies ; Purpura, Schoenlein-Henoch/drug therapy*

The development of new biomarkers or therapeutic targets for cancer immunotherapies requires deep under-standing of T cells. To date, the complete landscape and systematic characterization of long noncoding RNAs (lncRNAs) in T cells in cancer immunity are lacking. Here, by systematically analyzing full-length single-cell RNA sequencing (scRNA-seq) data of more than 20,000 libraries of T cells across three cancer types, we provided the first comprehensive catalog and the functional repertoires of lncRNAs in human T cells. Specifically, we developed a custom pipeline for de novo transcriptome assembly and obtained a novel lncRNA catalog containing 9433 genes. This increased the number of current human lncRNA catalog by 16％and nearly doubled the number of lncRNAs expressed in T cells. We found that a portion of expressed genes in single T cells were lncRNAs which had been overlooked by the majority of previous studies. Based on metacell maps constructed by the MetaCell algorithm that partitions scRNA-seq datasets into disjointed and homogenous groups of cells (metacells), 154 signature lncRNA genes were identified. They were associated with effector, exhausted, and regulatory T cell states. Moreover, 84 of them were functionally annotated based on the co-expression networks, indicating that lncRNAs might broadly participate in the regulation of T cell functions. Our findings provide a new point of view and resource for investigating the mechanisms of T cell regulation in cancer immunity as well as for novel cancer-immune biomarker development and cancer immunotherapies.