BACKGROUND: T-cell lymphoblastic lymphoma/acute lymphoblastic leukemia (T-LBL/ALL) is an aggressive form of hematological malignancy associated with poor prognosis in adult patients. Histone deacetylases (HDACs) are aberrantly expressed in T-LBL/ALL and are considered potential therapeutic targets. Here, we investigated the antitumor effect of a novel HDAC inhibitor, chidamide, on T-LBL/ALL. METHODS: HDAC1, HDAC2 and HDAC3 levels in T-LBL/ALL cell lines and patient samples were compared with those in normal controls. Flow cytometry, transmission electron microscopy, and lactate dehydrogenase release assays were conducted in Jurkat and MOLT-4 cells to assess apoptosis and pyroptosis. A specific forkhead box O1 (FOXO1) inhibitor was used to rescue pyroptosis and upregulated gasdermin E (GSDME) expression caused by chidamide treatment. The role of the FOXO1 transcription factor was evaluated by dual-luciferase reporter and chromatin immunoprecipitation assays. The efficacy of chidamide in vivo was evaluated in a xenograft mouse. RESULTS: The expression of HDAC1, HDAC2 and HDAC3 was significantly upregulated in T-LBL/ALL. Cell viability was obviously inhibited after chidamide treatment. Pyroptosis, characterized by cell swelling, pore formation on the plasma membrane and lactate dehydrogenase leakage, was identified as a new mechanism of chidamide treatment. Chidamide triggered pyroptosis through caspase 3 activation and GSDME transcriptional upregulation. Chromatin immunoprecipitation assays confirmed that chidamide led to the increased transcription of GSDME through a more relaxed chromatin structure at the promoter and the upregulation of FOXO1 expression. Moreover, we identified the therapeutic effect of chidamide in vivo . CONCLUSIONS This study suggested that chidamide exerts an antitumor effect on T-LBL/ALL and promotes a more inflammatory form of cell death via the FOXO1/GSDME axis, which provides a novel choice of targeted therapy for patients with T-LBL/ALL.
Humans ; Pyroptosis/drug effects* ; Forkhead Box Protein O1/genetics* ; Aminopyridines/pharmacology* ; Animals ; Mice ; Benzamides/pharmacology* ; Cell Line, Tumor ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy* ; Phosphate-Binding Proteins/metabolism* ; Histone Deacetylase Inhibitors/pharmacology* ; Jurkat Cells ; Histone Deacetylases/metabolism* ; Apoptosis/drug effects* ; Gasdermins

Objective To investigate the impact of Qizhi Jiangtang Capsule (QZJT) on renal damage in diabetic nephropathy (DN) mice via NOD like receptors family pyrin domain containing 3/caspase-1/ Gasdermin D (NLRP3/caspase-1/GSDMD) signaling pathway. Methods Mice were randomly allocated into six experimental groups: a normal control group (NC), a diabetic nephropathy model group (DN), a low-dose QZJT treatment group (L-QZJT), a high-dose QZJT treatment group (H-QZJT), a positive control group administered Shenqi Jiangtang Granules (SQJT), and an ML385 group (treated with an inhibitor of nuclear factor erythroid 2-related factor 2, Nrf2). Upon successful model induction, therapeutic interventions were commenced. Renal function impairment in the mice was evaluated through quantification of fasting blood glucose (FBG), 24-hour urinary albumin (UAlb), serum creatinine (SCr), blood urea nitrogen (BUN), and the kidney-to-body mass ratio (K/B). Renal tissue pathology was evaluated using HE and PAS staining. Serum levels of inflammatory cytokines IL-1β and IL-18 were quantified by ELISA. Levels of podocyte markers and proteins involved in relevant pathways were assessed using Western blot analysis. Results Compared with the NC group, FBG, 24 h UAlb, SCr, and BUN were increased in the DN group, and the K/B mass ratio was also increased. In contrast, compared with the DN group, FBG, 24 h UAlb, SCr, and BUN in both the low-dose (L-QZJT) and high-dose Quanzhou Jintang (H-QZJT) groups were decreased, and the K/B mass ratio was decreased as well. The therapeutic efficacy of H-QZJT was comparable to that of Shenqi Jiangtang Granules. QZJT ameliorated renal histopathological injury in DN mouse, increased the protein levels of Nephrin (a podocyte marker), and decreased the protein levels of NLRP3, apoptosis-associated speck-like protein containing CARD (ASC), pro-caspase-1, and GSDMD-N. After ML385 treatment, renal cells exhibited swelling and morphological changes, the inflammatory infiltrate area was enlarged, the protein levels of NLRP3, ASC, pro-caspase-1, and GSDMD-N were up-regulated, and the levels of IL-1β and IL-18 were increased. Conclusion QZJT may inhibit podocyte pyroptosis by acting on the Nrf2 to regulate the NLRP3/caspase-1/GSDMD pathway, thus improving renal damage in DN mouse.
Animals ; Diabetic Nephropathies/pathology* ; Podocytes/pathology* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Pyroptosis/drug effects* ; Drugs, Chinese Herbal/administration & dosage* ; Caspase 1/genetics* ; Signal Transduction/drug effects* ; Mice ; Phosphate-Binding Proteins/genetics* ; Male ; Intracellular Signaling Peptides and Proteins/metabolism* ; Mice, Inbred C57BL ; Kidney/pathology* ; Gasdermins

This study established a pyroptosis injury model by stimulating insulinoma cells(INS-1) of rats with high glucose(HG) and observed the impact of additional ethanol(ET) exposure on cell pyroptosis, as well as the intervention effect of salidroside(SAL). INS-1 cells were cultured and divided into a normal control group(NG), an HG group, an HG + ET(100 mmol·L~(-1)) group, and an HG + ET + SAL(1-100 μmol·L~(-1)) group. After 72 hours of treatment, cell viability was assessed using the cell counting kit-8(CCK-8) assay. The number of pyroptotic bodies was observed under a microscope. Western blot was used to detect changes in the intracellular Nod-like receptor protein 3(NLRP3)/gasdermin D(GSDMD) signaling pathway and adenosine monophosphate-activated protein kinase(AMPK) activity. A fluorescence probe was used to detect changes in intracellular reactive oxygen species(ROS) levels. Time-resolved fluorescence resonance energy transfer(TR-FRET) technology was employed to observe the effect of SAL on recombinant AMPK protein kinase activity in vitro. The results showed that compared to the NG group, HG exposure induced an increase in the number of pyroptotic bodies, elevated ROS levels, and activation of the NLRP3/GSDMD signaling pathway in INS-1 cells. Compared to the HG group, HG + ET exposure further exacerbated these changes. Compared to the HG + ET group, SAL dose-dependently increased cell viability, reduced the formation of pyroptotic bodies in INS-1 cells, and inhibited excessive ROS production, overactivation of the NLRP3/GSDMD signaling pathway, and the decrease in AMPK activity. TR-FRET experiments indicated that SAL could directly activate AMPK. When INS-1 cells were pretreated with an AMPK inhibitor, the effects of SAL on increasing cell viability, alleviating the formation of pyroptotic bodies, and inhibiting excessive ROS production were abolished. These results suggest that SAL can alleviate HG combined with ET-induced exacerbation of INS-1 pyroptosis by activating AMPK.
Pyroptosis/drug effects* ; Animals ; Rats ; Glucose/metabolism* ; Insulinoma/metabolism* ; Ethanol/pharmacology* ; Reactive Oxygen Species/metabolism* ; Glucosides/pharmacology* ; Phenols/pharmacology* ; Cell Line, Tumor ; Signal Transduction/drug effects* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Cell Survival/drug effects* ; AMP-Activated Protein Kinases/metabolism* ; Phosphate-Binding Proteins/genetics*

To investigate the therapeutic effect of Fuzheng Tongluo Granules on idiopathic pulmonary fibrosis(IPF) and its mechanism. Seventy-two SD rats were randomly divided into the control group, model group, pirfenidone group(162 mg·kg~(-1)), and low-, medium-and high-dose of Fuzheng Tongluo Granules groups(2.63, 5.25, 10.5 g·kg~(-1)). Rat model of IPF was induced by a single non-invasive tracheal intubation drip of bleomycin(BLM). The corresponding drugs were given daily by gavage after the 2nd day of modeling, and body mass was recorded. On the 28th day, the samples were collected and weighed, and the lung coefficients were calculated. The pathological changes in the lung tissue were observed by HE and Masson staining, and the hydroxyproline(HYP) content of the lung tissue was detected by alkaline hydrolysis. The contents of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), and interleukin-18(IL-18) of the lung tissue were determined by ELISA. The expression of collagen type Ⅰ(collagen Ⅰ) and α-smooth muscle actin(α-SMA) was observed by immunohistochemistry. The expression levels of NOD-, LRR-and pyrin domain-containing 3(NLRP3), cysteine-requiring aspartate protease type 1(caspase-1), gasdermin D-N(GSDMD-N), and apoptosis-associated speck-like protein containing a CARD(ASC) in the lung tissue were detected by Western blot. Immunofluorescence co-localization was used to observe the expression of GSDMD and CD68. The results show that compared with the control group, the model group showed increased lung coefficient, Ashcroft score, Szapiel score, HYP, TNF-α, IL-1β, and IL-18 content in the lung tissue and elevated protein expression levels of NLRP3, caspase-1, GSDMD-N, and ASC. The expression levels of GSDMD and CD68 were increased, and there was a high degree of co-localization between GSDMD and CD68. Compared with those in the model group, the lung coefficient, Ashcroft score, and Szapiel score decreased in all drug administration groups, and the content of HYP, TNF-α, IL-1β, and IL-18 decreased. The protein expression levels of NLRP3, caspase-1, GSDMD-N, and ASC decreased, and the expression levels of GSDMD and CD68 were reduced. There was a high degree of co-localization between GSDMD and CD68. In summary, Fuzheng Tongluo Granules can effectively reduce pulmonary fibrosis and inflammation levels in rats with IPF, and the mechanism may be related to the down-regulation of the NLRP3/caspase-1/GSDMD pathway to inhibit macrophage pyroptosis.
Animals ; Rats, Sprague-Dawley ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Pyroptosis/drug effects* ; Rats ; Drugs, Chinese Herbal/administration & dosage* ; Male ; Caspase 1/genetics* ; Disease Models, Animal ; Macrophages/metabolism* ; Signal Transduction/drug effects* ; Pulmonary Fibrosis/metabolism* ; Lung/metabolism* ; Phosphate-Binding Proteins/metabolism* ; Humans ; Intracellular Signaling Peptides and Proteins/genetics* ; Gasdermins

The transition from spermatogonia to spermatocytes and the initiation of meiosis are key steps in spermatogenesis and are precisely regulated by a plethora of proteins. However, the underlying molecular mechanism remains largely unknown. Here, we report that Src homology domain tyrosine phosphatase 2 (Shp2; encoded by the protein tyrosine phosphatase, nonreceptor type 11 [Ptpn11] gene) is abundant in spermatogonia but markedly decreases in meiotic spermatocytes. Conditional knockout of Shp2 in spermatogonia in mice using stimulated by retinoic acid gene 8 (Stra8)-cre enhanced spermatogonial differentiation and disturbed the meiotic process. Depletion of Shp2 in spermatogonia caused many meiotic spermatocytes to die; moreover, the surviving spermatocytes reached the leptotene stage early at postnatal day 9 (PN9) and the pachytene stage at PN11-13. In preleptotene spermatocytes, Shp2 deletion disrupted the expression of meiotic genes, such as disrupted meiotic cDNA 1 (Dmc1), DNA repair recombinase rad51 (Rad51), and structural maintenance of chromosome 3 (Smc3), and these deficiencies interrupted spermatocyte meiosis. In GC-1 cells cultured in vitro, Shp2 knockdown suppressed the retinoic acid (RA)-induced phosphorylation of extracellular-regulated protein kinase (Erk) and protein kinase B (Akt/PKB) and the expression of target genes such as synaptonemal complex protein 3 (Sycp3) and Dmc1. Together, these data suggest that Shp2 plays a crucial role in spermatogenesis by governing the transition from spermatogonia to spermatocytes and by mediating meiotic progression through regulating gene transcription, thus providing a potential treatment target for male infertility.
Animals ; Cell Cycle Proteins/genetics* ; Cell Line ; Cell Survival ; Chondroitin Sulfate Proteoglycans/genetics* ; Chromosomal Proteins, Non-Histone/genetics* ; Gene Expression Regulation ; Gene Knockdown Techniques ; Infertility, Male ; Male ; Meiosis/genetics* ; Mice ; Mice, Knockout ; Mice, Transgenic ; Phosphate-Binding Proteins/genetics* ; Protein Tyrosine Phosphatase, Non-Receptor Type 11/genetics* ; Rad51 Recombinase/genetics* ; Real-Time Polymerase Chain Reaction ; Spermatocytes/metabolism* ; Spermatogenesis/genetics* ; Spermatogonia/metabolism*

In recent years, human cancer genome projects provide unprecedented opportunities for the discovery of cancer genes and signaling pathways that contribute to tumor development. While numerous gene mutations can be identified from each cancer genome, what these mutations mean for cancer is a challenging question to address, especially for those from less understood putative new cancer genes. As a powerful approach, in silico bioinformatics analysis could efficiently sort out mutations that are predicted to damage gene function. Such an analysis of human large tumor suppressor genes, LATS1 and LATS2, has been carried out and the results support a role of hLATS1//2 as negative growth regulators and tumor suppressors.
Adaptor Proteins, Signal Transducing ; chemistry ; metabolism ; Animals ; Carrier Proteins ; chemistry ; metabolism ; Computational Biology ; Genes, Neoplasm ; Humans ; LIM Domain Proteins ; chemistry ; metabolism ; Mice ; Mutation ; Neoplasms ; genetics ; pathology ; Phosphoproteins ; chemistry ; metabolism ; Phosphorylation ; Protein Binding ; Protein Structure, Tertiary ; Protein-Serine-Threonine Kinases ; chemistry ; genetics ; metabolism ; Transferases (Other Substituted Phosphate Groups) ; chemistry ; metabolism ; Tumor Suppressor Proteins ; chemistry ; genetics ; metabolism

OBJECTIVETo Investigate the differences of sorbitol fermentation related genes and optimize molecular analysis method for distinguishing an epidemic with nonepidemic strains of Vibrio cholerae.

METHODSSequence analysis on four genes of sugar fermentation stimulation protein, periplasmic maltose-binding protein, periplasmic phosphate-binding protein and periplasmic amino acid-binding protein.

RESULTSIn this study, the following data was noticed: for O1 serogroup El Tor biotype V. cholerae, twenty-four epidemic and eight nonepidemic strains were chosen; For O139 serogroup V. cholerae, five epidemic and four nonepidemic strains were chosen. With those genes of sugar fermentation stimulation protein, there were three point mutations. The 106th, 150th, 378th oligonucleotide in epidemic strains were A, A and T, comparing to the nonepidemic strains which were G, G and C. When comparing the protein sequences, epidemic strains had a Threonine at 36th amino acid, whereas nonepidemic strains had an Alanine. The results in O139 serogroup were consistent with those in O1 serogroup El Tor biotype strains. Another two point mutations were found in the genes of periplasmic maltose-binding protein. The 999th, 1003rd oligonucleotides in epidemic strains were A and C, while in nonepidemic which were G and T. For the gene of periplasmic amino acid-binding protein, two point mutations were noticed. The 504th and 690th oligonucleotides in epidemic strains were T and C, but were C and T in nonepidemic. However, no amino acid differences were found in periplasmic maltose-binding protein and periplasmic amino acid-binding protein. For periplasmic amino acid-binding protein gene, there was no difference on oligonucleotide between epidemic and nonepidemic strains.

CONCLUSIONResults suggested that SNPs in these genes might serve as a useful tool to distinguish the epidemic strains from nonepidemic strains. The 36th amino acid mutation of sugar fermentation stimulation protein in epidemic and nonepidemic strains might change the activity of the protein which might be associated with sorbitol fermentation.

Amino Acid Sequence ; Bacterial Proteins ; genetics ; metabolism ; Base Sequence ; Carrier Proteins ; genetics ; metabolism ; Fermentation ; Maltose-Binding Proteins ; Molecular Sequence Data ; Periplasmic Binding Proteins ; genetics ; metabolism ; Phosphate-Binding Proteins ; genetics ; metabolism ; Point Mutation ; Sequence Analysis, Protein ; Sorbitol ; Vibrio cholerae ; genetics ; metabolism