ABSTRACT B cell activation is accompanied by dynamic metabolic reprogramming, supported by a multitude of nutrients that include glucose, amino acids and fatty acids. While several studies have indicated that fatty acid mitochondrial oxidation is critical for immune cell functions, contradictory findings have been reported. Carnitine palmitoyltransferase II (CPT2) is a critical enzyme for long-chain fatty acid oxidation in mitochondria. Here, we test the requirement of CPT2 for humoral immunity using a mouse model with a lymphocyte specific deletion of CPT2. Stable 13 C isotope tracing reveals highly reduced fatty acid-derived citrate production in CPT2 deficient B cells. Yet, CPT2 deficiency has no significant impact on B cell development, B cell activation, germinal center formation, and antibody production upon either thymus-dependent or –independent antigen challenges. Together, our findings indicate that CPT2 mediated fatty acid oxidation is dispensable for humoral immunity, highlighting the metabolic flexibility of lymphocytes.
Stimulation of TLR7/9 by their respective ligands leads to the activation of IκB kinase α (IKKα) and Interferon Regulatory Factor 1 (IRF-1) and results in interferon (IFN)-β production in conventional dendritic cells (cDC). However, which other signaling molecules are involved in IKKα and IRF-1 activation during TLR7/9 signaling pathway are not known. We and others have shown that Bruton's Tyrosine Kinase (BTK) played a part in TLR9-mediated cytokine production in B cells and macrophages. However, it is unclear if BTK participates in TLR7/9-induced IFN-β production in cDC. In this study, we show that BTK is required for IFN-β synthesis in cDC upon TLR7/9 stimulation and that stimulated BTK-deficient cDC are defective in the induction of IKKα/β phosphorylation and IRF-1 activation. In addition, we demonstrate that Protein Kinase C µ (PKCµ) is also required for TLR7/9-induced IRF-1 activation and IFN-β upregulation in cDC and acts downstream of BTK. Taken together, we have uncovered two new molecules, BTK and PKCµ, that are involved in TLR7/9-triggered IFN-β production in cDC.
SUMMARY Degradation by the 26S proteasome is an intricately regulated process fine-tuned by the precise nature of ubiquitin modifications attached to a protein substrate. By debranching ubiquitin chains composed of K48 linkages, the proteasome-associated ubiquitin C-terminal hydrolase UCHL5/UCH37 serves as a positive regulator of protein degradation. How UCH37 achieves specificity for K48 chains is unclear. Here, we use a combination of hydrogen-deuterium mass spectrometry, chemical crosslinking, small-angle X-ray scattering, NMR, molecular docking, and targeted mutagenesis to uncover a cryptic K48 ubiquitin chain specific binding site on the opposite face of UCH37 relative to the canonical S1 ubiquitin-binding site. Biochemical assays demonstrate the K48 chain-specific binding site is required for chain debranching and proteasome-mediated degradation of proteins modified with branched chains. Using quantitative proteomics, translation shutoff experiments, and linkage-specific affinity tools, we then identify specific proteins whose degradation depends on the debranching activity of UCH37. Our findings suggest that UCH37 and potentially other DUBs could use more than one S1 site to perform different biochemical functions.
Gastric cancer (GC) is one of the most malignant tumors that seriously threaten human health. Increased reports have indicated that long noncoding RNAs (lncRNAs) are associated with GC. This study aims to investigate the regulatory role of colon cancer-associated transcript-1 (CCAT1) in GC. The results exhibited the fact that CCAT1 was expressed higher in 57 GC tissue samples than in 57 paired adjacent normal tissue samples. The expression of CCAT1 was also increased in GC cell lines (MKN45, Hs746T, and SGC-7901) compared with the gastric epithelial cell line GES-1. Besides this, decreased cell proliferation with increased cell apoptosis was detected in SGC-7902 cells transfected with CCAT1 short hairpin RNA (shRNA). At the same time, a lower cell invasion ability was measured in SCG-7901 cells transfected with CCAT1 shRNA.In addition, miR-219-1 was predicted and convinced a direct target of CCAT1. The expression of miR-219-1 was decreased in GC tissues and GC cell lines. Further studies demonstrated that the roles of CCAT1 in cell proliferation, apoptosis, and invasion were inhibited by miR-219-1. Finally, in vivo experiment indicated that tumor growth of GC was suppressed through knockdown of CCAT1. In conclusion, these results suggested that CAT1 promotes the tumorigenesis and progression of GC by negatively regulating miR-219-1.
Background High expression of monoamine oxidase A (MAOA) in non‐small cell lung cancer (NSCLC) is related to epithelial‐mesenchymal transition (EMT) and the development of clinicopathological features of NSCLC. Nevertheless, the role of MAOA in drug resistance still remains unclear. Hence, the aim of this article was to evaluate a previously synthesized MAOA inhibitor (G11) on inhibiting paclitaxel‐resistant NSCLC metastasis and growth. Methods First, MAOA expression level was evaluated in several NSCLC cell lines. An MTT assay was used to validate the inhibitory effect of G11 on NSCLC cells in vitro. Second, gene expression in G11‐treated H460/PTX cells was analyzed by microarray gene expression. Third, transwell assay was performed to assess the invasion and metastasis of G11‐treated A549/PTX and H460/PTX cells and western blot assay used to analyze vital protein expression level in G11‐treated H460/PTX cells. Finally, the antimetastatic effect of G11 was tested in an NSCLC in vivo model. Results Our data revealed that G11 significantly inhibited the viability of paclitaxel (PTX)‐resistant NSCLC cell lines (A549/PTX and H460/PTX). G11 dramatically reduced the expression of MAOA in A549/PTX and H460/PTX cells, which exhibited relatively high MAOA expression levels. Additionally, G11 was found to hinder A549/PTX and H460/PTX cell migration and invasion. Furthermore, the in vivo study indicated that the coadministration of G11 and paclitaxel significantly suppressed tumor metastasis in H460/PTX lung metastasis models. Conclusions These findings indicated G11 showed a moderate inhibitory effect on paclitaxel‐resistant NSCLC metastasis and growth, and support further investigation on MAOA potentially as a promising therapeutic target for paclitaxel‐resistant NSCLC treatment. Key points Significant findings of the study Inhibition of MAOA might contribute to the suppression of metastasis and growth in PTX‐resistant NSCLC cells. What this study adds This study explored the potential function of MAOA in drug‐resistant NSCLC and might consider MAOA as a promising target for the treatment of drug‐resistant NSCLC.
Objective To strengthen the monitoring of multi-drug resistant bacteria in order to provide basis for clinically preventing and controlling nosocomial infections of multi-drug resistant bacteria.Methods The drug resistant phenotype of MRS,VRE,ESBLs Escherichia or K.pneumoniae and multi-drug resistant Acinetobacter baumannii were detected by disk diffusion according to the standard document of CLSI2008.The comprehensive measures of prevention and control were taken for infected patients of multi-drug resistant bacteria and colonized bacteria.Results The detectable rate of multi-drug resistant bacteria in 2008,2009,2010 and 2011 were MRS 56.0%,65.0%,56.5%,60.0%;VRE 0%,0%,0%,0%;ESBLs colon bacillus 69.5%,71.0%,61.0%,59.4%;ESBLs pneumonia kleber 44.5%,26.5%,11.0%,7.1%;and multi-drug resistant Acinetobacter baumannii 0%,0%,0%,0%.Conclusion The objective detection of multi-drug resistant bacteria should be strengthened and the comprehensive measures of prevention and control should be implemented by medical institution,in order to effectively prevent and control the spread of multi-drug resistant bacteria among patients in hospital.
Abstract MicroRNA-191 (miR-191) has been identified as being upregulated in several types of cancers, and plays the role of oncogene. The expression of miR-191 has been found to be upregulated in prostate cancer tissues as well as cell lines. In this study, we analyzed the correlation of miR-191 expression with clinicopathologic factors and prognosis in prostate cancer. Prostate cancer tissue samples and adjacent normal prostate tissue samples were collected from 146 patients who underwent laparoscopic radical prostatectomy between April 2013 and March 2018. Student two-tailed t -test was used for comparisons of 2 independent groups. The relationships between miR-191 expression and different clinicopathological characteristics were evaluated using the Chi-squared test. Kaplan–Meier survival plots and log-rank tests were used to assess the differences in overall survival of the different subgroups of prostate cancer patients. miR-191 expression was significantly higher in prostate cancer tissues compared with normal adjacent prostate tissues ( P < .001). miR-191 expression was observed to be significantly correlated with Gleason score ( P < .001), pelvic lymph node metastasis ( P = .006), bone metastases ( P < .001), and T stage ( P = .005). Kaplan–Meier analysis showed that patients with higher levels of miR-191 had significantly poorer survival than those with lower expression of this miRNA in prostate cancer patients (log rank test, P = .011). Multivariate analysis revealed that miR-191 expression (hazard ratio [HR] = 2.311, 95% confidence interval, [CI]: 1.666–9.006; P = .027) was independently associated with the overall survival of prostate cancer patients. Our results demonstrated that miR-191 might serve as an independent prognostic indicator for prostate cancer patients.
Immune cells can metabolize glucose, amino acids, and fatty acids (FAs) to generate energy. The roles of different FA species and their impacts on humoral immunity remain poorly understood. Here, we report that proliferating B cells require monounsaturated FAs (MUFAs) to maintain mitochondrial metabolism and mTOR activity and to prevent excessive autophagy and endoplasmic reticulum (ER) stress. Furthermore, B cell-extrinsic stearoyl-CoA desaturase (SCD) activity generates MUFA to support early B cell development and germinal center (GC) formation in vivo during immunization and influenza infection. Thus, SCD-mediated MUFA production is critical for humoral immunity.
Objective To investigate the antitumor efficacy of intratumoral administration of plasmid expressing Apoptin in athymic mouse T24 bladder tumor exnograft models grafted subcutaneously.Methods Established athymic mouse human bladder tumor transplantation tumor modelby injecting T24 cells subcutaneously;To observe the growth of athymic mouse transplantation tumor,when tumor diameter was about 0.5cm,Apoptin-EGFP plasmid parcled with liposome injected in it,empty plasmid group and physiological saline group were established as control groups,The tumor weight was measured and calculated continually,The mice were killed five weeks later,tumor suppression rate was evaluated and the apoptosis index of cells was detected by in situ TUNEL.Results Established athymic mouse exnograft tumor model successfully,the Apoptin-EGFP group could inhibit the growth of tumor obviously contrasted to the controls,the apoptosis rate was(23.24±6.12)%.Conclusion Apoptin could inhibit obviously the growth of athymic mouse human T24 bladder carcinoma transplantation tumor by inducing apoptosis.
ABSTRACT The development of many systemic autoimmune diseases, including systemic lupus erythematosus, is associated with overactivation of the type I interferon (IFN) pathway, lymphopenia, and increased follicular helper T (Tfh) cell differentiation. However, the cellular and molecular mechanisms underlying these immunological perturbations remain incompletely understood. Here we show that the mechanistic target of rapamycin complex 2 (mTORC2) promotes Tfh differentiation and disrupts Treg homeostasis. Inactivation of mTORC2 in total T cells, but not in Tregs, greatly ameliorated the immunopathology in a systemic autoimmunity mouse model. This was associated with reduced Tfh differentiation, B cell activation, and reduced T cell glucose metabolism. Finally, we show that type I IFN can synergize with TCR ligation to activate mTORC2 in T cells, which partially contributes to T cell lymphopenia. These data indicate that mTORC2 may act as downstream of type I IFN, TCR, and costimulatory receptor ICOS, to promote glucose metabolism, Tfh differentiation, and T cell lymphopenia, but not to suppress Treg function in systemic autoimmunity. Our results suggest that mTORC2 might be a rational target for systemic autoimmunity treatment.
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