Strong associations between fasting lipids and glucose concentrations and ALT levels strengthened with increasing ALT quantiles.

BACKGROUND: A persistent redox state and excessive reactive species involved in carbohydrate and lipid metabolism lead to oxidative damage in the liver, however, how fasting plasma concentrations of lipids and glucose are associated with fasting blood levels of alanine transaminase (ALT) and aspartate transaminase (AST) remains to be evaluated in large-scale population. METHODS: A cross-sectional study with 182,971 residents aged 18 to 92 years; multidimensional stratified analyses including quantile linear regression analysis and sex stratification were adopted to improve the quality of the evidence. RESULTS: The associations between the concentrations of non-HDL-C and triglyceride and ALT levels were positive, stronger in males in each quantile of ALT levels and the coefficients expanded with increasing ALT levels at slopes of 3.610 and 5.678 in males and 2.977 and 5.165 in females, respectively. The associations between the HDL-C concentrations and ALT levels were negative, also stronger in males in each quantile and the coefficients expanded with increasing ALT levels at slopes of -7.839 in females and - 5.797 in males. The associations between glucose concentrations and ALT levels were positive, but stronger in females in each quantile and the coefficients expanded with increasing ALT levels at slopes of 1.736 in males and 2.177 in females, respectively. Similar pattern consist of relatively weaker coefficients and slops were observed between concentrations of non-HDL-C, triglyceride and glucose and AST levels. The associations between albumin concentration and concentrations of blood lipids and glucose were relatively steady across all quantiles. CONCLUSIONS: The dose dependent effect between blood concentrations of lipids and glucose and liver function changes suggests that excessive carbohydrate and lipid metabolism may cause subclinical liver damage. Long term sustained primary and secondary inflammatory factors produced in the liver might be transmitted to adjacent organs, such as the heart, kidneys, and lungs, to cause and/or exacerbate pathological changes in these visceral organs.

Alteration in gut microbiota is associated with immune imbalance in Graves' disease.

Background: Graves' disease (GD), characterized by immune aberration, is associated with gut dysbiosis. Despite the growing interest, substantial evidence detailing the precise impact of gut microbiota on GD's autoimmune processes remains exceedingly rare. Objective: This study was designed to investigate the influence of gut microbiota on immune dysregulation in GD. Methods: It encompassed 52 GD patients and 45 healthy controls (HCs), employing flow cytometry and enzyme-linked immunosorbent assay to examine lymphocyte and cytokine profiles, alongside lipopolysaccharide (LPS) levels. Gut microbiota profiles and metabolic features were assessed using 16S rRNA gene sequencing and targeted metabolomics. Results: Our observations revealed a disturbed B-cell distribution and elevated LPS and pro-inflammatory cytokines in GD patients compared to HCs. Significant differences in gut microbiota composition and a marked deficit in short-chain fatty acid (SCFA)-producing bacteria, including ASV263(Bacteroides), ASV1451(Dialister), and ASV503(Coprococcus), were observed in GD patients. These specific bacteria and SCFAs showed correlations with thyroid autoantibodies, B-cell subsets, and cytokine levels. In vitro studies further showed that LPS notably caused B-cell subsets imbalance, reducing conventional memory B cells while increasing naïve B cells. Additionally, acetate combined with propionate and butyrate showcased immunoregulatory functions, diminishing cytokine production in LPS-stimulated cells. Conclusion: Overall, our results highlight the role of gut dysbiosis in contributing to immune dysregulation in GD by affecting lymphocyte status and cytokine production.

Expression, Purification, and Crystallization of the Vγ9Vδ2 T-cell Receptor Recognizing Protein/Peptide Antigens.

γδ T cells, especially Vγ9Vδ2 T cells, play an important role in mycobacterial infection. We have identified some Vγ9Vδ2 T cells that recognize protein/peptide antigens derived from mycobacteria, which may induce protective immune responses to mycobacterial infection. To clarify the structural basis of the molecular recognition mechanism, we tried many methods to express the Vγ9Vδ2 T-cell receptor (TCR). The Vγ9Vδ2 TCR was not expressed well in a prokaryotic expression system or a baculovirus expression system, even after extensive optimization. In a mammalian cell expression system, the Vγ9Vδ2 TCR was expressed in the form of a soluble heterodimer, which was suitable for crystal screening. Reduced-temperature cultivation (cold shock) increased the yield of the recombinant TCR. The recombinant purified TCR was used for crystal trials, and crystals that could be used for X-ray diffraction were obtained. Although we have not yet determined the crystal structure of the Vγ9Vδ2 TCR, we have established a procedure for Vγ9Vδ2 TCR expression and purification, which is useful for basic research and potentially for clinical application.

Inflammatory Cytokine Interleukin-6 (IL-6) Promotes the Proangiogenic Ability of Adipose Stem Cells from Obese Subjects via the IL-6 Signaling Pathway.

BACKGROUND: The prevalence of obesity, as well as obesity-induced chronic inflammatory diseases, is increasing worldwide. Chronic inflammation is related to the complex process of angiogenesis, and we found that adipose-derived stem cells from obese subjects (obADSCs) had proangiogenic features, including higher expression levels of interleukin-6 (IL-6), Notch ligands and receptors, and proangiogenic cytokines, than those from control subjects. We hypothesized that IL-6 and Notch signaling pathways are essential for regulating the proangiogenic characteristics of obADSCs. OBJECTIVE: This study aimed to investigate whether the inflammatory cytokine interleukin 6 (IL-6) promotes the proangiogenic capacity of adipose stem cells in obese subjects via the IL-6 signaling pathway. METHODS: We compared the phenotype analysis as well as cell doubling time, proliferation, migration, differentiation, and proangiogenic properties of ADSCs in vitro. Moreover, we used small interfering RNAs to inhibit the gene and protein expression of IL-6. RESULTS: We found that ADSCs isolated from control individuals (chADSCs) and obADSCs had similar phenotypes and growth characteristics, and chADSCs had a stronger differentiation ability than obADSCs. However, obADSCs were more potent in promoting EA.hy926 cell migration and tube formation than chADSCs in vitro. We confirmed that IL-6 siRNA significantly reduced the transcriptional level of IL-6 in obADSCs, thereby reducing the expression of vascular endothelial growth factor (VEGF)- A, VEGF receptor 2, transforming growth factor ß, and Notch ligands and receptors in obADSCs. CONCLUSION: The finding suggests that inflammatory cytokine interleukin-6 (IL-6) promotes the proangiogenic ability of obADSCs via the IL-6 signaling pathway.

Comparison of the biological characteristics of umbilical cord mesenchymal stem cells derived from the human heterosexual twins.

Mesenchymal stromal/stem cells (MSCs) are attracting more and more attention due to their tissue regenerative properties and immunomodulatory functions. MSCs may be the most acceptable, safe, and effective source for allogeneic cell therapy, and have been used in medical treatment. However, the similarities and differences between umbilical cord-derived MSCs (UC-MSCs) of heterosexual twins remain poorly understood. In this study, we compared the biological characteristics of UC-MSCs of heterosexual twins in vitro. We found that male fetal UC-MSCs and female fetal UC-MSCs share a similar phenotype and multi-lineage differentiation potential, and male fetal UC-MSCs show a significantly higher proliferation and adipogenic ability than female fetal UC-MSCs. UC-MSCs from heterosexual twins showed significant differences in the expression levels of NANOG, OCT4, TERT, and SOX2. In addition, male MSCs are more potent in the expression of inflammatory cytokines to lipopolysaccharide (LPS)-induced inflammation. In future clinical applications using MSCs for inflammation-related diseases, these biological characteristics differences with different genders will guide our clinical methods.

Transduction with Lentiviral Vectors Altered the Expression Profile of Host MicroRNAs.

RNA interference (RNAi) is widely used in gene knockdown analysis and as a tool to screen host genes involved in viral infection. Owing to the limitations of transducing cells with synthetic small interfering RNAs (siRNAs), lentiviral short hairpin RNA (shRNA) vectors are more widely used. However, we found that stable transduction with lentiviral shRNA vectors inhibited hepatitis C virus (HCV) propagation in human hepatoma cells. We found by microRNA (miRNA) microarray analysis that this inhibition was induced by the alteration of host miRNA expression. In addition to one miRNA (miR-196b-5p) previously reported to be involved in HCV infection, other miRNAs (miR-216a-5p, -216b-5p, 217, and -30b-5p) were found to influence HCV infection in this study. Further studies suggested that this effect was independent of the transcription of shRNAs. The lentiviral vector itself and the integration site of the lentiviral vector might determine the change in miRNA expression. Moreover, the upregulation of JUN contributed to the dysregulation of miR-216a-5p, -216b-5p, and -217 in stably transduced cells. Although the changes in miRNA expression were beneficial for inhibiting HCV infection in our study, this off-target effect should be considered when transduction with lentiviral vectors is performed for other purposes, especially in therapy.IMPORTANCE We found that stable transduction with lentiviral shRNA was able to nonspecifically inhibit HCV infection by the dysregulation of host miRNAs. Previous studies showed that the overexpression of shRNAs oversaturated the host miRNA pathways to inhibit HCV infection. In contrast, the miRNA machinery was not affected in our study. Knockout studies suggested that the nonspecific effect was independent of the transcription of shRNAs. The lentiviral vector itself and the integration sites in the host genome determined the changes in miRNAs. Stable transduction with lentiviral vectors was able to increase the expression of JUN, which in turn upregulated miR-216a-5p, miR-216b-5p, and miR-217. miR-216a-5p and miR-216b-5p might inhibit HCV by suppressing the host autophagic machinery. Our study suggested a novel nonspecific effect of lentiviral vectors, and this side effect should be considered when transduction with lentiviral vectors is performed for other purposes, especially in therapy.

Author Correction: Next generation sequencing reveals changes of the γδ T cell receptor repertoires in patients with pulmonary tuberculosis.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Next generation sequencing reveals changes of the γδ T cell receptor repertoires in patients with pulmonary tuberculosis.

Tuberculosis (TB) is a severe global threat to human health. The immune protection initiated by γδ T cells play an important role in mycobacterial infection. Vaccines for Mycobacterium tuberculosis (Mtb) based on γδ T cells provide a novel approach for TB control. In our previous studies, we found a preponderant complementarity-determining region 3 (CDR3) sequence of the γδ T cell receptor (TCR) in TB patients, and successfully identified a tuberculosis antigen that can effectively activate γδ T cells with a reverse genetic strategy. However, due to the throughput limitation of the method we used, the information we obtained about the γδ TCR repertoire and preponderant CDR3 sequences was limited. In this study, we introduced next generation sequencing (NGS) to study the γδ TCR CDR3 repertoires in TB patients. We found that the CDR3δ tended to be more polyclonal and CDR3γ tended to be longer in TB patients; the γδ T cells expressing CDR3 sequences using a Vγ9-JγP rearrangement expanded significantly during Mtb infection. We also identified new preponderant CDR3 sequences during Mtb infection. This study comprehensively characterized the γδ T cell receptor repertoire changes, and provides useful information for the development of new vaccines and adjuvants against TB.

Enterovirus 71 protease 2Apro and 3Cpro differentially inhibit the cellular endoplasmic reticulum-associated degradation (ERAD) pathway via distinct mechanisms, and enterovirus 71 hijacks ERAD component p97 to promote its replication.

Endoplasmic reticulum-associated degradation (ERAD) is an important function for cellular homeostasis. The mechanism of how picornavirus infection interferes with ERAD remains unclear. In this study, we demonstrated that enterovirus 71 (EV71) infection significantly inhibits cellular ERAD by targeting multiple key ERAD molecules with its proteases 2Apro and 3Cpro using different mechanisms. Ubc6e was identified as the key E2 ubiquitin-conjugating enzyme in EV71 disturbed ERAD. EV71 3Cpro cleaves Ubc6e at Q219G, Q260S, and Q273G. EV71 2Apro mainly inhibits the de novo synthesis of key ERAD molecules Herp and VIMP at the protein translational level. Herp differentially participates in the degradation of different glycosylated ERAD substrates α-1 antitrypsin Null Hong Kong (NHK) and the C-terminus of sonic hedgehog (SHH-C) via unknown mechanisms. p97 was identified as a host factor in EV71 replication; it redistributed and co-exists with the viral protein and other known replication-related molecules in EV71-induced replication organelles. Electron microscopy and multiple-color confocal assays also showed that EV71-induced membranous vesicles were closely associated with the endoplasmic reticulum (ER), and the ER membrane molecule RTN3 was redistributed to the viral replication complex during EV71 infection. Therefore, we propose that EV71 rearranges ER membranes and hijacks p97 from cellular ERAD to benefit its replication. These findings add to our understanding of how viruses disturb ERAD and provide potential anti-viral targets for EV71 infection.

Recombinant adenovirus expressing F and H fusion proteins of peste des petits ruminants virus induces both humoral and cell-mediated immune responses in goats.

Peste des petits ruminants (PPR) is an acute and contagious disease of some small ruminants caused by peste des petits ruminants virus (PPRV). Fusion (F) protein and hemagglutinin (H) protein are two glycoproteins of PPRV that might induce a protective immune response. In this study, three replication-defective recombinant adenoviruses were constructed and the immunogenicity was evaluated in goats (the natural host). The recombinant adenoviruses (rAds) expressing F, H, and F-H fusion protein were named rAd-F, rAd-H, and rAd-F-H, respectively. In vitro, the proteins expressed in AAV-293 cells infected with different rAds were identified by Western blotting and immunofluorescence. The results showed that the proteins could be expressed in vitro. Three groups of goats (6 goats per group) were inoculated subcutaneously twice at 3-week intervals with the rAds. As negative controls, two additional groups were inoculated with wild-type adenovirus (wtAd) or PBS. In vivo, goats immunized with the rAds developed PPRV-specific virus neutralizing antibody (VNA) by 3 weeks after primary immunization. Moreover, the seroconversions were maintained for approximately 21 weeks after primary immunization. Stronger lymphocyte proliferation responses were induced in goats immunized with the three rAds than in the negative controls (P<0.05). Notably, goats inoculated with rAd-F-H developed significantly higher VNA titers (P<0.05) and stronger cell-mediated immune responses than did goats inoculated with rAd-F or rAd-H alone. The results suggest that the three rAds might be attractive candidate differentiating infected from vaccinated animals (DIVA) vaccines for preventing PPRV infection. Notably, the rAd-F-H expressing F-H fusion protein is likely the most potent candidate of the rAds.