Reducing gut microbiome-driven adipose tissue inflammation alleviates metabolic syndrome.

BACKGROUND: The gut microbiota contributes to macrophage-mediated inflammation in adipose tissue with consumption of an obesogenic diet, thus driving the development of metabolic syndrome. There is a need to identify and develop interventions that abrogate this condition. The hops-derived prenylated flavonoid xanthohumol (XN) and its semi-synthetic derivative tetrahydroxanthohumol (TXN) attenuate high-fat diet-induced obesity, hepatosteatosis, and metabolic syndrome in C57Bl/6J mice. This coincides with a decrease in pro-inflammatory gene expression in the gut and adipose tissue, together with alterations in the gut microbiota and bile acid composition. RESULTS: In this study, we integrated and interrogated multi-omics data from different organs with fecal 16S rRNA sequences and systemic metabolic phenotypic data using a Transkingdom Network Analysis. By incorporating cell type information from single-cell RNA-seq data, we discovered TXN attenuates macrophage inflammatory processes in adipose tissue. TXN treatment also reduced levels of inflammation-inducing microbes, such as Oscillibacter valericigenes, that lead to adverse metabolic phenotypes. Furthermore, in vitro validation in macrophage cell lines and in vivo mouse supplementation showed addition of O. valericigenes supernatant induced the expression of metabolic macrophage signature genes that are downregulated by TXN in vivo. CONCLUSIONS: Our findings establish an important mechanism by which TXN mitigates adverse phenotypic outcomes of diet-induced obesity and metabolic syndrome. TXN primarily reduces the abundance of pro-inflammatory gut microbes that can otherwise promote macrophage-associated inflammation in white adipose tissue. Video Abstract.

Spontaneous mutation in the Cd79b gene leads to a block in B-lymphocyte development at the C' (early pre-B) stage.

We discovered B-lymphocyte-deficient mice within a group of B10.A-CD45.1 mice, and established that this deficiency was a recessively inherited trait. Gene mapping and sequence analysis showed a mutation in the third exon of the Cd79b gene (c.224G>A) that leads to the generation of a stop codon (W75X) in the mutant mouse. Fluorescent-activated cell sorting analysis of bone marrow cells showed that the mutant mice did not express the CD79B antigen. To establish where the block in development happens, we analyzed CD43(pos)B220(pos) B-lymphocyte precursors present in the mutant mice and found that the fraction C' (corresponding to early pre-B lymphocytes) was absent in the mutant mouse, whereas fractions B and C showed a relative accumulation. As expected, we found no IgG or IgA in mutant mice. These results suggest that this CD79b-mutant strain may be a useful tool for immunological research in human immunodeficiencies.

Expression of Fas, FasL, and soluble Fas mRNA in endomyocardial biopsies of human cardiac allografts.

Apoptosis mediated by the Fas/Fas ligand (FasL) has been implicated in rejection of solid organ allografts and it has been recently proposed that soluble forms of Fas could interfere with this interaction, blocking apoptosis. The purpose of this study was to analyze intragraft Fas, FasL, and soluble Fas mRNA levels in relation to acute rejection in cardiac allografts in humans. mRNA levels were determined by quantitative reverse transcriptase-polymerase chain reaction in 42 samples of endomyocardial biopsies obtained from 18 cardiac transplant recipients within the first 6 months after transplantation. FasL and Fas mRNA levels were higher in biopsies with rejection than in biopsies without rejection, and no difference was observed in soluble Fas mRNA. During rejection, there was a positive correlation between the mRNA levels of Fas-FasL, Fas-soluble Fas, and FasL-soluble Fas. During quiescent periods, however, the only correlation observed was between Fas and soluble Fas mRNA levels. In conclusion, our findings do not suggest a role for soluble Fas, confirm the heightened expression of FasL, and indicate, for the first time, an increased expression of Fas in acute rejection of cardiac allografts.

Characterization of CD28, CTLA4, and ICOS polymorphisms in three Brazilian ethnic groups.

Costimulatory molecules CD28, CTLA-4 (cytotoxic T-lymphocyte-associated antigen-4), and ICOS (inducible costimulator) genes lie within the 300-kb chromosome region 2q33. CD28, CTLA-4, and ICOS have been described to be important regulators of T-cell activation. With the objective to study ethnic variations in allelic frequencies and linkage disequilibrium (LD) patterns, we genotyped CD28 intron 3 (+17 T>C), CTLA4 promoter (-319 C>T), CTLA4 exon 1 (+49 A>G), and ICOS 3' UTR (1564 T>C) polymorphisms in white (n = 103), mulatto (n = 97), and black (n = 79) Brazilian healthy individuals. No significant deviations from Hardy-Weinberg equilibrium were found in any of the population samples. A higher frequency of CD28 +17 C allele was detected in white (27%) in comparison with mulatto (15%) and black (13%) (p = 0.005) populations. LD between CD28 +17 C and CTLA4 -319 T alleles was observed in whites (p < 0.0001), mulattos (p = 0.0001), and blacks (p = 0.0002).

Interleukin-2 gene polymorphism is associated with renal but not cardiac transplant outcome.

It was recently shown that IL-2 gene single nucleotide polymorphism (SNP) at position -330 (G-->T) is related to in vitro cytokine production levels, with the T/T and T/G genotypes being associated with low production and the G/G genotype associated with high production. The objective of this study was to investigate a possible influence of this polymorphism on renal and cardiac allograft outcomes. IL-2 SNP G-T (-330) was determined by PCR-RFLP in 67 recipients of heart allografts and in 63 recipients of renal grafts from HLA-haplo-identical, related donors. A higher frequency of the T/T genotype was observed in renal transplant patients who experienced at least one acute rejection episode during the first 3 months after transplantation than in those without rejection during this period (80% vs 49%, respectively, P <.05). Accordingly, the same genotype tended to be more frequent in renal recipients with a 6-month serum creatinine level above 1.5 mg/dL (median value for the whole group of kidney recipients) than in patients with lower creatinine levels (79% vs 45%, P <.08). Regarding cardiac transplant recipients, no associations were observed concerning acute rejection or graft survival. The finding of the association of T/T but not T/G genotype with acute kidney rejection was unexpected considering that both genotypes were shown to be associated with equal (low) IL-2 in vitro production. Further studies are necessary not only to dissect the nature of IL-2 T/T genotype association with kidney rejection, but also to explain why this genotype does not apparently influence cardiac allograft outcome.

Intragraft activation of genes encoding cytotoxic T lymphocyte effector molecules precedes the histological evidence of rejection in human cardiac transplantation.

BACKGROUND: The purpose of the present study was to investigate transcripts of perforin, granzyme B, and Fas ligand (FasL) in heart transplants undergoing rejection. METHODS: Quantitative reverse transcriptase-polymerase chain reaction was applied for mRNA detection in 29 endomyocardial biopsy specimens from 11 cardiac allograft recipients. RESULTS: The mRNA levels of granzyme B, perforin, and FasL were higher (P<0.05) in biopsy specimens with rejection than in biopsy specimens without rejection (granzyme B, 0.53 vs. 0.09; perforin, 0.34 vs. 0; FasL, 0.57 vs. 0.36). In prerejection biopsy specimens, granzyme B and FasL levels were significantly higher than in biopsy specimens without rejection. Any two of the three transcripts were increased in 100% of prerejection, in 92% of rejection, and in 36% of no rejection biopsy specimens (P<0.04). CONCLUSIONS: The assessment of intragraft levels of cytotoxic T lymphocyte effector molecule mRNA represents a valuable tool in the monitoring of cardiac allograft rejection, especially considering the predictive value for warning of impending acute rejection.

Expression of CD40 ligand, interferon-gamma and Fas ligand genes in endomyocardial biopsies of human cardiac allografts: correlation with acute rejection

The purpose of the present study was to investigate the expression (mRNA) of CD40 ligand (CD40L), interferon-gamma (IFN-gamma) and Fas ligand (FasL) genes in human cardiac allografts in relation to the occurrence of acute cardiac allograft rejection as well as its possible value in predicting acute rejection. The mRNA levels were determined by a semiquantitative reverse transcriptase-polymerase chain reaction method in 39 samples of endomyocardial biopsies obtained from 10 adult cardiac transplant recipients within the first six months after transplantation. Biopsies with ongoing acute rejection showed significantly higher CD40L, IFN-gamma and FasL mRNA expression than biopsies without rejection. The median values of mRNA expression in biopsies with and without rejection were 0.116 and zero for CD40L (P<0.003), 0.080 and zero for IFN-gamma (P<0.0009), and 0.156 and zero for FasL (P<0.002), respectively. In addition, the levels of IFN-gamma mRNA were significantly increased 7 to 15 days before the appearance of histological evidence of rejection (median of 0.086 in pre-rejection biopsies), i.e., they presented a predictive value. This study provides further evidence of heightened expression of immune activation genes during rejection and shows that some of these markers may present predictive value for the occurrence of acute rejection.

Expression of CD40 ligand, interferon-gamma and Fas ligand genes in endomyocardial biopsies of human cardiac allografts: correlation with acute rejection.

The purpose of the present study was to investigate the expression (mRNA) of CD40 ligand (CD40L), interferon-gamma (IFN-gamma) and Fas ligand (FasL) genes in human cardiac allografts in relation to the occurrence of acute cardiac allograft rejection as well as its possible value in predicting acute rejection. The mRNA levels were determined by a semiquantitative reverse transcriptase-polymerase chain reaction method in 39 samples of endomyocardial biopsies obtained from 10 adult cardiac transplant recipients within the first six months after transplantation. Biopsies with ongoing acute rejection showed significantly higher CD40L, IFN-gamma and FasL mRNA expression than biopsies without rejection. The median values of mRNA expression in biopsies with and without rejection were 0.116 and zero for CD40L (P<0.003), 0.080 and zero for IFN-gamma (P<0.0009), and 0.156 and zero for FasL (P<0.002), respectively. In addition, the levels of IFN-gamma mRNA were significantly increased 7 to 15 days before the appearance of histological evidence of rejection (median of 0.086 in pre-rejection biopsies), i.e., they presented a predictive value. This study provides further evidence of heightened expression of immune activation genes during rejection and shows that some of these markers may present predictive value for the occurrence of acute rejection.

Monitoring of intragraft and peripheral blood TIRC7 expression as a diagnostic tool for acute cardiac rejection in humans.

T-cell immune response cDNA 7 (TIRC7) is a recently described T-cell costimulatory molecule that exhibits a central role in T-cell activation in vitro and in vivo. The present study was undertaken to investigate association between intragraft and peripheral blood mononuclear cell (PBMC) TIRC7 mRNA levels and cardiac allograft rejection in humans. TIRC7 gene expression levels were determined by a quantitative-competitive reverse transcriptase-polymerase chain reaction (QC-RT-PCR) in endomyocardial biopsies and in PBMC from cardiac transplant recipients. Biopsies collected during rejection or up to 15 days before rejection showed heightened TIRC7 mRNA expression in comparison with biopsies without rejection. All prerejection and rejection biopsies showed TIRC7 mRNA upregulation, while this was present in only 30% of the biopsies without rejection. Regarding TIRC7 mRNA in PBMC, transplant recipients showed lower levels than healthy individuals and, in contrast to the results obtained in biopsies, the levels were lower during rejection than in rejection-free periods. In summary, TIRC7 mRNA expression levels increase in biopsies and decrease in peripheral blood during acute cardiac rejection. We conclude that intragraft detection of TIRC7 transcripts is a useful tool not only for the diagnosis but also for the prediction of acute heart allograft rejection episodes.