Andrey Santos

Linking Gut Microbiota and Inflammation to Obesity and Insulin Resistance

Obesity and insulin resistance are the major predisposing factors to comorbidities, such as Type 2 diabetes, nonalcoholic fatty liver disease, cardiovascular and neurodegenerative diseases, and several types of cancer. The prevalence of obesity is still increasing worldwide and now affects a large number of individuals. Here, we review the role of the gut microbiota in the pathophysiology of insulin resistance/obesity. The human intestine is colonized by ∼100 trillion bacteria, which constitute the gut microbiota. Studies have shown that lean and overweight rodents and humans may present differences in the composition of their intestinal flora. Over the past 10 years, data from different sources have established a causal link between the intestinal microbiota and obesity/insulin resistance. It is important to emphasize that diet-induced obesity promotes insulin resistance by mechanisms independent and dependent on gut microbiota. In this review, we present several mechanisms that contribute to explaining the link between intestinal flora and insulin resistance/obesity. The LPS from intestinal flora bacteria can induce a chronic subclinical inflammatory process and obesity, leading to insulin resistance through activation of TLR4. The reduction in circulating SCFA may also have an essential role in the installation of reduced insulin sensitivity and obesity. Other mechanisms include effects of bile acids, branched-chain amino acids (BCAA), and some other lesser-known factors. In the near future, this area should open new therapeutic avenues for obesity/insulin resistance and its comorbidities.

Blocking iNOS and endoplasmic reticulum stress synergistically improves insulin resistance in mice

Objective Recent data show that iNOS has an essential role in ER stress in obesity. However, whether iNOS is sufficient to account for obesity-induced ER stress and Unfolded Protein Response (UPR) has not yet been investigated. In the present study, we used iNOS knockout mice to investigate whether high-fat diet (HFD) can still induce residual ER stress-associated insulin resistance. Methods For this purpose, we used the intraperitoneal glucose tolerance test (GTT), euglycemic-hyperinsulinemic clamp, western blotting and qPCR in liver, muscle, and adipose tissue of iNOS KO and control mice on HFD. Results The results of the present study demonstrated that, in HFD fed mice, iNOS-induced alteration in insulin signaling is an essential mechanism of insulin resistance in muscle, suggesting that iNOS may represent an important target that could be blocked in order to improve insulin sensitivity in this tissue. However, in liver and adipose tissue, the insulin resistance induced by HFD was only partially dependent on iNOS, and, even in the presence of genetic or pharmacological blockade of iNOS, a clear ER stress associated with altered insulin signaling remained evident in these tissues. When this ER stress was blocked pharmacologically, insulin signaling was improved, and a complete recovery of glucose tolerance was achieved. Conclusions Taken together, these results reinforce the tissue-specific regulation of insulin signaling in obesity, with iNOS being sufficient to account for insulin resistance in muscle, but in liver and adipose tissue ER stress and insulin resistance can be induced by both iNOS-dependent and iNOS-independent mechanisms.

Probiotics modulate gut microbiota and improve insulin sensitivity in DIO mice

Obesity and type 2 diabetes are characterized by subclinical inflammatory process. Changes in composition or modulation of the gut microbiota may play an important role in the obesity-associated inflammatory process. In the current study, we evaluated the effects of probiotics (Lactobacillus rhamnosus, L. acidophilus and Bifidobacterium bifidumi) on gut microbiota, changes in permeability, and insulin sensitivity and signaling in high-fat diet and control animals. More importantly, we investigated the effects of these gut modulations on hypothalamic control of food intake, and insulin and leptin signaling. Swiss mice were submitted to a high-fat diet (HFD) with probiotics or pair-feeding for 5 weeks. Metagenome analyses were performed on DNA samples from mouse feces. Blood was drawn to determine levels of glucose, insulin, LPS, cytokines and GLP-1. Liver, muscle, ileum and hypothalamus tissue proteins were analyzed by Western blotting and real-time polymerase chain reaction. In addition, liver and adipose tissues were analyzed using histology and immunohistochemistry. The HFD induced huge alterations in gut microbiota accompanied by increased intestinal permeability, LPS translocation and systemic low-grade inflammation, resulting in decreased glucose tolerance and hyperphagic behavior. All these obesity-related features were reversed by changes in the gut microbiota profile induced by probiotics. Probiotics also induced an improvement in hypothalamic insulin and leptin resistance. Our data demonstrate that the intestinal microbiome is a key modulator of inflammatory and metabolic pathways in both peripheral and central tissues. These findings shed light on probiotics as an important tool to prevent and treat patients with obesity and insulin resistance.

Green propolis modulates gut microbiota, reduces endotoxemia and expression of TLR4 pathway in mice fed a high-fat diet

Due to the various beneficial effects attributed to propolis, which include anti-inflammatory and anti-bacterial infection properties, the objective of the study was to evaluate the effect of propolis supplementation on the composition of the intestinal microbiota and its anti-inflammatory action. Forty male C57BL/6 mice were fed either a standard diet (control), a high-fat (HF) diet, or a high-fat diet supplemented with 0.2% crude propolis (HFP) for 2 or 5weeks prior to sacrifice. Blood samples were collected for the determination of lipopolysaccharide (LPS) and classical biochemical parameters. Expression of the TLR4 pathway in muscle, and DNA sequencing for the 16S rRNA of the gut microbiota were performed. The HF diet increased the proportion of the phylum Firmicutes and inflammatory biomarkers, while supplementation with propolis for five weeks rendered the microbiota profile nearly normal. Consistently with the above, the supplementation reduced levels of circulating LPS and down-regulated the TLR4 pathway and inflammatory cytokine expressions in muscle. Moreover, propolis improved such biochemical parameters as serum triacylglycerols and glucose levels. The data suggest that propolis supplementation reduces inflammatory response and endotoxemia by preventing dysbiosis in mice challenged with a high-fat diet.

Dietary whey proteins shield murine cecal microbiota from extensive disarray caused by a high-fat diet

High-fat diets are used to induce adverse alterations in the intestinal microbiota, or dysbiosis, generalized inflammation and metabolic stress, which ultimately may lead to obesity. The influence of dietary whey proteins, whether intact or hydrolyzed, has been reported to improve glucose homeostasis and reduce stress. Therefore, the purpose of this work was to test if dietary milk-whey proteins, both in the intact form and hydrolyzed, could have an effect on the compositional changes of the cecal microbiota that can be induced in mice when receiving a high-fat diet in combination with the standard casein. Male C57BL/6 mice were fed a control casein diet (AIN 93-G); high-fat-casein (HFCAS); high-fat-whey protein concentrate (HFWPC) and high-fat whey-protein hydrolysate (HFWPH) for 9weeks. The intestinal microbiota composition was analyzed by 16S-rRNA of the invariant (V1-V3) gene, potentially endotoxemic lipopolysaccharide (LPS) release was determined colorimetrically, and liver fat infiltration assessed by light microscopy. The high-fat diet proved to induce dysbiosis in the animals by inverting the dominance of the phylum Firmicutes over Bacteroidetes, promoted the increase of LPS and resulted in liver fat infiltration. The whey proteins, whether intact or hydrolyzed, resisted the installation of dysbiosis, prevented the surge of circulating LPS and prevented fat infiltration in the liver. It is concluded that dietary whey proteins exert metabolic actions that tend to preserve the normal microbiota profile, while mitigating liver fat deposition in mice consuming a high-fat diet for nine weeks. Such beneficial effects were not seen when casein was the dietary protein. The hydrolyzed whey protein still differed from the normal whey protein by selectively protecting the Bacteroidetes phylum.

BCR-ABL1 Transcript Levels at 3 and 6 Months Are Better for Identifying Chronic Myeloid Leukemia Patients with Poor Outcome in Response to Second-Line Second-Generation Tyrosine Kinase Inhibitors after Imatinib Failure: A Report from a Single Institution.

Early reduction of BCR-ABL1 transcript levels has been associated with improved outcome in chronic myeloid leukemia (CML) treatment. We evaluated 54 chronic-phase CML patients treated with imatinib who switched therapy to dasatinib (n = 33) or nilotinib (n = 21). BCR-ABL1 transcript levels were measured in peripheral blood using real-time quantitative PCR (RQ-PCR) every 3 months from the start of second-line treatment. Patients with BCR-ABL transcript levels >10% at 3 months and >1% at 6 months had significantly inferior progression-free (PFS) and event-free survival (EFS) than patients with RQ-PCR <10% at 3 months and <1% at 6 months (66 vs. 100%, p = 0.01, and 33 vs. 73%, p = 0.02, respectively). Patients with RQ-PCR <10% at 3 months and >1% at 6 months also had inferior PFS and EFS than patients with RQ-PCR <10% at 3 months and <1% at 6 months (48 vs. 100%, p = 0.002, and 25 vs. 73%, p < 0.0001, respectively). Two measurements of BCR-ABL levels were better than a single one to stratify chronic-phase CML patients as failure after second-line therapy.

Novel mutations associated with pyruvate kinase deficiency in Brazil

Background Pyruvate kinase deficiency is a hereditary disease that affects the glycolytic pathway of the red blood cell, causing nonspherocytic hemolytic anemia. The disease is transmitted as an autosomal recessive trait and shows a marked variability in clinical expression. This study reports on the molecular characterization of ten Brazilian pyruvate kinase-deficient patients and the genotype–phenotype correlations. Method Sanger sequencing and in silico analysis were carried out to identify and characterize the genetic mutations. A non-affected group of Brazilian individuals were also screened for the most commonly reported variants (c.1456C>T and c.1529G>A). Results Ten different variants were identified in the PKLR gene, of which three are reported here for the first time: p.Leu61Gln, p.Ala137Val and p.Ala428Thr. All the three missense variants involve conserved amino acids, providing a rationale for the observed enzyme deficiency. The allelic frequency of c.1456C>T was 0.1% and the 1529G>A variant was not found. Conclusion This is the first comprehensive report on molecular characterization of pyruvate kinase deficiency from South America. The results allowed us to correlate the severity of the clinical phenotype with the identified variants.

Helminth infection in mice improves insulin sensitivity via modulation of gut microbiota and fatty acid metabolism

&NA; Intestinal helminths are prevalent in individuals who live in rural areas of developing countries, where obesity, type 2 diabetes, and metabolic syndrome are rare. In the present study, we analyzed the modulation of the gut microbiota in mice infected with the helminth Strongyloides venezuelensis, and fed either a standard rodent chow diet or high‐fat diet (HFD). To investigate the effects of the microbiota modulation on the metabolism, we analyzed the expression of tight‐junction proteins present in the gut epithelium, inflammatory markers in the serum and tissue and quantified glucose tolerance and insulin sensitivity and resistance. Additionally, the levels of lipids related to inflammation were evaluated in the feces and serum. Our results show that infection with Strongyloides venezuelensis results in a modification of the gut microbiota, most notably by increasing Lactobacillus spp. These modifications in the microbiota alter the host metabolism by increasing the levels of anti‐inflammatory cytokines, switching macrophages from a M1 to M2 pattern in the adipose tissue, increasing the expression of tight junction proteins in the intestinal cells (thereby reducing the permeability) and decreasing LPS in the serum. Taken together, these changes correlate with improved insulin signaling and sensitivity, which could also be achieved with HFD mice treated with probiotics. Additionally, helminth infected mice produce higher levels of oleic acid, which participates in anti‐inflammatory pathways. These results suggest that modulation of the microbiota by helminth infection or probiotic treatment causes a reduction in subclinical inflammation, which has a positive effect on the glucose metabolism of the host. Graphical abstract Figure. No caption available.

Impact of Trans-Fats on Heat-Shock Protein Expression and the Gut Microbiota Profile of Mice: Hydrogenated oil, HSPs & gut microbiota…

Partially hydrogenated oils are known to cause metabolic stress and dyslipidemia. This paper explores a new dimension about the interaction between dietary trans-fats and the defense heat-shock protein (HSP) system, inflammation, and the gut microbiota of mice consuming a hyperlipidic diet containing partially hydrogenated vegetable oil free of animal fat. Five diet groups were installed: control diet, 2 hyperlipidic-partially hydrogenated-oil diets, each containing either casein or whey-protein hydrolysate (WPH) as protein source, and 2 consuming hyperlipidic-unhydrogenated-oil diets containing either WPH or casein as a protein source. The partially hydrogenated oil inhibited c-Jun NH2 -terminal kinase phosphorylation in the casein diets, but without altering κ-B kinase. Neither the lipid nor the protein had an influence on the proinflammatory toll-like receptor 4 (TLR4) pathway, but the combination of the high-lipid content and WPH impaired glucose tolerance without altering insulin or glucose transporter-4 translocation. It was remarkable to observe that, contrary to the case of a common high-fat diet, the lard-free hyperlipidic diets were hardly able to invert the Bacteroidetes:Firmicutes phylum ratio. Our results suggest that, in the absence of lard, the intake of trans-fatty acids is less harmful than expected because it does not trigger TLR4-inflammation or pose great threat to the normal gut microbiota. WPH had the effect of promoting the expression of HSP90, HSP60, and HSP25, but did not prevent dysbiosis, when the diet contained the unhydrogenated oil. The partially hydrogenated oil also seemed to antagonize the ability of WPH to induce the expression of protective HSPs.

Bacterial remediation from effluent containing multi-walled carbon nanotubes

Multi-wall carbon nanotubes (MWCNT) were functionalized with functional groups containing oxygen, mainly carboxylic groups (-COOH), through reaction with a mixture of H2SO4/HNO3 (3:1 v/v). The oxidized multi-wall carbon nanotubes (MWCNTOOH) were used to prepare an effluent, 2 mg L−1 in a saline solution of NaCl (0.9%), to study of remediation of MWCNTOOH in aqueous suspension by utilization of Escherichia coli. The suspensions of E. coli (4.5 × 105 CFU mL−1 and 4.5 × 108 CFU mL−1) in test tubes with MWCNTOOH effluent caused the precipitation of a large amount of MWCNTOOH and supernatant clearing. The scanning electron microscopy (SEM) analysis of the precipitate and supernatant showed the adhesion and interlace of MWCNTOOH in bacteria surface. Although the precipitate consist of a large quantity of MWCNTOOH and bacteria, it was verified their presence in the supernatant. The spread plate technique showed that MWCNTOOH caused no cellular death of E. coli in the supernatant.