Acute Effects of Dietary Protein Consumption on the Postprandial Metabolic Response, Amino Acid Levels and Circulating MicroRNAs in Patients with Obesity and Insulin Resistance.

The post-nutritional intervention modulation of miRNA expression has been previously investigated; however, post-acute dietary-ingestion-related miRNA expression dynamics in individuals with obesity and insulin resistance (IR) are unknown. We aimed to determine the acute effects of protein ingestion from different dietary sources on the postprandial metabolic response, amino acid levels, and circulating miRNA expression in adults with obesity and IR. This clinical trial included adults with obesity and IR who consumed (1) animal-source protein (AP; calcium caseinate) or (2) vegetable-source protein (VP; soy protein isolate). Glycaemic, insulinaemic, and glucagon responses, amino acid levels, and exosomal microRNAs isolated from plasma were analysed. Post-AP ingestion, the area under the curve (AUC) of insulin (p = 0.04) and the plasma concentrations of branched-chain (p = 0.007) and gluconeogenic (p = 0.01) amino acids increased. The effects of different types of proteins on the concentration of miRNAs were evaluated by measuring their plasma circulating levels. Compared with the baseline, the AP group presented increased circulating levels of miR-27a-3p, miR-29b-3p, and miR-122-5p (p < 0.05). Subsequent analysis over time at 0, 30, and 60 min revealed the same pattern and differences between treatments. We demonstrated that a single dose of dietary protein has acute effects on hormonal and metabolic regulation and increases exosomal miRNA expression in individuals with obesity and IR.

Intermittent fasting, calorie restriction, and a ketogenic diet improve mitochondrial function by reducing lipopolysaccharide signaling in monocytes during obesity: A randomized clinical trial.

BACKGROUND: Mitochondrial dysfunction occurs in monocytes during obesity and contributes to a low-grade inflammatory state; therefore, maintaining good mitochondrial conditions is a key aspect of maintaining health. Dietary interventions are primary strategies for treating obesity, but little is known about their impact on monocyte bioenergetics. Thus, the aim of this study was to evaluate the effects of calorie restriction (CR), intermittent fasting (IF), a ketogenic diet (KD), and an ad libitum habitual diet (AL) on mitochondrial function in monocytes and its modulation by the gut microbiota. METHODS AND FINDINGS: A randomized controlled clinical trial was conducted in which individuals with obesity were assigned to one of the 4 groups for 1 month. Subsequently, the subjects received rifaximin and continued with the assigned diet for another month. The oxygen consumption rate (OCR) was evaluated in isolated monocytes, as was the gut microbiota composition in feces and anthropometric and biochemical parameters. Forty-four subjects completed the study, and those who underwent CR, IF and KD interventions had an increase in the maximal respiration OCR (p = 0.025, n2p = 0.159 [0.05, 0.27] 95% confidence interval) in monocytes compared to that in the AL group. The improvement in mitochondrial function was associated with a decrease in monocyte dependence on glycolysis after the IF and KD interventions. Together, diet and rifaximin increased the gut microbiota diversity in the IF and KD groups (p = 0.0001), enriched the abundance of Phascolarctobacterium faecium (p = 0.019) in the CR group and Ruminococcus bromii (p = 0.020) in the CR and KD groups, and reduced the abundance of lipopolysaccharide (LPS)-producing bacteria after CR, IF and KD interventions compared to the AL group at the end of the study according to ANCOVA with covariate adjustment. Spearman's correlation between the variables measured highlighted LPS as a potential modulator of the observed effects. In line with this findings, serum LPS and intracellular signaling in monocytes decreased with the three interventions (CR, p = 0.002; IF, p = 0.001; and KD, p = 0.001) compared to those in the AL group at the end of the study. CONCLUSIONS: We conclude that these dietary interventions positively regulate mitochondrial bioenergetic health and improve the metabolic profile of monocytes in individuals with obesity via modulation of the gut microbiota. Moreover, the evaluation of mitochondrial function in monocytes could be used as an indicator of metabolic and inflammatory status, with potential applications in future clinical trials. TRIAL REGISTRATION: This trial was registered with ClinicalTrials.gov (NCT05200468).

Ramon Flour (Brosimum alicastrum Swartz) Ameliorates Hepatic Lipid Accumulation, Induction of AMPK Phosphorylation, and Expression of the Hepatic Antioxidant System in a High-Fat-Diet-Induced Obesity Mouse Model.

Excessive consumption of fat and carbohydrates, together with a decrease in traditional food intake, has been related to obesity and the development of metabolic alterations. Ramon seed is a traditional Mayan food used to obtain Ramon flour (RF) with high biological value in terms of protein, fiber, micronutrients, and bioactive compounds such as polyphenols. However, few studies have evaluated the beneficial effects of RF. Thus, we aimed to determine the metabolic effects of RF consumption on a high-fat-diet-induced obesity mouse model. We divided male BALB/c mice into four groups (n = 5 each group) and fed them for 90 days with the following diets: Control (C): control diet (AIN-93), C + RF: control diet adjusted with 25% RF, HFD: high-fat diet + 5% sugar in water, and HFD + RF: high-fat diet adjusted with 25% RF + 5% sugar in water. The RF prevented the increase in serum total cholesterol (TC) and alanine transaminase (ALT) that occurred in the C and HFD groups. Notably, RF together with HFD increased serum polyphenols and antioxidant activity, and it promoted a decrease in the adipocyte size in white adipose tissue, along with lower hepatic lipid accumulation than in the HFD group. In the liver, the HFD + RF group showed an increase in the expression of ß-oxidation-related genes, and downregulation of the fatty acid synthase (Fas) gene compared with the HFD group. Moreover, the HFD + RF group had increased hepatic phosphorylation of AMP-activated protein kinase (AMPK), along with increased nuclear factor erythroid 2-related factor 2 (NRF2) and superoxide dismutase 2 (SOD2) protein expression compared with the HFD group. Thus, RF may be used as a nutritional strategy to decrease metabolic alterations during obesity.

Genistein-mediated thermogenesis and white-to-beige adipocyte differentiation involve transcriptional activation of cAMP response elements in the Ucp1 promoter.

Genistein is an isoflavone present in soybeans and is considered a bioactive compound due to its widely reported biological activity. We have previously shown that intraperitoneal genistein administration and diet supplementation activates the thermogenic program in rats and mice subcutaneous white adipose tissue (scWAT) under multiple environmental cues, including cold exposure and high-fat diet feeding. However, the mechanistic insights of this process were not previously unveiled. Uncoupling protein 1 (UCP1), a mitochondrial membrane polypeptide responsible for dissipating energy into heat, is considered the most relevant thermogenic marker; thus, we aimed to evaluate whether genistein regulates UCP1 transcription. Here we show that genistein administration to thermoneutral-housed mice leads to the appearance of beige adipocyte markers, including a sharp upregulation of UCP1 expression and protein abundance in scWAT. Reporter assays showed an increase in UCP1 promoter activity after genistein stimulation, and in silico analysis revealed the presence of estrogen (ERE) and cAMP (CRE) response elements as putative candidates of genistein activation. Mutation of the CRE but not the ERE reduced genistein-induced promoter activity by 51%. Additionally, in vitro and in vivo ChIP assays demonstrated the binding of CREB to the UCP1 promoter after acute genistein administration. Taken together, these data elucidate the mechanism of genistein-mediated UCP1 induction and confirm its potential applications in managing metabolic disorders.

Edible and medicinal mushrooms (Pleurotus ostreatus, Ustilago maydis, Ganoderma lucidum) reduce endoplasmic reticulum stress and inflammation in adipose tissue of obese Wistar rats fed with a high fat plus saccharose diet.

Obesity is an increasing global public health problem. A strategy to treat obesity is the use of functional foods. Edible and medicinal mushrooms contain diverse bioactive compounds showing important antihyperlipidemic, antioxidant, and prebiotic properties. We analysed the effects of adding (10%) of Pleurotus ostreatus (Po, basidiomata), Ganoderma lucidum (Gl, basidiomata), or Ustilago maydis (Um, galls), milled, to a high fat plus saccharose diet (HFD + S) for 6 months in a model of obesity with Wistar rats. We assessed weight gain, body composition, lipid parameters, endoplasmic reticulum stress (proteins and inflammatory markers: BiP, XBP-1, JNK, p-JNK, TNF-α), and adiponectin in subcutaneous adipose tissue (SAT). The consumption of edible and medicinal mushrooms decreased weight gain (-17.2-30.1%) and fat mass (-23.7-43.1%), maintained fat-free mass, reduced levels of serum biochemical parameters (TC: -40.1-44.1%, TG: -37.7-51.6%, LDL-C: -64.5-71.1%), and prevented adipocyte hypertrophy (-30.9-36.9%) and collagen deposition (-70.9-73.7%) in SAT. Compared with the HFD + S group, mushroom consumption by Wistar rats significantly reduced the expression of proteins associated with endoplasmic reticulum stress and inflammation (BiP: -72.2-88.2%; XBP-1: -71.5-81.8%; JNK: -71.2-90.0%; p-JNK: -37.3-81.0%; TNF-α: -80.7-91.5%), whereas significantly increased adiponectin protein expression (246.4-654.2%) in SAT. These effects outperformed those obtained through the commercial lipid-lowering drug atorvastatin, contributing synergistically to prevent further obesity-related dysfunctions, such as insulin resistance derived from inflammation and ER stress in adipose tissue. Bioactive compounds from edible, functional and medicinal mushrooms represent new emerging therapies for obesity treatments using natural products.

The Gut Microbiota-Brain Axis during Aging, Mild Cognitive Impairment and Dementia: Role of Tau Protein, ß-Amyloid and LPS in Serum and Curli Protein in Stool.

Currently, there is an increasing number of people with mild cognitive (MCI) impairment and dementia (D). In the present work we studied the role of tau protein, ß-amyloid, LPS (lipopolysaccharide), and curli protein of elderly adults with MCI or D and the contribution of gut microbiota. Four groups were studied: young subjects, healthy adults older than 60 years (A), elderly adults with MCI (MCI), and elderly adults with dementia (D). A preclinical study was conducted in old male Wistar rats to evaluate the impact of gut microbiota on curli protein abundance in feces and brain. The results showed that with increasing age, tau protein, ß-amyloid, and LPS significantly increased in serum during MCI and D, and this was associated with an increase in the abundance of E. coli that synthesize the amyloid protein curli, that may promote the aggregation of amyloid proteins. Rats showed a clear increase in the abundance of curli protein in the brain during aging. Thus, cognitive impairment and dementia are in part due to an alteration in the gut microbiota-brain axis via increase in curli protein and LPS leading to an increase in tau and ß-amyloid protein.

Association of BCAT2 and BCKDH polymorphisms with clinical, anthropometric and biochemical parameters in young adults.

BACKGROUND AND AIM: Circulating amino acids are modified by sex, body mass index (BMI) and insulin resistance (IR). However, whether the presence of genetic variants in branched-chain amino acid (BCAA) catabolic enzymes modifies circulating amino acids is still unknown. Thus, we determined the frequency of two genetic variants, one in the branched-chain aminotransferase 2 (BCAT2) gene (rs11548193), and one in the branched-chain ketoacid dehydrogenase (BCKDH) gene (rs45500792), and elucidated their impact on circulating amino acid levels together with clinical, anthropometric and biochemical parameters. METHODS AND RESULTS: We performed a cross-sectional comparative study in which we recruited 1612 young adults (749 women and 863 men) aged 19.7 ± 2.1 years and with a BMI of 24.9 ± 4.7 kg/m2. Participants underwent clinical evaluation and provided blood samples for DNA extraction and biochemical analysis. The single nucleotide polymorphisms (SNPs) were determined by allelic discrimination using real-time polymerase chain reaction (PCR). The frequencies of the less common alleles were 15.2 % for BCAT2 and 9.83 % for BCKDH. The subjects with either the BCAT2 or BCKDH SNPs displayed no differences in the evaluated parameters compared with subjects homozygotes for the most common allele at each SNP. However, subjects with both SNPs had higher body weight, BMI, blood pressure, glucose, and circulating levels of aspartate, isoleucine, methionine, and proline than the subjects homozygotes for the most common allele (P < 0.05, One-way ANOVA). CONCLUSION: Our findings suggest that the joint presence of both the BCAT2 rs11548193 and BCKDH rs45500792 SNPs induces metabolic alterations that are not observed in subjects without either SNP.

SIRT7 modulates the stability and activity of the renal K-Cl cotransporter KCC4 through deacetylation.

SIRT7 is a NAD+ -dependent deacetylase that controls important aspects of metabolism, cancer, and bone formation. However, the molecular targets and functions of SIRT7 in the kidney are currently unknown. In silico analysis of kidney transcripts of the BXD murine genetic reference population revealed a positive correlation between Sirt7 and Slc12a7 mRNA expression, suggesting a link between the corresponding proteins that these transcripts encode, SIRT7, and the K-Cl cotransporter KCC4, respectively. Here, we find that protein levels and activity of heterologously expressed KCC4 are significantly modulated depending on its acetylation status in Xenopus laevis oocytes. Moreover, SIRT7 interacts with KCC4 in a NAD+ -dependent manner and increases its stability and activity in HEK293 cells. Interestingly, metabolic acidosis increases SIRT7 expression in kidney, as occurs with KCC4. In contrast, total SIRT7-deficient mice present lower KCC4 expression and an exacerbated metabolic acidosis than wild-type mice during an ammonium chloride challenge. Altogether, our data suggest that SIRT7 interacts with, stabilizes and modulates KCC4 activity through deacetylation, and reveals a novel role for SIRT7 in renal physiology.

Genistein stimulates insulin sensitivity through gut microbiota reshaping and skeletal muscle AMPK activation in obese subjects.

OBJECTIVE: Obesity is associated with metabolic abnormalities, including insulin resistance and dyslipidemias. Previous studies demonstrated that genistein intake modifies the gut microbiota in mice by selectively increasing Akkermansia muciniphila, leading to reduction of metabolic endotoxemia and insulin sensitivity. However, it is not known whether the consumption of genistein in humans with obesity could modify the gut microbiota reducing the metabolic endotoxemia and insulin sensitivity. RESEARCH DESIGN AND METHODS: 45 participants with a Homeostatic Model Assessment (HOMA) index greater than 2.5 and body mass indices of ≥30 and≤40 kg/m2 were studied. Patients were randomly distributed to consume (1) placebo treatment or (2) genistein capsules (50 mg/day) for 2 months. Blood samples were taken to evaluate glucose concentration, lipid profile and serum insulin. Insulin resistance was determined by means of the HOMA for insulin resistance (HOMA-IR) index and by an oral glucose tolerance test. After 2 months, the same variables were assessed including a serum metabolomic analysis, gut microbiota, and a skeletal muscle biopsy was obtained to study the gene expression of fatty acid oxidation. RESULTS: In the present study, we show that the consumption of genistein for 2 months reduced insulin resistance in subjects with obesity, accompanied by a modification of the gut microbiota taxonomy, particularly by an increase in the Verrucomicrobia phylum. In addition, subjects showed a reduction in metabolic endotoxemia and an increase in 5'-adenosine monophosphate-activated protein kinase phosphorylation and expression of genes involved in fatty acid oxidation in skeletal muscle. As a result, there was an increase in circulating metabolites of ß-oxidation and ω-oxidation, acyl-carnitines and ketone bodies. CONCLUSIONS: Change in the gut microbiota was accompanied by an improvement in insulin resistance and an increase in skeletal muscle fatty acid oxidation. Therefore, genistein could be used as a part of dietary strategies to control the abnormalities associated with obesity, particularly insulin resistance; however, long-term studies are needed.

Insulin and SGK1 reduce the function of Na+/monocarboxylate transporter 1 (SMCT1/SLC5A8).

SMCTs move several important fuel molecules that are involved in lipid, carbohydrate, and amino acid metabolism, but their regulation has been poorly studied. Insulin controls the translocation of several solutes that are involved in energetic cellular metabolism, including glucose. We studied the effect of insulin on the function of human SMCT1 expressed in Xenopus oocytes. The addition of insulin reduced α-keto-isocaproate (KIC)-dependent 22Na+ uptake by 29%. Consistent with this result, the coinjection of SMCT1 with SGK1 cRNA decreased the KIC-dependent 22Na+ uptake by 34%. The reduction of SMCT1 activity by SGK1 depends on its kinase activity, and it was observed that the coinjection of SMCT1 with S442D-SGK1 (a constitutively active mutant) decreased the KIC-dependent 22Na+ uptake by 50%. In contrast, an SMCT1 coinjection with K127M-SGK1 (an inactive mutant) had no effect on the KIC-dependent Na+ uptake. The decreasing SMCT1 function by insulin or SGK1 was corroborated by measuring [1-14C]acetate uptake and the electric currents of SMCT1-injected oocytes. Previously, we found that SMCT2/Slc5a12-mRNA, but not SMCT1/Slc5a8-mRNA, is present in zebrafish pancreas (by in situ hybridization); however, SLC5a8 gene silencing was associated with the development of human pancreatic cancer. We confirmed that the mRNA and protein of both transporters were present in rat pancreas using RT-PCR with specific primers, Western blot analysis, and immunohistochemistry. Additionally, significant propionate-dependent 22Na+ uptake occurred in pancreatic islets and was reduced by insulin treatment. Our data indicate that human SMCT1 is regulated by insulin and SGK1 and that both SMCTs are present in the mammalian pancreas.

Prolactin and the dietary protein/carbohydrate ratio regulate the expression of SNAT2 amino acid transporter in the mammary gland during lactation.

The sodium coupled neutral amino acid transporter 2 (SNAT2/SAT2/ATA2) is expressed in the mammary gland (MG) and plays an important role in the uptake of alanine and glutamine which are the most abundant amino acids transported into this tissue during lactation. Thus, the aim of this study was to assess the amount and localization of SNAT2 before delivery and during lactation in rat MG, and to evaluate whether prolactin and the dietary protein/carbohydrate ratio might influence SNAT2 expression in the MG, liver and adipose tissue during lactation. Our results showed that SNAT2 protein abundance in the MG increased during lactation and this increase was maintained along this period, while 24 h after weaning it tended to decrease. To study the effect of prolactin on SNAT2 expression, we incubated MG explants or T47D cells transfected with the SNAT2 promoter with prolactin, and we observed in both studies an increase in the SNAT2 expression or promoter activity. Consumption of a high-protein/low carbohydrate diet increased prolactin concentration, with a concomitant increase in SNAT2 expression not only in the MG during lactation, but also in the liver and adipose tissue. There was a correlation between SNAT2 expression and serum prolactin levels depending on the amount of dietary protein/carbohydrate ratio consumed. These findings suggest that prolactin actively supports lactation providing amino acids to the gland through SNAT2 for the synthesis of milk proteins.