Daily Orange Consumption Reduces Hepatic Steatosis Prevalence in Patients with Metabolic Dysfunction-Associated Steatotic Liver Disease: Exploratory Outcomes of a Randomized Clinical Trial.

Background: Consumption of flavonoid-rich orange juice has been shown to reduce adiposity and liver steatosis in murine models of diet-induced obesity. However, little is known about the effects of whole orange intake, independent of body weight changes, on liver function and steatosis in individuals with metabolic dysfunction-associated steatotic liver disease (MASLD). The goal is to understand the direct impact of orange consumption on metabolic health. Methods: Sixty-two men and women aged 30-65 with MASLD (Controlled Attenuation Parameter, (CAP) > 275 dB/m) were randomly assigned to consume either 400 g of whole oranges or non-citrus fruits daily for 4 weeks. Baseline evaluations included medical assessments, blood tests, and body composition. Liver health was assessed using transient elastography (FibroScan®) for steatosis and fibrosis, conducted by blinded personnel. This clinical trial was registered at ClinicalTrials.gov (NCT05558592). Results: After 4 weeks of orange supplementation, liver steatosis decreased in the treatment group, with 70.9% showing steatosis compared to 100% in controls (p < 0.004), indicating a 30% reduction in liver disease prevalence. There were no significant changes in fibrosis or plasma liver enzymes, though plasma gamma glutaril transferase (GGT) levels decreased significantly. Body weight, waist circumference, body composition, lipid profile, fasting glucose, insulin, and C-reactive protein levels remained unchanged. Dietary analysis revealed no change in caloric intake, but vitamins C, A, thiamine, and riboflavin increased in the orange group. Conclusions: Our findings suggest that phytochemical-rich foods, especially whole fruits like oranges, may enhance liver function as an adjunct treatment for MASLD. The notable reduction in liver steatosis prevalence occurred independently of body weight changes. Further studies are needed to investigate the long-term effects of orange supplementation on steatosis and fibrosis progression and to identify the specific bioactive compounds and mechanisms involved.

Relationship between Markers of Gut Barrier Function and Erythrocyte Membrane PUFAs in Diarrhea-Predominant IBS Patients Undergoing a Low-FODMAP Diet.

Many patients with irritable bowel syndrome (IBS) have a compromised intestinal barrier associated with low-grade inflammation. Polyunsaturated fatty acids (PUFAs) are potential mediators of inflammation: omega-6 PUFAs are pro-inflammatory, while omega-3 PUFAs are antioxidant and anti-inflammatory. Zonulin is a potential biomarker for small intestinal permeability (s-IP). This study investigated the relationship between PUFAs and gastrointestinal (GI) barrier integrity in IBS patients with predominant diarrhea (IBS-D). We evaluated GI barrier function indicators in the urine and bloodstream and erythrocyte membrane PUFA composition in 38 IBS-D patients (5 men, 33 women, 44.11 ± 1.64 years), categorized at baseline by fecal zonulin levels into high (≥107 ng/mL, H-FZ) and normal (<107 ng/mL N-FZ) groups. Evaluations were conducted prior to and following a 12-week diet low in FODMAPs (LFD). At baseline, H-FZ patients had s-IP significantly higher than the reference value, lower n-3 PUFAs levels, and higher n-6/n-3 PUFAs and arachidonic acid (AA) to eicosapentaenoic acid (EPA) ratios than N-FZ. After LFD, H-FZ patients showed significant increases in n-3 PUFAs levels; decreases in n-6 PUFAs, n-6/n-3 PUFAs and AA/EPA ratios; and improved s-IP. The n-6/n-3 PUFAs ratio positively correlated with fecal zonulin levels in all subjects. These findings highlight the relationship between PUFAs and the intestinal barrier, suggesting their role in IBS-D pathophysiology and confirming the positive effects of LFD in managing IBS-D.

Significant Increase in Oxidative Stress Indices in Erythrocyte Membranes of Obese Patients with Metabolically-Associated Fatty Liver Disease.

Metabolic dysfunction-associated hepatic steatosis (MAFLD) indicates the metabolic risk associated with hepatic steatosis, overweight and obesity, and clinical evidence of metabolic dysregulation. Since MAFLD is one of the diseases that show a high frequency of alterations in the lipid content of cell membranes, the aim of this study was to evaluate the indices of oxidative damage of erythrocyte membranes in overweight and obese MAFLD subjects. The study was conducted on serum samples and red blood cell membranes of overweight and obese MAFLD subjects. For each patient, biochemical measurements and lipidomic analyses of erythrocytes membranes were performed. Significant differences in fatty acid profiles of RBC membranes were found between overweight and obese patients. In particular, the Peroxidation Index (PI) was higher in the erythrocyte membranes of obese subjects than in overweight subjects. The same behavior was observed for Unsaturation Index (UI) and Free Radical Stress Index (Free RSI), supporting the fact that the systemic increase in oxidative stress was associated with obesity. The study shows that there is a different susceptibility to erythrocyte membrane peroxidation for overweight and obese subjects, and the increased values of oxidative stress indices observed in the erythrocyte membranes of obese patients with MAFLD may be a possible indicator of pro-oxidative events occurring in obesity-related diseases.

Semaglutide Modulates Extracellular Matrix Production of LX-2 Cells via Exosomes and Improves Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD).

Metabolic dysfunction-associated steatotic liver disease (MASLD) is closely related to some metabolic disorders, such as central obesity and type 2 diabetes (T2D). Glucagon-like peptide 1 receptor agonists (GLP-1RAs), such as semaglutide, may have therapeutic roles in MASLD associated with T2D. This study aims to investigate the molecular mechanisms underlying the effectiveness of semaglutide on MASLD in terms of progression from liver steatosis to fibrosis. We characterized exosomes from ten patients with type 2 diabetes (T2D) before (T0) and after 12 months (T12) of treatment with once-weekly subcutaneous semaglutide. Six of ten patients were considered responders to therapy (R) based on MASLD severity downgrading by at least one class according to a validated ultrasonographic (US) score. Normal hepatocytes (HEPA-RG) and stellate (LX-2) cells were challenged with exosomes from R and NR patients, isolated before and after 12 months of therapy. Exosomes from both R and NR patients isolated at T0 significantly affected LX-2 viability. After 12 months of treatment, only those isolated from R patients restored cell viability, whereas those from NR patients did not. No effects were observed on HEPA-RG cells. Exosomes at T12 from R but not from NR patients significantly decreased the production of α-SMA, a marker of LX-2 activation, a liver stellate cell model, and ph-SMAD2 and CTGF, involved in fibrosis processes. TGF-ß1 was not modulated by the exosomes of R and NR patients. As a downstream effect, Vimentin, Collagen 1A1, and Fibronectin extracellular matrix components were also downregulated, as measured by droplets digital PCR. In conclusion, these results shed light on the potential effectiveness of semaglutide in improving liver fibrosis in MASLD.

Serum Cytokine and miRNA Levels Are Differently Expressed in Right- and Left-Sided Colon Cancer.

The tumor location in colorectal cancer (right- or left-sided colon cancer) is a key factor in determining disease progression. Right- and left-sided colon tumors are different in their clinical and molecular characteristics. Dysregulation of serum levels of proinflammatory cytokines, such as Transforming Growth Factor ß (TGF-ß) and Tumor Necrosis Factor-α (TNF-α), and Peroxisome Proliferator Activated Receptor-γ (PPAR-γ), known to be a growth-limiting and differentiation-promoting factor, as well as changes in miRNAs expression, are the major signaling pathways involved in the pathogenesis of this neoplasia. In the serum from 60 colorectal cancer (CRC) patients, we compared the differences in the expression of the levels of TGF-ß, TNF-α, and PPAR-γ and in the expression of the main human miRNAs between right and left CRC. A significant over-expression in the TGF-ß and TNF-α levels was observed in the serum from right-sided colon cancer patients. For the PPAR-γ, the patients with CRC located on the right-side showed lower levels than those detected in the serum from left-sided CRC subjects. Furthermore, significant differences also existed in the expression of specific circulating miRNAs between right- and left-sided CRC. In particular, the right upregulated miRNAs were all involved in the cell growth and proliferation related pathways. These findings confirm that the analysis of circulating levels of TGF-ß, TNF-α, and PPAR-γ, as well as the study of the specific miRNAs in the serum, are able to identify specific characteristics of CRC patients, useful for choosing a personalized treatment protocol.

The oleic/palmitic acid imbalance in exosomes isolated from NAFLD patients induces necroptosis of liver cells via the elongase-6/RIP-1 pathway.

Excessive toxic lipid accumulation in hepatocytes underlies the development of non-alcoholic fatty liver disease (NAFLD), phenotypically characterized by necrosis and steato-fibrosis, whose molecular mechanism is not yet fully understood. Patients with NAFLD display an imbalanced palmitic (PA) to oleic acid (OA) ratio. Moreover, increasing experimental evidence points out a relevant involvement of the exosomal content in disease progression. Aim of the study was to highlight the PA/OA imbalance within circulating exosomes, the subsequent intracellular alterations, and the impact on NALFD. Liver cells were challenged with exosomes isolated from both healthy subjects and NAFLD patients. The exosomal PA/OA ratio was artificially modified, and biological effects were evaluated. A NAFLD-derived exosomal PA/OA imbalance impacts liver cell cycle and cell viability. OA-modified NAFLD-derived exosomes restored cellular viability and proliferation, whereas the inclusion of PA into healthy subjects-derived exosomes negatively affected cell viability. Moreover, while OA reduced the phosphorylation and activation of the necroptosis marker, Receptor-interacting protein 1 (phospho-RIP-1), PA induced the opposite outcome, alongside increased levels of stress fibers, such as vimentin and fibronectin. Administration of NAFLD-derived exosomes led to increased expression of Elongase 6 (ELOVL6), Stearoyl-CoA desaturase 1 (SCD1), Tumor necrosis factor α (TNF-α), Mixed-lineage-kinase-domain-like-protein (MLKL) and RIP-1 in the hepatocytes, comparable to mRNA levels in the hepatocytes of NAFLD patients reported in the Gene Expression Omnibus (GEO) database. Genetic and pharmacological abrogation of ELOVL6 elicited a reduced expression of downstream molecules TNF-α, phospho-RIP-1, and phospho-MLKL upon administration of NAFLD-derived exosomes. Lastly, mice fed with high-fat diet exhibited higher phospho-RIP-1 than mice fed with control diet. Targeting the Elongase 6-RIP-1 signaling pathway offers a novel therapeutic approach for the treatment of the NALFD-induced exosomal PA/OA imbalance.

Bioaccessibility and Bioavailability of Diet Polyphenols and Their Modulation of Gut Microbiota.

It is generally accepted that diet-derived polyphenols are bioactive compounds with several potentially beneficial effects on human health. In general, polyphenols have several chemical structures, and the most representative are flavonoids, phenolic acids, and stilbenes. It should be noted that the beneficial effects of polyphenols are closely related to their bioavailability and bioaccessibility, as many of them are rapidly metabolized after administration. Polyphenols-with a protective effect on the gastrointestinal tract-promote the maintenance of the eubiosis of the intestinal microbiota with protective effects against gastric and colon cancers. Thus, the benefits obtained from dietary supplementation of polyphenols would seem to be mediated by the gut microbiota. Taken at certain concentrations, polyphenols have been shown to positively modulate the bacterial component, increasing Lactiplantibacillus spp. and Bifidobacterium spp. involved in the protection of the intestinal barrier and decreasing Clostridium and Fusobacterium, which are negatively associated with human well-being. Based on the diet-microbiota-health axis, this review aims to describe the latest knowledge on the action of dietary polyphenols on human health through the activity of the gut microbiota and discusses micro-encapsulation of polyphenols as a strategy to improve the microbiota.

Plasma-Derived Exosomes from NAFLD Patients Modulate the Cannabinoid Receptors' Expression in Cultured HepaRG Cells.

Exosomes produced by hepatocytes upon lipotoxic insult play a relevant role in pathogenesis of nonalcoholic fatty liver disease (NAFLD), suggesting an inflammatory response by the activation of monocytes and macrophages and accelerating the disease progression. In the pathogenesis of NAFLD and liver fibrosis, the endogenous cannabinoids and their major receptors CB1 and CB2 appear to be highly involved. This study aimed at evaluating the expression of cannabinoids receptors (CB1R and CB2R) in plasma-derived exosomes extracted from patients with NAFLD, as well as investigating the in vitro effects of the circulating exosomes in cultured human HepaRG cells following their introduction into the culture medium. The results demonstrated that plasma-derived exosomes from NAFLD patients are vehicles for the transport of CB1R and are able to modulate CB receptors' expression in HepaRG cells. In particular, circulating exosomes from NAFLD patients are inflammatory drivers for HepaRG cells, acting through CB1R activation and the downregulation of CB2R. Moreover, CB1R upregulation was associated with increased expression levels of PPAR-γ, a well-known mediator of liver tissue injury. In conclusion, this study provides evidence for CB1R transport by exosomes and suggests that the in vitro effects of circulating exosomes from NAFLD patients are mediated by the expression of cannabinoid receptors.

Anti-Proliferative and Pro-Apoptotic Effects of Digested Aglianico Grape Pomace Extract in Human Colorectal Cancer Cells.

Grape pomace (GP)-the major by-product of winemaking processes-still contains bioactive molecules with known beneficial properties for human health, such as an antiradical scavenging activity or an antiproliferative activity of tumors. In vitro studies have demonstrated that GP polyphenols specifically influence colon cancer cell proliferation. In addition to previously published work, we tested the phenolic compounds of Aglianico GP following an in vitro simulated gastrointestinal digestion on colorectal cancer cell lines at different degrees of differentiation. Our experiments, using HT29 and SW480 cells, confirmed the anti-proliferative effect of GP gastrointestinal digested extract and provided intriguing insights on the way it influences the cancer cell features (i.e., viability, proliferation, and apoptosis). We observed that Aglianico GP extract showed a great ability to affect cell proliferation and apoptosis. Interestingly, both HT29 and SW480 cells produced a significant increase in Bax, and a significant increase in the Bax/Bcl-2 ratio and caspase-3. The gastrointestinal digested GP extract was previously characterized both for antioxidant activity and phenolic composition. As a result, the TPC and the antioxidant activity reached high values in the Aglianico GP digested extract, and the main compounds assessed by UHPLC-DAD were anthocyanins, phenolic acids, and flavonoids. This work shed light on the use of digested GP extract as a dietary ingredient, a very sustainable source of nutritional compounds with potential health benefits for colon cancer cell proliferation.

Beneficial Effects of Table Grape Use on Serum Levels of Omega-3 Index and Liver Function: A Randomized Controlled Clinical Trial.

This clinical trial was aimed to investigate the effects of fresh table grape intake on the serum levels of the Omega-3 index, defined as the sum of eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA) levels. Forty consecutive healthy subjects were randomly assigned to the control group, receiving only dietary recommendations, and the grape group receiving a daily dose of 5 g of fresh table grape per kg of body weight, for 21 days. Compared with baseline, the grape treatment produced no significant difference in the serum levels of glucose, liver transaminase, and triglycerides, with the exception of cholesterol value, which was significantly reduced in both control and grape group (180.5 ± 20.32 vs. 196.1 ± 30.0 and 181.4 ± 21.9 vs. 194.3 ± 37.5, respectively). After 4 weeks from the end of grape treatment, the analysis of single fatty acids showed a significant increase in oleic acid content (14.15 ± 1.8 vs. 12.85 ± 1.6, p < 0.05) and a significant induction of the Omega-3 index (8.23 ± 1.9 vs. 6.09 ± 1.2, p < 0.05), associated with increased serum levels of adiponectin (24.09 ± 1.08 vs. 8.8 ± 0.7, p < 0.001). In contrast, the expression of fibroblast growth factor 21 (FGF21), a molecule associated with metabolic syndrome and liver disease, was significantly reduced (37.9 ± 6.8 vs. 107.8 ± 10.1, p < 0.001). The data suggest that the intake of fresh grape improves the Omega-3 index in the serum and exerts beneficial effects on liver function through the overexpression of adiponectin and the reduction in FGF21 levels.