Nanoparticles Formulation Improves the Antifibrogenic Effect of Quercetin on an Adenine-Induced Model of Chronic Kidney Disease.

Renal fibrosis is the final stage of chronic kidney injury characterized by glomerulosclerosis and tubulointerstitial fibrosis with parenchymal destruction. Quercetin belongs to the most studied flavonoids with antioxidant, anti-inflammatory, antifibrogenic, and antitumor activity. It modifies the TGF-ß/Smad signaling pathway, decreasing profibrogenic expression molecules and inducing the expression of antioxidant, anti-inflammatory, and antifibrogenic molecules. However, quercetin exhibits poor water solubility and low absorption and bioavailability. This limitation was solved by developing a nanoparticles formulation that improves the solubility and bioavailability of several bioactive compounds. Therefore, we aimed to investigate the in vivo antifibrogenic effect of a quercetin nanoparticles formulation. Male C57BL/6 mice were induced into chronic renal failure with 50 mg/kg of adenine for four weeks. The animals were randomly grouped and treated with 25, 50, or 100 mg/kg of quercetin, either macroparticles or nanoparticles formulation. We performed biochemical, histological, and molecular analyses to evaluate and compare the effect of macroparticles versus nanoparticles formulation on kidney damage. Here, we demonstrated that smaller doses of nanoparticles exhibited the same beneficial effect as larger doses of macroparticles on preventing kidney damage. This finding translates into less quercetin consumption reaching the desired therapeutic effect.

Methodological and Statistical Considerations for Cross-Sectional, Case-Control, and Cohort Studies.

Epidemiological studies are essential in medicine and public health as they help identify risk factors and causes of diseases. Additionally, they are key to planning, implementing, and evaluating health interventions aimed at preventing and controlling the spread of diseases. Among these studies, analytical observational studies, such as cross-sectional, case-control, and cohort studies, are the most used. The validity of their results largely depends on the robustness of the design, execution, and statistical analysis. Objective: The objective of this study is to examine the most common errors in the selection of methodological design and statistical tests in analytical observational studies and to provide recommendations to correct them. Methodology: A comprehensive review of the available literature on methodology in epidemiological observational studies was conducted, focusing on cross-sectional, case-control, and cohort studies. Common errors in the selection of designs and statistical tests were identified and analyzed. Results and Conclusions: Errors in the selection of methodological design and statistical tests are common in epidemiological observational studies. Based on the identified errors, a series of recommendations is provided to improve the selection of methodological design and statistical tests, thereby increasing the reliability of the results in cross-sectional, case-control, and cohort studies.

Intestinal Dysbiosis in Subjects with Obesity from Western Mexico and Its Association with a Proinflammatory Profile and Disturbances of Folate (B9) and Carbohydrate Metabolism.

Obesity is a public health problem with a growing prevalence worldwide. In Mexico, it is estimated that one out of three adults suffer from obesity. In these patients, the intestinal microbiota (IM) undergoes pathological changes that are associated with a dysbiotic state; however, the microbiota profile of adult subjects with obesity from western Mexico has not been described. To assess this, fecal samples were obtained from 65 participants (Obese = 38; Control = 27). The microbial composition was characterized by 16S rRNA amplicon sequencing. The IM of the group with obesity revealed a clear decrease in richness and diversity (p < 0.001), as well as a significant increase in proinflammatory bacterial groups, mainly genera belonging to the Negativicutes class, Escherichia/Shigella, and Prevotella. Likewise, an increase in short-chain fatty acid-producing bacteria was found, especially the genus Lachnoclostridium. Additionally, PICRUSt2 analysis showed a depletion of vitamin B9 metabolism and an increase in saccharolytic pathways. The IM of patients with obesity possesses a dysbiotic, proinflammatory environment, possibly contributing to lipogenesis and adiposity. Thus, assessing the IM will allow for a better understanding of the pathophysiology of metabolic diseases of high prevalence, such as obesity. These findings are described for the first time in the adult population of western Mexico.

Automated Computer-Assisted Image Analysis for the Fast Quantification of Kidney Fibrosis.

Chronic kidney disease (CKD) is a common and worldwide health problem and one of the most important causes of morbidity and mortality. Most primary research on this disease requires evaluating the fibrosis index in animal model kidneys, specifically using Masson's trichrome stain. Different programs are used to calculate the percentage of fibrosis; however, the analysis is time-consuming since one image must be performed at a time. CellProfiler™ is a program designed to analyze data obtained from biological samples and can process multiple images through pipelines, and the results can be exported to databases. This article explains how CellProfiler™ can be used to automatically analyze kidney histology photomicrographs from samples stained with Masson's trichrome stain to assess the percentage of fibrosis in an experimental animal model of CKD. A pipeline was created to analyze Masson's trichrome-stained slides in a model of CDK induced by adenine at doses of 50 mg/kg and 100 mg/kg, in addition to samples with the vehicle (75% glycerin). The results were compared with those obtained by ImageJ, and no significant differences were found between both programs. The CellProfiler™ pipeline made here is a reliable, fast, and easy alternative for kidney fibrosis analysis and quantification in experimental animal models.

Pirfenidone Is an Agonistic Ligand for PPARα and Improves NASH by Activation of SIRT1/LKB1/pAMPK.

Nonalcoholic steatohepatitis (NASH) is recognized by hepatic lipid accumulation, inflammation, and fibrosis. No studies have evaluated the prolonged-release pirfenidone (PR-PFD) properties on NASH features. The aim of this study is to evaluate how PR-PFD performs on metabolic functions, and provide insight on a mouse model of human NASH. Male C57BL/6J mice were fed with either normo diet or high-fat/carbohydrate diet for 16 weeks and a subgroup also fed with PR-PFD (300 mg/kg/day). An insulin tolerance test was performed at the end of treatment. Histological analysis, determination of serum hormones, adipocytokines measurement, and evaluation of proteins by western blot was performed. Molecular docking, in silico site-directed mutagenesis, and in vitro experiments using HepG2 cultured cells were performed to validate PR-PFD binding to peroxisome proliferator-activated receptor alpha (PPAR-α), activation of PPAR-α promoter, and sirtuin 1 (SIRT1) protein expression. Compared with the high-fat group, the PR-PFD-treated mice displayed less weight gain, cholesterol, very low density lipoprotein and triglycerides, and showed a significant reduction of hepatic macrosteatosis, inflammation, hepatocyte ballooning, fibrosis, epididymal fat, and total adiposity. PR-PFD restored levels of insulin, glucagon, adiponectin, and resistin along with improved insulin resistance. Noteworthy, SIRT1-liver kinase B1-phospho-5' adenosine monophosphate-activated protein kinase signaling and the PPAR-α/carnitine O-palmitoyltransferase 1/acyl-CoA oxidase 1 pathway were clearly induced in high fat + PR-PFD mice. In HepG2 cells incubated with palmitate, PR-PFD induced activation and nuclear translocation of both PPARα and SIRT1, which correlated with increased SIRT1 phosphorylated in serine 47, suggesting a positive feedback loop between the two proteins. These results were confirmed with both synthetic PPAR-α and SIRT1 activators and inhibitors. Finally, we found that PR-PFD is a true agonist/ligand for PPAR-α. Conclusions: PR-PFD provided an anti-steatogenic effect and protection for inflammation and fibrosis.

Improved effects of viral gene delivery of human uPA plus biliodigestive anastomosis induce recovery from experimental biliary cirrhosis.

Gene therapy may represent a new avenue for the development of multimodal treatment for diverse forms of cirrhosis. This study explores the potential benefits of combining adenovirus-mediated human urokinase-plasminogen activator (AdHuPA) gene delivery and biliodigestive anastomosis to enhance the therapeutic efficacy of each treatment alone for cholestatic disorders resulting in secondary biliary cirrhosis. In an experimental model of secondary biliary cirrhosis, application of 6 x 10(11) vp/kg AdHuPA adenovirus vector resulted in 25.8% liver fibrosis reduction and some improvement in liver histology. The relief of bile cholestasis by a surgical procedure (biliodigestive anastomosis) combined with AdHuPA hepatic gene delivery rendered a synergistic effect, with a substantial 56.9 to 42.9% fibrosis decrease. AdHuPA transduction resulted in clear-cut expression of human uPA protein detected by immunohistochemistry and induction of up-regulation in the expression of metalloproteinases MMP-3, MMP-9, and MMP-2. Importantly, functional hepatic tests, specifically direct bilirubin, were improved. Also, hepatic cell regeneration, rearrangement of hepatic architecture, ascites, and gastric varices improved in cirrhotic rats treated with AdHuPA but not in counterpart AdGFP cirrhotic animals. We believe this might represent a novel therapeutic strategy for human cholestatic diseases.