Pablo José Giraudi

Background/Objective: Pediatric obesity represents a paramount problem for global health that affects children’s actual quality of life, dramatically compromising their future health. Obesity has been demonstrated to alter skeletal muscle, a highly plastic tissue with dimorphic characteristics, which changes during development, homeostasis, adaptive responses, and diseases. Since it is responsible for 80% of insulin-stimulated glucose uptake, its alteration establishes a vicious circle profoundly impacting overall metabolism. Although obesogenic diets are known to distinctly affect adult male and female muscles, scarce information is available on growing individuals. Therefore, we performed an exploratory analysis on the early effects of an obesogenic diet in murine muscles during maturation. Methods: We analyzed the transcriptome of tibialis anterior from C57BL/6J male and female mice, fed on a high-fat, high-sugar diet (HFHS) for 4 and 20 weeks starting at 3 weeks of age. Results: Experimental data revealed different HFHS-associated transcriptional patterns in female and male mice, within a background of sex-associated transcriptional differences across muscle maturation. Conclusions: These preliminary observations lay the groundwork for further investigation of the mechanisms underlying the differences in diet-associated transcriptional patterns between female and male skeletal muscle, with the ultimate goal of addressing obesity and its comorbidities.

Chronic low-grade inflammation is a key feature of obesity-associated dysglycemia, yet substantial heterogeneity exists in inflammatory responses among individuals with normoglycemia, prediabetes, and type 2 diabetes mellitus (T2DM). Whether circulating inflammatory protein profiles define distinct patient phenotypes beyond conventional glycemic classification remains incompletely understood. In this cross-sectional analysis of 142 individuals with severe obesity, plasma inflammatory proteins were quantified using Olink proximity extension assay technology. Subjects were stratified by glycemic status (noDM, normoglycemia; PreDM, prediabetes and T2DM) while maintaining comparable distributions of metabolic dysfunction-associated steatotic liver disease. Differential expression analyses were performed across glycemic groups, and unsupervised topological data analysis (TDA) was applied to identify inflammatory protein-based patient subgroups. Several inflammatory proteins were significantly upregulated in T2DM and PreDM compared with noDM, with interleukin-8 (IL-8), Fms-relatedlike tyrosine kinase 3 ligand (Flt3L), and CUB domain containing protein (CDCP1) showing the largest significant differences. NPX distributions of these proteins exhibited gradual increases across glycemic stages with substantial inter-individual variability. TDA identified seven clusters defined by distinct inflammatory protein signatures. One cluster was enriched for individuals with T2DM and characterized by coordinated upregulation of IL-8, Flt3L, CDCP1, and additional immune- and cytokine-related proteins, whereas other clusters displayed alternative inflammatory profiles that were not explained by glycemic status alone. Inflammatory proteomic profiling in severe obesity reveals both glycemia-associated protein changes and distinct inflammatory phenotypes that transcend conventional clinical classification. Integration of differential expression analysis with TDA highlights heterogeneity in inflammatory states, supporting a hypothesis-generating framework for future studies aimed at validating these proteomic patterns and clarifying their longitudinal relevance in obesity-related dysglycemia.

Introduction and Objectives: Metabolic dysfunction associated steatotic liver disease (MASLD) affects about one quarter of adults worldwide, and liver fibrosis is its strongest predictor of liver related morbidity and mortality. Using combined in-silico screening and clinical validation, we aimed to identify circulating biomarkers associated with fibrosis progression. Fibulin 3 was identified, and its diagnostic performance was evaluated in biopsy proven MASLD cohorts. Materials and Methods: The GSE125251 RNAseq dataset was reanalyzed to compare liver transcriptomes from MASLD subjects with minimal (F0/F1) versus moderate to advanced fibrosis (F2/F3, F4). Differentially expressed genes (DEGs) were filtered to retain plasma secreted, protein coding candidates. Top ranked genes were evaluated in liver biopsies from a morbidly obese cohort (n = 65) stratified by fibrosis stage, and their plasma levels were measured via ELISA in an independent bariatric cohort (n = 225). Results: Among 106 DEGs, 22 encoded plasma circulating proteins. Six top candidates (EFEMP1, LTBP2, LUM, DPT, CHI3L1, CCL20) were prioritized. EFEMP1 (Fibulin 3) showed the strongest association with fibrosis, with significantly higher hepatic mRNA and protein expression in F2/F3, F4 versus F0 F1 (p < 0.005). Plasma Fibulin 3 levels correlated with fibrosis stage (ρ = 0.40, p < 0.0001), increasing from 9.4 ng/mL in F0F1 to 21.7 ng/mL in F2/F3,F4. Its diagnostic performance for F ≥ 2 (AUROC = 0.78) exceeded that of APRI, FIB-4, NFS, and HSI. A combined index including Fibulin 3, HSI, platelets, and GGT increased the AUROC to 0.87 (CI: 0.79 0.92). Conclusions: Plasma Fibulin 3 is notably higher in individuals with advanced MASLD and represents a promising non-invasive biomarker for liver fibrosis stratification in metabolically unhealthy obese populations.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is closely linked to comorbidities like obesity, type 2 diabetes, and cardiovascular disease. Given that liver biopsy is the diagnostic gold standard, there is a critical need for minimally invasive tests, particularly for the inflammatory form, metabolic dysfunction-associated steatohepatitis (MASH). In this discovery study, we investigated the plasma proteome to identify blood biomarkers for MASH and explored their potential tissue sources, the liver and visceral adipose tissue. Plasma low-abundance proteome profiling was performed on samples from a cohort of morbidly obese MASLD subjects (n = 90; 40 with MASH, 50 without) using Olink^® panels. Paired liver and visceral adipose biopsies were also analyzed. Data showed 34 significantly different plasma proteins between the two groups, including Pleiotrophin (PTN), Fibroblast growth factor-21 (FGF-21), and Hepatocyte growth factor (HGF), among others. While plasma-tissue correlation was only found for STX8, PTN and FGF-21 demonstrated the strongest associations with the histopathological features of MASH. A diagnostic model combining PTN, FGF-21, and AST achieved a robust AUC of 0.88 (95% CI: 0.84–0.97) for distinguishing MASH. Based on this discovery pilot study, circulating PTN and FGF-21 emerge as promising non-invasive biomarkers for improving patient stratification and supporting therapeutic evaluation in MASH, warranting validation in independent cohorts and future studies.

Metabolic and bariatric surgery (MBS) is an effective treatment for severe obesity, though individual responses vary widely, partly due to genetic predisposition. This study investigates the association of a body mass index (BMI) polygenic score (PGS) with weight loss and metabolic outcomes following surgery. A cohort of 225 patients undergoing MBS was analyzed at baseline (T₀), six (T₆), and twelve (T₁₂) months, with anthropometric and biochemical parameters recorded at each time point. Total weight loss (TWL) and excess weight loss (EWL) percentages were calculated. PGS was computed using the LDpred-grid Bayesian method. The mean age was 45.9 ± 9.4 years. Males had a higher baseline prevalence of type 2 diabetes (T2D) and comorbidities (p < 0.001). Linear regression analysis confirmed an association between PGS and baseline BMI (p = 0.012). Moreover, mediation analysis revealed that baseline BMI mediated the effect of the PGS on %TWL at T₁₂, with an indirect effect (p-value = 0.018). In contrast, high-density lipoprotein-cholesterol (HDL-C) at T₆ and triglycerides (TG) at T₁₂ showed direct associations with the PGS (p-value = 0.004 and p-value = 0.08, respectively), with no significant mediation by BMI. This study showed a BMI-mediated association of PGS with %TWL and a direct association with lipid changes, suggesting its potential integration into personalized obesity treatment.

Background: Prader–Willi syndrome (PWS) is a rare genetic disorder characterized by distinctive physical, cognitive, and behavioral manifestations, coupled with profound alterations in appetite regulation, leading to severe obesity and metabolic dysregulation. These clinical features arise from disruptions in neurodevelopment and neuroendocrine regulation, yet the molecular intricacies of PWS remain incompletely understood. Methods: This study aimed to comprehensively profile circulating neuromodulatory factors in the serum of 53 subjects with PWS and 34 patients with non-syndromic obesity, utilizing a proximity extension assay with the Olink Target 96 neuro-exploratory and neurology panels. The ANOVA p-values were adjusted for multiple testing using the Benjamani–Hochberg method. Protein–protein interaction networks were generated in STRING V.12. Corrplots were calculated with R4.2.2 by using the Hmisc, Performance Analytics, and Corrplot packages Results: Our investigation explored the potential genetic underpinnings of the circulating protein signature observed in PWS, revealing intricate connections between genes in the PWS critical region and the identified circulating proteins associated with impaired oxytocin, NAD metabolism, and sex-related neuromuscular impairment involving, CD38, KYNU, NPM1, NMNAT1, WFIKKN1, and GDF-8/MSTN. The downregulation of CD38 in individuals with PWS (p < 0.01) indicates dysregulation of oxytocin release, implicating pathways associated with NAD metabolism in which KYNU and NMNAT1 are involved and significantly downregulated in PWS (p < 0.01 and p < 0.05, respectively). Sex-related differences in the circulatory levels of WFIKKN1 and GDF-8/MSTN (p < 0.05) were also observed. Conclusions: This study highlights potential circulating protein biomarkers associated with impaired oxytocin, NAD metabolism, and sex-related neuromuscular impairment in PWS individuals with potential clinical implications.

Purpose: Childhood obesity, a pressing global health issue, significantly increases the risk of metabolic complications, including metabolic dysfunction associated with steatotic liver disease (MASLD). Accurate non-invasive tests for early detection and screening of steatosis are crucial. In this study, we explored the serum proteome, identifying proteins as potential biomarkers for inclusion in non-invasive steatosis diagnosis tests. Methods: Fifty-nine obese adolescents underwent ultrasonography to assess steatosis. Serum samples were collected and analyzed by targeted proteomics with the Proximity Extension Assay technology. Clinical and biochemical parameters were evaluated, and correlations among them, the individuated markers, and steatosis were performed. Receiver operating characteristic (ROC) curves were used to determine the steatosis diagnostic performance of the identified candidates, the fatty liver index (FLI), and their combination in a logistic regression model. Results: Significant differences were observed between subjects with and without steatosis in various clinical and biochemical parameters. Gender-related differences in the serum proteome were also noted. Five circulating proteins, including Cathepsin O (CTSO), Cadherin 2 (CDH2), and Prolyl endopeptidase (FAP), were identified as biomarkers for steatosis. CDH2, CTSO, Leukocyte Immunoglobulin Like Receptor A5 (LILRA5), BMI, waist circumference, HOMA-IR, and FLI, among others, significantly correlated with the steatosis degree. CDH2, FAP, and LDL combined in a logit model achieved a diagnostic performance with an AUC of 0.91 (95% CI 0.75–0.97, 100% sensitivity, 84% specificity). Conclusions: CDH2 and FAP combined with other clinical parameters, represent useful tools for accurate diagnosis of fatty liver, emphasizing the importance of integrating novel markers into diagnostic algorithms for MASLD.

In the context of the increasing number of obese individuals, a major problem is represented by obesity and malnutrition in children. This condition is mainly ascribable to unbalanced diets characterized by high intakes of fat and sugar. Childhood obesity and malnutrition are not only associated with concurrent pathologies but potentially compromise adult life. Considering the strict correlation among systemic metabolism, obesity, and skeletal muscle health, we wanted to study the impact of juvenile malnutrition on the adult skeletal muscle. To this aim, 3-week-old C56BL/6 female and male mice were fed for 20 weeks on a high-fat. high-sugar diet, and their muscles were subjected to a histological evaluation. MyHCs expression, glycogen content, intramyocellular lipids, mitochondrial activity, and capillary density were analyzed on serial sections to obtain the metabolic profile. Our observations indicate that a high-fat, high-sugar diet alters the metabolic profile of skeletal muscles in a sex-dependent way and induces the increase in type II fibers, mitochondrial activity, and lipid content in males, while reducing the capillary density in females. These data highlight the sex-dependent response to nutrition, calling for the development of specific strategies and for a systematic inclusion of female subjects in basic and applied research in this field.