Biology subjects

Perugini, J.

Publications and source records attributed to Perugini, J..

6 recordsLinked to original sources

bioRxiv2026

Ciliary neurotrophic factor curbs palmitate-induced inflammatory and metabolic changes in NPY/AgRP hypothalamic neurons

Introduction: In both humans and experimental animals, obesity is associated with hypothalamic inflammation and neuronal metabolic dysfunction, which contribute to disrupting body energy homeostasis. Although ciliary neurotrophic factor (CNTF) is a neuroprotective cytokine with recognized anti-obesity effects in peripheral metabolic tissues, its role in obesity-associated hypothalamic dysfunction is still poorly characterized. We investigated whether CNTF exerts any direct effects on mHypoE-N41 (N41) hypothalamic neurons and whether it modulates the effects induced by palmitate (PA), an established model of obesity-associated neuronal metabolic dysfunction. Methods: N41 cells were administered CNTF alone or combined with PA to assess whether CNTF affects PA-induced lipotoxic stress. Gene and protein expression was assessed by qRT-PCR, western blotting, and immunofluorescence analysis. Results: In N41 cells, CNTF activated the STAT3, ERK and AMPK signaling pathways. It also upregulated genes involved in mitochondrial metabolism and fatty acid oxidation, including Ucp2, Cpt1 and Pgc1, as well as the proliferation-associated markers Ki-67 and PCNA. PA treatment induced inflammatory and endoplasmic reticulum stress responses and impaired insulin signaling. In PA-treated N41 cells, CNTF significantly attenuated the expression of pro-inflammatory markers Tnf, Mcp1 and Nfkb, restored insulin-induced AKT phosphorylation, and partially reversed PA-induced changes in the expression of metabolic genes Ucp2 and Cpt1. Conclusions: Collectively, these findings indicate that CNTF exerts direct protective metabolic effects on hypothalamic N41 neurons subjected to lipotoxic stress and supports a role for it in the preservation of neuronal metabolic function under obesogenic conditions.

molecular biology↗

bioRxiv2026

Plasma oxytocin measured by LC-MS/MS varies with life stage, sex, and obesity in mice

ObjectiveOxytocin (Oxt) assessment in plasma is challenging, and available data are contradictory. We aimed to assess circulating Oxt in mice by a validated nano-liquid chromatography/mass-spectrometry (nLC-MS/MS) protocol, combined with Oxt hypothalamic expression in different sex, life stages, and in diet-induced obesity. MethodsWe assessed plasma Oxt by nLC-MS/MS, Oxt hypothalamic expression by qPCR, and Oxt-immunoreactive neuron and fiber densities by immunohistochemistry and morphometric analyses in C57BL/6 mice at 21 and 60 days of life (p21 and p60, respectively). Mice in normo-fed condition and following 12 weeks of high-fat diet (HFD) were studied alongside food intake and hypothalamic expression of its regulators. ResultsCirculating Oxt does not vary based on sex at p21 and p60 but increases with aging. While hypothalamic Oxt mRNA expression followed the same trend across both sexes, Oxt neuron and fiber densities exhibited a similar trend only in females. Plasma vasopressin (Avp) followed Oxt trend in females but was opposite in males and was not mirrored by Avp mRNA hypothalamic expression. HFD-fed females were more resistant to weight gain compared to males and displayed higher Oxt plasma levels and hypothalamic expression. Sex dimorphism in food intake and hypothalamic expression of Avp and of key anorexigenic and orexigenic neuropeptides was detected. ConclusionsOxt plasma levels are higher in adulthood compared to weaning in mice of both sexes who displayed similar concentrations. Oxt plasma levels are mirrored by Oxt hypothalamic expression. In obesity, females display a lower increase in body weight but higher Oxt plasma levels than males. SignificanceThis study delineates the pattern of Oxt plasma levels across various age, sex and in obesity in mice. We overcome some limitations of previous research, which were often confounded by numerous factors, by reliably measuring circulating Oxt and integrating this with parallel analyses of Oxt gene expression, and Oxt neuron and immunoreactive fiber densities. Critically, this is among the few studies to employ LC-MS/MS specifically in mice. This novel, integrated perspective is relevant given the compelling evidence for Oxts role as an anti-obesity agent and its potential as a therapeutic target. A precise understanding of Oxt regulation in different conditions, backed by reliable methodologies, is indispensable for the rational development of effective treatments for obesity and related endocrine diseases.

neuroscience↗

bioRxiv2026

The ciliary neurotrophic factor induces Stat3 phosphorylation in distinctive cytotypes of organs involved in body metabolism: an immunohistochemical study

Administration of ciliary neurotrophic factor (CNTF) reduces food intake and body weight in both humans and experimental animals, where it also ameliorates hyperglycemia, hyperinsulinemia, and dyslipidemia. To exert its anti-obesogenic and anti-diabetogenic effects, CNTF targets brain feeding centers as well as multiple peripheral organs inducing the phosphorylation of the transcription factor signal transducer and activator of transcription 3 (p-STAT3). However, data showing which peripheral cytotypes are specifically targeted by exogenous CNTF in vivo in metabolically relevant organs are currently lacking. Here, we first evaluated the gene expression levels of the subunits of the tripartite CNTF receptor (Cntfr) complex, i.e., the Cntfr, the leukemia inhibitory factor receptor {beta} (Lifr{beta}) and the glycoprotein 130 (gp130), by quantitative real-time PCR in metabolically relevant organs of adult male mice: gastrointestinal (GI) tract, pancreas, liver, visceral and subcutaneous white (WAT) and interscapular brown adipose tissue (iBAT), skeletal muscle and the sciatic nerve. We then quantified p-STAT3 by Western blotting in these organs after intraperitoneal administration of CNTF (0.3 mg/kg) or saline. Finally, we mapped CNTF-responsive cells by immunohistochemistry, followed by morphometric quantification and confocal microscopy in both CNTF- and saline-treated mice. Lifr{beta} and gp130 were ubiquitously detected across all the investigated organs; the Cntfr showed the highest expression levels in the skeletal muscle, sciatic nerve, and iBAT, whereas it was found to be expressed to a lesser extent in the other sites. Administration of CNTF led to a significant increase of p-STAT3/STAT3 protein ratio in all organs examined, except the duodenum, and induced a distinctive pattern of cell nuclear p-STAT3 immunoreactivity. Notably, along the analyzed GI tract CNTF induced nuclear STAT3 phosphorylation in neurons of the submucosal and myenteric plexuses of the enteric nervous system and in contractile cells of the muscularis externa, where the response peaked in the mesenteric gut and colon. In the pancreas, CNTF triggered a higher activation within the endocrine component compared to the exocrine parenchyma. In the liver, CNTF induced STAT3 phosphorylation not only in parenchymal cells but also in sinusoids and resident macrophages. The cytokine activated p-STAT3 in subcutaneous and visceral white adipocytes, but also in brown adipocytes, with a prominent response observed in the beige subcutaneous adipocytes; adipose resident macrophages and endothelial cells of numerous blood vessels were also CNTF-responsive. Lastly, in skeletal muscle, a major site for glucose/lipid utilization, CNTF induced widespread nuclear p-STAT3 immunoreactivity in muscle fibers and in connective and Schwann cells of the peripheral nerves, including the sciatic nerve, supplying the gastrocnemius. In conclusion, our data indicate that CNTF acts across diverse cytotypes within metabolically relevant organs and tissues, likely fostering its peripheral metabolic effects through this cellular heterogeneity.

cell biology↗

bioRxiv2026

A2 and A1B in vitro milk digests: effects on in vitro leaky gut model and adipose cells

Obesity is associated with chronic low-grade systemic inflammation of adipose tissue and is often linked to intestinal epithelial barrier (IEB) dysfunction. The present study aimed to evaluate the effects of in vitro gastrointestinal digests of bovine milk containing A1B or A2 {beta}-casein variants on leaky IEB and adipocyte inflammation. Digests of A1B (DA1B) and A2 (DA2) milk were administered to an in vitro Caco-2/HT-29 intestinal cell co-culture mimicking a leaky gut. Intestinal absorbed fractions derived from A1B (MA1B) and A2 (MA2) were administered to hMADS adipocytes. DA1B and DA2 did not modify intestinal permeability, either in the absence or the presence of inflammation. DA1B reduced Claudin-1 mRNA, as well as zonula occludens-1 mRNA and protein expression. Both DA1B and DA2 increased interleukin-8 expression, but only DA1B increased tumor necrosis factor-. In human adipocytes, MA1B, and to a lesser extent MA2, increased the expression of pro-inflammatory markers monocyte chemoattractant protein-1 and interleukin-6, while reducing adiponectin levels. DA2 preserved in vitro leaky IEB integrity and exhibited a lower inflammatory potential in both leaky gut and adipocytes compared to DA1B. This study is the first to establish a link among A2 milk, leaky gut syndrome, and obesity.

molecular biology↗

bioRxiv2025

Role of Hypertrophic Adipocytes, Collagen VI and CD38 in Fat Fibrosis of Patients with Obesity

Fat fibrosis correlates to metabolic consequences in patients with obesity, and is due to three types of collagen: I and III (fibrillar) and VI (non-fibrillar). In this sudy the extent of fibrosis in obese patients (n 50) was significant only in visceral parenchymal fat (4.7% vs 2.5% in controls (n 15) P<0.0001) and not in subcutaneous fat. Electron microscopy, in vivo and in vitro data, suggested that obese adipocytes are responsible for fibrillar collagen (I and III) production. COL6 (gene producing the non fibrillar form) resulted less expressed. In line, patients with COL6 mutations, showed increased fibrotic tissue even in subcutaneous fat: about 6.5 times vs controls in the patient with the severe form (Ullrich) and 2.8 times in two patients with the milder form (Bethlem). Approximately 15% of obese adipocytes were dead (perilipin1 negative), and consequent infiltrating macrophages showed hyperexpression of CD38, an ectoenzyme implicated in systemic fibrosis. Correlations with gene expression confirmed the importance also of myofibroblasts and the extracellular matrix peptidase D. All together our data support a role for obese adipocytes in the fibrillar collagen production and evidentiate collagen VI and CD38 as new molecular determinants, reinforcing the idea of a multi-factorial origin of fat fibrosis.

cell biology↗

bioRxiv2021

Visceral Fat Inflammation and Fat Embolism are associated with Lung's Lipidic Hyaline Membranes in COVID-19 patients

BackgroundVisceral obesity is a critical determinant of severe coronavirus disease-2019 (COVID-19). Methods: In this study, we performed a comprehensive histomorphologic analysis of autoptic visceral adipose tissues (VAT), lungs and livers of 19 COVID-19 and 23 non-COVID-19 subjects. ResultsAlthough there were no between-groups differences in body-mass-index and adipocytes size, higher prevalence of CD68+ macrophages in COVID-19 subjects VAT was detected (p=0.005) and accompanied by crown-like structures presence, signs of adipocytes stress and death. Consistently, human adipocytes were successfully infected by SARS-CoV2 in vitro and displayed lower cell viability. Being VAT inflammation associated with lipids spill-over from dead adipocytes, we studied lipids distribution employing Oil-Red-O staining (ORO). Lipids were observed within lungs and livers interstitial spaces, macrophages, endothelial cells, and vessels lumen, features suggestive of fat embolism syndrome, more prevalent among COVID-19 individuals (p<0.001). Notably, signs of fat embolism were more prevalent among obese (p=0.03) independently of COVID-19 diagnosis, suggesting that such condition may be an obesity complication, exacerbated by SARS-CoV2 infection. Importantly, all infected subjects lungs presented lipids-rich (ORO+) hyaline membranes, formations associated with COVID-19-related pneumonia, present only in one control with non-COVID-19 pneumonia. ConclusionsThis study describes for the first time novel COVID-19-related features possibly underlying the unfavorable prognosis in obese SARS-CoV2-infected-subjects.

pathology↗