Anna Negroni

Electrospinning is an advanced manufacturing strategy used to create innovative medical devices from continuous nanoscale fibers that is endowed with tunable biological, chemical, and physical properties. Innovative medical patches manufactured entirely by electrospinning are discussed in this paper, using a specific plant-derived formulation “1 Primary Wound Dressing©” (1-PWD) as an active pharmaceutical ingredient (API). 1-PWD is composed of neem oil (Azadirachta indica A. Juss.) and the oily extracts of Hypericum perforatum (L.) flowers, according to the formulation patented by the ENEA of proven therapeutic efficacy as wound dressings. The goal of this work is to encapsulate this API and demonstrate that its slow release from an engineered electrospun patch can increase the therapeutic efficacy for wound healing. The prototyped patch is a three-layer core–shell membrane, with a core made of fibers from a 1-PWD-PEO blend, enveloped within two external layers made of medical-grade polycaprolactone (PCL), ensuring mechanical strength and integrity during manipulation. The system was characterized via electron microscopy (SEM) and chemical and contact angle tests. The encapsulation, release, and efficacy of the API were confirmed by FTIR and LC-HRMS and were validated via in vitro toxicology and scratch assays.

Extracellular High-mobility group box 1 (HMGB1) contributes to the pathogenesis of inflammatory disorders, including inflammatory bowel diseases (IBD). Poly (ADP-ribose) polymerase 1 (PARP1) has been recently reported to promote HMGB1 acetylation and its secretion outside cells. In this study, the relationship between HMGB1 and PARP1 in controlling intestinal inflammation was explored. C57BL6/J wild type (WT) and PARP1^−/− mice were treated with DSS to induce acute colitis, or with the DSS and PARP1 inhibitor, PJ34. Human intestinal organoids, which are originated from ulcerative colitis (UC) patients, were exposed to pro-inflammatory cytokines (INFγ + TNFα) to induce intestinal inflammation, or coexposed to cytokines and PJ34. Results show that PARP1^−/− mice develop less severe colitis than WT mice, evidenced by a significant decrease in fecal and serum HMGB1, and, similarly, treating WT mice with PJ34 reduces the secreted HMGB1. The exposure of intestinal organoids to pro-inflammatory cytokines results in PARP1 activation and HMGB1 secretion; nevertheless, the co-exposure to PJ34, significantly reduces the release of HMGB1, improving inflammation and oxidative stress. Finally, HMGB1 release during inflammation is associated with its PARP1-induced PARylation in RAW264.7 cells. These findings offer novel evidence that PARP1 favors HMGB1 secretion in intestinal inflammation and suggest that impairing PARP1 might be a novel approach to manage IBD.

A tight relationship between gut-liver diseases and brain functions has recently emerged. Bile acid (BA) receptors, bacterial-derived molecules and the blood-brain barrier (BBB) play key roles in this association. This study was aimed to evaluate how non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) impact the BA receptors Farnesoid X receptor (FXR) and Takeda G-protein coupled receptor 5 (TGR5) expression in the brain and to correlate these effects with circulating BAs composition, BBB integrity and neuroinflammation. A mouse model of NAFLD was set up by a high-fat and sugar diet, and NASH was induced with the supplementation of dextran-sulfate-sodium (DSS) in drinking water. FXR, TGR5 and ionized calcium-binding adaptor molecule 1 (Iba-1) expression in the brain was detected by immunohistochemistry, while Zonula occludens (ZO)-1, Occludin and Plasmalemmal Vesicle Associated Protein-1 (PV-1) were analyzed by immunofluorescence. Biochemical analyses investigated serum BA composition, lipopolysaccharide-binding protein (LBP) and S100β protein (S100β) levels. Results showed a down-regulation of FXR in NASH and an up-regulation of TGR5 and Iba-1 in the cortex and hippocampus in both treated groups as compared to the control group. The BA composition was altered in the serum of both treated groups, and LBP and S100β were significantly augmented in NASH. ZO-1 and Occludin were attenuated in the brain capillary endothelial cells of both treated groups versus the control group. We demonstrated that NAFLD and NASH provoke different grades of brain dysfunction, which are characterized by the altered expression of BA receptors, FXR and TGR5, and activation of microglia. These effects are somewhat promoted by a modification of circulating BAs composition and by an increase in LBP that concur to damage BBB, thus favoring neuroinflammation.

Crohn’s disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract. Chronic inflammation is the main factor leading to intestinal fibrosis, resulting in recurrent stenosis, especially in CD patients. Currently, the underlying molecular mechanisms of fibrosis are still unclear. ZNF281 is a zinc-finger transcriptional regulator that has been characterized as an epithelial-to-mesenchymal transition (EMT)-inducing transcription factor, suggesting its involvement in the regulation of pluripotency, stemness, and cancer. The aim of this study is to investigate in vivo and in vitro the role of ZNF281 in intestinal fibrogenesis. Intestinal fibrosis was studied in vivo in C57BL/6J mice with chronic colitis induced by two or three cycles of administration of dextran sulfate sodium (DSS). The contribution of ZNF281 to gut fibrosis was studied in vitro in the human colon fibroblast cell line CCD-18Co, activated by the pro-fibrotic cytokine TGFβ1. ZNF281 was downregulated by siRNA transfection, and RNA-sequencing was performed to identify genes regulated by TGFβ1 in activated colon fibroblasts via ZNF281. Results showed a marked increase of ZNF281 in in vivo murine fibrotic colon as well as in in vitro human colon fibroblasts activated by TGFβ1. Moreover, abrogation of ZNF281 in TGFβ1-treated fibroblasts affected the expression of genes belonging to specific pathways linked to fibroblast activation and differentiation into myofibroblasts. We demonstrated that ZNF281 is a key regulator of colon fibroblast activation and myofibroblast differentiation upon fibrotic stimuli by transcriptionally controlling extracellular matrix (ECM) composition, remodeling, and cell contraction, highlighting a new role in the onset and progression of gut fibrosis.

Microalgae are natural sources of valuable bioactive compounds, such as polyunsaturated fatty acids (PUFAs), that show antioxidant, anti-inflammatory, anticancer and antimicrobial activities. The marine microalga Isochrysis galbana (I. galbana) is extremely rich in ω3 PUFAs, mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Probiotics are currently suggested as adjuvant therapy in the management of diseases associated with gut dysbiosis. The Lactobacillus reuteri (L. reuteri), one of the most widely used probiotics, has been shown to produce multiple beneficial effects on host health. The present study aimed to present an innovative method for growing the probiotic L. reuteri in the raw seaweed extracts from I. galbana as an alternative to the conventional medium, under conditions of oxygen deprivation (anaerobiosis). As a result, the microalga I. galbana was shown for the first time to be an excellent culture medium for growing L. reuteri. Furthermore, the gas-chromatography mass-spectrometry analysis showed that the microalga-derived ω3 PUFAs were still available after the fermentation by L. reuteri. Accordingly, the fermented compound (FC), obtained from the growth of L. reuteri in I. galbana in anaerobiosis, was able to significantly reduce the adhesiveness and invasiveness of the harmful adherent-invasive Escherichia coli to intestinal epithelial cells, due to a cooperative effect between L. reuteri and microalgae-released ω3 PUFAs. These findings open new perspectives in the use of unicellular microalgae as growth medium for probiotics and in the production of biofunctional compounds.

Necroptosis is a caspases-independent programmed cell death displaying intermediate features between necrosis and apoptosis. Albeit some physiological roles during embryonic development such tissue homeostasis and innate immune response are documented, necroptosis is mainly considered a pro-inflammatory cell death. Key actors of necroptosis are the receptor-interacting-protein-kinases, RIPK1 and RIPK3, and their target, the mixed-lineage-kinase-domain-like protein, MLKL. The intestinal epithelium has one of the highest rates of cellular turnover in a process that is tightly regulated. Altered necroptosis at the intestinal epithelium leads to uncontrolled microbial translocation and deleterious inflammation. Indeed, necroptosis plays a role in many disease conditions and inhibiting necroptosis is currently considered a promising therapeutic strategy. In this review, we focus on the molecular mechanisms of necroptosis as well as its involvement in human diseases. We also discuss the present developing therapies that target necroptosis machinery.

Necroptosis is a caspases-independent form of programmed cell death exhibiting intermediate features between necrosis and apoptosis. Albeit some physiological roles during embryonic development, tissue homeostasis and innate immune response are documented, necroptosis is mainly considered a pro-inflammatory cell death. Key actors of necroptosis are the receptor-interacting-protein-kinases, RIPK1 and RIPK3, and their target, the mixed-lineage-kinase-domain-like protein, MLKL. The intestinal epithelium has one of the highest rates of cellular turnover in a process that is tightly regulated. Altered necroptosis at the intestinal epithelium leads to uncontrolled microbial translocation and deleterious inflammation. Indeed, necroptosis has been associated to chronic inflammatory diseases and cancer. Drugs that inhibit necroptosis could, therefore, be used therapeutically for the treatment of these diseases, and researches to develop such inhibitors are already underway. In this Review, we outline pathways for necroptosis and its role in chronic inflammation and cancer. We also discuss current and developing therapies that target necroptosis machinery.

Although the cystic fibrosis (CF) lung microbiota has been characterized in several studies, little is still known about the temporal changes occurring at the whole microbiome level using untargeted metagenomic analysis. The aim of this study was to investigate the taxonomic and functional temporal dynamics of the lower airway microbiome in a cohort of CF patients. Multiple sputum samples were collected over 15 months from 22 patients with advanced lung disease regularly attending three Italian CF Centers, given a total of 79 samples. DNA extracted from samples was subjected to shotgun metagenomic sequencing allowing both strain-level taxonomic profiling and assessment of the functional metagenomic repertoire. High inter-patient taxonomic heterogeneity was found with short-term compositional changes across clinical status. Each patient exhibited distinct sputum microbial communities at the taxonomic level, and strain-specific colonization of both traditional and atypical CF pathogens. A large core set of genes, including antibiotic resistance genes, were shared across patients despite observed differences in clinical status, and consistently detected in the lung microbiome of all subjects independently from known antibiotic exposure. In conclusion, an overall stability in the microbiome-associated genes was found despite taxonomic fluctuations of the communities.

Objectives: The gut‐liver axis has been recently investigated in depth in relation to intestinal and hepatic diseases. Key actors are bile acid (BA) receptors, as farnesoid‐X‐receptor (FXR), pregnane‐X‐receptor (PXR), and G‐protein‐coupled‐receptor (GPCR; TGR5), that control a broad range of metabolic processes as well as inflammation and fibrosis. The present study aims to investigate the impact of intestinal inflammation on liver health with a focus on FXR, PXR, and TGR5 expression. The strategy to improve liver health by reducing gut inflammation is also considered. Modulation of BA receptors in the inflamed colonic tissues of inflammatory bowel disease (IBD) pediatric patients is analyzed.Methods: A dextran sodium sulphate (DSS) colitis animal model was built. Co‐cultures with Caco2 and HepG2 cell lines were set up. Modulation of BA receptors in biopsies of IBD pediatric patients was assessed by real‐time PCR and immunohistochemistry.Results: Histology showed inflammatory cell infiltration in the liver of DSS mice, where FXR and PXR were significantly decreased and oxidative stress was increased. Exposure of Caco2 to inflammatory stimuli resulted in the reduction of BA receptor expression in HepG2. Caco2 treatment with dipotassium glycyrrhizate (DPG) reduced these effects on liver cells. Inflamed colon of patients showed altered FXR, PXR, and TGR5 expression.Conclusions: This study strongly suggests that gut inflammation affects hepatic cells by altering BA receptor levels as well as increasing the production of pro‐inflammatory cytokines and oxidative stress. Hence, reducing gut inflammation is needed not only to improve the intestinal disease but also to protect the liver.