Materials derived from the human elastin-like polypeptide fusion with an antimicrobial peptide strongly promote cell adhesion

Protein and peptide materials have attracted great interest in recent years, especially for biological applications, in light of their possibility to easily encode bioactivity whilst maintaining cytocompatibility and biodegradability. Heterologous recombinant expression to produce antimicrobial peptides is increasingly considered a convenient alternative for the transition from conventional methods to more sustainable production systems. The human elastin-like polypeptide (HELP) has proven to be a valuable fusion carrier, and due to its cutting-edge properties, biomimetic materials with antimicrobial capacity have been successfully developed. In this work, we have taken advantage of this platform to produce a difficult-to-synthesise sequence as that of the human β-defensin 1 (hBD1), an amphipathic cationic peptide with structural folding constraints relevant to its bioactivity. In the design of the gene, highly specific endoproteinases recognition sites were introduced to release the active forms of hBD1. After the expression and purification of the new fusion construct, its biological activity was evaluated. It was found that both the fusion biopolymer and the released active forms can inhibit the growth of Escherichia coli in redox environments. Remarkably, 2D and 3D materials derived from the biopolymer showed a strong cell adhesion-promoting activity. These results suggest that HELP represents a multitasking platform that not only facilitates the production of bioactive domains and derived materials but could also pave the way for the development of new approaches to study biological interactions at the molecular level

Physicochemical Characterization of a Biomimetic, Elastin-Inspired Polypeptide with Enhanced Thermoresponsive Properties and Improved Cell Adhesion

Genetic engineering allows fine-tuning and controlling protein properties, thus exploiting the new derivatives to obtain novel materials and systems with improved capacity to actively interact with biological systems. The elastin-like polypeptides are tunable recombinant biopolymers that have proven to be ideal candidates for realizing bioactive interfaces that can interact with biological systems. They are characterized by a thermoresponsive behavior that is strictly related to their peculiar amino acid sequence. We describe here the rational design of a new biopolymer inspired by elastin and the comparison of its physicochemical properties with those of another already characterized member of the same protein class. To assess the cytocompatibility, the behavior of cells of different origins toward these components was evaluated. Our study shows that the biomimetic strategy adopted to design new elastin-based recombinant polypeptides represents a versatile and valuable tool for the development of protein-based materials with improved properties and advanced functionality

Palmitate-induced lipotoxicity alters acetylation of multiple proteins in clonal β cells and human pancreatic islets

Type 2 diabetes is characterized by progressive β cell dysfunction, with lipotoxicity playing a possible pathogenetic role. Palmitate is often used to examine the direct effects of lipotoxicity and it may cause mitochondrial alterations by activating protein acetylation. However, it is unknown whether palmitate influences protein acetylation in β cells. We investigated lysine acetylation in mitochondrial proteins from INS-1E β cells (INS-1E) and in proteins from human pancreatic islets (HPI) after 24 h palmitate exposure. First, we confirmed that palmitate damages β cells and demonstrated that chemical inhibition of deacetylation also impairs INS-1E function and survival. Then, by 2-D gel electrophoresis, Western Blot and Liquid Chromatography-Mass Spectrometry we evaluated the effects of palmitate on protein acetylation. In mitochondrial preparations from palmitate-treated INS-1E, 32 acetylated spots were detected, with 13 proteins resulting over-acetylated. In HPI, 136 acetylated proteins were found, of which 11 were over-acetylated upon culture with palmitate. Interestingly, three proteins, glutamate dehydrogenase, mitochondrial superoxide dismutase, and SREBP-1, were over-acetylated in both INS-1E and HPI. Therefore, prolonged exposure to palmitate induces changes in β cell protein lysine acetylation and this modification could play a role in causing β cell damage. Dysregulated acetylation may be a target to counteract palmitate-induced β cell lipotoxicity

4-(3-Phenyl-4-(3,4,5-trimethoxybenzoyl)-1H-pyrrol-1-yl)benzenesulfonamide, a Novel Carbonic Anhydrase and Wnt/β-Catenin Signaling Pathway Dual-Targeting Inhibitor with Potent Activity against Multidrug Resistant Cancer Cells

We synthesized new pyrrole and indole derivatives as human carbonic anhydrase (hCA) inhibitors with the potential to inhibit the Wnt/β-catenin signaling pathway. The presence of both N1-(4-sulfonamidophenyl) and 3-(3,4,5-trimethoxyphenyl) substituents was essential for strong hCA inhibitors. The most potent hCA XII inhibitor 15 (Ki = 6.8 nM) suppressed the Wnt/β-catenin signaling pathway and its target genes MYC, Fgf20, and Sall4 and exhibited the typical markers of apoptosis, cleaved poly(ADP-ribose)polymerase, and cleaved caspase-3. Compound 15 showed strong inhibition of viability in a panel of cancer cells, including colorectal cancer and triple-negative breast cancer cells, was effective against the NCI/ADR-RES DOX-resistant cell line, and restored the sensitivity to doxorubicin (DOX) in HT29/DX and MDCK/P-gp cells. Compound 15 is a novel dual-targeting compound with activity against hCA and Wnt/β-catenin. It thus has a broad targeting spectrum and is an anticancer agent with specific potential in P-glycoprotein overexpressing cell lines

Calprotectin in nasal secretion: a new biomarker of non-type 2 inflammation in CRSwNP

We analysed calprotectin in sinonasal secretions of different chronic rhinosinusitis with nasal polyps (CRSwNP) endotypes to assess its role as a biomarker of non-type 2 inflammation.We included primary diffuse CRSwNP patients (n = 41) and three different control groups [non-allergic rhinitis (NAR) (n = 13), non-allergic eosinophilic syndrome (NARES) (n = 10) and healthy subjects (n = 12)]. Calprotectin levels were detected in nasal secretions using a chemiluminescent immunoassay (CLIA).Calprotectin levels in nasal secretions were significantly higher in all non-type 2 endotypes of CRSwNP compared to healthy controls (p0.05). In contrast, in type-2 CRSwNP calprotectin was significantly lower compared to controls (p0.05). A significant correlation between calprotectin levels and neutrophilic count/HPF was found in CRSwNP (p0.01). Clinically, mean levels of calprotectin and neutrophilia were significantly higher in patients who previously underwent 3 or more endoscopic sinus surgeries (p0.05).Calprotectin in nasal secretions may be a biomarker of non-type 2 inflammation. Low levels of calprotectin are indicative of a type-2 immune response in both CRSwNP and non-allergic rhinitis. We observed that calprotectin levels significantly increased based on the number of previous surgeries.Calprotectina nel secreto nasale: nuovo biomarker di infiammazione non-tipo 2 nella poliposi nasale.Abbiamo analizzato i livelli di calprotectina nelle secrezioni nasali di diversi endotipi di CRSwNP, per valutarne il ruolo di marcatore di infiammazione non-tipo 2.Abbiamo incluso pazienti con CRSwNP diffusa primaria (n = 41) e tre diversi gruppi di controllo [rinite non allergica (NAR) (n = 13), rinite eosinofila non allergica (NARES) (n = 10) e soggetti sani (n = 12)]. I livelli di calprotectina sono stati dosati mediante test di chemoluminescenza.I livelli di calprotectina sono risultati significativamente più alti in tutti gli endotipi non-tipo 2 di CRSwNP, rispetto ai controlli sani (p0,05). Al contrario, nella CRSwNP tipo 2 essa è risultata inferiore rispetto ai controlli (p0,05). È stata riscontrata una correlazione significativa tra i livelli di calprotectina e la conta dei neutrofili/HPF (p0.01). I livelli medi di calprotectina aumentano inoltre nei pazienti in precedenza sottoposti a 3 o più ESS (p0,05).La calprotectina nelle secrezioni nasali può essere considerata un biomarcatore di infiammazione non-tipo 2 mentre bassi livelli di calprotectina sono indicativi di immunoflogosi di tipo 2. Abbiamo infine osservato che i livelli di calprotectina aumentano significativamente in relazione al numero di interventi chirurgici pregressi

Assessing the pathogenicity of BRCA1/2 variants of unknown significance: Relevance and challenges for breast cancer precision medicine

IntroductionBreast cancer (BC) is the leading cause of cancer-related death in women worldwide. Pathogenic variants in BRCA1 and BRCA2 genes account for approximately 50% of all hereditary BC, with 60-80% of patients characterized by Triple Negative Breast Cancer (TNBC) at an early stage phenotype. The identification of a pathogenic BRCA1/2 variant has important and expanding roles in risk-reducing surgeries, treatment planning, and familial surveillance. Otherwise, finding unclassified Variants of Unknown Significance (VUS) limits the clinical utility of the molecular test, leading to an “imprecise medicine”.MethodsWe reported the explanatory example of the BRCA1 c.5057A>C, p.(His1686Pro) VUS identified in a patient with TNBC. We integrated data from family history and clinic-pathological evaluations, genetic analyses, and bioinformatics in silico investigations to evaluate the VUS classification.ResultsOur evaluation posed evidences for the pathogenicity significance of the investigated VUS: 1) association of the BRCA1 variant to cancer-affected members of the family; 2) absence of another high-risk mutation; 3) multiple indirect evidences derived from gene and protein structural analysis.DiscussionIn line with the ongoing efforts to uncertain variants classification, we speculated about the relevance of an in-depth assessment of pathogenicity of BRCA1/2 VUS for a personalized management of patients with BC. We underlined that the efficient integration of clinical data with the widest number of supporting molecular evidences should be adopted for the proper management of patients, with the final aim of effectively guide the best prognostic and therapeutic paths

Broadly directed virus-specific CD4+ T cell responses are primed during acute hepatitis C infection, but rapidly disappear from human blood with viral persistence

Vigorous proliferative CD4+ T cell responses are the hallmark of spontaneous clearance of acute hepatitis C virus (HCV) infection, whereas comparable responses are absent in chronically evolving infection. Here, we comprehensively characterized the breadth, specificity, and quality of the HCV-specific CD4+ T cell response in 31 patients with acute HCV infection and varying clinical outcomes. We analyzed in vitro T cell expansion in the presence of interleukin-2, and ex vivo staining with HCV peptide-loaded MHC class II tetramers. Surprisingly, broadly directed HCV-specific CD4+ T cell responses were universally detectable at early stages of infection, regardless of the clinical outcome. However, persistent viremia was associated with early proliferative defects of the HCV-specific CD4+ T cells, followed by rapid deletion of the HCV-specific response. Only early initiation of antiviral therapy was able to preserve CD4+ T cell responses in acute, chronically evolving infection. Our results challenge the paradigm that HCV persistence is the result of a failure to prime HCV-specific CD4+ T cells. Instead, broadly directed HCV-specific CD4+ T cell responses are usually generated, but rapid exhaustion and deletion of these cells occurs in the majority of patients. The data further suggest a short window of opportunity to prevent the loss of CD4+ T cell responses through antiviral therapy

The clinical effectiveness of an integrated multidisciplinary evidence-based program to prevent intraoperative pressure injuries in high-risk children undergoing long-duration surgical procedures: a quality improvement study

The prevention of hospital-acquired pressure injuries (HAPIs) in children undergoing long-duration surgical procedures is of critical importance due to the potential for catastrophic sequelae of these generally preventable injuries for the child and their family. Long-duration surgical procedures in children have the potential to result in high rates of HAPI due to physiological factors and the difficulty or impossibility of repositioning these patients intraoperatively. We developed and implemented a multi-modal, multi-disciplinary translational HAPI prevention quality improvement program at a large European Paediatric University Teaching Hospital. The intervention comprised the establishment of wound prevention teams, modified HAPI risk assessment tools, specific education, and the use of prophylactic dressings and fluidized positioners during long-duration surgical procedures. As part of the evaluation of the effectiveness of the program in reducing intraoperative HAPI, we conducted a prospective cohort study of 200 children undergoing long-duration surgical procedures and compared their outcomes with a matched historical cohort of 200 children who had undergone similar surgery the previous year. The findings demonstrated a reduction in HAPI in the intervention cohort of 80% (p < 0.01) compared to the comparator group when controlling for age, pathology, comorbidity, and surgical duration. We believe that the findings demonstrate that it is possible to significantly decrease HAPI incidence in these highly vulnerable children by using an evidence-based, multi-modal, multidisciplinary HAPI prevention strategy

How future surgery will benefit from SARS-COV-2-related measures: a SPIGC survey conveying the perspective of Italian surgeons

COVID-19 negatively affected surgical activity, but the potential benefits resulting from adopted measures remain unclear. The aim of this study was to evaluate the change in surgical activity and potential benefit from COVID-19 measures in perspective of Italian surgeons on behalf of SPIGC. A nationwide online survey on surgical practice before, during, and after COVID-19 pandemic was conducted in March-April 2022 (NCT:05323851). Effects of COVID-19 hospital-related measures on surgical patients' management and personal professional development across surgical specialties were explored. Data on demographics, pre-operative/peri-operative/post-operative management, and professional development were collected. Outcomes were matched with the corresponding volume. Four hundred and seventy-three respondents were included in final analysis across 14 surgical specialties. Since SARS-CoV-2 pandemic, application of telematic consultations (4.1% vs. 21.6%; p < 0.0001) and diagnostic evaluations (16.4% vs. 42.2%; p < 0.0001) increased. Elective surgical activities significantly reduced and surgeons opted more frequently for conservative management with a possible indication for elective (26.3% vs. 35.7%; p < 0.0001) or urgent (20.4% vs. 38.5%; p < 0.0001) surgery. All new COVID-related measures are perceived to be maintained in the future. Surgeons' personal education online increased from 12.6% (pre-COVID) to 86.6% (post-COVID; p < 0.0001). Online educational activities are considered a beneficial effect from COVID pandemic (56.4%). COVID-19 had a great impact on surgical specialties, with significant reduction of operation volume. However, some forced changes turned out to be benefits. Isolation measures pushed the use of telemedicine and telemetric devices for outpatient practice and favored communication for educational purposes and surgeon-patient/family communication. From the Italian surgeons' perspective, COVID-related measures will continue to influence future surgical clinical practice