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Background:Human serum albumin (HSA), the most abundant protein in plasma, is a monomeric multi-domain macromolecule with at least nine binding sites for endogenous and exogenous ligands. HSA displays an extraordinary ligand binding capacity as a depot and carrier for many compounds including most acidic drugs. Consequently, HSA has the potential to influence the pharmacokinetics and pharmacodynamics of drugs.Objective:In this review, the structural determinants of drug binding to the multiple sites of HSA are analyzed and discussed in detail. Moreover, insight into the allosteric and competitive mechanisms underpinning drug recognition, delivery, and efficacy are analyzed and discussed.Conclusion:As several factors can modulate drug binding to HSA (e.g., concurrent administration of drugs competing for the same binding site, ligand binding to allosteric-coupled clefts, genetic inherited diseases, and post-translational modifications), ligand binding to HSA is relevant not only under physiological conditions, but also in the pharmacological therapy management.

Joint flexion to diminish the gap and avoid nerve grafts fell into disuse for decades, but recently attention for using this technique was regained. We report a case series of nerve suture under joint flexion, ultrasound monitoring, and physiotherapy. Our main objective was to determine how effective this multimodality treatment is.A retrospective review of 8 patients treated with direct repair with joint flexion was done. Depending on the affected nerve, either the knee or the elbow was flexed intraoperatively to determine if direct suturing was possible. After surgery, the limb was held immobilized. Through serial ultrasounds and a physiotherapy program, the limb was fully extended. If a nerve repair rupture was observed, the patient was re-operated and grafts were used.Of the eight nerve sutures analyzed, four sustained a nerve rupture revealed by US at an early stage, while four did not show any sign of dehiscence. In the patients in whom the nerve suture was preserved, an early and very good response was observed. Ultrasound was 100% accurate at identifying nerve suture preservation. Early detection of nerve failure permitted early re-do surgery using grafts without flexion, ultimately determining good final results.We observed a high rate of dehiscence in our group of patients treated with direct repair and joint flexion. We believe this was due to an incorrect use of the immobilization device, excessive movement, or a broken device. In opposition to this, we observed that applying direct nerve sutures and joint flexion offers unusually good and fast results. If this technique is employed, it is mandatory to closely monitor suture status with US, together with physiotherapy providing progressive, US-guided extension of the flexed joint. If nerve rupture occurs, the close monitoring dictated by this protocol should ensure the timely application of a successful graft repair.

The circadian gene Timeless (TIM) provides a molecular bridge between circadian and cell cycle/DNA replication regulatory systems and has been recently involved in human cancer development and progression. However, its functional role in colorectal cancer (CRC), the third leading cause of cancer-related deaths worldwide, has not been fully clarified yet. Here, the analysis of two independent CRC patient cohorts (total 1159 samples) reveals that loss of TIM expression is an unfavorable prognostic factor significantly correlated with advanced tumor stage, metastatic spreading, and microsatellite stability status. Genome-wide expression profiling, in vitro and in vivo experiments, revealed that TIM knockdown induces the activation of the epithelial-to-mesenchymal transition (EMT) program. Accordingly, the analysis of a large set of human samples showed that TIM expression inversely correlated with a previously established gene signature of canonical EMT markers (EMT score), and its ectopic silencing promotes migration, invasion, and acquisition of stem-like phenotype in CRC cells. Mechanistically, we found that loss of TIM expression unleashes ZEB1 expression that in turn drives the EMT program and enhances the aggressive behavior of CRC cells. Besides, the deranged TIM-ZEB1 axis sets off the accumulation of DNA damage and delays DNA damage recovery. Furthermore, we show that the aggressive and genetically unstable ‘CMS4 colorectal cancer molecular subtype’ is characterized by a lower expression of TIM and that patients with the combination of low-TIM/high-ZEB1 expression have a poorer outcome. In conclusion, our results as a whole suggest the engagement of an unedited TIM-ZEB1 axis in key pathological processes driving malignant phenotype acquisition in colorectal carcinogenesis. Thus, TIM-ZEB1 expression profiling could provide a robust prognostic biomarker in CRC patients, supporting targeted therapeutic strategies with better treatment selection and patients’ outcomes.

Background Challenges exist in the clinical treatment of luminal estrogen receptor α (ERα)-positive breast cancers (BCs) both to prevent resistance to endocrine therapy (ET) and to treat ET-resistant metastatic BCs (MBC). Therefore, we evaluated if kinases could be new targets for the treatment of luminal primary and MBCs. Methods ~ 170 kinase inhibitors were applied to MCF-7 cells either with adaptative or genetic resistance to ET drugs and both ERα levels and cell proliferation were measured. Robust-Z-score calculation identified AZD7762 (CHK1/CHK2 inhibitor) as a positive hit. Subsequently, Kaplan–Meier analyses of CHK1 and CHK2 impact on ERα-positive BC patients relapse-free-survival (RFS), bioinformatic evaluations of CHK1 and CHK2 expression and activation status as a function of ERα activation status as well as drug sensitivity studies in ERα-positive BC cell lines, validation of the impact of the ATR:CHK1 and ATM:CHK2 pathways on the control of ERα stability and BC cell proliferation via inhibitor- and siRNA-based approaches, identification of the molecular mechanism required for inhibitor-dependent ERα degradation in BC and the impact of CHK1 and CHK2 inhibition on the 17β-estradiol (E2):ERα signaling, synergy proliferation studies between ET-drugs and clinically relevant CHK1 inhibitors in different luminal BC cell lines, were performed. Results A reduced CHK1 expression correlates with a longer RFS in women with ERα-positive BCs. Interestingly, women carrying luminal A BC display an extended RFS when expressing low CHK1 levels. Accordingly, CHK1 and ERα activations are correlated in ERα-positive BC cell lines, and the ATR:CHK1 pathway controls ERα stability and cell proliferation in luminal A BC cells. Mechanistically, the generation of DNA replication stress rather than DNA damage induced by ATR:CHK1 pathway inhibition is a prerequisite for ERα degradation. Furthermore, CHK1 inhibition interferes with E2:ERα signaling to cell proliferation, and drugs approved for clinical treatment of primary and MBC (4OH-tamoxifen and the CDK4/CDK6 inhibitors abemaciclib and palbociclib) exert synergic effects with the CHK1 inhibitors in clinical trials for the treatment of solid tumors (AZD7762, MK8776, prexasertib) in preventing the proliferation of cells modeling primary and MBC. Conclusions CHK1 could be considered as an appealing novel pharmacological target for the treatment of luminal primary and MBCs.

Thalassemias (α, β, γ, δ, δβ, and εγδβ) are the most common genetic disorders worldwide and constitute a heterogeneous group of hereditary diseases characterized by the deficient synthesis of one or more hemoglobin (Hb) chain(s). This leads to the accumulation of unstable non-thalassemic Hb chains, which precipitate and cause intramedullary destruction of erythroid precursors and premature lysis of red blood cells (RBC) in the peripheral blood. Non-thalassemic Hbs display high oxygen affinity and no cooperativity. Thalassemias result from many different genetic and molecular defects leading to either severe or clinically silent hematologic phenotypes. Thalassemias α and β are particularly diffused in the regions spanning from the Mediterranean basin through the Middle East, Indian subcontinent, Burma, Southeast Asia, Melanesia, and the Pacific Islands, whereas δβ-thalassemia is prevalent in some Mediterranean regions including Italy, Greece, and Turkey. Although in the world thalassemia and malaria areas overlap apparently, the RBC protection against malaria parasites is openly debated. Here, we provide an overview of the historical, geographic, genetic, structural, and molecular pathophysiological aspects of thalassemias. Moreover, attention has been paid to molecular and epigenetic pathways regulating globin gene expression and globin switching. Challenges of conventional standard treatments, including RBC transfusions and iron chelation therapy, splenectomy and hematopoietic stem cell transplantation from normal donors are reported. Finally, the progress made by rapidly evolving fields of gene therapy and gene editing strategies, already in pre-clinical and clinical evaluation, and future challenges as novel curative treatments for thalassemia are discussed.

Serum albumin (SA) is the most abundant protein in plasma and represents the main carrier of endogenous and exogenous compounds. Several evidence supports the notion that SA binds single and double-stranded deoxynucleotides and ribonucleotides at two sites, with values of the dissociation equilibrium constant (i.e., K-d) ranging from micromolar to nanomolar values. This can be relevant from a physiological and pathological point of view, as in human plasma circulates cell-free nucleic acids (cfNAs), released by different tissues via apoptosis, necrosis, and secretions, circulates as single and double-stranded NAs. Albeit SA shows low hydrolytic reactivity toward DNA and RNA, the high plasma concentration of this protein and the occurrence of several SA receptors may be pivotal for sequestering and hydrolyzing cfNAs. Therefore, pathological conditions like cancer, characterized by altered levels of human SA or by altered SA post-translational modifications, may influence cfNAs distribution and metabolism. Besides, the stability, solubility, biocompatibility, and low immunogenicity make SA a golden share for biotechnological applications related to the delivery of therapeutic NAs (TNAs). Indeed, pre-clinical studies report the therapeutic potential of SA:TNAs complexes in precision cancer therapy. Here, the molecular and biotechnological implications of SA:NAs interaction are discussed, highlighting new perspectives on SA plasmatic functions.

The lockdown due to the coronavirus outbreak in 2019 (COVID‐19) has caused psychological distress and cognitive discomfort for emerging adults, who have experienced increased rumination, intrusive thoughts and cognitive failures. States of mindfulness and being in the moment can prevent anxiety and fear associated with the lockdown and alleviate ruminative and automatic negative thinking. This longitudinal study investigated the role of mindfulness before and during the COVID‐19 lockdown in protecting a sample of emerging adults from experiencing cognitive failures, intrusive thoughts and rumination and examined how lockdown‐related variables—emotions, socio‐economic status and housing conditions—influenced this mindfulness profile. The results showed overall more cognitive failures and rumination during lockdown, especially in participants whose mindfulness status diminished. However, these signs of cognitive distress remained stable or decreased among participants who reported improved changes in mindfulness. Financial difficulties and a reduced sense of privacy associated with the lockdown predicted lower stability in mindfulness profiles. The state of being fully aware of what is happening in the present moment may be helpful in reducing cognitive discomfort and psychological maladjustment, especially during stressful times such as lockdowns.

Background and aims Precise evaluation of resting metabolic rate (RMR) is critical, especially for seniors in syndromes conditions. The study aimed to compare different methods and devices to evaluate the resting metabolic rate and assess them’ reliability in Brazilian women with metabolic syndrome. Methods A single-center prospective study with forty elderly postmenopausal women was performed to verify the reliability of indirect calorimetry (IC) versus Bioimpedance (BIA) on RMR fluctuations for an interval length of six months. Results Measurements showed a high correlation between devices at baseline [BIA vs IC, intraclass correlation coefficient (ICC) = 0.906 (0.822–0.950)]. Surprisingly, a high correlation was kept between BIA and IC after six months [BIA vs. IC, ICC = 0.909 (0.829–0.952)]. The results suggest that both BIA and IC are excellent strategies to measure RMR in elderly postmenopausal women and with metabolic syndrome. Conclusions However, the BIA method presents greater convenience, optimizes patients’ time, and does not require prolonged fasting to obtain good reliable results compared to IC.

Human serum albumin (HSA) is the most abundant protein in plasma, contributing actively to oncotic pressure maintenance and fluid distribution between body compartments. HSA acts as the main carrier of fatty acids, recognizes metal ions, affects pharmacokinetics of many drugs, provides the metabolic modification of some ligands, renders potential toxins harmless, accounts for most of the anti-oxidant capacity of human plasma, and displays esterase, enolase, glucuronidase, and peroxidase (pseudo)-enzymatic activities. HSA-based catalysis is physiologically relevant, affecting the metabolism of endogenous and exogenous compounds including proteins, lipids, cholesterol, reactive oxygen species (ROS), and drugs. Catalytic properties of HSA are modulated by allosteric effectors, competitive inhibitors, chemical modifications, pathological conditions, and aging. HSA displays anti-oxidant properties and is critical for plasma detoxification from toxic agents and for pro-drugs activation. The enzymatic properties of HSA can be also exploited by chemical industries as a scaffold to produce libraries of catalysts with improved proficiency and stereoselectivity for water decontamination from poisonous agents and environmental contaminants, in the so called “green chemistry” field. Here, an overview of the intrinsic and metal dependent (pseudo-)enzymatic properties of HSA is reported to highlight the roles played by this multifaced protein.

Purpose: Recently, it has been demonstrated that Raman spectroscopy is able to differentiate between healthy parathyroid tissues and parathyroid adenoma based on the basis of a specific molecular fingerprint. However, to our knowledge, no previous studies have been performed to evaluate the metabolic profile of parathyroid adenoma. Therefore, we designed a proof of concept study aimed to investigate the glucose/fatty acid metabolisms, in addition to the mitochondrial changes, in solitary parathyroid adenoma and in healthy parathyroid glands. Methods: Nine females with primary hyperparathyroidism due to a solitary parathyroid adenoma and formal surgical indication for parathyroidectomy have been enrolled. At the time of surgery, the removed specimens were immediately submitted unfixed and a tissue slice of about 0.5 cm in diameter was obtained from the nodular lesion. The expression of selected metabolic enzymes and proteins has been evaluated by western blot analysis, using human parathyroid whole tissue lysates as control. Results: Data obtained highlighted an increase, compared with the healthy group, of: (i) the glucose uptake by the GLUT-1 receptor and its phosphorylation by hexokinase II (HXKII); (ii) the expression of 3-phosphoglycerate dehydrogenase (3-PGDH) and glucose-6-phosphate dehydrogenase (G6PD); (iii) lipids biosynthesis; and (iv) cytochrome c expression. Conclusions: Our findings highlight for the first time the parathyroid adenoma metabolic hallmarks that could represent potential molecular targets usable for the development of new pharmacological treatments, allowing to reduce surgical parathyroidectomy.

Background The phrenic nerve has been extensively reported to be a very powerful source of transferable axons in brachial plexus injuries. The most used technique used is supraclavicular sectioning of this nerve. More recently, video-assisted thoracoscopic techniques have been reported as a good alternative, since harvesting a longer phrenic nerve avoids the need of an interposed graft. Objective To compare grafting vs phrenic nerve transfer via thoracoscopy with respect to mean elbow strength at final follow-up. Methods A retrospective analysis was conducted among patients who underwent phrenic nerve transfer for elbow flexion at 2 centers from 2008 to 2017. All data analysis was performed in order to determine statistical significance among the analyzed variables. Results A total of 32 patients underwent supraclavicular phrenic nerve transfer, while 28 underwent phrenic nerve transfer via video-assisted thoracoscopy. Demographic characteristics were similar in both groups. A statistically significant difference in elbow flexion strength recovery was observed, favoring the supraclavicular phrenic nerve section group against the intrathoracic group (P = .036). A moderate though nonsignificant difference was observed favoring the same group in mean elbow flexion strength. Also, statistical differences included patient age (P = .01) and earlier time from trauma to surgery (P = .069). Conclusion Comparing supraclavicular sectioning of the nerve vs video-assisted, intrathoracic nerve sectioning to restore elbow flexion showed that the former yielded statistically better results than the latter, in terms of the percentage of patients who achieve at least level 3 MRC strength at final follow-up. Furthermore, larger scale prospective studies assessing the long-term effects of phrenic nerve transfers remain necessary.

Background: In 2010, the European Working Group on Sarcopenia in Older People (EWGSOP1) issued its first operational definition to diagnose sarcopenia. This was updated in 2019 with a revised sequence of muscle mass and muscle strength (EWGSOP2). The aim of the study was to investigate the impact of these different operational definitions on sarcopenia prevalence in a representative population-based sample. Methods: For each algorithm, the prevalence of sarcopenia-related categories was calculated and related to sociodemographic and lifestyle variables, anthropometric parameters, and laboratory biomarkers. The present analysis used data from the Salus in Apulia Study (Italy, 740 subjects, mean age 75.5 ± 5.9 years, 54% women). Results: The application of the EWGSOP1 adapted algorithm resulted in 85% [95% confidence intervals (CI): 82–88%] non-sarcopenic subjects, 10% (95% CI: 8–12%) pre-sarcopenic subjects, and 5% (95% CI: 3–7%) sarcopenic/severe sarcopenic subjects. The sarcopenia-related categories were inversely related to weight and body mass index (BMI), particularly in overweight/obese subjects, and these categories showed favorable metabolic biomarkers. The EWGSOP2 algorithm yielded 73% (95% CI: 69–76%) non-sarcopenic subjects, 24% (95% CI: 21–27%) probably sarcopenic subjects, and 4% (95% CI: 2–5%) sarcopenic subjects. Conclusions: The present study identified BMI as a potential confounder of the prevalence estimates of sarcopenia-related categories in population-based settings with different EWGSOP operational definitions.

The O2-mediated oxidation of all-β-barrel ferrous nitrosylated nitrobindin from Arabidopsis thaliana (At-Nb(II)-NO), Mycobacterium tuberculosis (Mt-Nb(II)-NO), and Homo sapiens (Hs-Nb(II)-NO) to ferric derivative (At-Nb(III), Mt-Nb(III), and Hs-Nb(III), respectively) has been investigated at pH 7.0 and 20.0 °C. Unlike ferrous nitrosylated horse myoglobin, human serum heme-albumin and human hemoglobin, the process in Nb(II)-NO is mono-exponential and linearly dependent on the O2 concentration, displaying a bimolecular behavior, characterized by kon = (6.3 ± 0.8) × 103 M−1 s−1, (1.4 ± 0.2) × 103 M−1 s−1, and (3.9 ± 0.5) × 103 M−1 s−1 for At-Nb(II)-NO, Mt-Nb(II)-NO, and Hs-Nb(II)-NO, respectively. No intermediate is detected, indicating that the O2 reaction with Nb(II)-NO is the rate-limiting step and that the subsequent conversion of the heme-Fe(III)-N(O)OO− species (i.e., N-bound peroxynitrite to heme-Fe(III)) to heme-Fe(III) and NO3− is much faster. A similar mechanism can be invoked for ferrous nitrosylated human neuroglobin and rabbit hemopexin, in which the heme-Fe(III)-N(O)OO− species is formed as well, although the rate-limiting step seems represented by the reshaping of the six-coordinated heme-Fe(III) complex. Although At-Nb(II)-NO and Mt-Nb(II)-NO are partially (while Hs-Nb(II)-NO is almost completely) penta-coordinated, density functional theory (DFT) calculations rule out that the cleavage of the proximal heme-Fe-His bond in Nb(II)-NO is responsible for the more stable heme-Fe(III)-N(O)OO− species. Moreover, the oxidation of the penta-coordinated heme-Fe(II)-NO adduct does not depend on O2 binding at the proximal side of the metal center. These features may instead reflect the peculiarity of Nb folding and of the heme environment, with a reduced steric constraint for the formation of the heme-Fe(III)-N(O)OO− complex.

Acute promyelocytic leukemia (APL) is a hematological disease characterized by a balanced reciprocal translocation that leads to the synthesis of the oncogenic fusion protein PML-RAR&alpha, APL is mainly managed by a differentiation therapy based on the administration of all-trans retinoic acid (ATRA) and arsenic trioxide (ATO). However, therapy resistance, differentiation syndrome, and relapses require the development of new low-toxicity therapies based on the induction of blasts differentiation. In keeping with this, we reasoned that a better understanding of the molecular mechanisms pivotal for ATRA-driven differentiation could definitely bolster the identification of new therapeutic strategies in APL patients. We thus performed an in-depth high-throughput transcriptional profile analysis and metabolic characterization of a well-established APL experimental model based on NB4 cells that represent an unevaluable tool to dissect the complex mechanism associated with ATRA-induced granulocytic differentiation. Pathway-reconstruction analysis using genome-wide transcriptional data has allowed us to identify the activation/inhibition of several cancer signaling pathways (e.g., inflammation, immune cell response, DNA repair, and cell proliferation) and master regulators (e.g., transcription factors, epigenetic regulators, and ligand-dependent nuclear receptors). Furthermore, we provide evidence of the regulation of a considerable set of metabolic genes involved in cancer metabolic reprogramming. Consistently, we found that ATRA treatment of NB4 cells drives the activation of aerobic glycolysis pathway and the reduction of OXPHOS-dependent ATP production. Overall, this study represents an important resource in understanding the molecular &ldquo, portfolio&rdquo, pivotal for APL differentiation, which can be explored for developing new therapeutic strategies.

Graphical abstract, In the Fall of 2019 a sudden and dramatic outbreak of a pulmonary disease (Coronavirus Disease COVID-19), due to a new Coronavirus strain (i.e., SARS-CoV-2), emerged in the continental Chinese area of Wuhan and quickly diffused throughout the world, causing up to now several hundreds of thousand deaths. As for common viral infections, the crucial event for the viral life cycle is the entry of genetic material inside the host cell, realized by the spike protein of the virus through its binding to host receptors and its activation by host proteases; this is followed by translation of the viral RNA into a polyprotein, exploiting the host cell machinery. The production of individual mature viral proteins is pivotal for replication and release of new virions. Several proteolytic enzymes either of the host and of the virus act in a concerted fashion to regulate and coordinate specific steps of the viral replication and assembly, such as (i) the entry of the virus, (ii) the maturation of the polyprotein and (iii) the assembly of the secreted virions for further diffusion. Therefore, proteases involved in these three steps are important targets, envisaging that molecules which interfere with their activity are promising therapeutic compounds. In this review, we will survey what is known up to now on the role of specific proteolytic enzymes in these three steps and of most promising compounds designed to impair this vicious cycle.

The anticoagulant therapy is widely used to prevent and treat thromboembolic events. Until the last decade, vitamin K antagonists were the only available oral anticoagulants; recently, direct oral anticoagulants (DOACs) have been developed. Since 55% to 95% of DOACs are bound to plasma proteins, the in silico docking and ligand-binding properties of drugs apixaban, betrixaban, dabigatran, edoxaban, and rivaroxaban and of the prodrug dabigatran etexilate to human serum albumin (HSA), the most abundant plasma protein, have been investigated. DOACs bind to the fatty acid (FA) site 1 (FA1) of ligand-free HSA, whereas they bind to the FA8 and FA9 sites of heme-Fe(III)- and myristic acid-bound HSA. DOACs binding to the FA1 site of ligand-free HSA has been validated by competitive inhibition of heme-Fe(III) recognition. Values of the dissociation equilibrium constant for DOACs binding to the FA1 site (ie, calc KDOAC ) derived from in silico docking simulations (ranging between 1.2 × 10-8 M and 1.4 × 10-6 M) agree with those determined experimentally from competitive inhibition of heme-Fe(III) binding (ie, exp KDOAC ; ranging between 2.5 × 10-7 M and 2.2 × 10-6 M). In addition, this study highlights the inequivalence of rivaroxaban binding to mammalian serum albumin. Given the HSA concentration in vivo (~7.5 × 10-4 M), values of KDOAC here determined indicate that the formation of the HSA:DOACs complexes in the absence and presence of FAs and heme-Fe(III) may occur in vivo. Therefore, HSA appears to be an important determinant for DOACs transport.

In the present paper, the biological effects of three different naphthalene diimides (NDIs) G-quadruplex (G4) ligands (H-NDI-Tyr, H-NDI-NMe2, and tetra-NDI-NMe2) were comparatively evaluated to those exerted by RHPS4, a well-characterized telomeric G4-ligand, in an in vitro model of glioblastoma. Data indicated that NDIs were very effective in blocking cell proliferation at nanomolar concentrations, although displaying a lower specificity for telomere targeting compared to RHPS4. In addition, differently from RHPS4, NDIs failed to enhance the effect of ionizing radiation, thus suggesting that additional targets other than telomeres could be involved in the strong NDI-mediated anti-proliferative effects. In order to test telomeric off-target action of NDIs, a panel of genes involved in tumor progression, DNA repair, telomere maintenance, and cell-cycle regulation were evaluated at transcriptional and translational level. Specifically, the compounds were able to cause a marked reduction of TERT and BCL2 amounts as well as to favor the accumulation of proteins involved in cell cycle control. A detailed cytofluorimetric analysis of cell cycle progression by means of bromodeoxyuridine (BrdU) incorporation and staining of phospho-histone H3 indicated that NDIs greatly reduce the progression through S-phase and lead to G1 accumulation of BrdU-positive cells. Taken together, these data indicated that, besides effects on telomeres and oncogenes such as Tert and Bcl2, nanomolar concentrations of NDIs determined a sustained block of cell proliferation by slowing down cell cycle progression during S-phase. In conclusion, our data indicate that NDIs G4-ligands are powerful antiproliferative agents, which act through mechanisms that ultimately lead to altered cell-cycle control.

Objective: The present study was aimed at analyzing the effect of physical activity on motor coordination in children with ASD. Methods: On 28 June 2021, a systematic review with meta-analysis was performed using the following databases: MEDLINE, SciELO, LILACS, PEDro, SPORTDiscus, CINAHL, SCOPUS, Web of Science, Cochrane, and Science Direct. We analyzed the methodological quality and risk of bias using the Jadad scale and Cochrane tool, respectively. Motor coordination results were meta-analyzed using the RevMan program. Two independent researchers used the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) tool to assess the level of evidence from the meta-analysis. Results: We found four studies in the listed databases and five randomized clinical trials were included in the meta-analysis that included 109 children with ASD. Children with ASD who performed physical activity did not present significantly better motor coordination than control children (p = 0.12). Conclusions: Considering the clinical importance of physical activity for children with ASD, this systematic review with meta-analysis showed that physical activity had no statistically significant effects on coordination in individuals with ASD.

Background Irritable bowel syndrome (IBS) is characterised by gastrointestinal (GI) and psychological symptoms (e.g., depression, anxiety, and somatization). Depression and anxiety, but not somatization, have already been associated with altered intestinal barrier function, increased LPS, and dysbiosis. The study aimed to investigate the possible link between somatization and intestinal barrier in IBS with diarrhoea (IBS-D) patients. Methods Forty-seven IBS-D patients were classified as having low somatization (LS = 19) or high somatization (HS = 28) according to the Symptom Checklist-90-Revised (SCL-90-R), (cut-off score = 63). The IBS Severity Scoring System (IBS-SSS) and the Gastrointestinal Symptom Rating Scale (GSRS) questionnaires were administered to evaluate GI symptoms. The intestinal barrier function was studied by the lactulose/mannitol absorption test, faecal and serum zonulin, serum intestinal fatty-acid binding protein, and diamine oxidase. Inflammation was assessed by assaying serum Interleukins (IL-6, IL-8, IL-10), and tumour necrosis factor-α. Dysbiosis was assessed by the urinary concentrations of indole and skatole and serum lipopolysaccharide (LPS). All data were analysed using a non-parametric test. Results The GI symptoms profiles were significantly more severe, both as a single symptom and as clusters of IBS-SSS and GSRS, in HS than LS patients. This finding was associated with impaired small intestinal permeability and increased faecal zonulin levels. Besides, HS patients showed significantly higher IL-8 and lowered IL-10 concentrations than LS patients. Lastly, circulating LPS levels and the urinary concentrations of indole were higher in HS than LS ones, suggesting a more pronounced imbalance of the small intestine in the former patients. Conclusions IBS is a multifactorial disorder needing complete clinical, psychological, and biochemical evaluations. Trial registration: https://clinicaltrials.gov/ct2/show/NCT03423069.

Kawasaki disease is an acute systemic vascular disease, generally self-limited and typically affecting children

Over the last 50 years, the incidence of human thyroid cancer disease has seen a significative increment. This comes along with an even higher increment of surgery, since, according to the international guidelines, patients are sometimes addressed to surgery also when the fine needle aspiration gives undetermined cytological diagnosis. As a matter of fact, only 30% of the thyroid glands removed for diagnostic purpose have a post surgical histological report of malignancy: this implies that about 70% of the patients have suffered an unnecessary thyroid removal. Here we show that Raman spectroscopy investigation of thyroid tissues provides reliable cancer diagnosis. Healthy tissues are consistently distinguished from cancerous ones with an accuracy of $$\sim $$ ∼ 90%, and the three cancer typology with highest incidence are clearly identified. More importantly, Raman investigation has evidenced alterations suggesting an early stage of transition of adenoma tissues into cancerous ones. These results suggest that Raman spectroscopy may overcome the limits of current diagnostic tools.

The pathogenicity of group AStreptococcus(GAS) is mediated by direct bacterial invasivity and toxin-associated damage. Among the extracellular products, the exotoxin streptolysin O (SLO) is produced by almost all GAS strains. SLO is a pore forming toxin (PFT) hemolitically active and extremely toxicin vivo. Recent evidence suggests that human serum albumin (HSA), the most abundant protein in plasma, is a player in the innate immunity “orchestra.” We previously demonstrated that HSA acts as a physiological buffer, partially neutralizingClostridioides difficiletoxins that reach the bloodstream after being produced in the colon. Here, we report thein vitroandex vivocapability of HSA to neutralize the cytotoxic and hemolytic effects of SLO. HSA binds SLO with high affinity at a non-conventional site located in domain II, which was previously reported to interact also withC. difficiletoxins. HSA:SLO recognition protects HEp-2 and A549 cells from cytotoxic effects and cell membrane permeabilization induced by SLO. Moreover, HSA inhibits the SLO-dependent hemolytic effect in red blood cells isolated from healthy human donors. The recognition of SLO by HSA may have a significant protective role in human serum and sustains the emerging hypothesis that HSA is an important constituent of the innate immunity system.

Heterochromatin Protein 1 (HP1) and the Mre11-Rad50-Nbs1 (MRN) complex are conserved factors that play crucial role in genome stability and integrity. Despite their involvement in overlapping cellular functions, ranging from chromatin organization, telomere maintenance to DNA replication and repair, a tight functional relationship between HP1 and the MRN complex has never been elucidated. Here we show that the Drosophila HP1a protein binds to the MRN complex through its chromoshadow domain (CSD). In addition, loss of any of the MRN members reduces HP1a levels indicating that the MRN complex acts as regulator of HP1a stability. Moreover, overexpression of HP1a in nbs (but not in rad50 or mre11) mutant cells drastically reduces DNA damage associated with the loss of Nbs suggesting that HP1a and Nbs work in concert to maintain chromosome integrity in flies. We have also found that human HP1α and NBS1 interact with each other and that, similarly to Drosophila, siRNA-mediated inhibition of NBS1 reduces HP1α levels in human cultured cells. Surprisingly, fibroblasts from Nijmegen Breakage Syndrome (NBS) patients, carrying the 657del5 hypomorphic mutation in NBS1 and expressing the p26 and p70 NBS1 fragments, accumulate HP1α indicating that, differently from NBS1 knockout cells, the presence of truncated NBS1 extends HP1α turnover and/or promotes its stability. Remarkably, an siRNA-mediated reduction of HP1α in NBS fibroblasts decreases the hypersensitivity to irradiation, a characteristic of the NBS syndrome. Overall, our data provide an unanticipated evidence of a close interaction between HP1 and NBS1 that is essential for genome stability and point up HP1α as a potential target to counteract chromosome instability in NBS patient cells.

Background Glioblastoma is the most aggressive and most lethal primary brain tumor in the adulthood. Current standard therapies are not curative and novel therapeutic options are urgently required. Present knowledge suggests that the continued glioblastoma growth and recurrence is determined by glioblastoma stem-like cells (GSCs), which display self-renewal, tumorigenic potential, and increased radio- and chemo-resistance. The G-quadruplex ligand RHPS4 displays in vitro radiosensitizing effect in GBM radioresistant cells through the targeting and dysfunctionalization of telomeres but RHPS4 and Ionizing Radiation (IR) combined treatment efficacy in vivo has not been explored so far. Methods RHPS4 and IR combined effects were tested in vivo in a heterotopic mice xenograft model and in vitro in stem-like cells derived from U251MG and from four GBM patients. Cell growth assays, cytogenetic analysis, immunoblotting, gene expression and cytofluorimetric analysis were performed in order to characterize the response of differentiated and stem-like cells to RHPS4 and IR in single and combined treatments. Results RHPS4 administration and IR exposure is very effective in blocking tumor growth in vivo up to 65 days. The tumor volume reduction and the long-term tumor control suggested the targeting of the stem cell compartment. Interestingly, RHPS4 treatment was able to strongly reduce cell proliferation in GSCs but, unexpectedly, did not synergize with IR. Lack of radiosensitization was supported by the GSCs telomeric-resistance observed as the total absence of telomere-involving chromosomal aberrations. Remarkably, RHPS4 treatment determined a strong reduction of CHK1 and RAD51 proteins and transcript levels suggesting that the inhibition of GSCs growth is determined by the impairment of the replication stress (RS) response and DNA repair. Conclusions We propose that the potent antiproliferative effect of RHPS4 in GSCs is not determined by telomeric dysfunction but is achieved by the induction of RS and by the concomitant depletion of CHK1 and RAD51, leading to DNA damage and cell death. These data open to novel therapeutic options for the targeting of GSCs, indicating that the combined inhibition of cell-cycle checkpoints and DNA repair proteins provides the most effective means to overcome resistance of GSC to genotoxic insults. Electronic supplementary material The online version of this article (10.1186/s13046-019-1293-x) contains supplementary material, which is available to authorized users.

Chronic inflammation increases the risk of developing certain types of cancer, such as colorectal cancer (CRC). The oxidative metabolism of polyunsaturated fatty acids (PUFAs) has a strong effect on colonic tumorigenesis and the levels of arachidonic acid (AA) and eicosapentaenoic acid (EPA) can contribute to the development of an inflammatory microenvironment. Aim of this study was to evaluate the possible differences in the AA/EPA ratio tissue levels between CRC patients with and without synchronous metastases. Moreover, the expression of the most important inflammatory enzymes and mediators, linked with the AA/EPA ratio, have been also assessed. Sixty-eight patients with CRC were enrolled in the study, of which 33 patients with synchronous metastasis. Fatty acid profile analysis in tissue samples was done to examine the levels of AA and EPA. High levels of the AA/EPA ratio were detected in tumor tissue of patients with metastatic CRC. Moreover, an increase of expression of the main enzymes and mediators involved in inflammation was also detected in the same samples. The lipidomic approach of inflammation allows to evaluate lipid homeostasis changes that occur in cancer and in its metastatic process, in order to identify new biomarkers to be introduced into clinical practice. © 2019 by the authors. Licensee MDPI, Basel, Switzerland.

Haptoglobin related protein (Hpr) is a component of the trypanosome lytic factor (TLF), a complex acting in the innate immune response against African trypanosomes. Like haptoglobin (Hp), Hpr binds hemoglobin (Hb) in the blood, but unlike Hp, Hpr does not bind the CD163 receptor. Moreover, unlike Hp, Hpr retains the

C.difficile infection (CDI) is not a merely “gut-confined” disease as toxemia could drive the development of CDI-related extra-intestinal effects. These effects could explain the high CDI-associated mortality, not just justified by diarrhea and dehydration. Here, the extra-intestinal effects of toxin A (TcdA) and B (TcdB) produced by C. difficile have been studied in vivo using the zebrafish embryo model. Noteworthy, protective properties of human serum albumin (HSA) towards toxins-induced extra-intestinal effects were also addressed. Zebrafish embryos were treated with TcdA, TcdB and/or HSA at 24 h post-fertilization. Embryos were analyzed for 48 h after treatment to check vital signs and morphological changes. Markers related to cardio-vascular damage and inflammation were evaluated by Real-Time quantitative PCR and/or western blotting. Both toxins induced cardiovascular damage in zebrafish embryos by different mechanisms: (i) direct toxicity (i.e., pericardial edema, cardiac chambers enlargement, endothelial alteration); (ii) increased hormonal production and release (i.e., atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP)), (iii) alteration of the vascular system through the increase of the vascular endothelial growth factor (VEGF-A) levels, as well as of its receptors, (iv) pro-inflammatory response through high cytokines production (i.e., CXCL8, IL1B, IL6 and TNFα) and (v) cell-mediated damage due to the increase in neutrophils number. In addition to cardiovascular damage, we observe skin alteration and inflammation. Finally, our data indicate a protective effect of HSA toward the toxins induced extra-intestinal effects. Together, our findings can serve as a starting point for humans’ studies to substantiate and understand the extra-intestinal effects observed in CDI patients.

Ferric nitrobindins (Nbs) selectively bind NO and catalyze the conversion of peroxynitrite to nitrate. In this study, we show that NO scavenging occurs through the reductive nitrosylation of ferric Mycobacterium tuberculosis and Homo sapiens nitrobindins (Mt-Nb(III) and Hs-Nb(III), respectively). The conversion of Mt-Nb(III) and Hs-Nb(III) to Mt-Nb(II)-NO and Hs-Nb(II)-NO, respectively, is a monophasic process, suggesting that over the explored NO concentration range (between 2.5 × 10−5 and 1.0 × 10−3 M), NO binding is lost in the mixing time (i.e., NOkon ≥ 1.0 × 106 M−1 s−1). The pseudo-first-order rate constant for the reductive nitrosylation of Mt-Nb(III) and Hs-Nb(III) (i.e., k) is not linearly dependent on the NO concentration but tends to level off, with a rate-limiting step (i.e., klim) whose values increase linearly with [OH−]. This indicates that the conversion of Mt-Nb(III) and Hs-Nb(III) to Mt-Nb(II)-NO and Hs-Nb(II)-NO, respectively, is limited by the OH−-based catalysis. From the dependence of klim on [OH−], the values of the second-order rate constant kOH− for the reductive nitrosylation of Mt-Nb(III)-NO and Hs-Nb(III)-NO were obtained (4.9 (±0.5) × 103 M−1 s−1 and 6.9 (±0.8) × 103 M−1 s−1, respectively). This process leads to the inactivation of two NO molecules: one being converted to HNO2 and another being tightly bound to the ferrous heme-Fe(II) atom.

Background Traumatic brachial plexus injuries cause long-term maiming of patients. The major target function to restore in complex brachial plexus injury is elbow flexion. Objective To retrospectively analyze the correlation between the length of the nerve graft and the strength of target muscle recovery in extraplexual and intraplexual nerve transfers. Methods A total of 51 patients with complete or near-complete brachial plexus injuries were treated with a combination of nerve reconstruction strategies. The phrenic nerve (PN) was used as axon donor in 40 patients and the spinal accessory nerve was used in 11 patients. The recipient nerves were the anterior division of the upper trunk (AD), the musculocutaneous nerve (MC), or the biceps branches of the MC (BBs). An index comparing the strength of elbow flexion between the affected and the healthy arms was correlated with the choice of target nerve recipient and the length of nerve grafts, among other parameters. The mean follow-up was 4 yr. Results Neither the choice of MC or BB as a recipient nor the length of the nerve graft showed a strong correlation with the strength of elbow flexion. The choice of very proximal recipient nerve (AD) led to axonal misrouting in 25% of the patients in whom no graft was employed. Conclusion The length of the nerve graft is not a negative factor for obtaining good muscle recovery for elbow flexion when using PN or spinal accessory nerve as axon donors in traumatic brachial plexus injuries.

A robust and highly specific sensor based on electroactive molecularly imprinted polymer nanoparticles (nanoMIP) was developed. The nanoMIP tagged with a redox probe, combines both recognition and reporting capabilities. The developed nanoMIP replaces enzyme-mediator pairs used in traditional biosensors thus, offering enhanced molecular recognition for insulin, improving performance in complex biological samples, and yielding high stability. Also, most of existing sensors show poor performance after storage. To improve costs of the logistics and avoid the need of cold storage in the chain supply, we developed an alternative to biorecognition system that relies on nanoMIP. NanoMIP were computationally designed using “in-silico” insulin epitope mapping and synthesized by solid phase polymerisation. The characterisation of the polymer nanoparticles was performed by transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier-transform Infrared (FT-IR) and surface plasmon resonance (SPR). The electrochemical sensor was developed by chemical immobilisation of the nanoMIP on screen printed platinum electrodes. The insulin sensor displayed satisfactory performances and reproducible results (RSD = 4.2%; n = 30) using differential pulse voltammetry (DPV) in the clinically relevant concentration range from 50 to 2000 pM. The developed nanoMIP offers the advantage of large number of specific recognition sites with tailored geometry, as the resultant, the sensor showed high sensitivity and selectivity to insulin with a limit of detection (LOD) of 26 and 81 fM in buffer and human plasma, respectively, confirming the practical application for point of care monitoring. Moreover, the nanoMIP showed adequate storage stability of 168 days, demonstrating the robustness of sensor for several rounds of insulin analysis.

The purpose of this study was to assess the results of elbow flexion strength fatigue, rather than the maximal power of strength, after brachial plexus re-innervation with phrenic and spinal accessory nerves. We designed a simple but specific test to study whether statistical differences were observed among those two donor nerves. We retrospectively reviewed patients with severe brachial plexus palsy for which either phrenic nerve (PN) or spinal accessory nerve (SAN) to musculocutaneous nerve (MCN) transfer was performed. A dynamometer was used to determine the maximal contraction strength. One and two kilograms circular weights were utilized to measure isometrically the duration of submaximal and near-maximal contraction time. Statistical analysis was performed between the two groups. Twenty-eight patients were included: 21 with a PN transfer while 7 with a SAN transfer for elbow flexion. The mean time from trauma to surgery was 7.1 months for spinal accessory nerve versus 5.2 for phrenic nerve, and the mean follow-up was 57.7 and 38.6 months, respectively. Statistical analysis showed a quicker fatigue for the PN, such that patients with the SAN transfer could hold weights of 1 kg and 2 kg for a mean of 91.0 and 61.6 s, respectively, while patients with transfer of the phrenic nerve could hold 1 kg and 2 kg weights for just a mean of 41.7 and 19.6 s, respectively. Both differences were statistically significant (at p = 0.006 and 0.011, respectively). Upon correlation analysis, endurances at 1 kg and 2 kg were strongly correlated, with r = 0.85 (p

Background: The pathogenic effects of Clostridium difficile are primarily attributable to the production of the large protein toxins (C difficile toxins [Tcd]) A (TcdA) and B (TcdB). These toxins monoglucosylate Rho GTPases in the cytosol of host cells, causing destruction of the actin cytoskeleton with cytotoxic effects. Low human serum albumin (HSA) levels indicate a higher risk of acquiring and developing a severe C difficile infection (CDI) and are associated with recurrent and fatal disease. Methods: We used a combined approach based on docking simulation and biochemical analyses that were performed in vitro on purified proteins and in human epithelial colorectal adenocarcinoma cells (Caco-2), and in vivo on stem cell-derived human intestinal organoids and zebrafish embryos. Results: Our results show that HSA specifically binds via its domain II to TcdA and TcdB and thereby induces their autoproteolytic cleavage at physiological concentrations. This process impairs toxin internalization into the host cells and reduces the toxin-dependent glucosylation of Rho proteins. Conclusions: Our data provide evidence for a specific HSA-dependent self-defense mechanism against C difficile toxins and provide an explanation for the clinical correlation between CDI severity and hypoalbuminemia.

Septic cardiopathy and bacterial exotoxins (letter)

This study aimed to assess the predictors of acute kidney injury (AKI) during colistin therapy in a cohort of patients with bloodstream infections (BSI) due to colistin-susceptible Gram-negative bacteria, focusing on the role of serum albumin levels. The study consisted of two parts: (1) a multicentre retrospective clinical study to assess the predictors of AKI during colistin therapy, defined according to the Kidney Disease: Improving Global Outcomes (KDIGO) criteria; and (2) bioinformatic and biochemical characterization of the possible interaction between human serum albumin and colistin. Among the 170 patients included in the study, 71 (42%), 35 (21%), and 11 (6%) developed KDIGO stage 1 (K1-AKI), KDIGO stage 2 (K2-AKI), and KDIGO stage 3 (K3-AKI), respectively. In multivariable analyses, serum albumin

Structural and functional properties of ferrous Mycobacterium tuberculosis (Mt-Nb) and human (Hs-Nb) nitrobindins (Nbs) were investigated. At pH 7.0 and 25.0 °C, the unliganded Fe(II) species is penta-coordinated and unlike most other hemoproteins no pH-dependence of its coordination was detected over the pH range between 2.2 and 7.0. Further, despite a very open distal side of the heme pocket (as also indicated by the vanishingly small geminate recombination of CO for both Nbs), which exposes the heme pocket to the bulk solvent, their reactivity toward ligands, such as CO and NO, is significantly slower than in most hemoproteins, envisaging either a proximal barrier for ligand binding and/or crowding of H2O molecules in the distal side of the heme pocket which impairs ligand binding to the heme Fe-atom. On the other hand, liganded species display already at pH 7.0 and 25 °C a severe weakening (in the case of CO) and a cleavage (in the case of NO) of the proximal Fe-His bond, suggesting that the ligand-linked movement of the Fe(II) atom onto the heme plane brings about a marked lengthening of the proximal Fe-imidazole bond, eventually leading to its rupture. This structural evidence is accompanied by a marked enhancement of both ligands dissociation rate constants. As a whole, these data clearly indicate that structural–functional relationships in Nbs strongly differ from what observed in mammalian and truncated hemoproteins, suggesting that Nbs play a functional role clearly distinct from other eukaryotic and prokaryotic hemoproteins.

Haptoglobin (Hp) binds human hemoglobin (Hb), contributing to prevent extra-erythrocytic Hb-induced damage. Hp forms preferentially complexes with αβ dimers, displaying heme-based reactivity. Here, kinetics and thermodynamics of fluoride and azide binding to ferric human Hb (Hb(III)) complexed with the human Hp phenotypes 1-1 and 2-2 (Hp1-1:Hb(III) and Hp2-2:Hb(III), respectively) are reported (pH 7.0 and 20.0 °C). Fluoride binds to Hp1-1:Hb(III) and Hp2-2:Hb(III) with a one-step kinetic and equilibrium behavior. In contrast, kinetics of azide binding to and dissociation from Hp1-1:Hb(III)(–N3−) and Hp2-2:Hb(III)(–N3−) follow a two-step process. However, azide binding to Hp1-1:Hb(III) and Hp2-2:Hb(III) is characterized by a simple equilibrium, reflecting the compensation of kinetic parameters. The fast and the slow step of azide binding to Hp1-1:Hb(III) and Hp2-2:Hb(III) should reflect azide binding to the ferric β and α chains, respectively, as also proposed for the similar behavior observed in Hb(III). Present results highlight the ligand-dependent kinetic inequivalence of Hb subunits in the ferric form, reflecting structural differences between the two subunits in the interaction with some ferric ligands.

BACKGROUND Among other factors, like the time from trauma to surgery or the number of axons that reach the muscle target, a patient's age might also impact the final results of brachial plexus surgery. OBJECTIVE To identify (1) any correlations between age and the 2 outcomes: elbow flexion strength and shoulder abduction range; (2) whether childhood vs adulthood influences outcomes; and (3) other baseline variables associated with surgical outcomes. METHODS Twenty pediatric patients (under age 20 yr) who had sustained a traumatic brachial plexus injury were compared against 20 patients, 20 to 29 yr old, and 20 patients, 30 yr old or older. Univariate, univariate trend, and correlation analyses were conducted with patient age, time to surgery, type of injury, and number of injured roots included as independent variables. RESULTS A statistically significant trend toward decreasing mean strength in elbow flexion, progressing from the youngest to oldest age group, was observed. This linear trend persisted when subjects were subdivided into 4 age groups (

Background The management of locally advanced rectal cancer (RC) is an evolving clinical field where the multidisciplinary approach can reach its best, and liquid biopsy for obtaining tumor-derived component such as circulating tumor DNA (ctDNA) might provide complementary informations. Methods A systematic review of studies available in literature of liquid biopsy in non-metastatic RC has been performed according to PRISMA criteria to assess the role of ctDNA as a diagnostic, predictive and prognostic biomarker in this setting. Results Twenty-five publications have been retrieved, of which 8 full-text articles, 7 abstracts and 10 clinical trials. Results have been categorized into three groups: diagnostic, predictive and prognostic. Few but promising data are available about the use of liquid biopsy for early diagnosis of RC, with the main limitation of sensitivity due to low concentrations of ctDNA in this setting. In terms of prediction of response to chemoradiation, still inconclusive data are available about the utility of a pre-treatment liquid biopsy, whereas some studies report a positive correlation with a dynamic (pre/post-treatment) monitoring. The presence of minimal residual disease by ctDNA was consistently associated with worse prognosis across studies. Conclusions The use of liquid biopsy for monitoring response to chemoradiation and assess the risk of disease recurrence are the most advanced potential applications for liquid biopsy in RC, with implications also in the context of non-operative management strategies.

Human serum albumin (HSA) represents an important determinant of plasma oncotic pressure and a relevant factor that modulates fluid distribution between the body compartments. Moreover, HSA (i) represents the depot and transporter of several compounds, both endogenous and exogenous, (ii) affects the pharmacokinetics of many drugs, (iii) regulates chemical modifications of some ligands, (iv) shows (pseudo-)enzymatic properties, (v) inactivates some toxic compounds, and (vi) displays anti-oxidant properties. HSA binding and (pseudo-)enzymatic properties are regulated competitively, allosterically, and by covalent modifications. While competitive inhibition of HSA binding properties is evident, allosteric mechanisms and covalent modifications affecting HSA reactivity are less clear. In several pathological conditions in which free heme-Fe levels increase, the buffering capacity of plasma hemopexin is overwhelmed and most of heme-Fe binds to the fatty acid site 1 of HSA. HSA-heme-Fe displays globin-like properties; in turn, heme-Fe modulates competitively and allosterically HSA binding and reactivity properties. Remarkably, heme-Fe-mediated HSA properties are time-dependent, representing a case for "chronosteric effects". Here, we review the drug-based modulation of (i) heme-Fe-recognition by HSA and (ii) heme-Fe-mediated reactivity.

Classical all α-helical globins are present in all living organisms and are ordered in three lineages: (i) flavohemoglobins and single domain globins, (ii) protoglobins and globin coupled sensors and (iii) truncated hemoglobins, displaying the 3/3 or the 2/2 all α-helical fold. However, over the last two decades, all β-barrel and mixed α-helical-β-barrel heme-proteins displaying heme-based functional properties (e.g. ligand binding, transport and sensing) closely similar to those of all α-helical globins have been reported. Monomeric nitrophorins (NPs) and α1-microglobulin (α1-m), belonging to the lipocalin superfamily and nitrobindins (Nbs) represent prototypical heme-proteins displaying the all β-barrel and mixed α-helical-β-barrel folds. NPs are confined to the Reduviidae and Cimicidae families of Heteroptera, whereas α1-m and Nbs constitute heme-protein families spanning bacteria toHomo sapiens. The structural organization and the reactivity of the stable ferric solvent-exposed heme-Fe atom suggest that NPs and Nbs are devoted to NO transport, storage and sensing, whereasHs-α1-m participates in heme metabolism. Here, the structural and functional properties of NPs and Nbs are reviewed in parallel with those of sperm whale myoglobin, which is generally taken as the prototype of monomeric globins.

Several studies have demonstrated that for complex surgical procedures, surgeons who treat more patients have better outcomes than their lower-volume counterparts. The aim of this paper is to review the experience with video-assisted thoracic surgery (VATS) lobectomies in our small thoracic unit (group A), to understand whether our short-term results were different to the outcomes obtained by the same surgeon previously working in a high-volume unit (group B). 37 patients underwent VATS lobectomy. Hospital stay was on average 4.5 days (group A) versus 4.1 days (group B). Operative time and the number of ‘frozen sections’ were higher in group A. Hospital mortality was 0. VATS lobectomies are a safe approach in a low-volume unit formed by a single surgeon with a previous high-volume experience.

Clostridium difficile infection (CDI) has significant clinical impact especially on the elderly and/or immunocompromised patients. The pathogenicity of Clostridium difficile is mainly mediated by two exotoxins: Toxin A (TcdA) and toxin B (TcdB). These toxins primarily disrupt the cytoskeletal structure and the tight junctions of target cells causing cell rounding and ultimately cell death. Detectable C. difficile toxemia is strongly associated with fulminant disease. However, besides the well-known intestinal damage, recent animal and in vitro studies have suggested a more far-reaching role for these toxins activity including cardiac, renal, and neurologic impairment. The creation of C. difficile strains with mutations in the genes encoding toxin A and B indicate that toxin B plays a major role in overall CDI pathogenesis. Novel insights, such as the role of a regulator protein (TcdE) on toxin production and binding interactions between albumin and C. difficile toxins, have recently been discovered and will be described. Our review focuses on the toxin-mediated pathogenic processes of CDI with an emphasis on recent studies.

Background Infections caused by multidrug-resistant Enterobacteriaceae are hard to treat and life-threatening due to reduced therapeutic options. Systemic infections caused by Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae strains have increased in many European regions, becoming frequent in many clinical settings, and are associated with high mortality. The co-formulation of ceftazidime, a third-generation cephalosporin, with avibactam, a new suicide inhibitor beta-lactamase inhibitor able to block most Klebsiella pneumoniae carbapenemases, has been recently licensed, with promising results in patients with limited or absent therapeutic options. Little is known, however, as to the efficacy of such a combination in patients with soft tissue infections caused by multidrug-resistant Klebsiella pneumoniae carbapenemase-producing strains of Klebsiella pneumoniae. Case presentation A Caucasian 53-year-old man with paraplegia suffered multiple vertebral fractures due to a car crash. He was treated with external fixators that became infected early after insertion and were repeatedly and inefficiently treated with multiple antibiotics. He suffered repeated septic episodes caused by Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae strains with a multidrug-resistant profile. Meropenem, tigecycline, and colistin combinations allowed only temporary improvements, but septic shock episodes recurred, in spite of removal of infected external fixators. After approval of pre-marketing prescription by our local Ethics Committee, full clinical resolution was obtained with a compassionate treatment using meropenem and ceftazidime/avibactam in combination for 16 days. Conclusions Our experience provides additional evidence that ceftazidime/avibactam, possibly in combination with meropenem rescued by avibactam, may be an efficacious treatment option also for complicated skin and soft tissue infections caused by multidrug-resistant strains of Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae.

Idiopathic pulmonary fibrosis is a peripheral subpleural interstitial lung disorder limited to the lung not involving the airways. It has a poor prognosis (survival less than 5 years) and commonly an interstitial pneumonia radiological pattern. Patients complain of a chronic dry cough in 80% of cases. A cough is often the first symptom of this rare disease, preceding dyspnea by years, and is associated with a poor prognosis, high dyspnea scores and low FVC percentages. The pathogenetic mechanisms leading to coughing in IPF are unclear. This review focuses on recent evidence of cough pathophysiology in this disease. Gastroesophageal reflux may promote coughing in IPF patients; bile salts and pepsin may be abundant in BAL of these patients, inducing overproduction of TGF-β by airway epithelial cells and mesenchymal transition with fibroblast recruitment/activation and extracellular matrix deposition. Patients have an enhanced cough reflex to capsaicin and substance P with respect to control subjects. Moreover, patients with the MUC5B polymorphism show more severe coughing as MUC5B encodes for the dominant mucin in the honeycomb cysts of IPF patients. Comorbidities, including asthma, gastroesophageal reflux, hypersensitivity pneumonitis, bronchiectasis, chronic obstructive pulmonary disease and emphysema, can induce coughing in IPF patients. There is no clear explanation of the causes of coughing in IPF. Further research into the pathophysiology of IPF and the pathogenetic mechanisms of coughing is necessary to improve survival and quality of life.

Acute myeloid leukemia (AML) is a genetically heterogeneous malignancy characterized by the expansion of hematopoietic stem/progenitor cells (HPCs) blocked at different stages of maturation/differentiation. The poor outcome of AMLs necessitates therapeutic improvement. In AML, genes encoding for myeloid transcription factors, signaling receptors regulating cell proliferation, and epigenetic modifiers can be mutated by somatically acquired genetic mutations or altered by chromosomal translocations. These mutations modify chromatin organization at genes sites regulating HPCs proliferation, terminal differentiation, and DNA repair, contributing to the development and progression of the disease. The reversibility of the epigenetic modifications by drug treatment makes epigenetic changes attractive targets for AML therapeutic intervention. Recent findings underline increased DNA damage and abnormalities in the DNA damage response (DDR) as a critical feature of AML blasts. The DDR preserves cell integrity and must be tightly coordinated with DNA methylation and chromatin remodeling to ensure the accessibility to the DNA of transcription factors and repair enzymes. A crucial role in these events is played by the ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related protein (ATR) kinases, which are hyperactive in AML. Based on these findings, we hypothesize the inhibition of DNA damage kinases as an alternative or complementary strategy for the differentiation treatment of AML as it leads to a reduced ability to repair the DNA damage, and to the inhibition of specific epigenetic modifiers whose function is altered in leukemic cells. © 2018 IUBMB Life, 70(11):1057-1066, 2018.

Controversy exists surrounding the use of the phrenic nerve for transfer in severe brachial plexus injuries. The objectives of this study are: (1) to present the experience of the authors using the phrenic nerve in a single institution; and (2) to thoroughly review the existing literature to date.Adult patients with C5-D1 and C5-C8 lesions and a phrenic nerve transfer were retrospectively included. Patients with follow-up shorter than 18 months were excluded. The MRC muscle strength grading system was used to rate the outcome. Clinical repercussions relating to sectioning of the phrenic nerve were studied. An intense rehabilitation program was started after surgery, and compliance to this program was monitored using a previously described scale. Statistical analysis was performed with the obtained data.Fifty-one patients were included. The mean time between trauma and surgery was 5.7 months. Three-quarters of the patients had C5-D1, with the remainder C5-C8. Mean post-operative follow-up was 32.5 months A MRC of M4 was achieved in 62.7% patients, M3 21.6%, M2 in 3.9%, and M1 in 11.8%. The only significant differences between the two groups were in graft length (9.8 vs. 15.1 cm, p = 0.01); and in the rehabilitation compliance score (2.86 vs. 2.00, p = 0.01).Results of phrenic nerve transfer are predictable and good, especially if the grafts are short and the rehabilitation is adequate. It may adversely affect respiratory function tests, but this rarely correlates clinically. Contraindications to the use of the phrenic nerve exist and should be respected.

Background The independent prognostic impact of diabetes mellitus ( DM ) and prediabetes mellitus (pre‐ DM ) on survival outcomes in patients with chronic heart failure has been investigated in observational registries and randomized, clinical trials, but the results have been often inconclusive or conflicting. We examined the independent prognostic impact of DM and pre‐ DM on survival outcomes in the GISSI ‐HF (Gruppo Italiano per lo Studio della Sopravvivenza nella Insufficienza Cardiaca‐Heart Failure) trial. Methods and Results We assessed the risk of all‐cause death and the composite of all‐cause death or cardiovascular hospitalization over a median follow‐up period of 3.9 years among the 6935 chronic heart failure participants of the GISSI ‐ HF trial, who were stratified by presence of DM (n=2852), pre‐ DM (n=2013), and non‐ DM (n=2070) at baseline. Compared with non‐ DM patients, those with DM had remarkably higher incidence rates of all‐cause death (34.5% versus 24.6%) and the composite end point (63.6% versus 54.7%). Conversely, both event rates were similar between non‐ DM patients and those with pre‐ DM . Cox regression analysis showed that DM , but not pre‐ DM , was associated with an increased risk of all‐cause death (adjusted hazard ratio, 1.43; 95% CI , 1.28–1.60) and of the composite end point (adjusted hazard ratio, 1.23; 95% CI , 1.13–1.32), independently of established risk factors. In the DM subgroup, higher hemoglobin A1c was also independently associated with increased risk of both study outcomes (all‐cause death: adjusted hazard ratio, 1.21; 95% CI , 1.02–1.43; and composite end point: adjusted hazard ratio, 1.14; 95% CI , 1.01–1.29, respectively). Conclusions Presence of DM was independently associated with poor long‐term survival outcomes in patients with chronic heart failure. Clinical Trial Registration URL : http://www.clinicaltrials.gov . Unique identifier: NCT 00336336.

The molecular chaperone heat shock protein 90 (Hsp90α) regulates cell proteostasis and mitigates the harmful effects of endogenous and exogenous stressors on the proteome. Indeed, the inhibition of Hsp90α ATPase activity affects the cellular response to ionizing radiation (IR). Although the interplay between Hsp90α and several DNA damage response (DDR) proteins has been reported, its role in the DDR is still unclear. Here, we show that ataxia-telangiectasia-mutated kinase (ATM) and nibrin (NBN), but not 53BP1, RAD50, and MRE11, are Hsp90α clients as the Hsp90α inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) induces ATM and NBN polyubiquitination and proteosomal degradation in normal fibroblasts and lymphoblastoid cell lines. Hsp90α-ATM and Hsp90α-NBN complexes are present in unstressed and irradiated cells, allowing the maintenance of ATM and NBN stability that is required for the MRE11/RAD50/NBN complex-dependent ATM activation and the ATM-dependent phosphorylation of both NBN and Hsp90α in response to IR-induced DNA double-strand breaks (DSBs). Hsp90α forms a complex also with ph-Ser1981-ATM following IR. Upon phosphorylation, NBN dissociates from Hsp90α and translocates at the DSBs, while phThr5/7-Hsp90α is not recruited at the damaged sites. The inhibition of Hsp90α affects nuclear localization of MRE11 and RAD50, impairs DDR signaling (e.g., BRCA1 and CHK2 phosphorylation), and slows down DSBs repair. Hsp90α inhibition does not affect DNA-dependent protein kinase (DNA-PK) activity, which possibly phosphorylates Hsp90α and H2AX after IR. Notably, Hsp90α inhibition causes H2AX phosphorylation in proliferating cells, this possibly indicating replication stress events. Overall, present data shed light on the regulatory role of Hsp90α on the DDR, controlling ATM and NBN stability and influencing the DSBs signaling and repair.