Search

Salmonella enterica serovar Rissen ( S. Rissen) is an emerging causative agent of foodborne diseases. The current emergence of antibiotic resistance makes necessary alternative therapeutic strategies. In this study, we investigated the potential of a phage-resistant strain of S. Rissen (R R ) as a tool for developing an effective lipopolysaccharide (LPS)-based vaccine. The LPS O-antigen is known to play critical roles in protective immunity against Salmonella . However, the high toxicity of the LPS lipid A moiety limits its use in vaccines. Here, we demonstrated that the acquisition of bacteriophage resistance by S. Rissen leads to structural modifications in the LPS structure. Using NMR and mass spectrometry, we characterized the LPS from phage-resistant strains as a smooth variant bearing under-acylated Lipid A portions (penta- and tetra-acylated forms). We then combined RT-qPCR and NMR-based metabolomics to explore the effects of phage resistance and LPS modification on bacterial fitness and virulence. Finally, we conducted in vivo studies to determine whether lysogeny-induced remodeling of LPS affects the host immune response. Results revealed that the under-acylated variant of LPS from R R attenuates the inflammatory response in BALB/c mice, while eliciting a specific antibody response that protects against S. Rissen (R W ) infection. In conclusion, our findings suggest that phage resistance, through lipid A modification, may offer a novel strategy for reducing LPS toxicity, highlighting its potential as a promising biological approach for developing LPS-based vaccines against Salmonella infections., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Cuomo, Medaglia, Casillo, Gentile, Fruggiero, Corsaro and Capparelli.)

Preclinical studies have shown that the combination of Cistus × incanus L. and Scutellaria lateriflora L. extracts exerts beneficial effects on oral health against gingivitis. Thus, this study aimed to assess the tolerability of a chewing gum and its efficacy on gingivitis in a double-blind, placebo-controlled clinical trial. Enrolled subjects ( n = 60, 18-70 years) were randomized to receive two chewing gums or a placebo daily for 3 months. At baseline (t0) and monthly (t1, t2, and t3) timepoints, the Quantitative Gingival Bleeding Index (QGBI), the Modified Gingival Index (MGI), and the Oral Health 15 items (OH-15)] were employed to assess potential improvements in gingivitis. Pain was self-quantified via the Visual Analogue Scale (VAS), and the Clinical Global Impression Scale for Severity of illness (CGI-S) helped in evaluating the oral general conditions. This study is listed on the ISRCTN registry. At t3, the QGBI, MGI, OH-15, VAS, and CGI-S values decreased in the treated but not in the placebo group (β = 0.6 ± 0.1, t 176 = 3.680, p < 0.001; β = 0.87 ± 0.21, t 115 = 4.263, p < 0.001; β = 5.3 ± 2.5, t 172 = 2.086, p = 0.038; β = 3.16 ± 0.51, t 88 = 6.253, p < 0.001; and β = 1.09 ± 0.32, t 83 = 3.419, p < 0.001, respectively). A significant improvement in gingival health occurred after a 3-month intervention with the chewing gums containing S. lateriflora and C. incanus extracts.

The role of the Toll-like receptor 4 (TLR4) is of recognising intracellular and extracellular pathogens and of activating the immune response. This process can be compromised by single nucleotide polymorphisms (SNPs) which might affect the activity of several TLRs. The aim of this study is of ascertaining whether SNPs in the TLR4 of Bubalus bubalis infected by Brucella abortus, compromise the protein functionality. For this purpose, a computational analysis was performed. Next, computational predictions were confirmed by performing genotyping analysis. Finally, NMR-based metabolomics analysis was performed to identify potential biomarkers for brucellosis. The results indicate two SNPs (c. 672 A > C and c. 902 G > C) as risk factor for brucellosis in Bubalus bubalis, and three metabolites (lactate, 3-hydroxybutyrate and acetate) as biological markers for predicting the risk of developing the disease. These metabolites, together with TLR4 structural modifications in the MD2 interaction domain, are a clear signature of the immune system alteration during diverse Gram-negative bacterial infections. This suggests the possibility to extend this study to other pathogens, including Mycobacterium tuberculosis. In conclusion, this study combines multidisciplinary approaches to evaluate the biological and structural effects of SNPs on protein function., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Plastic pollution is an important environmental problem, and microplastics have been shown to have harmful effects on human and animal health, affecting immune and metabolic physiological functions. Further, microplastics can interfere with commensal microorganisms and exert deleterious effects on exposure to pathogens. Here, we compared the effects of 1 µm diameter polystyrene microplastic (PSMPs) on Candida albicans infection in both in vitro and in vivo models by using HT29 cells and Galleria mellonella larvae, respectively. The results demonstrated that PSMPs could promote Candida infection in HT29 cells and larvae of G. mellonella , which show immune responses similar to vertebrates. In this study, we provide new experimental evidence for the risk to human health posed by PSMPs in conjunction with Candida infections.

Staphylococcus aureus is a Gram-positive bacterium, which can be found, as a commensal microorganism, on the skin surface or in the nasal mucosa of the human population. However, S. aureus may become pathogenic and cause severe infections, especially in hospitalized patients. As an opportunistic pathogen, in fact, S. aureus interferes with the host Ca 2+ signaling, favoring the spread of the infection and tissue destruction. The identification of novel strategies to restore calcium homeostasis and prevent the associated clinical outcomes is an emerging challenge. Here, we investigate whether harzianic acid, a bioactive metabolite derived from fungi of the genus Trichoderma , could control S. aureus -induced Ca 2+ movements. First, we show the capability of harzianic acid to complex calcium divalent cations, using mass spectrometric, potentiometric, spectrophotometric, and nuclear magnetic resonance techniques. Then, we demonstrate that harzianic acid significantly modulates Ca 2+ increase in HaCaT (human keratinocytes) cells incubated with S. aureus . In conclusion, this study suggests harzianic acid as a promising therapeutical alternative against diseases associated with Ca 2+ homeostasis alteration.

Helicobacter pylori (H. pylori) is a common human pathogen causing inflammation. Recent studies have suggested a sophisticated interplay between mitochondria, innate immunity and inflammatory response, thus proposing mitochondrial disfunction as the hallmark of severe inflammatory disorders. In this study, humic substances isolated from composted fennel residues (HS-FEN) were tested as potential therapeutical strategy to restore the mitochondrial physiology and control the inflammation associated with H. pylori infection. The molecular features of HS-FEN were characterized by infrared spectrometry, thermochemolysis-GC/MS, NMR spectroscopy, and high-performance size-exclusion chromatography (HPSEC), which revealed the presence of aromatic polyphenolic components arranged in a rather stable conformation. In vitro results showed antioxidant and anti-inflammatory properties of HS-FEN, that was found to increase the expression level of OPA-1 and SOD-2 genes and in AGS cells stimulated with H. pylori culture filtrate (Hpcf) and concomitantly decrease the expression level of Drp-1 gene and IL-12, IL-17 and G-CSF proteins. The hydrophobic features of HS, their conformational arrangement and large content of bioactive molecules may explain the beneficial effects of HS-FEN, that may potentially become an interesting source of anti-inflammatory agents capable to counteract or prevent the H. pylori-related inflammatory disorders., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Verrillo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Gut microbiota regulates essential processes of host metabolism and physiology: synthesis of vitamins, digestion of foods non-digestible by the host (such as fibers), and-most important-protects the digestive tract from pathogens. In this study, we focus on the CRISPR/Cas9 technology, which is extensively used to correct multiple diseases, including liver diseases. Then, we discuss the non-alcoholic fatty liver disease (NAFLD), affecting more than 25% of the global population; colorectal cancer (CRC) is second in mortality. We give space to rarely discussed topics, such as pathobionts and multiple mutations. Pathobionts help to understand the origin and complexity of the microbiota. Since several types of cancers have as target the gut, it is vital extending the research of multiple mutations to the type of cancers affecting the gut-liver axis.

Studies carried out in the last ten years have shown that the metabolites made up from the gut microbiota are essential for multiple functions, such as the correct development of the immune system of newborns, interception of pathogens, and nutritional enrichment of the diet. Therefore, it is not surprising that alteration of the gut microbiota is the starting point of gastrointestinal infection, obesity, type 2 diabetes, inflammatory bowel disease, colorectal cancer, and lung cancer. Diet changes and antibiotics are the major factors damaging the gut microbiota. Early exposure of the newborns to antibiotics may prevent their correct development of the immune system, exposing them to pathogen infections, allergies, and chronic inflammatory diseases. We already know much on how host genes, microbiota, and the environment interact, owing to experiments in several model animals, especially in mice; advances in molecular technology; microbiota transplantation; and comparative metagenomic analysis. However, much more remains to be known. Longitudinal studies on patients undergoing to therapy, along with the identification of bacteria prevalent in responding patients may provide valuable data for improving therapies.

Life-threatening hyperammonemia occurs in both inherited and acquired liver diseases affecting ureagenesis, the main pathway for detoxification of neurotoxic ammonia in mammals. Protein O-GlcNAcylation is a reversible and nutrient-sensitive post-translational modification using as substrate UDP-GlcNAc, the end-product of hexosamine biosynthesis pathway. Here we show that increased liver UDP-GlcNAc during hyperammonemia increases protein O-GlcNAcylation and enhances ureagenesis. Mechanistically, O-GlcNAcylation on specific threonine residues increased the catalytic efficiency for ammonia of carbamoyl phosphate synthetase 1 (CPS1), the rate-limiting enzyme in ureagenesis. Pharmacological inhibition of O-GlcNAcase, the enzyme removing O-GlcNAc from proteins, resulted in clinically relevant reductions of systemic ammonia in both genetic (hypomorphic mouse model of propionic acidemia) and acquired (thioacetamide-induced acute liver failure) mouse models of liver diseases. In conclusion, by fine-tuned control of ammonia entry into ureagenesis, hepatic O-GlcNAcylation of CPS1 increases ammonia detoxification and is a novel target for therapy of hyperammonemia in both genetic and acquired diseases., (© 2022. The Author(s).)

Aims: The proximal tibia (PT) is the anatomical site most frequently affected by primary bone tumours after the distal femur. Reconstruction of the PT remains challenging because of the poor soft-tissue cover and the need to reconstruct the extensor mechanism. Reconstructive techniques include implantation of massive endoprosthesis (megaprosthesis), osteoarticular allografts (OAs), or allograft-prosthesis composites (APCs)., Methods: This was a retrospective analysis of clinical data relating to patients who underwent proximal tibial arthroplasty in our regional bone tumour centre from 2010 to 2018., Results: A total of 76 patients fulfilled the inclusion criteria and were included in the study. Mean age at surgery was 43.2 years (12 to 86 (SD 21)). The mean follow-up period was 60.1 months (5.4 to 353). In total 21 failures were identified, giving an overall failure rate of 27.6%. Prosthesis survival at five years was 75.5%, and at ten years was 59%. At last follow-up, mean knee flexion was 89.8° (SD 36°) with a mean extensor lag of 18.1° (SD 24°). In univariate analysis, factors associated with better survival of the prosthesis were a malignant or metastatic cancer diagnosis (versus benign), with a five- and ten-year survival of 78.9% and 65.7% versus 37.5% (p = 0.045), while in-hospital length of stay longer than nine days was also associated with better prognosis with five- and ten-year survival rates at 84% and 84% versus 60% and 16% (p < 0.001). In multivariate analysis, only in-hospital length of stay was associated with longer survival (hazard ratio (HR) 0.23, 95% confidence interval (CI) 0.08 to 0.66)., Conclusion: We have shown that proximal tibial arthroplasty with endoprosthesis is a safe and reliable method for reconstruction in patients treated for orthopaedic oncological conditions. Either modular or custom implants in this series performed well.Cite this article: Bone Jt Open  2022;3(9):733-740.

Branched-chain amino acids (BCAAs) include leucine, isoleucine, and valine. Mammalians cannot synthesize these amino acids de novo and must acquire them through their diet. High levels of BCAAs are associated with insulin resistance; type 2 diabetes; obesity; and non-metabolic diseases, including several forms of cancer. BCAAs-in particular leucine-activate the rapamycin complex1 mTORC1, which regulates cell growth and metabolism, glucose metabolism and several more essential physiological processes. Diets rich in BCAAs are associated with metabolic diseases (listed above), while diets low in BCAAs are generally reported to promote metabolic health. As for the dysregulation of the metabolism caused by high levels of BCAAs, recent studies propose that the accumulation of acyl-carnitine and diacyl-CoA in muscles alters lipid metabolism. However, this suggestion is not broadly accepted. On clinical grounds, pre- and post-operative metabolic profiles of candidate patients for bariatric surgery are being used to select the optimal procedure for each individual patient.

Chronic obstructive pulmonary disease (COPD) is characterized by different phenotypes and clinical presentations. Therefore, a single strategy of pulmonary rehabilitation (PR) does not always yield the expected clinical outcomes as some individuals respond excellently, others discreetly, or do not respond at all. Fifty consecutive COPD patients were enrolled. Of them, 35 starting a 5-week PR program were sampled at admission (T 0 ), after 2 (T 2W ) and 5 (T 5W ) weeks, while 15 controls not yet on PR were tested at T 0 and T 5W . Nuclear magnetic resonance (NMR) profiling of exhaled breath condensate (EBC) and multivariate statistical analysis were applied to investigate the relationship between biomarkers and clinical parameters. The model including the three classes correctly located T 2W between T 0 and T 5W , but 38.71% of samples partially overlapped with T 0 and 32.26% with T 5W , suggesting that for some patients PR is already beneficial at T 2W (32.26% overlapping with T 5W ), while for others (38.71% overlapping with T 0 ) more time is required. Rehabilitated patients presented several altered biomarkers. In particular, methanol from T 0 to T 5W decreased in parallel with dyspnea and fatigue, while the walk distance increased. Methanol could be ascribed to lung inflammation. We demonstrated that the metabolic COPD phenotype clearly evolves during PR, with a strict relationship between clinical and molecular parameters. Methanol, correlating with clinical parameters, represents a useful biomarker for monitoring personalized outcomes and establishing more targeted protocols.

Non-typhoid Salmonella (NTS) represents one of the major causes of foodborne diseases, which are made worse by the increasing emergence of antibiotic resistance. Thus, NTS are a significant and common public health concern. The purpose of this study is to investigate whether selection for phage-resistance alters bacterial phenotype, making this approach suitable for candidate vaccine preparation. We therefore compared two strains of Salmonella enterica serovar Rissen : R R (the phage-resistant strain) and R W (the phage-sensitive strain) in order to investigate a potential cost associated with the bacterium virulence. We tested the ability of both R R and R W to infect phagocytic and non-phagocytic cell lines, the activity of virulence factors associated with the main Type-3 secretory system (T3SS), as well as the canonic inflammatory mediators. The mutant R R strain-compared to the wildtype R W strain-induced in the host a weaker innate immune response. We suggest that the mitigated inflammatory response very likely is due to structural modifications of the lipopolysaccharide (LPS). Our results indicate that phage-resistance might be exploited as a means for the development of LPS-based antibacterial vaccines.

The identification of novel strategies to control Helicobacter pylori ( Hp )-associated chronic inflammation is, at present, a considerable challenge. Here, we attempt to combat this issue by modulating the innate immune response, targeting formyl peptide receptors (FPRs), G-protein coupled receptors that play key roles in both the regulation and the resolution of the innate inflammatory response. Specifically, we investigated, in vitro, whether Caulerpin-a bis-indole alkaloid isolated from algae of the genus Caulerpa -could act as a molecular antagonist scaffold of FPRs. We showed that Caulerpin significantly reduces the immune response against Hp culture filtrate, by reverting the FPR2-related signaling cascade and thus counteracting the inflammatory reaction triggered by Hp peptide Hp(2-20). Our study suggests Caulerpin to be a promising therapeutic or adjuvant agent for the attenuation of inflammation triggered by Hp infection, as well as its related adverse clinical outcomes.

Soybeans ( Glycine max ) are an excellent source of dietary proteins and peptides with potential biological activities, such as antihypertensive, anti-cholesterol, and antioxidant activity; moreover, they could prevent cancer. Also, soy contains all the essential amino acids for nutrition; therefore, it represents an alternative to animal proteins. The goal of this paper was the comprehensive characterization of medium-sized and short peptides (two to four amino acids) obtained from simulated gastrointestinal digestion. Two different analytical approaches were employed for peptide characterization, namely a common peptidomic analysis for medium-sized peptides and a suspect screening analysis for short peptides, employing an inclusion list of exact m / z values of all possible amino acid combinations. Moreover, fractionation by preparative reversed-phase liquid chromatography was employed to simplify the starting protein hydrolysate. Six fractions were collected and tested for antioxidative activity by an innovative antioxidant assay on human gastric adenocarcinoma AGS cell lines. The two most active fractions (2 and 3) were then characterized by a peptidomic approach and database search, as well as by a suspect screening approach, in order to identify potential antioxidant amino acid sequences. Some of the peptides identified in these two fractions have been already reported in the literature for their antioxidant activity.

Background: Extra-abdominal desmoid tumor fibromatosis (DTF) is a rare, locally aggressive soft tissue tumour. The best treatment modality for this patient cohort is still object of debate., Questions/purpose: This paper aimed to (1) to compare the outcomes of DTF after different treatment modalities, (2) to assess prognostic factors for recurrence following surgical excision, and (3) to assess prognostic factors for progression during observation., Methods: This was a retrospective multicenter study under the patronage of the European Musculoskeletal Oncology Society (EMSOS). All seven centres involved were tertiary referral centres for soft tissue tumours. Baseline demographic data was collected for all patients as well as data on the diagnosis, tumour characteristics, clinical features, treatment modalities and whether they had any predisposing factors for DTF., Results: Three hundred eighty-eight patients (240 female, 140 male) with a mean age of 37.6 (±18.8 SD, range: 3-85) were included in the study. Two hundred fifty-seven patients (66%) underwent surgical excision of ADF, 70 patients (18%) were observed without therapy, the residual patients had different conservative treatments. There were no significant differences in terms of tumour recurrence or progression between the different treatment groups. After surgical excision, younger age, recurrent disease and larger tumour size were risk factors for recurrence, while tumours around the shoulder girdle and painful lesions were at risk of progression in the observational group., Conclusion: Local recurrence rate after surgery was similar to progression rates under observation. Hence, observation in DTF seems to be justified, considering surgery in case of dimensional progression in 2 consecutive controls (3 and 6 months) and in painful lesions, with particular attention to lesions around the shoulder girdle.

Formyl peptide receptors (FPRs) are cell surface pattern recognition receptors (PRRs), belonging to the chemoattractant G protein-coupled receptors (GPCRs) family. They play a key role in the innate immune system, regulating both the initiation and the resolution of the inflammatory response. FPRs were originally identified as receptors with high binding affinity for bacteria or mitochondria N-formylated peptides. However, they can also bind a variety of structurally different ligands. Among FPRs, formyl peptide receptor-like 1 (FPRL1) is the most versatile, recognizing N-formyl peptides, non-formylated peptides, and synthetic molecules. In addition, according to the ligand nature, FPRL1 can mediate either pro- or anti-inflammatory responses. Hp(2-20), a Helicobacter pylori -derived, non-formylated peptide, is a potent FPRL1 agonist, participating in Helicobacter pylori -induced gastric inflammation, thus contributing to the related site or not-site specific diseases. The aim of this review is to provide insights into the role of FPRs in H. pylori -associated chronic inflammation, which suggests this receptor as potential target to mitigate both microbial and sterile inflammatory diseases.

Urea cycle disorders (UCD) are inherited defects in clearance of waste nitrogen with high morbidity and mortality. Novel and more effective therapies for UCD are needed. Studies in mice with constitutive activation of autophagy unravelled Beclin-1 as druggable candidate for therapy of hyperammonemia. Next, we investigated efficacy of cell-penetrating autophagy-inducing Tat-Beclin-1 (TB-1) peptide for therapy of the two most common UCD, namely ornithine transcarbamylase (OTC) and argininosuccinate lyase (ASL) deficiencies. TB-1 reduced urinary orotic acid and improved survival under protein-rich diet in spf-ash mice, a model of OTC deficiency (proximal UCD). In Asl Neo/Neo mice, a model of ASL deficiency (distal UCD), TB-1 increased ureagenesis, reduced argininosuccinate, and improved survival. Moreover, it alleviated hepatocellular injury and decreased both cytoplasmic and nuclear glycogen accumulation in Asl Neo/Neo mice. In conclusion, Beclin-1-dependent activation of autophagy improved biochemical and clinical phenotypes of proximal and distal defects of the urea cycle., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

Aim To evaluate tunnel positioning on radiographs in singlebundle (SB) and double-bundle (DB) anterior cruciate ligament (ACL) reconstruction, to evaluate if measurement is accurate and reproducible. Methods Radiographs of 30 SB and 30 DB ACL reconstruction were reviewed by two examiners who measured tunnel positioning with the quadrant method on the femur (a=depth, b=height) and the Amis and Jakob method on the tibia. Intra- and inter-observer reliability were evaluated with intra-class correlation coefficient (ICC). Results A radiographic analysis was completed in all patients in a SB-group and in 27 in a DB-group (p>0.05). Intra-observer reliability was almost perfect on femoral (ICC: a=0.85, b=0.83) and tibial (ICC=0.87) side in the SB-group. In the DB-group, it was almost perfect for tibial anteromedial (AM) and posterolateral (PL) bundles (ICC: AM=0.84, PL=0.81) and for femoral PL bundle (ICC: a=0.83, b=0.82), and substantial for femoral AM bundle (ICC: a=0.78, b=0.74). Inter-observer reliability was almost perfect on tibial (ICC=0.81) and femoral (ICC: a=0.81, b=0.87) side in the SB-group, and substantial on tibial (ICC: AM=0.71, PL=0.77) and femoral (ICC: AM a=0.73, b=0.78; PL a=0.74, b=0.76) side in the DB-group. Standard deviation (SD) was low (±9%) with respect to the centre of tunnel(s). Conclusion The quadrant method and the Amis and Jakob method are accurate and reproducible measurement methods. Also, as SD was low, an outside-in approach with a front-entry guide, which is free-hand positioned, can be postulated as a reliable method to locate the femoral tunnel in SB reconstruction and the AM bundle in DB reconstruction., (Copyright© by the Medical Assotiation of Zenica-Doboj Canton.)

Helicobacter pylori ( Hp ) is a Gram-negative bacterium colonizing the human stomach. Nuclear Magnetic Resonance (NMR) analysis of intracellular human gastric carcinoma cells (MKN-28) incubated with the Hp cell filtrate ( Hpcf ) displays high levels of amino acids, including the branched chain amino acids (BCAA) isoleucine, leucine, and valine. Polymerase chain reaction (PCR) Array Technology shows upregulation of mammalian Target Of Rapamycin Complex 1 (mTORC1), inflammation, and mitochondrial dysfunction. The review of literature indicates that these traits are common to type 2 diabetes, obesity, Alzheimer's diseases, and cardiometabolic disease. Here, we demonstrate how Hp may modulate these traits. Hp induces high levels of amino acids, which, in turn, activate mTORC1, which is the complex regulating the metabolism of the host. A high level of BCAA and upregulation of mTORC1 are, thus, directly regulated by Hp . Furthermore, Hp modulates inflammation, which is functional to the persistence of chronic infection and the asymptomatic state of the host. Finally, in order to induce autophagy and sustain bacterial colonization of gastric mucosa, the Hp toxin VacA localizes within mitochondria, causing fragmentation of these organelles, depletion of ATP, and oxidative stress. In conclusion, our in vitro disease model replicates the main traits common to the above four diseases and shows how Hp may potentially manipulate them.

The Toll-interleukin 1 receptor superfamily includes the genes interleukin 1 receptor-like 1 (IL1RL1), Toll like receptors (TLRs), myeloid differentiation primary-response 88 (MyD88), and MyD88 adaptor-like (TIRAP). This study describes the interaction between MyD88, TIRAP and IL1RL1 against Helicobacter pylori infection. Cases and controls were genotyped at the polymorphic sites MyD88 rs6853, TIRAP rs8177374 and IL1RL1 rs11123923. The results show that specific combinations of IL1RL1-TIRAP (AA-CT; P: 2,8 × 10 -17 ) and MyD88-TIRAP-IL1RL1 (AA-CT-AA; P: 1,4 × 10 -8 ) - but not MyD88 alone-act synergistically against Helicobacter pylori. Nuclear magnetic resonance (NMR) clearly discriminates cases from controls by highlighting significantly different expression levels of several metabolites (tyrosine, tryptophan, phenylalanine, branched-chain amino acids, short chain fatty acids, glucose, sucrose, urea, etc.). NMR also identifies the following dysregulated metabolic pathways associated to Helicobacter pylori infection: phenylalanine and tyrosine metabolism, pterine biosynthesis, starch and sucrose metabolism, and galactose metabolism. Furthermore, NMR discriminates between the cases heterozygous at the IL1RL1 locus from those homozygous at the same locus. Heterozygous patients are characterized by high levels of lactate, and IL1RL1-both associated with anti-inflammatory activity-and low levels of the pro-inflammatory molecules IL-1β, TNF-α, COX-2, and IL-6.

Helicobacter pylori ( H. pylori ) is a Gram-negative bacterium which colonizes the human stomach. The ability of H. pylori to evade the host defense system and the emergence of antibiotic resistant strains result in bacteria persistence and chronic inflammation, which leads to both severe gastric and extra-gastric diseases. Consequently, innovative approaches able to overcome H. pylori clinical outcomes are needed. In this work, we develop a novel non-toxic therapy based on the synergistic action of H. pylori phage and lactoferrin adsorbed on hydroxyapatite nanoparticles, which effectively impairs bacteria colonization and minimizes the damage of the host pro-inflammatory response.

Bacterial biofilm provides bacteria with resistance and protection against conventional antimicrobial agents and the host immune system. Bacteriophages are known to move across the biofilm to make it permeable to antimicrobials. Mineral hydroxyapatite (HA) can improve the lytic activity of bacteriophages, and, together with eicosanoic acid (C20:0), can destroy the biofilm structure. Here, we demonstrate the efficacy of the combined use of phage, HA and C20:0 against Xanthomonas campestris pv campestris ( Xcc ) biofilm. We used nuclear magnetic resonance (NMR)-based metabolomics to investigate the molecular determinants related to the lytic action, aiming at identifying the metabolic pathways dysregulated by phage treatment. Furthermore, we identified specific markers (amino acids, lactate and galactomannan) which are involved in the control of biofilm stability. Our data show that Xccφ1, alone or in combination with HA and C20:0, interferes with the metabolic pathways involved in biofilm formation. The approach described here might be extended to other biofilm-producing bacteria.

Introduction: Pidotimod, a synthetic dipeptide molecule with biological and immunological activities, is used to reduce the number of exacerbations or pneumonitis in patients with inflammatory diseases. In the present study, we investigated whether Pidotimod modifies the metabolomic pathways measured in the exhaled breath condensate (EBC) of non-cystic fibrosis bronchiectatic patients (NCFB). Materials and Methods: We analyzed 40 adult patients affected by NCFB. They were randomly selected to receive Pidotimod 800 mg b/d for 21 consecutive days (3 weeks) per month for 6 months (20 patients, V 1 group) or no drug (20 patients, V 0 group), with a 1:1 criterion and then followed as outpatients. Results: EBC samples were collected from all patients at baseline and after 6 months. They were investigated by combined nuclear magnetic resonance (NMR) spectroscopy and multivariate statistical analysis to uncover metabolic differences between EBC from NCFB patients before and after therapy with Pidotimod. Pulmonary function test and pulmonary exacerbations were analyzed at baseline and at the end of Pidotimod therapy. The EBC metabolites were all identified, and through statistical evaluation, we were able to discriminate the two samples' classes, with acetate, acetoin, lactate, and citrate as statistically significant discriminatory metabolites. The model vas validated by using a blind set of 20 NCFB samples, not included in the primary analysis. No differences were observed in PFT after 6 months. At the end of the study, there was a significant decrease of exacerbation rate in V 1 group as compared with V 0 group, with a substantial reduction of the number of mild or severe exacerbations ( p < 0.001). Discussion: Pidotimod modifies the respiratory metabolic phenotype ("metabotype") of NCFB patients and reduces the number of exacerbations., (Copyright © 2019 D’Amato, Paris, Molino, Cuomo, Fulgione, Sorrentino, Palomba, Maniscalco and Motta.)

Background: Although multidisciplinary therapies have improved local control and overall survival in Ewing sarcoma (ES), the prognosis of pelvic lesions remains markedly worse than that of limb ES., Methods: We retrospectively evaluated the influence of the type of local treatment, margins, necrosis and sacrum involvement on overall survival (OS) and disease-free survival (DFS) in a series of 21 non-metastatic pelvic ES., Results: The average follow-up was 46.3 months (range 3-156). Only one patient had recurrence, at 11 months after surgery. Eight patients showed pulmonary metastasis and five showed bone metastases. Necrosis was the only significant prognostic factor for overall survival at 5 years (p=0.0132) and disease-free survival (p=0.0086). Overall survival at 5 years was 40.1%., Conclusion: Local control in pelvic Ewing sarcoma is comparable for patients treated with surgery (S), surgery plus radiotherapy (S/RT), or definitive radiotherapy (RT). The combination of surgery plus radiotherapy could be indicated in cases of large tumor, a poor necrosis response (< 90%), or an inadequate margin with involvement of the sacrum. A poor response to neoadjuvant therapy is a significant risk factor for both local control and overall survival.

Background and Objectives: Liposarcoma (LPS) is a malignant mesenchymal tumor and the most common soft tissue sarcoma. Four different subtypes are described: well differentiated (WD) LPS or atypical lipomatous tumor (ALT), dedifferentiated (DD) LPS, myxoid LPS, and pleomorphic LPS (PLS). The objective of the study was to investigate prognostic factors and clinical outcome of liposarcoma., Methods: We retrospectively examined the clinico-pathological features of a series of 307 patients affected by Liposarcoma at a mean follow-up of 69 months (range 6-257). ALT/WD LPS were analyzed separately. The influence of site, size, type of presentation, grading, histotype and local recurrence on local and systemic control and survival was assessed., Results: The statistical analysis indicated that only surgical margins represented a significant prognostic factor for local recurrence in ALT/WD LPS (P = 0.0007) and other subtypes of LPS (P = 0.0055). In myxoid, PLS and DD LPS, significant prognostic factors for metastasis free survival (MFS) were surgical margins (P = 0.0009), size of the tumor (P = 0.0358), histology (P = 0.0117) and local recurrence (P = 0.0015). In multivariate analysis, surgical margins (0.0180), size (0.0432) and local recurrence (0.0288) correlated independently with MFS. Margins (P = 0.0315), local recurrence (P = 0.0482) and metastases (P < 0.0001) were prognostic factors for overall survival (OS)., Conclusion: Marginal surgery can be an accepted treatment for ALT/WD LPS. In other liposarcoma subtypes (Myxoid, DD, PLS) wide or radical surgery is recommended as the margins significantly influence local recurrence-free survival (LRFS), metastasis-free survival (MFS) and overall survival (OS). Local recurrence and metastases were significant prognostic factors for OS., (Copyright © 2018 The Japanese Orthopaedic Association. Published by Elsevier B.V. All rights reserved.)

Background: Contact sensitization (CS) and allergic contact dermatitis (ACD) were once thought to be rare in children. The aim of our study was to assess CS and ACD in children affected by eczematous conditions referring to our contact and occupational dermatoses service, and to compare the obtained data to the results of a similar study performed in our Department in late Eighties., Methods: A retrospective analysis of Patch Test data from the Database of Contact Allergy of the Department of Dermatology, University Federico II of Naples, was performed. Moreover results were compared to previous data recorded and analyzed in our center., Results: Of the 416 examined patients, tested during 2005-2010 with the pediatric series, 37.5% developed at least one positive reaction, 15.4% was diagnosed with ACD. The most common allergen was nickel sulphate (16.8%) and the most frequently involved body sites were head and neck (16.3%). Compared to the 1989 study data, our current results highlighted an increase of CS (P<0.001) and ACD diagnosis (P<0.01)., Conclusions: ACD and CS are significantly increased during the last 20 years in our young patients population. The widespread chemical use and premature adult-like attitude may be responsible for earlier and extended exposure to sensitizers.

Introduction: Soft tissue tumors around the knee joint still pose problems for the excision and subsequent reconstruction., Methods: In the 6 included patients the soft tissue sarcoma has its base on the anterior surface of the extensor mechanism and expands towards the skin. The entire extensor apparatus (quadriceps tendon, patella, and patellar tendon) was resected and replaced by a fresh-frozen allograft., Results: The mean follow-up was 6.7 years (range: 2-12.4 years). In two patients a local recurrence occurred, resulting in a 5-year local recurrence-free rate of 66.7% (95% CI: 19.5%-90.4%). Distant metastases were found in 4 patients resulting in a 5-year metastasis-free rate of 33.3% (95% CI: 4.6%-67.5%). Two patients underwent at least one revision surgery, including one patient in whom the allograft had to be removed. According to the ISOLS function score 24.7 points (range: 19-28 points) were achieved at the last follow-up. The mean active flexion of the knee joint was 82.5° (range: 25-120°) and a mean extension lag of 10° (range: 0-30°) was observed., Conclusions: The replacement of the extensor mechanism by an allograft is a reasonable option, allowing wide margins and restoration of active extension in most patients., Trial Registration: The presented study is listed on the ISRCTN registry with trial number ISRCTN63060594.

Background: Reconstruction of large bone defects around the elbow joint is surgically demanding due to sparse soft tissue coverage, complex biomechanics and the close proximity to neurovascular structures. Modular megaprostheses are established reconstruction tools for the elbow, but only small case series have been reported in the literature., Methods: Thirty-six patients who underwent reconstruction of the elbow joint with a modular megaprosthesis were reviewed retrospectively. In 31 patients (86.1%), elbow replacement was performed after resection of a bone tumour, whereas five non-oncological patients (13.9%) underwent surgery because of a previous failed elbow reconstruction. Functional outcome, rate of complications and oncological results were considered as primary endpoints., Results: The mean follow-up was 25 months. The average achieved Mayo Elbow Performance Score (MEPS) was 77.08 (range 40-95) and the average Musculoskeletal Tumor Society (MSTS) score was 22.9 (range 8-30). Six complications (16.7%) were observed: two radial palsies, one temporary radial nerve dysfunction, one ulnar palsy, one disassembling of the articular prosthesis component and one deep infection necessitating the only implant removal. The overall 5-year survival rate of the patients was poor (25.1%) because of rapid systemic progression of the oncological disease in patients with metastatic lesion. However, the 5-year survival rate of the implant was very satisfactory (93%)., Conclusions: Modular megaprosthesis is a reliable and effective reconstruction tool in large bone defects around the elbow joint. The complication rates are lower than seen in osteoarticular allografts and allograft-prosthesis composites while the functional outcome is equal. In palliative situations with metastatic disease involving the elbow, modular megaprosthesis enables rapid recovery and pain relief and preserves elbow function., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Modular megaprosthesis (MP) and allograft-prosthetic composite (APC) are the most commonly used reconstructions for large bone defects of the proximal tibia. The primary objective of this study was to compare the two different techniques in terms of failures and functional results. A total of 42 consecutive patients with a mean age of 39.6 years (range 15-81 years) who underwent a reconstruction of the proximal tibia between 2001 and 2012 were included. Twenty-three patients were given an MP, and 19 patients received an APC. There were nine reconstruction failures after an average follow-up of 62 months: five in the MP group and four in the APC group (p=0.957). The 10-year implant survival rate was 78.8% for the MP and 93.7% for the APC (p=0.224). There were no relevant differences between the two groups in functional results. Both MP and APC are valid and satisfactory reconstructive options for massive bone defects in the proximal tibia. In high-demanding patients with no further risk factors, an APC should be considered to provide the best possible functional result for the extensor mechanism., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The use of modular endoprostheses is a viable option to manage both tumor resection and severe bone loss due to nonneoplastic conditions such as fracture sequelae, failed osteoarticular grafts, arthroplasty revisions, and periprosthetic fractures. We sought to investigate both midterm complications and failures occurred in 87 patients who underwent a megaprosthetic reconstruction in a nonneoplastic setting. After a mean follow-up of 58 (1-167) months, overall failure-free survival was 91.5% at 1 year, 80% at 2 years, 71.6% at 5 years, and 69.1% at 5 and 10 years. There was no significant difference in the survival rate according to the diagnosis at the index procedure ( p = 0.921), nor to the reconstruction site ( p = 0.402). The use of megaprostheses in a postneoplastic setting did not affect survival rate in comparison with endoprosthetic reconstruction of pure nonneoplastic conditions ( p = 0.851). Perimegaprosthetic infection was the leading complication, occurring in 10 (11.5%) patients and implying a megaprosthetic revision in all but one case. Physicians should consider these results when discussing with patients desired outcomes of endoprosthetic reconstructions of a nonneoplastic disease., Competing Interests: The authors declare that there is no conflict of interests regarding the publication of this paper.

Needle biopsy is the main standard method used for diagnosis of musculoskeletal tumors of the limbs and superficial trunk. Pain control during this procedure is through the use of Local Anaestetic (L.A.). In order to achieve a complete pain control in our cases, recently we started using diclofenac HPβCD 50 mg via s.c. preoperativly. We present the clinical results of a non-randomized study of two eterogeneous groups of patients: "Experimental" Group (1): diclofenac HPβCD 50 mg via s.c. one hour before surgical procedure, local anesthesia and ev. diclofenac HPβCD 50 mg via s.c. 12 hours postoperative; "Conventional" Group (2): local anesthesia and ev. postoperative tramadol 100 mg via oral for pain control. In October 2014, at the Department of Orthopedic Oncology and Reconstructive Surgery of Florence, 37 musculoskeletal biopsies for a bone or a soft tissue lesion were performed. Exclusion criteria for this study were: known allergies to lidocaine, diclofenac, tramadol; known gastric or duodenal ulcers; known gastrointestinal bleed or perforation; refusal of the patients to collaborate. For one or more of these reasons, 6 patients were excluded from this study. In the Group 1, 10 patients (59%) referred no pain during the surgical procedure (8/14 biopsies on soft tissue and 2/3 on bone). In 5 cases (29%) no exacerbation of previous chronic pain, and in 2 cases (12%) a progression of local pain after biopsy (average 1 points higher in the VAS). In Group 2, only 6 patients (42%) did not have any pain during the procedure, 4 (29%) no exacerbation of previous chronic pain and 4 (29%) a progression of local pain (average 2 points higher in the VAS). Despite similar results in both Groups, Group 1 seemed to have a mild better control of perioperative pain. The use of diclofenac HPβCD 50 mg preoperative seems to be a rational approach for minimizing perioperative pain and the preliminary data of our study seem encouraging. Obviously many bias are present in this study (small numbers of cases, heterogeneity of diseases, association with local anesthetic, non-randomized study, comparison between preoperative versus postoperative treatment) and this cannot absolutely be considerate as definitive conclusions.

Background: Double-bundle ACL reconstruction popularity is increasing with the aim to reproduce native ACL anatomy and improve ACL reconstruction outcome. However, to date, only a few randomized clinical studies have been published., Purpose: The aim of this study was to prospectively compare the clinical results of single- and double-bundle ACL reconstruction., Study Design: Randomized controlled clinical trial; Level of evidence, 1., Methods: Seventy patients with a chronic unilateral ACL rupture who underwent arthroscopically assisted ACL reconstruction using a hamstring graft were randomized to receive a single- (SB) or double-bundle (DB) reconstruction. Both groups were comparable with regard to preoperative data. A double-incision surgical technique was adopted in both groups. The graft was fixed by looping the hamstring tendons around a bony (DB) or a metallic (SB) bridge on the tibial side and with interference screws reinforced with a staple on the femur. The same rehabilitation protocol was adopted. Outcome assessment was performed by a blinded, independent observer using the visual analog scale (VAS) score, the new International Knee Documentation Committee (IKDC) form, the Knee Injury and Osteoarthritis Outcome Score (KOOS), and KT-1000 arthrometer evaluation., Results: All the patients reached a minimum follow-up of 2 years. No differences between the 2 groups were observed in terms of KOOS and IKDC subjective score. A statistically significant difference in favor of the DB group was found with the VAS (P < .03). The objective IKDC final scores showed statistically significantly more "normal knees" in the DB group than in the SB group (P = .03). There was 1 stability failure in the DB group and 3 in the SB group. The KT-1000 arthrometer data showed a statistically significant decrease in the average anterior tibial translation in the DB group (1.2 mm DB vs 2.1 mm SB; P < .03). The incidence of a residual pivot-shift glide was 14% in DB and 26% in SB (P = .08)., Conclusion: In the 2-year minimum follow-up, DB ACL reconstructions showed better VAS, anterior knee laxity, and final objective IKDC scores than SB. However, longer follow-up and accurate instrumented in vivo rotational stability assessment are needed.

Proper femoral and tibial component rotational positioning in TKA is critical for outcomes. Several rotational landmarks are frequently used with different advantages and limitations. We wondered whether coronal axes in the tibia and femur based on the transepicondylar axis in the femur would correlate with anteroposterior deformity. We obtained computed tomography scans of 100 patients with arthritis before they underwent TKA. We measured the posterior condylar angle on the femoral side and the angle between Akagi's line and perpendicular to the projection of the femoral transepicondylar axis on the tibial side. On the femoral side, we found a linear relationship between the posterior condylar angle and coronal deformity with valgus knees having a larger angle than varus knees, ie, gradual external rotation increased with increased coronal deformity from varus to valgus. On the tibial side, the angle between Akagi's line and the perpendicular line to the femoral transepicondylar axis was on average approximately 0 degrees , but we observed substantial interindividual variability without any relationship to gender or deformity. A preoperative computed tomography scan was a useful, simple, and relatively inexpensive tool to identify relevant anatomy and to adjust rotational positioning. We do not, however, recommend routine use because on the femoral side, we found a relationship between rotational landmarks and coronal deformity.

Background: Double-bundle anterior cruciate ligament reconstruction replicates the 2 functional bundles of the native ligament, the posterolateral and the anteromedial, to control anteroposterior and rotational laxity., Hypothesis: Double-bundle anterior cruciate ligament reconstruction laxity should be affected by the way grafts are tensioned., Study Design: Controlled laboratory study., Methods: Fourteen intact cadaveric knees were instrumented in a 6 degree of freedom rig, and kinematics throughout flexion-extension were recorded with an electromagnetic system under a 90-N anterior force or a 5-N.m internal rotation torque. Anteromedial and posterolateral bundle bovine extensor tendon grafts were fixed to load cells on the tibia, and tension was adjusted to match the intact knee anteroposterior laxity with 3 different protocols: (1) anteromedial bundle first and then posterolateral bundle at 90 degrees and 20 degrees of flexion, respectively; (2) posterolateral bundle first and then anteromedial bundle at 20 degrees and 90 degrees of flexion, respectively; and (3) both bundles together at 20 degrees of flexion. Finally, a single-bundle graft positioned at 10 o'clock was tensioned at 20 degrees of flexion., Results: Lower graft tensions were required to match intact knee laxity in double-bundle anterior cruciate ligament reconstruction. Tension patterns with knee flexion were independent from the tensioning protocol. Protocols 1 and 2 overconstrained anteroposterior laxity, whereas protocol 3 matched intact knee anteroposterior laxity throughout the range of motion. The single-bundle reconstructions had excess anteroposterior laxity in flexion. Rotations were better restored with double-bundle protocols 2 and 3., Conclusion: Knee laxity after double-bundle anterior cruciate ligament reconstruction is affected by the sequence in which the grafts are tensioned., Clinical Relevance: Double-bundle anterior cruciate ligament reconstruction ensures better laxity restoration than does single bundle when both bundles are fixed together.

Unlabelled: Valgus deformity correction with total knee arthroplasty is challenging. We hypothesized selective release of the tight lateral structures (pie-crusting technique), and of the lateral retinaculum in case of patellar maltracking, would obtain and maintain correction of the frontal plane deformity, restore patellar tracking and function, and avoid the complications of the extensive releases, including lateral condyle avascularity and residual lateral instability. We followed 48 patients with 53 valgus knees who underwent TKA and were followed a minimum of 5 years (mean, 8 years; range, 5-12 years). Soft tissue balancing of the lateral structures was performed with the pie-crusting technique. We employed either a fixed posterior stabilized or a mobile implant. A lateral release was performed in 67% of the cases. We observed one postoperative complication, a transient postoperative peroneal nerve palsy that spontaneously completely recovered. In 51 of the 53 knees (96%) we achieved alignment within 5 degrees from neutral. One patient had varus instability in extension. No component was revised. The pie-crusting technique reliably corrects moderate to severe fixed valgus deformities with a low complication rate and reasonable mid-term results. The multiple punctures allow gradual stretching of the lateral soft tissues and preservation of the popliteus tendon reducing the risk of posterolateral instability., Level of Evidence: Level IV, case series. See the Guidelines for Authors for a complete description of levels of evidence.

Purpose: The aim of this study was to examine whether a double-bundle anterior cruciate ligament (ACL) reconstruction with a transtibial approach could position the tibial and femoral tunnels accurately in the native bundle attachments., Methods: In 21 fresh-frozen knees the tibial and femoral attachments of the anteromedial (AM) and posterolateral (PL) bundles were outlined. The AM tibial tunnel guidewire was drilled with the 65 degree Howell tibial guide (Arthrotek, Warsaw, IN) located against the femur in the extended knee. The PL tibial wire was drilled through a prototype attachment to the Howell guide. Of the knees, 14 were available for the femoral part of the study. The AM femoral guidewire used an aimer offset 3 mm from the over-the-top position. The PL wire was drilled transtibially at 70 degrees of flexion, with external rotation and posterior drawer loads being applied. The plateaus and condyles were photographed and the wire positions measured., Results: With regard to the tibia, 17 of 21 AM wires were in the AM bundle attachment (at 61% and 36% of the natural ACL posteroanterior and mediolateral length, respectively) and 19 of 21 PL wires were in the PL bundle attachment (at 28% and 36% of the posteroanterior and mediolateral length, respectively). With regard to the femur, 12 of 14 AM wires and 9 of 14 PL wires were in the correct native bundle attachment. The AM wire was 3% more shallow than the center of the AM attachment (P = .03) and 6% more superior (P < .001), where 100% was the diameter of the posterior lateral condyle. The PL wire was 4% more shallow than the center of the PL attachment (P = .026) and 6% more superior (P < .001)., Conclusions: Anatomic and reproducible tibial guidewire positioning was achieved. Femoral wires were reproducibly positioned, but both were superior to and more shallow than the natural ACL bundle attachments, so further development or a different approach is appropriate., Clinical Relevance: The double-bundle reconstruction aims to restore anterior drawer and rotational stability. This technique ensures anatomic tibial positioning. Further improvements are needed with regard to the femur.

Double-bundle anterior cruciate ligament (ACL) reconstruction is intended to replicate the anatomy and the function of the anteromedial and posterolateral bundles of the native ACL to improve patients' satisfaction and knee stability. We prospectively assigned 75 consecutive patients with an isolated ACL lesion to one of three sequential groups of 25 patients each. Group I received a single-bundle, single-incision transtibial ACL reconstruction. Groups II and III received a double-bundle reconstruction with a single-incision transtibial technique or a double-bundle, twoincision outside-in technique, respectively. We obtained subjective International Knee Documentation Committee and Knee Injury and Osteoarthritis Outcome Score evaluations and objective International Knee Documentation Committee scores and KT-1000 measurements preoperatively and at followup. All patients reached a minimum followup of 2 years. KT side-to-side difference in Groups I, II, and III were 2.4, 1.6 and 1.4 mm, respectively. Group III had fewer patients with a positive pivot shift than Group I. The double-bundle double-incision outside-in ACL reconstruction resulted in improved anteroposterior stability and less residual pivot shift than single-incision single-bundle technique.

Background and Objective: In the perivascular sheath of the brachial plexus, the volume of anaesthetic solution determines the quality of anaesthetic cover. Fibrous septa may divide the perivascular space into compartments, leading to inadequate diffusion of the anaesthetic solution. The aim of our study was to obtain good anaesthesia and less complications using high volume of low concentration anaesthetic solution, overcoming the obstacle of the septa with a double approach to the scalene sheath., Methods: Sixty patients scheduled for shoulder capsuloplasty received both Winnie interscalene brachial plexus block and Pippa proximal cranial needle approach. The patients were randomly assigned to two groups. A constant dose of local anaesthetic was administered to each group: Group I (30 patients) received high volume (60 mL of anaesthetic solution) and Group II (30 patients) received low volume of solution (30 mL of anaesthetic solution). Sensory and motor block in the upper limb and complications were evaluated., Results: In all the patients the quality of anaesthesia obtained at the surgical site was excellent. In Group I also the areas supplied by the medial cutaneous nerves of the arm and forearm, ulnar, median and radial nerves were blocked (P < 0.002). Complications were only observed in Group II and consisted of bradycardia and hypotension (66% of the patients) and phrenic nerve paresis (27% of the patients)., Conclusions: The lower concentration of the anaesthetic solution avoids complications while increased volume provides good analgesic cover. The combination of the Winnie interscalene plexus block and the Pippa proximal cranial needle approach should contribute to fill up the scalene sheath overcoming the septa obstacles.

Endoscopic anterior cruciate ligament (ACL) reconstruction is one of the most popular orthopaedic procedures. Correct tunnel positioning is a prerequisite to success. Current surgical techniques are unable to duplicate the complex anatomy and function of the native ACL. Surgery mainly aims at restoring anteroposterior laxity. The ACL is not isometric and only a few fibers are nearly isometric over the full range of motion. However, a nearly isometric behaviour of the ACL graft is desirable. Isometry is mainly influenced by femoral attachment; thus the femoral tunnel position has a greater effect than the tibial on graft length changes. The purpose of this article is to describe the anatomy of the femoral ACL insertion and to discuss the surgical techniques used to replicate it.

Purpose: To study in cadaver knees the position of the tibial tunnel in anterior cruciate ligament (ACL) reconstruction using the 65 degrees Howell guide (Arthrotek, Ontario, CA)., Type of Study: Controlled laboratory study in vitro., Methods: Twenty-one fresh-frozen cadaver knees were used. The ACL was resected and its tibial attachment was demarcated. To drill the guidewire, we used the Howell 65 degrees tibial guide, which references off of the intercondylar roof in extension to avoid impingement. The intra-articular position of the wire was digitized with a digital camera and referred to a transverse axis passing through the over-the-back position and a sagittal axis passing through the lateral aspect of the medial spine. The percentage position of the wire within the ACL attachment was also calculated, taking the posterior and medial limits as the 0% positions., Results: All the wires were within the ACL attachment: 17 were in the ACL posterolateral bundle attachment and the other 4 in the anteromedial. The average distance of the wire from the transverse and sagittal axes was 12 mm (SD, 3 mm) anterior and 1 mm (SD, 1 mm) lateral, respectively. The wire was positioned at 38% (SD, 16%) of the length of the ACL attachment and at 40% (SD, 17%) of the width. Eighty percent of the wires were positioned at between 35% and 48% of the attachment length., Conclusions: The 65 degrees Howell guide, which positions the tibial tunnel in extension to avoid roof impingement, ensures anatomic positioning of the graft on the tibial side and reproducibility can be expected., Clinical Relevance: This study proves that a commonly used drill guide succeeds in placing the ACL graft in the tibial anatomic attachment.

Forty-three patients who had undergone an anterior cruciate ligament (ACL) reconstruction using a doubled semitendinosus and gracilis graft were prospectively reviewed at 5-year follow-up. All had suffered subacute or chronic tears of the ligament. At surgery, the femoral tunnel was drilled first through the antero-medial portal. The correct position of the femoral and tibial guide wire was checked fluoroscopically. A cortical fixation to the bone was achieved in the femur with a Mitek anchor, directly passing the two tendons in the slot of the anchor, and in the tibia with an RCI screw, supplemented with a spiked washer and bicortical screw. Rehabilitation was aggressive, controlled and without braces. The International Knee Documentation Committee (IKDC) form, KT-1000 arthrometer, and Cybex dynamometer were employed for clinical evaluation. A radiographic study was also performed. At the 5-year follow-up all the patients had recovered full range of motion and 2% of them complained of pain during light sports activities. Four patients (9.5%) reported giving-way symptoms. The KT-1000 side-to-side difference was on average 2.1 mm at 30 lb, and 68% of the knees were within 2 mm. The final IKDC score showed 90% satisfactory results. There was no difference between the 2-year and 5-year evaluations in terms of stability. Extensor and flexor muscle strength recovery was almost complete (maximum deficit 5%). Radiographic study showed a tunnel widening in 32% of the femurs and 40% of the tibias. A correlation was found between the incidence of tibial tunnel widening and the distance of the RCI screw from the joint (the closer the screw to the joint, the lower the incidence of widening). In conclusion, we can state that, using a four-strand hamstring graft and a cortical fixation at both ends, we were able to achieve satisfactory 5-year results in 90% of the patients.

In the biotechnology field, researchers often face complex moral and social issues while pursuing their scientific goals. Congress may regulate much of that research by debating the issues and placing restraints on funding granted by the government. In modern times, our presidents have also made some of those funding decisions through the issuance of executive orders. Presidents may issue executive orders with binding legal effect, despite having little constitutional authority to do so. Unless a new president takes office, only Congress or the courts can invalidate an executive order. In the regulation-sensitive biotechnology industry, presidents may exert profound influence by announcing executive orders. Former President Clinton and President George W. Bush are just the latest presidents who have used such orders to influence biotechnology as they have deemed appropriate. In doing so, those presidents have assumed a degree of unilateral lawmaking power over an industry that is fundamentally important to every citizen. This article stresses the need for the other branches of our government to discontinue granting unconstitutional legislative power to the president, and bring more social and scientific balance to important biotechnology policy decisions.

Therapeutic use of type I IFN (IFN-alpha/beta) has become common. Many of the diverse diseases targeted are marked by pathogenetic abnormalities in cell-mediated immunity (CMI), these cellular immune responses either causing injury to the host, lacking sufficient vigor for virus or tumor clearance, or both. In general, therapeutic efficacy is limited. It is thus notable that the pleiotropic effects of type I IFN on CMI remain poorly understood. We characterized the effects of type I IFN on the production of IL-12, the central immunoregulatory cytokine of the CD4(+) T cell arm of CMI. We show that type I IFN are potent inhibitors of IL-12 production by human monocytes/macrophages. The underlying mechanism involves transcriptional inhibition of the IL-12p40 gene, marked by down-regulation of PU.1 binding activity at the upstream Ets site of the IL-12p40 promoter. Type I IFN have previously been shown to be able to substitute for IL-12 in driving IFN-gamma production from T and NK cells. The ability of IFN-alpha/beta to suppress IL-12 production while up-regulating IFN-gamma production suggests a possible mechanistic basis for the difficulties of employing these cytokines in diseases involving abnormalities of CMI.

The prevalence and severity of allergic asthma continue to rise, lending urgency to the search for environmental triggers and genetic substrates. Using microarray analysis of pulmonary gene expression and single nucleotide polymorphism-based genotyping, combined with quantitative trait locus analysis, we identified the gene encoding complement factor 5 (C5) as a susceptibility locus for allergen-induced airway hyperresponsiveness in a murine model of asthma. A deletion in the coding sequence of C5 leads to C5-deficiency and susceptibility. Interleukin 12 (IL-12) is able to prevent or reverse experimental allergic asthma. Blockade of the C5a receptor rendered human monocytes unable to produce IL-12, mimicking blunted IL-12 production by macrophages from C5-deficient mice and providing a mechanism for the regulation of susceptibility to asthma by C5. The role of complement in modulating susceptibility to asthma highlights the importance of immunoregulatory events at the interface of innate and adaptive immunity in disease pathogenesis.

Endotoxin tolerance, the down-regulation of a subset of endotoxin-driven responses after an initial exposure to endotoxin, may provide protection from the uncontrolled immunological activation of acute endotoxic shock. Recent data suggest, however, that the inhibition of monocyte/macrophage function associated with endotoxin tolerance can lead to an inability to respond appropriately to secondary infections in survivors of endotoxic shock. IL-12 production by antigen-presenting cells is central to the orchestration of both innate and acquired cell-mediated immune responses to many pathogens. IL-12 has also been shown to play an important role in pathological responses to endotoxin. We therefore examined the regulation of IL-12 during endotoxin tolerance. Priming doses of lipopolysaccharide ablate the IL-12 productive capacity of primary human monocytes. This suppression of IL-12 production is primarily transcriptional. Unlike the down-regulation of TNF-alpha under such conditions, the mechanism of IL-12 suppression during endotoxin tolerance is not dependent upon IL-10 or transforming growth factor-beta, nor is IL-12 production rescued by IFN-gamma or granulocyte-macrophage colony-stimulating factor. Of note, human dendritic cells also undergo endotoxin tolerance, with potent down-regulation of IL-12 production. Endotoxin tolerance-related suppression of IL-12 production provides a likely mechanism for the anergy seen during the immunological paralysis which follows septic shock.

Glutaraldehyde-cross-linked microtubules were investigated as substrates for kinesin motility. Microtubules, formed in vitro from chicken brain tubulin, were stabilized with Taxol and chemically fixed with glutaraldehyde. The degree of tubulin monomer cross-linking as a function of time and glutaraldehyde concentration was characterized using polyacrylamide gel electrophoresis. Atomic force microscopy of fixed microtubules indicated that the cross-linking is sufficient to stabilize the gross structure of the microtubules against air drying or a distilled water challenge. Kinesin movement on immobilized, fixed microtubules was determined using a kinesin-coated bead motility assay observed with differential interference contrast microscopy. Within measurement error, kinesin bead movement velocities were independent of the degree of microtubule cross-linking. Binding affinity, however, decreased with increased cross-linking. Although air- and water-challenged microtubules did not support kinesin motility, a dilute suspension of glutaraldehyde-fixed microtubules in buffer supported kinesin motility for at least 2 days without any substantial degradation of activity. Fixed microtubules may be useful for several applications, including affinity purification of microtubule-associated proteins and motility measurements under extreme conditions of temperature and other variables.

The mechanisms underlying the profound suppression of cell-mediated immunity (CMI) accompanying measles are unclear. Interleukin-12 (IL-12), derived principally from monocytes and macrophages, is critical for the generation of CMI. Measles virus (MV) infection of primary human monocytes specifically down-regulated IL-12 production. Cross-linking of CD46, a complement regulatory protein that is the cellular receptor for MV, with antibody or with the complement activation product C3b similarly inhibited monocyte IL-12 production, providing a plausible mechanism for MV-induced immunosuppression. CD46 provides a regulatory link between the complement system and cellular immune responses.