Efficacy of balneotherapy on pain, function, and sleep quality in patients with chronic low-back pain: a prospective observational study.

The aim of this prospective observational study was to evaluate the efficacy of a cycle of sulfate-arsenical-ferruginous mud bath therapy on pain, function, and sleep quality in patients with chronic low-back pain (CLBP). One hundred twenty-three patients treated at Levico Spa Center (Italy) with 12 daily local mud packs and generalized thermal baths were included in this experience. General medical assessments were performed before starting the therapy, at the end of the treatment, and 3 months later. Pain intensity and stiffness were measured by a 0 to 10 cm visual analogue scale (VAS); the range of mobility of the lumbar spine was evaluated using the Schober test and functional disability by the Roland-Morris Disability Questionnaire (RMDI). The Pittsburgh Sleep Quality Index (PSQI) was used to assess the sleep quality. The VAS score for pain and stiffness showed a significant reduction (p < 0.0001) at the end of the treatment persisting for 3 months of follow-up. Similarly, a significant improvement was found for the Schober test and RMDQ. Finally, we showed a significant decrease of PSQI score at the end of the cycle of mud bath and at 3 months of follow-up. The Spearman analysis showed a significant positive correlation between the score of PSQI and VAS pain, VAS stiffness, and RMDQ. In conclusion, this preliminary study confirms the beneficial and long-term efficacy of balneotherapy on pain and function and, for the first one, shows the positive effect on quality of sleep in patients with CLBP treated with a cycle of mud bath therapy.

Spa therapy and peripheral serotonin and dopamine function: a systematic review.

Spa therapy consists of multiple techniques based on the healing effects of water, including hydrotherapy, balneotherapy, and mud therapy, often combined with therapeutic exercises, massage, or physical therapy. Balneotherapy is a clinically effective complementary approach in the treatment of low-grade inflammation- and stress-related pathologies, especially rheumatic conditions due to its anti-inflammatory properties. The main objective of this investigation was to conduct a systematic review analyzing the available evidence on the effect of spa therapy on serotonin and dopamine function. The databases PubMed, Web of Science, Scopus, and Cochrane Central Register of Controlled Trials (CENTRAL) were used from June to July 2023. Exclusion criteria were (1) articles not written in English, (2) full text not available, (3) article not related to the objective of the review. JADAD scale was used for methodological quality evaluation. Four studies were included in the systematic review. Two studies were related to serotonin in healthy individuals, one to serotonin in fibromyalgia, and one to dopamine in healthy individuals. One of the studies evaluated hydrotherapy, another one balneotherapy and mud-bath therapy, and the other two assessed balneotherapy interventions. Studies were very heterogeneous, and their methodological quality was low, making it difficult to draw clear conclusions regarding the effect of spa therapy on peripheral serotonin and dopamine function. The findings of this review highlight the lack of studies evaluating these neurotransmitters and hormones in the context of spa therapy. Further research is needed to evaluate the potential effects of these therapies on serotonin or dopamine function.

Structure and function of the EA1 surface layer of Bacillus anthracis.

The Gram-positive spore-forming bacterium Bacillus anthracis is the causative agent of anthrax, a deadly disease mostly affecting wildlife and livestock, as well as representing a bioterrorism threat. Its cell surface is covered by the mutually exclusive S-layers Sap and EA1, found in early and late growth phases, respectively. Here we report the nanobody-based structural characterization of EA1 and its native lattice contacts. The EA1 assembly domain consists of 6 immunoglobulin-like domains, where three calcium-binding sites structure interdomain contacts that allow monomers to adopt their assembly-competent conformation. Nanobody-induced depolymerization of EA1 S-layers results in surface defects, membrane blebbing and cell lysis under hypotonic conditions, indicating that S-layers provide additional mechanical stability to the cell wall. Taken together, we report a complete model of the EA1 S-layer and present a set of nanobodies that may have therapeutic potential against Bacillus anthracis.

MicroRNA as Possible Mediators of the Synergistic Effect of Celecoxib and Glucosamine Sulfate in Human Osteoarthritic Chondrocyte Exposed to IL-1ß.

This study investigated the role of a pattern of microRNA (miRNA) as possible mediators of celecoxib and prescription-grade glucosamine sulfate (GS) effects in human osteoarthritis (OA) chondrocytes. Chondrocytes were treated with celecoxib (1.85 µM) and GS (9 µM), alone or in combination, for 24 h, with or without interleukin (IL)-1ß (10 ng/mL). Cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, apoptosis and reactive oxygen species (ROS) by cytometry, nitric oxide (NO) by Griess method. Gene levels of miRNA, antioxidant enzymes, nuclear factor erythroid (NRF)2, and B-cell lymphoma (BCL)2 expressions were analyzed by quantitative real time polymerase chain reaction (real time PCR). Protein expression of NRF2 and BCL2 was also detected at immunofluorescence and western blot. Celecoxib and GS, alone or in combination, significantly increased viability, reduced apoptosis, ROS and NO production and the gene expression of miR-34a, -146a, -181a, -210, in comparison to baseline and to IL-1ß. The transfection with miRNA specific inhibitors significantly counteracted the IL-1ß activity and potentiated the properties of celecoxib and GS on viability, apoptosis and oxidant system, through nuclear factor (NF)-κB regulation. The observed effects were enhanced when the drugs were tested in combination. Our data confirmed the synergistic anti-inflammatory and chondroprotective properties of celecoxib and GS, suggesting microRNA as possible mediators.

Autoantibody status according to multiparametric assay accurately estimates connective tissue disease classification and identifies clinically relevant disease clusters.

OBJECTIVE: Assessment of circulating autoantibodies represents one of the earliest diagnostic procedures in patients with suspected connective tissue disease (CTD), providing important information for disease diagnosis, identification and prediction of potential clinical manifestations. The purpose of this study was to evaluate the ability of multiparametric assay to correctly classify patients with multiple CTDs and healthy controls (HC), independent of clinical features, and to evaluate whether serological status could identify clusters of patients with similar clinical features. METHODS: Patients with systemic lupus erythematosus (SLE), systemic sclerosis (SSc), Sjogren's syndrome (SjS), undifferentiated connective tissue disease (UCTD), idiopathic inflammatory myopathies (IIM) and HC were enrolled. Serum was tested for 29 autoantibodies. An XGBoost model, exclusively based on autoantibody titres was built and classification accuracy was evaluated. A hierarchical clustering model was subsequently developed and clinical/laboratory features compared among clusters. RESULTS: 908 subjects were enrolled. The classification model showed a mean accuracy of 60.84±4.05% and a mean area under the receiver operator characteristic curve of 88.99±2.50%, with significant discrepancies among groups. Cluster analysis identified four clusters (CL). CL1 included patients with typical features of SLE. CL2 included most patients with SjS, along with some SLE and UCTD patients with SjS-like features. CL4 included anti-Jo1 patients only. CL3 was the largest and most heterogeneous, including all the remaining subjects, overall characterised by low titre or lower-prevalence autoantibodies. CONCLUSION: Extended multiparametric autoantibody assay allowed an accurate classification of CTD patients, independently of clinical features. Clustering according to autoantibody titres is able to identify clusters of CTD subjects with similar clinical features, independently of their final diagnosis.

Improvement of Pain Symptoms in Musculoskeletal Diseases After Multimodal Spa Therapy in the Austrian Gastein Valley-A Study Based on Longitudinal Registry Data.

Objective: The study aim was to investigate the course of pain in rest and motion in seven different rheumatic diseases (RMD), prior and after multimodal spa therapy including low-dose radon treatment and at 3-, 6-; and 9-month follow up. Methods: Complete data from the radon indication registry including information on 561 subjects with RMD were analysed to explore the association of timepoint of measurement with pain in rest and motion. For this purpose, linear regression models adjusted for RMD-type, age, sex and body mass index (BMI) were applied. Results: The mean age of the sample was 55 years, the average body mass index was 26.8, and 275 subjects were women. Pain scores were significantly improved at all-time points compared to baseline. Pain courses were different for each RMD with the largest improvement seen in fibromyalgia. Conclusion: Timing spa facility visits according to RMD-specific pain courses may result in sustained pain reduction.

An update on the use of conventional and biological disease-modifying anti-rheumatic drugs in hand osteoarthritis.

Osteoarthritis (OA) is a highly prevalent condition worldwide associated with pain, progressive disability, reduced participation in social activities, and impaired quality of life. Despite its growing burden, the therapeutic options are still limited and almost exclusively addressed to symptoms' management, while no disease-modifying OA drugs able to prevent or retard disease progression are actually available. For these reasons, in the last decades, relevant efforts to find new potential therapeutic targets in OA have been made and a number of existing conventional and biological disease-modifying anti-rheumatic drugs (DMARDs), including hydroxychloroquine (HCQ), methotrexate (MTX), tumor necrosis factor (TNF)-α, interleukin (IL)-1, and IL-6 inhibitors, commonly used to treat inflammatory rheumatic diseases, have been repurposed for the treatment of OA and explored also in hand osteoarthritis (HOA). The current narrative review was aimed to provide a comprehensive and updated understanding of the possibilities and the criticisms related to the treatment of HOA with conventional and biological DMARDs. Unfortunately, therapy with conventional and biologic drugs in HOA has not achieved the expected success, despite a rationale for their use exists. Thus, our findings outline the urgent need to enhance the exploration of HOA basic molecular mechanisms to find new potential therapeutic targets, personalized for each patient, and appropriate for the different subsets of HOA and for the different phases of disease.

SARS CoV-2 spike protein-guided exosome isolation facilitates detection of potential miRNA biomarkers in COVID-19 infections.

OBJECTIVES: Nearly three years into the pandemic, SARS-CoV-2 infections are occurring in vaccinated and naturally infected populations. While humoral and cellular responses in COVID-19 are being characterized, novel immune biomarkers also being identified. Recently, an increase in angiotensin-converting enzyme 2 expressing (aka, ACE2 positive) circulating exosomes (ExoACE2) were identified in the plasma of COVID-19 patients (El-Shennawy et al.). In this pilot study, we describe a method to characterize the exosome-associated microRNA (exo-miRNA) signature in ACE2-positive and ACE2-negative exosomal populations (non-ExoACE2). METHODS: We performed a sorting protocol using the recombinant biotin-conjugated SARS CoV-2 spike protein containing the receptor binding domain (RBD) on plasma samples from six patients. Following purification, exo-miRNA were characterized for ACE2-positive and ACE2-negative exosome subpopulations by RT-PCR. RESULTS: We identified differential expression of several miRNA. Specifically let-7g-5p and hsa-miR-4454+miR-7975 were upregulated, while hsa-miR-208a-3p and has-miR-323-3p were downregulated in ExoACE2 vs. non-ExoACE2. CONCLUSIONS: The SARS CoV-2 spike-protein guided exosome isolation permits isolation of ExoACE2 exosomes. Such purification facilitates detailed characterization of potential biomarkers (e.g. exo-miRNA) for COVID-19 patients. This method could be used for future studies to further the understanding mechanisms of host response against SARS CoV-2.

Multiparametric autoantibody analysis: a new paradigm for the diagnosis of connective tissue diseases.

BACKGROUND: In patients affected by connective tissue diseases (CTDs), the identification of wide autoantibody profiles may prove useful in early diagnosis, in the evaluation of prognosis (risk stratification), and in predicting response to therapy. The aim of the present study was to evaluate the utility of multiparametric autoantibody analysis performed by a new fully automated particle-based multi-analyte technology (PMAT) digital system in a large multicenter cohort of CTD patients and controls. METHODS: Serum samples from 787 patients with CTD (166 systemic lupus erythematosus; 133 systemic sclerosis; 279 Sjögren's syndrome; 106 idiopathic inflammatory myopathies; 103 undifferentiated CTD), 339 patients with other disorders (disease controls) (118 infectious diseases, 110 organ-specific autoimmune diseases, 111 other rheumatic diseases), and 121 healthy subjects were collected in 13 rheumatologic centers of the FIRMA group. Sera were analyzed with the Aptiva-PMAT instrument (Inova Diagnostics) for a panel of 29 autoantibodies. RESULTS: Multiparametric logistic regression showed that enlarged antibody profiles have a higher diagnostic efficiency than that of individual antibodies or of antibodies that constitute classification criteria for a given disease and that probability of disease increases with multiple positive autoantibodies. CONCLUSIONS: This is the first study that analyzes the clinical and diagnostic impact of autoantibody profiling in CTD. The results obtained with the new Aptiva-PMAT method may open interesting perspectives in the diagnosis and sub-classification of patients with autoimmune rheumatic diseases.

Possible synergic action of non-steroidal anti-inflammatory drugs and glucosamine sulfate for the treatment of knee osteoarthritis: a scoping review.

BACKGROUND: Several studies have reported that glucosamine sulfate (GS) can improve knee osteoarthritis (OA) symptomatology. In parallel, the disease-modifying effects of non-steroidal anti-inflammatory drugs (NSAIDs) in knee OA have also been investigated. However, limited literature has reported the combined effect of GS and NSAIDs. The aim of this scoping review is to describe the scope and volume of the literature investigating the potential benefits and synergistic effect of a combination of GS and NSAIDs in patients with knee OA. METHODS: PubMed and Embase were searched for studies published from inception through April 2022, evaluating the effects of the combination of GS and NSAIDs in OA patients, versus either treatment alone. Data are reported narratively. RESULTS: Five studies were included in this review; 4 were randomized control trials and one was a prospective observational study. The duration of combination treatment was 6 to 12 weeks. The combination was compared to celecoxib in 2 studies, meloxicam in 1, etoricoxib in 1, and a conventional NSAID in 1 (ibuprofen or piroxicam). All 5 studies reported that in patients with knee OA, the combination of GS plus NSAID yielded a significantly greater benefit than single-agent therapy, in terms of outcomes including pain reduction, function, joint stiffness, and markers of inflammatory activity and cartilage degradation. CONCLUSION: The 5 studies included in this scoping review all report a significantly greater clinical benefit with a combination of GS plus NSAID compared to either treatment alone. The evidence supports efficacy in reducing pain, improving function, and possibly regulating joint damage. However, further randomized trials with larger sample sizes are warranted to confirm these findings.

Multimodal Multidisciplinary Management of Patients with Moderate to Severe Pain in Knee Osteoarthritis: A Need to Meet Patient Expectations.

Knee osteoarthritis (OA) is one of the most common and disabling medical conditions. In the case of moderate to severe pain, a single intervention may not be sufficient to allay symptoms and improve quality of life. Examples include first-line, background therapy with symptomatic slow-acting drugs for OA (SYSADOAs) or non-steroidal anti-inflammatory drugs (NSAIDs). Therefore, the European Society for Clinical and Economic Aspects of Osteoporosis, Osteoarthritis and Musculoskeletal Diseases (ESCEO) performed a review of a multimodal/multicomponent approach for knee OA therapy. This strategy is a particularly appropriate solution for the management of patients affected by knee OA, including those with pain and dysfunction reaching various thresholds at the different joints. The multimodal/multicomponent approach should be based, firstly, on different combinations of non-pharmacological and pharmacological interventions. Potential pharmacological combinations include SYSADOAs and NSAIDs, NSAIDs and weak opioids, and intra-articular treatments with SYSADOAs/NSAIDs. Based on the available evidence, most combined treatments provide benefit beyond single agents for the improvement of pain and other symptoms typical of knee OA, although further high-quality studies are required. In this work, we have therefore provided new, patient-centered perspectives for the management of knee OA, based on the concept that a multimodal, multicomponent, multidisciplinary approach, applied not only to non-pharmacological treatments but also to a combination of the currently available pharmacological options, will better meet the needs and expectations of patients with knee OA, who may present with various phenotypes and trajectories.

Synovial Fluid Regulates the Gene Expression of a Pattern of microRNA via the NF-κB Pathway: An In Vitro Study on Human Osteoarthritic Chondrocytes.

Synovial fluid (SF) represents the primary source of nutrients of articular cartilage and is implicated in maintaining cartilage metabolism. We investigated the effects of SF, from patients with osteoarthritis (OA), rheumatoid arthritis (RA), and controls, on a pattern of microRNA (miRNA) in human OA chondrocytes. Cells were stimulated with 50% or 100% SF for 24 h and 48 h. Apoptosis and superoxide anion production were detected by cytometry; miRNA (34a, 146a, 155, 181a), cytokines, metalloproteinases (MMPs), type II collagen (Col2a1), antioxidant enzymes, B-cell lymphoma (BCL)2, and nuclear factor (NF)-κB by real-time PCR. The implication of the NF-κB pathway was assessed by the use of NF-κB inhibitor (BAY-11-7082). RA and OA SF up-regulated miR-34a, -146a, -155, -181a, interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, MMP-1, MMP-13, and ADAMTs-5 gene expression, while it down-regulated Col2a1. Pathological SF also induced apoptosis, reduced viability, and decreased BCL2 mRNA, whereas it increased superoxide anions, the expression of antioxidant enzymes, p65 and p50 NF-κB. Opposite and positive results were obtained with 100% control SF. Pre-incubation with BAY-11-7082 counteracted SF effects on miRNA. We highlight the role of the SF microenvironment in regulating some miRNA involved in inflammation and cartilage degradation during OA and RA, via the NF-κB pathway.

Crosstalk between MicroRNA and Oxidative Stress in Physiology and Pathology 2.0.

MicroRNAs (miRNAs) are a class of small non-coding RNAs around 22 nucleotides long that regulate gene expression by binding specific sequences within target messenger RNA (mRNA) [...].

Prescription-grade crystalline glucosamine sulfate as an add-on therapy to conventional treatments in erosive osteoarthritis of the hand: results from a 6-month observational retrospective study.

OBJECTIVE: To evaluate the efficacy of prescription-grade Crystalline Glucosamine Sulfate (pCGS) as an add-on treatment to conventional therapy, compared to usual therapy alone, in patients with erosive osteoarthritis of the hand (EHOA). METHODS: This 6-month retrospective case-control study included patients with concomitant knee osteoarthritis and symptomatic EHOA. Participants were stratified into two groups based on whether or not pCGS (1500 mg/day) was added to the conventional therapy (education and training in ergonomic principles, exercise and use on-demand of symptomatic drugs) for hand osteoarthritis. Patients were evaluated at baseline, after 3 and 6 months. Primary outcomes were the change from baseline to month 6 in Visual Analogue Scale (VAS) hand pain and in Functional Index for Hand Osteoarthritis (FIHOA) score. A set of secondary parameters was also evaluated. RESULTS: 123 patients were included as follows: 67 treated with pCGS in addition to conventional therapy (pCGS Group) and 56 with conventional therapy alone (Control Group). After 6 months a significant difference in VAS and in FIHOA score (p < 0.01 and p < 0.001, respectively) was observed in favor of pCGS Group. Similar results were found for morning stiffness duration (p < 0.05), health assessment questionnaire (p < 0.01) and physical and mental component score of 36-item short form (p < 0.05 and p < 0.001, respectively). A significant reduction of symptomatic drug consumption at 3 and 6 months was reported in the pCGS Group (p < 0.001). No serious adverse event was recorded in both groups. CONCLUSIONS: Despite all the limitations inherent to an observational study, our results suggest the potential effectiveness of pCGS, when used in combination with conventional therapy in EHOA. Further randomized placebo-controlled trials are needed to confirm these positive findings. TRIAL REGISTRATION: ClinicalTrials.gov, http://www. CLINICALTRIALS: gov , date of registration: February 2, 2022, NCT05237596. The present trial was retrospectively registered.

Clinical Delphi on aPL Negativization: Report from the APS Study Group of the Italian Society for Rheumatology (SIR-APS).

BACKGROUND: The rate of antiphospholipid antibody (aPL) negativization in antiphospholipid syndrome (APS) patients is uncertain, but it is estimated to be as high as 8%. Currently, a consensus definition of aPL negativization is lacking, as well as international recommendations on how to approach treatment in patients with a persistent aPL-negative seroconversion. AIM: The aim of the Delphi survey was to evaluate the clinical approach and level of consensus among experts from the APS Study Group of the Italian Society for Rheumatology (SIR-APS) in different clinical scenarios. METHODS: Experts of SIR-APS were contacted using a survey methodology. RESULTS: A structured survey was circulated among 30 experts. Up to 90% of the interviewed experts agreed on defining aPL negativization as the presence of two negative determinations, 1 year apart (90%). Almost full consensus exists among experts in some clinical settings, including: (1) the role of aPL negativization in the management of a thrombotic event determined by concomitant presence of cardiovascular risk factors, both modifiable and not modifiable (90%); (2) approach to young patients with triple aPL positivity who experienced pulmonary arterial thrombotic events and tested negative for aPL detection after 5 years of vitamin K antagonist (VKA) treatment (90%); (3) the use of "extra criteria" aPL antibody testing before pondering VKA suspension (93%). CONCLUSION: A substantial agreement exists among experts on how to define aPL negativization. VKA suspension should be embraced with extreme caution, particularly in case of previous thrombotic events and/or triple aPL positivity. Nevertheless, VKA cessation might be considered when risk factors are carefully monitored/treated and the presence of "extra criteria" aPL is ruled out.

Circulating Mir-140 and leptin improve the accuracy of the differential diagnosis between psoriatic arthritis and rheumatoid arthritis: a case-control study.

The differential diagnosis of psoriatic arthritis (PsA) and rheumatoid arthritis (RA) is difficult because of the lack of diagnostic clinical signs and reliable biomarkers. This study investigated microRNAs (miRNA) and adipokines as potential additional markers to discriminate PsA from RA. The expression profile of miRNA (miR-21, miR-140, miR-146a, miR-155, miR-181b, miR-223, miR-let-7e) and inflammatory cytokines (IL-1ß, IL-6, IL-17a, IL-23a, TNF-α) from peripheral blood mononuclear cells of PsA and RA patients compared to healthy controls (HC) were evaluated by real-time PCR, and serum adipokines (adiponectin, chemerin, leptin, resistin, visfatin) and cytokines by ELISA assay. Univariable binary logistic regression was used to find the association between PsA and potential predictors. The gene expression of miRNA and cytokines and the serum levels of adipokines were found significantly different in PsA and RA patients compared to HC, as well as in PsA versus RA. MiR-140 gene expression resulted up-regulated in PsA patients and reduced in RA in comparison to HC, and, for the first time, significantly higher in PsA compared with RA. Serum levels of IL-23a and leptin were significantly increased in PsA and RA populations than in HC, as well as in PsA versus RA. Furthermore, circulating TNF-α was up-regulated in PsA and RA in comparison to controls, while resulted higher in RA than in PsA. Univariable binary logistic regression analysis found the above-mentioned markers associated to PsA versus RA. Our results first demonstrated an increased expression of circulating miR-140 and serum leptin in PsA patients compared to RA, which were identified as potential additional biomarkers to discriminate PsA from RA. Since the differential diagnosis of PsA and RA poses challenges in clinical practice, our data may help to enhance the diagnostic performance of PsA in daily practice.

Balneotherapy year in review 2021: focus on the mechanisms of action of balneotherapy in rheumatic diseases.

Balneotherapy (BT) is one of the most commonly used non-pharmacologic complementary therapies for different rheumatic diseases. Its beneficial properties probably derived from a combination of mechanical, thermal, and chemical effects, but the exact mechanism of action is not elucidated. This review aimed at summarizing the current knowledge about the effects of BT, and identifying its possible mechanism of action in different rheumatic diseases. Pubmed and Scopus were used to perform a search of the literature to extract articles including terms related to BT and rheumatic diseases published in the period from 2010 to 2021. We selected pre-clinical studies, randomized controlled trials, and clinical trials. The results of clinical studies confirmed the beneficial properties on different mediators and factors of inflammation, oxidative stress, cartilage metabolism, and humoral and cellular immune responses in patients affected by chronic degenerative musculoskeletal disorders. The data derived from OA and RA-induced murine models revealed the efficacy of different BT treatments in decreasing pain, inflammation, and improving mobility, as well as in reducing the expression of matrix-degrading enzymes and markers of oxidative stress damage. Different in vitro studies analyzed the potential effect of a mineral water, as a whole, or of a mineral element, demonstrating their anti-inflammatory, antioxidant, and chondroprotective properties in OA cartilage, synoviocytes and chondrocytes, and osteoblast and osteoclast cultures. The presented data are promising and confirm BT as an effective complementary approach in the management of several low-grade inflammation, degenerative, and stress-related pathologies, as rheumatic diseases.

The Bacillus anthracis S-layer is an exoskeleton-like structure that imparts mechanical and osmotic stabilization to the cell wall.

Surface layers (S-layers) are 2D paracrystalline protein monolayers covering the cell envelope of many prokaryotes and archaea. Proposed functions include a role in cell support, as scaffolding structure, as molecular sieve, or as virulence factor. Bacillus anthracis holds two S-layers, composed of Sap or EA1, which interchange in early and late exponential growth phase. We previously found that acute disruption of B. anthracis Sap S-layer integrity, by means of nanobodies, results in severe morphological cell surface defects and cell collapse. Remarkably, this loss of function is due to the destruction of the Sap lattice structure rather than detachment of monomers from the cell surface. Here, we combine force nanoscopy and light microscopy observations to probe the contribution of the S-layer to the mechanical, structural, and functional properties of the cell envelope, which have been so far elusive. Our experiments reveal that cells with a compromised S-layer lattice show a decreased compressive stiffness and elastic modulus. Furthermore, we find that S-layer integrity is required to resist cell turgor under hypotonic conditions. These results present compelling experimental evidence indicating that the S-layers can serve as prokaryotic exoskeletons that support the cell wall in conferring rigidity and mechanical stability to bacterial cells.

A Combination of Celecoxib and Glucosamine Sulfate Has Anti-Inflammatory and Chondroprotective Effects: Results from an In Vitro Study on Human Osteoarthritic Chondrocytes.

This study investigated the possible anti-inflammatory and chondroprotective effects of a combination of celecoxib and prescription-grade glucosamine sulfate (GS) in human osteoarthritic (OA) chondrocytes and their possible mechanism of action. Chondrocytes were treated with celecoxib (1.85 µM) and GS (9 µM), alone or in combination with IL-1ß (10 ng/mL) and a specific nuclear factor (NF)-κB inhibitor (BAY-11-7082, 1 µM). Gene expression and release of some pro-inflammatory mediators, metalloproteinases (MMPs), and type II collagen (Col2a1) were evaluated by qRT-PCR and ELISA; apoptosis and mitochondrial superoxide anion production were assessed by cytometry; B-cell lymphoma (BCL)2, antioxidant enzymes, and p50 and p65 NF-κB subunits were analyzed by qRT-PCR. Celecoxib and GS alone or co-incubated with IL-1ß significantly reduced expression and release of cyclooxygenase (COX)-2, prostaglandin (PG)E2, IL-1ß, IL-6, tumor necrosis factor (TNF)-α, and MMPs, while it increased Col2a1, compared to baseline or IL-1ß. Both drugs reduced apoptosis and superoxide production; reduced the expression of superoxide dismutase, catalase, and nuclear factor erythroid; increased BCL2; and limited p50 and p65. Celecoxib and GS combination demonstrated an increased inhibitory effect on IL-1ß than that observed by each single treatment. Drugs effects were potentiated by pre-incubation with BAY-11-7082. Our results demonstrated the synergistic effect of celecoxib and GS on OA chondrocyte metabolism, apoptosis, and oxidative stress through the modulation of the NF-κB pathway, supporting their combined use for the treatment of OA.