The life-saving benefit of dexamethasone in severe COVID-19 is linked to a reversal of monocyte dysregulation.

Dexamethasone is a life-saving treatment for severe COVID-19, yet its mechanism of action is unknown, and many patients deteriorate or die despite timely treatment initiation. Here, we identify dexamethasone treatment-induced cellular and molecular changes associated with improved survival in COVID-19 patients. We observed a reversal of transcriptional hallmark signatures in monocytes associated with severe COVID-19 and the induction of a monocyte substate characterized by the expression of glucocorticoid-response genes. These molecular responses to dexamethasone were detected in circulating and pulmonary monocytes, and they were directly linked to survival. Monocyte single-cell RNA sequencing (scRNA-seq)-derived signatures were enriched in whole blood transcriptomes of patients with fatal outcome in two independent cohorts, highlighting the potential for identifying non-responders refractory to dexamethasone. Our findings link the effects of dexamethasone to specific immunomodulation and reversal of monocyte dysregulation, and they highlight the potential of single-cell omics for monitoring in vivo target engagement of immunomodulatory drugs and for patient stratification for precision medicine approaches.

Severe COVID-19 Is Marked by a Dysregulated Myeloid Cell Compartment.

Coronavirus disease 2019 (COVID-19) is a mild to moderate respiratory tract infection, however, a subset of patients progress to severe disease and respiratory failure. The mechanism of protective immunity in mild forms and the pathogenesis of severe COVID-19 associated with increased neutrophil counts and dysregulated immune responses remain unclear. In a dual-center, two-cohort study, we combined single-cell RNA-sequencing and single-cell proteomics of whole-blood and peripheral-blood mononuclear cells to determine changes in immune cell composition and activation in mild versus severe COVID-19 (242 samples from 109 individuals) over time. HLA-DRhiCD11chi inflammatory monocytes with an interferon-stimulated gene signature were elevated in mild COVID-19. Severe COVID-19 was marked by occurrence of neutrophil precursors, as evidence of emergency myelopoiesis, dysfunctional mature neutrophils, and HLA-DRlo monocytes. Our study provides detailed insights into the systemic immune response to SARS-CoV-2 infection and reveals profound alterations in the myeloid cell compartment associated with severe COVID-19.

Early IFN-α signatures and persistent dysfunction are distinguishing features of NK cells in severe COVID-19.

Longitudinal analyses of the innate immune system, including the earliest time points, are essential to understand the immunopathogenesis and clinical course of coronavirus disease (COVID-19). Here, we performed a detailed characterization of natural killer (NK) cells in 205 patients (403 samples; days 2 to 41 after symptom onset) from four independent cohorts using single-cell transcriptomics and proteomics together with functional studies. We found elevated interferon (IFN)-α plasma levels in early severe COVD-19 alongside increased NK cell expression of IFN-stimulated genes (ISGs) and genes involved in IFN-α signaling, while upregulation of tumor necrosis factor (TNF)-induced genes was observed in moderate diseases. NK cells exert anti-SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) activity but are functionally impaired in severe COVID-19. Further, NK cell dysfunction may be relevant for the development of fibrotic lung disease in severe COVID-19, as NK cells exhibited impaired anti-fibrotic activity. Our study indicates preferential IFN-α and TNF responses in severe and moderate COVID-19, respectively, and associates a prolonged IFN-α-induced NK cell response with poorer disease outcome.

Swarm Learning for decentralized and confidential clinical machine learning.

Fast and reliable detection of patients with severe and heterogeneous illnesses is a major goal of precision medicine1,2. Patients with leukaemia can be identified using machine learning on the basis of their blood transcriptomes3. However, there is an increasing divide between what is technically possible and what is allowed, because of privacy legislation4,5. Here, to facilitate the integration of any medical data from any data owner worldwide without violating privacy laws, we introduce Swarm Learning-a decentralized machine-learning approach that unites edge computing, blockchain-based peer-to-peer networking and coordination while maintaining confidentiality without the need for a central coordinator, thereby going beyond federated learning. To illustrate the feasibility of using Swarm Learning to develop disease classifiers using distributed data, we chose four use cases of heterogeneous diseases (COVID-19, tuberculosis, leukaemia and lung pathologies). With more than 16,400 blood transcriptomes derived from 127 clinical studies with non-uniform distributions of cases and controls and substantial study biases, as well as more than 95,000 chest X-ray images, we show that Swarm Learning classifiers outperform those developed at individual sites. In addition, Swarm Learning completely fulfils local confidentiality regulations by design. We believe that this approach will notably accelerate the introduction of precision medicine.

Tocilizumab in Patients Hospitalized with Covid-19 Pneumonia.

BACKGROUND: Coronavirus disease 2019 (Covid-19) pneumonia is often associated with hyperinflammation. Despite the disproportionate incidence of Covid-19 among underserved and racial and ethnic minority populations, the safety and efficacy of the anti-interleukin-6 receptor antibody tocilizumab in patients from these populations who are hospitalized with Covid-19 pneumonia are unclear. METHODS: We randomly assigned (in a 2:1 ratio) patients hospitalized with Covid-19 pneumonia who were not receiving mechanical ventilation to receive standard care plus one or two doses of either tocilizumab (8 mg per kilogram of body weight intravenously) or placebo. Site selection was focused on the inclusion of sites enrolling high-risk and minority populations. The primary outcome was mechanical ventilation or death by day 28. RESULTS: A total of 389 patients underwent randomization, and the modified intention-to-treat population included 249 patients in the tocilizumab group and 128 patients in the placebo group; 56.0% were Hispanic or Latino, 14.9% were Black, 12.7% were American Indian or Alaska Native, 12.7% were non-Hispanic White, and 3.7% were of other or unknown race or ethnic group. The cumulative percentage of patients who had received mechanical ventilation or who had died by day 28 was 12.0% (95% confidence interval [CI], 8.5 to 16.9) in the tocilizumab group and 19.3% (95% CI, 13.3 to 27.4) in the placebo group (hazard ratio for mechanical ventilation or death, 0.56; 95% CI, 0.33 to 0.97; P = 0.04 by the log-rank test). Clinical failure as assessed in a time-to-event analysis favored tocilizumab over placebo (hazard ratio, 0.55; 95% CI, 0.33 to 0.93). Death from any cause by day 28 occurred in 10.4% of the patients in the tocilizumab group and 8.6% of those in the placebo group (weighted difference, 2.0 percentage points; 95% CI, -5.2 to 7.8). In the safety population, serious adverse events occurred in 38 of 250 patients (15.2%) in the tocilizumab group and 25 of 127 patients (19.7%) in the placebo group. CONCLUSIONS: In hospitalized patients with Covid-19 pneumonia who were not receiving mechanical ventilation, tocilizumab reduced the likelihood of progression to the composite outcome of mechanical ventilation or death, but it did not improve survival. No new safety signals were identified. (Funded by Genentech; EMPACTA ClinicalTrials.gov number, NCT04372186.).

Identification and characterization of a hepatic IL-13-producing ILC3-like population potentially involved in liver fibrosis.

BACKGROUND AND AIMS: Human innate lymphoid cells (ILCs) are critically involved in the modulation of homeostatic and inflammatory processes in various tissues. However, only little is known about the composition of the intrahepatic ILC pool and its potential role in chronic liver disease. Here, we performed a detailed characterization of intrahepatic ILCs in both healthy and fibrotic livers. APPROACH AND RESULTS: A total of 50 livers (nonfibrotic = 22, and fibrotic = 29) were analyzed and compared with colon and tonsil tissue (each N = 14) and peripheral blood (N = 32). Human intrahepatic ILCs were characterized ex vivo and on stimulation using flow cytometry and single-cell RNA sequencing. ILC differentiation and plasticity were analyzed by both bulk and clonal expansion experiments. Finally, the effects of ILC-derived cytokines on primary human HSteCs were studied. Unexpectedly, we found that an "unconventional" ILC3-like cell represented the major IL-13-producing liver ILC subset. IL-13 + ILC3-like cells were specifically enriched in the human liver, and increased frequencies of this cell type were found in fibrotic livers. ILC3-derived IL-13 production induced upregulation of proinflammatory genes in HSteCs, indicating a potential role in the regulation of hepatic fibrogenesis. Finally, we identified KLRG1-expressing ILC precursors as the potential progenitor of hepatic IL-13 + ILC3-like cells. CONCLUSIONS: We identified a formerly undescribed subset of IL-13-producing ILC3-like cells that is enriched in the human liver and may be involved in the modulation of chronic liver disease.

Profiling Cell Heterogeneity and Fructose Transporter Expression in the Rat Nephron by Integrating Single-Cell and Microdissected Tubule Segment Transcriptomes.

Single-cell RNA sequencing (scRNAseq) is a crucial tool in kidney research. These technologies cluster cells based on transcriptome similarity, irrespective of the anatomical location and order within the nephron. Thus, a transcriptome cluster may obscure the heterogeneity of the cell population within a nephron segment. Elevated dietary fructose leads to salt-sensitive hypertension, in part, through fructose reabsorption in the proximal tubule (PT). However, the organization of the four known fructose transporters in apical PTs (SGLT4, SGLT5, GLUT5, and NaGLT1) remains poorly understood. We hypothesized that cells within each subsegment of the proximal tubule exhibit complex, heterogeneous fructose transporter expression patterns. To test this hypothesis, we analyzed rat kidney transcriptomes and proteomes from publicly available scRNAseq and tubule microdissection databases. We found that microdissected PT-S1 segments consist of 81% ± 12% cells with scRNAseq-derived transcriptional characteristics of S1, whereas PT-S2 express a mixture of 18% ± 9% S1, 58% ± 8% S2, and 19% ± 5% S3 transcripts, and PT-S3 consists of 75% ± 9% S3 transcripts. The expression of all four fructose transporters was detectable in all three PT segments, but key fructose transporters SGLT5 and GLUT5 progressively increased from S1 to S3, and both were significantly upregulated in S3 vs. S1/S2 (Slc5a10: 1.9 log2FC, p < 1 × 10-299; Scl2a5: 1.4 log2FC, p < 4 × 10-105). A similar distribution was found in human kidneys. These data suggest that S3 is the primary site of fructose reabsorption in both humans and rats. Finally, because of the multiple scRNAseq transcriptional phenotypes found in each segment, our findings also imply that anatomical labels applied to scRNAseq clusters may be misleading.

HIV-Associated Microbial Translocation May Affect Cytokine Production of CD56bright NK Cells via Stimulation of Monocytes.

The mechanisms involved in HIV-associated natural killer (NK) cell impairment are still incompletely understood. We observed HIV infection to be associated with increased plasma levels of IFABP, a marker for gut epithelial barrier dysfunction, and LBP, a marker for microbial translocation. Both IFABP and LBP plasma concentrations were inversely correlated with NK cell interferon-γ production, suggesting microbial translocation to modulate NK cell functions. Accordingly, we found lipopolysaccharide to have an indirect inhibitory effect on NK cells via triggering monocytes' transforming growth factor-ß production. Taken together, our data suggest increased microbial translocation to be involved in HIV-associated NK cell dysfunction.

Natural Killer Cell-Mediated Antibody-Dependent Cellular Cytotoxicity Against SARS-CoV-2 After Natural Infection Is More Potent Than After Vaccination.

We compared the ability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike-specific antibodies to induce natural killer cell-mediated antibody-dependent cellular cytotoxicity (ADCC) in patients with natural infection and vaccinated persons. Analyzing plasma samples from 39 coronavirus disease 2019 (COVID-19) patients and 11 vaccinated individuals, significant induction of ADCC could be observed over a period of more than 3 months in both vaccinated and recovered individuals. Although plasma antibody concentrations were lower in recovered patients, we found antibodies elicited by natural infection induced a significantly stronger ADCC response compared to those induced by vaccination, which may affect protection conferred by vaccination.

Mitochondrial Dysfunction Contributes to Impaired Cytokine Production of CD56bright Natural Killer Cells From Human Immunodeficiency Virus-Infected Individuals Under Effective Antiretroviral Therapy.

Human immunodeficiency virus (HIV) infection is associated with impaired natural killer (NK) cell activity, which is only incompletely restored under antiretroviral therapy. Analyzing the bioenergetics profiles of oxygen consumption, we observed that several parameters were significantly reduced in HIV+ NK cells, indicating a mitochondrial defect. Accordingly, we found HIV+ CD56bright NK cells to display a decreased mitochondrial membrane potential and mitochondrial mass. Both parameters were positively correlated with interferon gamma (IFN-γ) production of NK cells. Finally, we demonstrated that stimulation of HIV+ NK cells with MitoTEMPO, a mitochondria-targeting antioxidant, significantly improved IFN-γ production. We identified mitochondrial dysfunction as a mechanism that contributes to impaired NK cell function.

Vascular hyperacetylation is associated with vascular smooth muscle dysfunction in a rat model of non-obese type 2 diabetes.

BACKGROUND: Advanced type 2 diabetes mellitus (T2DM) accelerates vascular smooth muscle cell (VSMC) dysfunction which contributes to the development of vasculopathy, associated with the highest degree of morbidity of T2DM. Lysine acetylation, a post-translational modification (PTM), has been associated with metabolic diseases and its complications. Whether levels of global lysine acetylation are altered in vasculature from advanced T2DM remains undetermined. We hypothesized that VSMC undergoes dysregulation in advanced T2DM which is associated with vascular hyperacetylation. METHODS: Aged male Goto Kakizaki (GK) rats, a non-obese murine model of T2DM, and age-matched male Wistar rats (control group) were used in this study. Thoracic aortas were isolated and examined for measurement of global levels of lysine acetylation, and vascular reactivity studies were conducted using a wire myograph. Direct arterial blood pressure was assessed by carotid catheterization. Cultured human VSMCs were used to investigate whether lysine acetylation participates in high glucose-induced reactive oxygen species (ROS), a crucial factor triggering diabetic vascular dysfunction. RESULTS: The GK rats exhibited marked glucose intolerance as well as insulin resistance. Cardiovascular complications in GK rats were confirmed by elevated arterial blood pressure and reduced VSMC-dependent vasorelaxation. These complications were correlated with high levels of vascular global lysine acetylation. Human VSMC cultures incubated under high glucose conditions displayed elevated ROS levels and increased global lysine acetylation. Inhibition of hyperacetylation by garcinol, a lysine acetyltransferase and p300/CBP association factor (PCAF) inhibitor, reduced high glucose-induced ROS production in VSMC. CONCLUSION: This study provides evidence that vascular hyperacetylation is associated with VSMC dysfunction in advanced T2DM. Understanding lysine acetylation regulation in blood vessels from diabetics may provide insight into the mechanisms of diabetic vascular dysfunction, and opportunities for novel therapeutic approaches to treat diabetic vascular complications.

A Multicenter Analysis of the Early Impact of COVID-19 on Junior Resident Operative Case Volume.

INTRODUCTION: Institutions have reported decreases in operative volume due to COVID-19. Junior residents have fewer opportunities for operative experience and COVID-19 further jeopardizes their operative exposure. This study quantifies the impact of the COVID-19 pandemic on resident operative exposure using resident case logs focusing on junior residents and categorizes the response of surgical residency programs to the COVID-19 pandemic. MATERIALS AND METHODS: A retrospective multicenter cohort study was conducted; 276,481 case logs were collected from 407 general surgery residents of 18 participating institutions, spanning 2016-2020. Characteristics of each institution and program changes in response to COVID-19 were collected via surveys. RESULTS: Senior residents performed 117 more cases than junior residents each year (P < 0.001). Prior to the pandemic, senior resident case volume increased each year (38 per year, 95% confidence interval 2.9-74.9) while junior resident case volume remained stagnant (95% confidence interval 13.7-22.0). Early in the COVID-19 pandemic, junior residents reported on average 11% fewer cases when compared to the three prior academic years (P = 0.001). The largest decreases in cases were those with higher resident autonomy (Surgeon Jr, P = 0.03). The greatest impact of COVID-19 on junior resident case volume was in community-based medical centers (246 prepandemic versus 216 during pandemic, P = 0.009) and institutions which reached Stage 3 Program Pandemic Status (P = 0.01). CONCLUSIONS: Residents reported a significant decrease in operative volume during the 2019 academic year, disproportionately impacting junior residents. The long-term consequences of COVID-19 on junior surgical trainee competence and ability to reach cases requirements are yet unknown but are unlikely to be negligible.

Genetic Variation of SAMM50 Is Not an Independent Risk Factor for Alcoholic Hepatocellular Carcinoma in Caucasian Patients.

Hepatocellular carcinoma (HCC) is a severe complication of advanced alcoholic liver disease, which is modulated by genetic predisposition. Identifying new genetic loci might improve screening. Genetic variation of SAMM50 was linked to HCC. We aimed to validate this finding in a large cohort of patients with advanced alcoholic liver disease (ALD). A large, well-characterised cohort of patients with alcoholic cirrhosis without (n = 674) and with (n = 386) HCC, as well as controls with HCC due to viral hepatitis (n = 134), controls with heavy alcohol abuse without liver disease (n = 266) and healthy subjects (n = 237), were genotyped for SAMM50 rs3827385 and rs3761472 and for PNPLA3 rs738409. Genotype frequencies were compared between patients with alcohol-associated cirrhosis with and without HCC by uni- and multivariate analysis. Minor variants in both SAMM50 rs3827385 and rs3761472 were significantly more frequent in patients with alcoholic HCC versus alcoholic cirrhosis and versus the control cohorts. An even stronger association was noted for PNPLA3 rs738409. The univariate analysis resulted in an odds ratio (OR) of 1.8 for carriers of at least one minor variant of SAMM50 rs3827385 and rs3761472 (each p < 0.001), but this association was lost in multivariate analysis with age (OR 1.1/year), male sex (OR 3.2), diabetes (OR 1.9) and carriage of PNPLA3 148M (OR 2.1) remaining in the final model. Although minor variants of both SAMM50 loci are strongly associated with alcoholic HCC, this association is not independent of carriage of the well-known risk variant PNPLA3 148M.

A genetic variant in toll-like receptor 5 is linked to chemokine levels and hepatocellular carcinoma in steatohepatitis.

BACKGROUND & AIMS: Bacterial translocation drives liver disease progression. We investigated whether functional genetic variants in toll-like receptor 5 (TLR5), the receptor for bacterial flagellin, affect the risk for hepatocellular carcinoma (HCC). METHODS: Healthy controls (n = 212), patients with alcohol abuse without liver disease (n = 382), and patients from a discovery cohort of alcohol-associated cirrhosis (n = 372 including 79 HCC cases), a validation cohort of alcohol-associated cirrhosis (n = 355 including 132 HCC cases), and a cohort of cirrhosis due to nonalcoholic steatohepatitis (NASH) (n = 145 including 62 HCC cases) were genotyped for the TLR5 rs5744174 and rs5744168 polymorphisms. Chemokine levels were measured by ELISA in patients' sera and supernatants of flagellin-stimulated healthy monocytes. RESULTS: Frequency of the TLR5 rs5744174 TT genotype was similar in healthy controls (33%), controls with alcohol abuse (34%), and patients with alcohol-associated cirrhosis in the discovery (28%), validation (33%), and NASH cohort (31%). The TT genotype was enriched in patients with versus without HCC in the discovery, validation, and NASH cohort (41% vs 25%; 39% vs 29%; 40% vs 24%; p < .05 each). This genotype remained a risk factor for HCC (OR = 1.9; p = .01) after multivariate correction for age, gender, diabetes, and carriage of the PNPLA3 148M variant. Interleukin-8 induction in monocytes from healthy controls and serum levels of interleukin-8 and CXCL1 from cirrhotic patients with the TT genotype were significantly increased versus C allele carriers. CONCLUSION: The TLR5 rs5744174 polymorphism, affecting immune response to flagellin, is linked to occurrence of HCC in cirrhosis caused by steatohepatitis.

Role of regulatory T cells and checkpoint inhibition in hepatocellular carcinoma.

Immune checkpoint inhibition suggests promising progress for the treatment of advanced hepatocellular carcinoma (HCC). However, the underlying cellular mechanisms remain unclear because liver cancer cells apparently do not upregulate inhibitory checkpoint molecules. Here, we analysed whether regulatory T cells (Tregs) can alternatively trigger checkpoint inhibition pathways in HCC. Using flow cytometry we analysed expression of checkpoint molecules (PD-1, PD-L1, CTLA-4, GITR, Tim-3) on peripheral CD4+CD25+Foxp3+ Tregs and their secretion of inhibitory mediators (IL-10, IL-35, TGF-beta, galectin-9) in 116 individuals (50 patients with HCC, 41 non-tumour bearing liver disease controls, 25 healthy controls). Functional activity of Tregs on T effector cells (IFN-gamma production, cytotoxicity) was characterized in vitro using a lectin-dependent cellular cytotoxicity (LDCC) assay against checkpoint inhibitor-negative P815 target cells. Unlike liver patients without malignancy and healthy controls, the frequency of checkpoint inhibitor-positive Tregs inversely correlated to age of patients with HCC (PD-L1, p = 0.0080; CTLA-4, p = 0.0029) and corresponded to enhanced numbers of Tregs producing IL-10 and IL-35 (p < 0.05 each). Tregs inhibited IFN-gamma secretion and cytotoxicity of CD8+ T cells when added to LDCC against P815 cells. Treg-induced inhibition of IFN-gamma secretion could be partially blocked by neutralizing PD-1 and PD-L1 antibodies specifically in HCC patients. In HCC peripheral Tregs upregulate checkpoint inhibitors and contribute to systemic immune dysfunction and antitumoural activity by several inhibitory pathways, presumably facilitating tumour development at young age. Blocking PD-L1/PD-1 interactions in vitro selectively interfered with inhibitory Treg -T effector cell interactions in the patients with HCC and resulted in improved antitumoural activity also against checkpoint inhibitor-negative tumour cells.

Compartment-specific distribution of human intestinal innate lymphoid cells is altered in HIV patients under effective therapy.

Innate lymphocyte cells (ILCs), a novel family of innate immune cells are considered to function as key orchestrators of immune defences at mucosal surfaces and to be crucial for maintaining an intact intestinal barrier. Accordingly, first data suggest depletion of ILCs to be involved in human immunodeficiency virus (HIV)-associated damage of the intestinal mucosa and subsequent microbial translocation. However, although ILCs are preferentially localized at mucosal surfaces, only little is known regarding distribution and function of ILCs in the human gastrointestinal tract. Here, we show that in HIV(-) individuals composition and functional capacity of intestinal ILCs is compartment-specific with group 1 ILCs representing the major fraction in the upper gastrointestinal (GI) tract, whereas ILC3 are the predominant population in ileum and colon, respectively. In addition, we present first data indicating that local cytokine concentrations, especially that of IL-7, might modulate composition of gut ILCs. Distribution of intestinal ILCs was significantly altered in HIV patients, who displayed decreased frequency of total ILCs in ileum and colon owing to reduced numbers of both CD127(+)ILC1 and ILC3. Of note, frequency of colonic ILC3 was inversely correlated with serum levels of I-FABP and sCD14, surrogate markers for loss of gut barrier integrity and microbial translocation, respectively. Both expression of the IL-7 receptor CD127 on ILCs as well as mucosal IL-7 mRNA levels were decreased in HIV(+) patients, especially in those parts of the GI tract with reduced ILC frequencies, suggesting that impaired IL-7 responses of ILCs might contribute to incomplete reconstitution of ILCs under effective anti-retroviral therapy. This is the first report comparing distribution and function of ILCs along the intestinal mucosa of the entire human gastrointestinal tract in HIV(+) and HIV(-) individuals.

Uptake and localization of fluorescently-labeled Karenia brevis metabolites in non-toxic marine microbial taxa.

Brevetoxin (PbTx) is a neurotoxic secondary metabolite of the dinoflagellate Karenia brevis. We used a novel, fluorescent BODIPY-labeled conjugate of brevetoxin congener PbTx-2 (B-PbTx) to track absorption of the metabolite into a variety of marine microbes. The labeled toxin was taken up and brightly fluoresced in lipid-rich regions of several marine microbes including diatoms and coccolithophores. The microzooplankton (20-200 µm) tintinnid ciliate Favella sp. and the rotifer Brachionus rotundiformis also took up B-PbTx. Uptake and intracellular fluorescence of B-PbTx was weak or undetectable in phytoplankton species representative of dinoflagellates, cryptophytes, and cyanobacteria over the same (4 h) time course. The cellular fate of two additional BODIPY-conjugated K. brevis associated secondary metabolites, brevenal (B-Bn) and brevisin (B-Bs), were examined in all the species tested. All taxa exhibited minimal or undetectable fluorescence when exposed to the former conjugate, while most brightly fluoresced when treated with the latter. This is the first study to observe the uptake of fluorescently-tagged brevetoxin conjugates in non-toxic phytoplankton and zooplankton taxa, demonstrating their potential in investigating whether marine microbes can serve as a significant biological sink for algal toxins. The highly variable uptake of B-PbTx observed among taxa suggests some may play a more significant role than others in vectoring lipophilic toxins in the marine environment.

Western diet triggers Toll-like receptor 4 signaling-induced endothelial dysfunction in female Wistar rats.

Overconsumption of a diet rich in fat and carbohydrates, called the Western diet, is a major contributor to the global epidemic of cardiovascular disease. Despite previously documented cardiovascular protection exhibited in female rats, this safeguard may be lost under certain metabolic stressors. We hypothesized that female Wistar rats challenged by a Western diet composed of 21% fat and 50% carbohydrate (34.1% sucrose) for 17 wk would develop endothelial dysfunction via endothelial Toll-like receptor 4 (TLR4) signaling. Western diet-fed female rats exhibited dysregulation of metabolism, revealing increased body weight and abdominal fat, decreased expression of adiponectin in white adipose tissue, glucose intolerance, and impaired insulin sensitivity. Western diet exposure increased hepatic triglycerides and cholesterol alongside hepatic steatosis, categorizing nonalcoholic fatty liver disease. Moreover, a Western diet negatively affected vascular function, revealing hypertension, impaired endothelium-dependent vasorelaxation, aortic remodeling, and increased reactive oxygen species (ROS) production. Aortic protein expression of TLR4 and its downstream proteins were markedly increased in the Western diet-fed group in association with elevated serum levels of free fatty acids. In vitro experiments were conducted to test whether free fatty acids contribute to vascular ROS overproduction via the TLR4 signaling pathway. Cultured endothelial cells were stimulated with palmitate in the presence of TAK-242, a TLR4 signaling inhibitor. Palmitate-induced overgeneration of ROS in endothelial cells was abolished in the presence of TAK-242. Our data show that a Western diet induced endothelial dysfunction in female rats and suggest that endothelial TLR4 signaling may play a key role in abolishing female cardiovascular protection. NEW & NOTEWORTHY A Western diet induced elevated levels of free fatty acids, produced nonalcoholic fatty liver disease, and provoked endothelial dysfunction in female rats in association with Toll-like receptor 4 signaling-mediated vascular reactive oxygen species production. Limited consumption of a Western diet in premenopausal women may decrease their risk of cardiovascular complications.

Inspiratory muscle training does not improve clinical outcomes in 3-week COPD rehabilitation: results from a randomised controlled trial.

The value of inspiratory muscle training (IMT) in pulmonary rehabilitation in chronic obstructive pulmonary disease (COPD) is unclear. The RIMTCORE (Routine Inspiratory Muscle Training within COPD Rehabilitation) randomised controlled trial examined the effectiveness of IMT added to pulmonary rehabilitation.In total, 611 COPD patients (Global Initiative for Chronic Obstructive Lung Disease stage II-IV) received a 3-week inpatient pulmonary rehabilitation, of which 602 patients were included in the intention-to-treat analyses. The intervention group (n=300) received highly intensive IMT and the control group (n=302) received sham IMT. The primary outcome was maximal inspiratory pressure (PImax). The secondary outcomes were 6-min walk distance, dyspnoea, quality of life and lung function. Outcomes were assessed pre- and post-pulmonary rehabilitation. ANCOVA was used.The intervention group showed higher effects in PImax (p<0.001) and forced inspiratory volume in 1 s (p=0.013). All other outcomes in both study groups improved significantly, but without further between-group differences. Sex and pulmonary rehabilitation admission shortly after hospitalisation modified quality of life effects.IMT as an add-on to a 3-week pulmonary rehabilitation improves inspiratory muscle strength, but does not provide additional benefits in terms of exercise capacity, quality of life or dyspnoea. A general recommendation for COPD patients to add IMT to a 3-week pulmonary rehabilitation cannot be made.