Worsened Disease Outcome Research Articles

Effects of inflammatory stimuli on the development of Mycoplasma bovis pneumonia in experimentally challenged calves

Many cattle infected with Mycoplasma bovis remain healthy while others develop severe chronic respiratory disease. We hypothesized that inflammatory stimuli such as co-pathogens worsen disease outcomes in M. bovis-infected calves. Calves (n=24) were intrabronchially inoculated with M. bovis and either killed bacterial lysate, transient M. haemolytica infection, or saline. Caseonecrotic lesions developed in 7/7 animals given M. haemolytica and M. bovis compared to 2/8 given M. bovis with no inflammatory stimulus, and 6/9 animals given bacterial lysate and M. bovis (P=0.01). Animals receiving M. haemolytica and M. bovis had more caseonecrotic foci in lungs than those receiving M. bovis with no inflammatory stimulus (median = 21 vs 0; P = 0.01), with an intermediate response (median = 5) in animals given bacterial lysate. In addition to caseonecrotic foci, infected animals developed neutrophilic bronchiolitis that appeared to develop into caseonecrotic foci, peribronchiolar lymphocytic cuffs that were not associated with the other lesions, and 4 animals with bronchiolitis obliterans. The data showed that transient lung inflammation at the time of M. bovis infection provoked the development of caseonecrotic bronchopneumonia, and the severity of inflammation influenced the number of caseonecrotic foci that developed. In contrast, caseonecrotic lesions were few or absent in M. bovis-infected calves without a concurrent inflammatory stimulus. These studies provide insight into how caseonecrotic lesions develop within the lung of M. bovis-infected calves. This and other studies suggest that controlling co-pathogens and harmful inflammatory responses in animals infected with M. bovis could potentially minimize development of M. bovis caseonecrotic bronchopneumonia.

Abstract PS04-09: Patient-Reported Benefit of Resources Designed to Assist with the Financial Toxicity of Breast Cancer

Abstract Background: A breast cancer (BC) diagnosis can cause financial stress, which may worsen disease outcomes and create financial instability. Tailored interventions are urgently needed, but the patient perspective on the utility of financial resource options is limited. Methods: From 6-7/2022, Breastcancer.org community members consented to an online survey in English or Spanish. Eligibility included:US resident, age ³ 18, and BC diagnosis within 10 years. Survey assessed burden of out-of-pocket expenses and perceived benefit of available (1 “Not at all” to 5 “Very much”) and potential (ranked choice) resources. Results: 1,437 participated. Mean age and time since diagnosis was 46 and 2 years, respectively; 75% were in active treatment for non-metastatic (89%) or metastatic (11%) disease. Patients were 60% White, 27% Hispanic, 8% Black, and 4% other race; 94% were women. 47% had either “significant” or “catastrophic” financial burden due to BC, with higher burdens in those with metastatic disease (61% vs 45%, p< 0.001). Direct assistance programs (e.g., reduced cost medications, grants, transportation) were rated the most helpful to respondents (Table). A grant list, a provider question checklist, and access to professional advice were most often ranked as 1st choice for potential resources. There were significant differences by race/ethnicity and disease status. Conclusions: Almost one-half of surveyed patients had significant financial burden due to BC. Patient-centered solutions to lessen financial burdens and optimize patient outcomes should consider direct assistance and patient education to improve knowledge, communication, and self-advocacy. How helpful were/are each of these resources in helping you cope with the financial burdens of BC? Overall Mean Disease Status Race/Ethnicity Non-metastatic Metastatic White Black Hispanic Other Patient assistance programs offered by pharma or medical test companies 2.9 2.9 3.1* 2.9* 2.6 3.1* 2.6 (n=1,107) (n=975) (n=132) (n=618) (n=94) (n=357) (n=38) Nonprofits that provide grants to help cover expenses of people with cancer 2.8 2.8 2.7 2.8 2.6 3.1* 2.4 (n=1,113) (n=991) (n=122) (n=611) (n=100) (n=358) (n=44) Financial assistance department at the cancer center or hospital 2.8 2.8 2.7 2.7* 2.7 3.1* 2.2 (n=1,176) (n=1,046) (n=130) (n=669) (n=103) (n=362) (n=42) Coupons and savings cards to reduce the cost of prescription drugs 2.8 2.8 2.7 2.7 2.5 3.0* 2.5 (n=1,140) (n=1,008) (n=132) (n=648) (n=92) (n=358) (n=42) Programs that provide free or low-cost transportation to medical appointments 2.7 2.8* 2.4 2.6 2.5 3.0* 2.3 (n=1,042) (n=934) (n=108) (n=556) (n=93) (n=356) (n=37) Clinical trials 2.7 2.7* 2.5 2.6* 2.3 3.0* 2.1 (n=1,032) (n=928) (n=104) (n=557) (n=84) (n=357) (n=34) Professional medical billing advocates 2.7 2.7* 2.3 2.5* 2.4 3.0* 2.1 (n=1,105) (n=994) (n=111) (n=612) (n=94) (n=358) (n=41) Social worker or case manager 2.7 2.7 2.6 2.6* 2.3 2.9* 2.4 (n=1,146) (n=1,027) (n=119) (n=644) (n=99) (n=360) (n=43) Nonprofits that provide advice and education about financial issues 2.7 2.7 2.5 2.6* 2.3 3.0* 2.0 (n=1,100) (n=983) (n=117) (n=607) (n=96) (n=359) (n=38) Financial navigator 2.6 2.6 2.5 2.5 2.5 2.9* 2.1 (n=1,087) (n=976) (n=111) (n=598) (n=94) (n=356) (n=39) Which of the following programs and resources would have been (or would be) most useful to you in navigating the financial burdens of BC? Total Non-metastatic Metastatic White Black Hispanic Other (n=1,282) (n=155) (n=686) (n=121) (n=384) (n=64) A list of financial grants people diagnosed with BC can apply for 18% 18% 19% 19%* 24%* 14% 25%* A checklist of questions to ask your healthcare team to help you minimize the financial burdens of BC 15% 16% 12% 16% 17% 15% 9% Access to free or reduced-price professionals to help with medical billing, financial, benefits, and/or legal issues 13% 13% 8% 11% 8% 15%* 20%* * p< 0.001 Citation Format: Fumiko Chino, Sam Meske, Muath Giaddui, Donna-Marie Manasseh, Bridgette Thom, Marisa Weiss. Patient-Reported Benefit of Resources Designed to Assist with the Financial Toxicity of Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS04-09.

The identification of small molecule inhibitors with anthelmintic activities that target conserved proteins among ruminant gastrointestinal nematodes.

Gastrointestinal nematode (GIN) infections are a major concern for the ruminant industry worldwide and result in significant production losses. Naturally occurring polyparasitism and increasing drug resistance that potentiate disease outcomes are observed among the most prevalent GINs of veterinary importance. Within the five major taxonomic clades, clade Va represents a group of GINs that predominantly affect the abomasum or small intestine of ruminants. However, the development of effective broad-spectrum anthelmintics against ruminant clade Va GINs has been challenged by a lack of comprehensive druggable genome resources. Here, we first assembled draft genomes for three clade Va species (Cooperia oncophora, Trichostrongylus colubriformis, and Ostertagia ostertagi) and compared them with closely related ruminant GINs. Genome-wide phylogenetic reconstruction showed a relationship among ruminant GINs structured by taxonomic classification. Orthogroup (OG) inference and functional enrichment analyses identified 220 clade Va-specific and Va-conserved OGs, enriched for functions related to cell cycle and cellular senescence. Further transcriptomic analysis identified 61 taxonomically and functionally conserved clade Va OGs that may function as drug targets for new broad-spectrum anthelmintics. Chemogenomic screening identified 11 compounds targeting homologs of these OGs, thus having potential anthelmintic activity. In in vitro phenotypic assays, three kinase inhibitors (digitoxigenin, K-252a, and staurosporine) exhibited broad-spectrum anthelmintic activities against clade Va GINs by obstructing the motility of exsheathed L3 (xL3) or molting of xL3 to L4. These results demonstrate valuable applications of the new ruminant GIN genomes in gaining better insights into their life cycles and offer a contemporary approach to discovering the next generation of anthelmintics.IMPORTANCEGastrointestinal nematode (GIN) infections in ruminants are caused by parasites that inhibit normal function in the digestive tract of cattle, sheep, and goats, thereby causing morbidity and mortality. Coinfection and increasing drug resistance to current therapeutic agents will continue to worsen disease outcomes and impose significant production losses on domestic livestock producers worldwide. In combination with ongoing therapeutic efforts, advancing the discovery of new drugs with novel modes of action is critical for better controlling GIN infections. The significance of this study is in assembling and characterizing new GIN genomes of Cooperia oncophora, Ostertagia ostertagi, and Trichostrongylus colubriformis for facilitating a multi-omics approach to identify novel, biologically conserved drug targets for five major GINs of veterinary importance. With this information, we were then able to demonstrate the potential of commercially available compounds as new anthelmintics.

The antiseizure medication valproate increases hemichannel activity found in brain cells, which could worsen disease outcomes.

Glial cells play relevant roles in neuroinflammation caused by epilepsy. Elevated hemichannel (HC) activity formed by connexins (Cxs) or pannexin1 (Panx1) largely explains brain dysfunctions commonly caused by neuroinflammation. Glia express HCs formed by Cxs 43, 30, or 26, while glia and neurons both express HCs formed by Panx1. Cx43 HCs allow for the influx of Ca2+, which promotes glial reactivity, enabling the release of the gliotransmitters that contribute to neuronal over-stimulation. Valproate (VPA), an antiseizure medication, has pleiotropic actions on neuronal molecular targets, and their action on glial cell HCs remains elusive. We used HeLa cells transfected with Cx43, Cx30, Cx26, or Panx1 to determine the effect of VPA on HC activity in the brain. VPA slightly increased HC activity under basal conditions, but significantly enhanced it in cells pre-exposed to conditions that promoted HC activity. Furthermore, VPA increased ATP release through Cx43 HCs. The increased HC activity caused by VPA was resistant to washout, being consistent with in silico studies, which predicted the binding site for VPA and Cx43, as well as for Panx1 HCs on the intracellular side, suggesting that VPA first enters through HCs, after which their activity increases.

Liberation of host heme by Clostridioides difficile-mediated damage enhances Enterococcus faecalis fitness during infection.

Clostridioides difficile and Enterococcus faecalis are two pathogens of great public health importance. Both bacteria colonize the human gastrointestinal tract where they are known to interact in ways that worsen disease outcomes. We show that the damage associated with C. difficile infection (CDI) releases nutrients that benefit E. faecalis. One particular nutrient, heme, allows E. faecalis to use oxygen to generate energy and grow better in the gut. Understanding the mechanisms of these interspecies interactions could inform therapeutic strategies for CDI.

The Immunopathology of Covid-19

SARS-CoV-2 is a virus causing Covid-19, with a genome encoding proteins for replication. The innate immune system responds rapidly, employing pattern recognition receptors (PRRs) detecting viral presence. Mucosal tissues offer initial defense, with interferons inhibiting viral replication. Inflammatory cytokines and chemokines attract immune cells, sometimes resulting in cytokine storm. However, excessive inflammation may worsen disease outcomes. The adaptive immune system, comprising CD4+ T cells, CD8+ T cells, and B cells, organize targeted responses. CD4+ T cells activate immune cells and coordinate responses, while CD8+ T cells eliminate infected cells using cytotoxic molecules. B cells produce antibodies that neutralize the virus. Memory B and T cells provide long-term immunity, aiding rapid response upon reinfection. Recovered individuals often exhibit reduced reinfection risks due to memory immunity. Vaccines, like mRNA-based and viral vector-based types, stimulate similar immune responses, bolstering immunity and reducing severe illness.

Survival Advantage Associated with Heterozygous Factor V Leiden Mutation in Sars-Cov-2-Infected K18-hACE2 Mice

The pathogenesis of coronavirus disease 2019 (COVID-19) complications in critically ill patients involves virus-mediated cellular damage in infected tissue and secondary organ damage associated with a hyperinflammatory response. The proinflammatory milieu can result in endothelial dysfunction, predisposing patients to thrombosis leading to a life-threatening condition known as COVID-19-associated coagulopathy (CAC) which is driven by cellular and molecular mechanisms that are still unclear. We developed a mouse model based on the K18-hACE2 transgenic mouse recapitulating many of the CAC-associated pathological findings observed in human patients offering a reliable animal model for the study of SARS-CoV-2 pathogenesis. Plasma analysis revealed virus-induced upregulation of endothelial activation markers and alterations of all the measured coagulation factors' activities, resulting first in increased vascular leakage in the lung on day 5 post-infection, followed by the appearance of focal hemorrhagic lesions potentially associated with local thrombotic events on day 7. The infection-associated procoagulant phenotype appeared as early as day 3 post-infection (p.i.). Prothrombin time showed a 1/3 reduction in the normal clotting time in males. The overall procoagulant state was also reflected in the alteration of Factor(F) VIII and FX activity increasing to ~150% compared to uninfected controls. FIX showed the most significant alteration and was ~200% of the normal activity on day 3 p.i. and gradually increased over time, reaching ~350%. We also observed an increase in the activity of FV starting 3 days p.i. at ~210% of the normal level and persisting unchanged up to day 7 p.i. As reported during human infections, FV activity increased without significant changes in protein concentration or FVa levels. Natural anticoagulant and fibrinolytic mechanisms were dysregulated during virus infection resulting in a profound imbalance in thrombus formation and resolution. Infected K18-hACE2 mice showed downregulation of plasma protein C concentration which was not associated with any alteration of liver function, while plasma protein S concentration remained unaltered. Given the procoagulant environment generated after SARS-CoV-2 infection, reducing activated protein C (APC) anticoagulant and cytoprotective activities by blocking APC's active site with the SPC-54 monoclonal antibody worsened disease outcome and brought the mortality rate from 50% to 90%. In this observed procoagulant pathological context, where we observe increased activity of FV and where a deficiency of APC functions resulted in increased mortality, it wouldn't be surprising to observe an even more severe phenotype with the addition of APC resistance. To test this idea, we generated K18-hACE2 mice harboring the homologous human FV Leiden (FV L) mutation (R504Q). The Leiden mutation in FV eliminates one of several sites in FVa that are substrates for proteolysis by APC, resulting in diminished inactivation of FVa by APC and preventing the formation of the anticoagulant form of FV that serves as a cofactor for protein S-dependent inactivation of FVIIIa by APC. Upon SARS-CoV-2 infection, the presence of the homozygous FV L mutation in K18-hACE2 mice didn't affect the severity of the disease as measured by body weight loss during the course of the infection, and homozygous mutated mice had the same mortality rate as FV WT mice. Surprisingly, K18-hACE2 male mice carrying a heterozygous FV L mutation were extremely protected from SARS-CoV-2 infection, as their mortality dropped from 50% to only 15%. Histological analysis reveals how heterozygous FV L mice displayed a great reduction in immune infiltrate-associated fibrin deposition, suggesting an unexpected protective role of this prothrombotic mutation in the development of CAC. The protective effect of FV L heterozygosity may be attributable to enhanced thrombin formation which boosts inflammation and enhanced APC generation controlling proinflammatory and prothrombotic signaling, thus showing a delicate balance achieved by FV L heterozygosity that was lost for FV L homozygosity. As seen also in mouse models of bacterial sepsis, such favorable effects associated with FV L support the somewhat counterintuitive speculation that a partial increase in thrombin production in the early phase of the infection may support the resolution of COVID-19 complications.

CYP19A1 mediates severe SARS-CoV-2 disease outcome in males

Male sex represents one of the major risk factors for severe COVID-19 outcome. However, underlying mechanisms that mediate sex-dependent disease outcome are as yet unknown. Here, we identify the CYP19A1 gene encoding for the testosterone-to-estradiol metabolizing enzyme CYP19A1 (also known as aromatase) as a host factor that contributes to worsened disease outcome in SARS-CoV-2-infected males. We analyzed exome sequencing data obtained from a human COVID-19 cohort (n= 2,866) using a machine-learning approach and identify a CYP19A1-activity-increasing mutation to be associated with the development of severe disease in men but not women. We further analyzed human autopsy-derived lungs (n= 86) and detect increased pulmonary CYP19A1 expression at the time point of death in men compared with women. In the golden hamster model, we show that SARS-CoV-2 infection causes increased CYP19A1 expression in the lung that is associated with dysregulated plasma sex hormone levels and reduced long-term pulmonary function in males but not females. Treatment of SARS-CoV-2-infected hamsters with a clinically approved CYP19A1 inhibitor (letrozole) improves impaired lung function and supports recovery of imbalanced sex hormones specifically in males. Our study identifies CYP19A1 as a contributor to sex-specific SARS-CoV-2 disease outcome in males. Furthermore, inhibition of CYP19A1 by the clinically approved drug letrozole may furnish a new therapeutic strategy for individualized patient management and treatment.

Potent broadly neutralizing antibody VIR-3434 controls hepatitis B and D virus infection and reduces HBsAg in humanized mice

Chronic hepatitis B is a global public health problem, and coinfection with hepatitis delta virus (HDV) worsens disease outcome. Here, we describe a hepatitis B virus (HBV) surface antigen (HBsAg)-targeting monoclonal antibody (mAb) with the potential to treat chronic hepatitis B and chronic hepatitis D. HBsAg-specific mAbs were isolated from memory B cells of HBV vaccinated individuals. Invitro neutralization was determined against HBV and HDV enveloped with HBsAg representing eight HBV genotypes. Human liver-chimeric mice were treated twice weekly with a candidate mAb starting 3 weeks post HBV inoculation (spreading phase) or during stable HBV or HBV/HDV coinfection (chronic phase). From a panel of human anti-HBs mAbs, VIR-3434 was selected and engineered for pre-clinical development. VIR-3434 targets a conserved, conformational epitope within the antigenic loop of HBsAg and neutralized HBV and HDV infection with higher potency than hepatitis B immunoglobulins invitro. Neutralization was pan-genotypic against strains representative of HBV genotypes A-H. In the spreading phase of HBV infection in human liver-chimeric mice, a parental mAb of VIR-3434 (HBC34) prevented HBV dissemination and the increase in intrahepatic HBV RNA and covalently closed circular DNA. In the chronic phase of HBV infection or co-infection with HDV, HBC34 treatment decreased circulating HBsAg by >1 log and HDV RNA by >2 logs. The potently neutralizing anti-HBs mAb VIR-3434 reduces circulating HBsAg and HBV/HDV viremia in human liver-chimeric mice. VIR-3434 is currently in clinical development for treatment of patients with chronic hepatitis B or D. Chronic infection with hepatitis B virus and co-infection with hepatitis D virus place approximately 290 million individuals worldwide at risk of severe liver disease and cancer. Available treatments result in low rates of functional cure or require lifelong therapy that does not eliminate the risk of liver disease. We isolated and characterized a potent human antibody that neutralizes hepatitis B and D viruses and reduces infection in a mouse model. This antibody could provide a new treatment for patients with chronic hepatitis B and D.

COLITIS-ASSOCIATED COLON CANCER: EXPLORING THE DCLK1-S-MMP13-P53 AXIS

Chronic intestinal inflammation in patients with inflammatory bowel diseases (IBD) significantly increases the risk of colorectal cancer (CRC). Tuft cells in the intestine marked by doublecortin-like kinase-1 (DCLK1), are long-lived chemosensory cells with newly identified roles in colitis as epithelial ablation of DCLK1 worsens disease outcome. DCLK1 exists in long and short isoforms (DCLK1-L and DCLK1-S). Hypermethylation of DCLK1 alpha promoter encoding DCLK1-L particularly in CRC, allows switching to the DCLK1-S isoform that confers a more invasive tumor phenotype. In an infectious colitis model, we recently discovered that DCLK1-S is predominantly expressed in Ly6G+;MHCII- neutrophils that coincides with elevated levels of CXCL1/KC and MMP13 expression in the colon. In an unbiased molecular docking study, DCLK1’s kinase domain was found to interact with the catalytic domain of MMP13. MMP13 plays a crucial role in the modulation of extracellular matrix (ECM) degradation and cell-matrix interactions and wild type and mutant p53 differentially regulate MMP13 gene expression. Mutations in TP53 (tumor suppressor p53) are crucial for neoplasia initiation in IBD patients and are observed in 50-85% of colitis-associated colon cancers (CACCs). We further observed that p53 mutation status dictates DCLK1-S expression in colon cancer cells and in mice mutant for p53, expression of DCLK1-S and MMP13 correlated with neutrophilic inflammation and crypt hyperplasia. We will expand on these novel discoveries by employing high fidelity proteomic and phosphoproteomic studies and decipher how DCLK1-S-MMP13-p53 axes stimulate fibrosis, tumorigenesis, and tumor progression. These studies are expected to lead to therapeutic avenues to target CACCs.

COLITIS-ASSOCIATED COLON CANCER: EXPLORING THE DCLK1-S-MMP13-P53 AXIS

Abstract Chronic intestinal inflammation in patients with inflammatory bowel diseases (IBD) significantly increases the risk of colorectal cancer (CRC). Tuft cells in the intestine marked by doublecortin-like kinase-1 (DCLK1), are long-lived chemosensory cells with newly identified roles in colitis as epithelial ablation of DCLK1 worsens disease outcome. DCLK1 exists in long and short isoforms (DCLK1-L and DCLK1-S). Hypermethylation of DCLK1 alpha promoter encoding DCLK1-L particularly in CRC, allows switching to the DCLK1-S isoform that confers a more invasive tumor phenotype. In an infectious colitis model, we recently discovered that DCLK1-S is predominantly expressed in Ly6G+;MHCII- neutrophils that coincides with elevated levels of CXCL1/KC and MMP13 expression in the colon. In an unbiased molecular docking study, DCLK1’s kinase domain was found to interact with the catalytic domain of MMP13. MMP13 plays a crucial role in the modulation of extracellular matrix (ECM) degradation and cell-matrix interactions and wild type and mutant p53 differentially regulate MMP13 gene expression. Mutations in TP53 (tumor suppressor p53) are crucial for neoplasia initiation in IBD patients and are observed in 50-85% of colitis-associated colon cancers (CACCs). We further observed that p53 mutation status dictates DCLK1-S expression in colon cancer cells and in mice mutant for p53, expression of DCLK1-S and MMP13 correlated with neutrophilic inflammation and crypt hyperplasia. We will expand on these novel discoveries by employing high fidelity proteomic and phosphoproteomic studies and decipher how DCLK1-S-MMP13-p53 axes stimulate fibrosis, tumorigenesis, and tumor progression. These studies are expected to lead to therapeutic avenues to target CACCs.

Effect of Comorbid Diabetes on Clinical Characteristics of COVID-19 Patients Infected by the Wild-Type or Delta Variant of SARS-CoV-2.

BackgroundDiabetes is one of the most common comorbidities in COVID-19 patients that pertains to disease severity, but the causal mechanism regarding its negative impact on COVID-19 outcome has yet been uncovered.MethodsWe retrospectively analyzed 459 COVID-19 patients admitted in early 2020 and 336 COVID-19 patients admitted in August 2021, with their demographic information, medical history, vaccination status (if applied), and laboratory data reported.ResultsAmong COVID-19 patients, compared to the non-diabetic group, the diabetic group exhibited elder age, higher proportion of patients with other major comorbidities, more severe dysfunction of innate immune cells, more refractory blood coagulopathy and more detrimental organ damage. For the wild-type SARS-CoV-2 infection, diabetic comorbidity was associated with COVID-19 severity but not mortality, and the glycemic levels in the non-diabetic group upon infection experienced high and analogous to those in the diabetic group. Besides, infected by the delta variant of SARS-CoV-2, the non-diabetic patients did not demonstrate hyperglycemia, and despite different vaccination statuses, the diabetic patients exhibited comparable antibody responses to non-diabetic, showing the robustness of acquired immunity.ConclusionsSARS-CoV-2 infection may superimpose the deterioration of innate immune systems in diabetic patients, which contributes to their worsened disease outcome, but timely COVID-19 immunization could provide adequate protection in diabetic population that leads to favored prognosis.

A228 A FERMENTABLE FIBRE FREE DIET ACCELERATES SPONTANEOUS COLITIS DEVELOPMENT AND ALTERS THE GUT MICROBIOME IN MUC2-/- MICE

Abstract Background A key feature of ulcerative colitis (UC) is a defective colonic mucus layer. As Muc2-/- mice lack intestinal mucus, they’re often used to model UC. In these mice, luminal bacteria translocate from the gut lumen, triggering spontaneous colitis by 12–16 weeks of age. Notably, mucus defects are also seen in wildtype mice fed a fermentable fibre free (FF-free) diet. Aside from thinner mucus, FF-free fed mice show increased susceptibility to bacterial infections and a microbiome shift from fibre-utilizing to mucus-degrading bacteria. Indeed, a recent clinical study found that a microbiome shift towards proteolytic bacteria preceded clinical diagnosis of UC Aims To determine whether feeding newly weaned Muc2-/- mice a FF-free versus chow or inulin-enriched diet leads to - more rapid development of spontaneous colitis - shifts in the gut microbiota towards protein fermentation Methods Newly weaned Muc2-/- mice (3–4 weeks old) were fed 1 of 3 diets: 1) usual chow (Chow; 14.7% fibre mix), 2) FF-free (10% cellulose – poorly fermented) and 3) inulin-enriched (IN; same nutritional content as FF-free, with 10% inulin instead of cellulose). Body weight, diet intake and disease activity were monitored daily. By day 10, mice in the FF-free group developed severe colitis requiring euthanization. At this time, cecal contents (short-chain fatty acid [SCFA] analysis) and colonic tissue from all mice were collected for alcian blue/PAS staining and histopathological scoring. Stool was collected for 16S rRNA sequencing Results FF-free fed mice consumed fewer calories (p=0.0073), experienced less body weight gain (p=0.002), had colonic shortening (p<0.0001) and higher disease activity scores (p<0.0001) than mice fed the IN diet. Histopathological scores were higher in FF-free versus Chow (p<0.0001) and IN (p<0.0001) fed mice. Goblet cell mucin staining was reduced in FF-free compared to Chow and IN fed mice. Beta-diversity analysis showed clear differences in microbiome composition between groups. Acetate, product of fibre fermentation, was higher in the Chow (p<0.0001) and IN (p=0.0195) groups whereas branched-chain fatty acids (BCFA), product of protein fermentation, was higher in the FF-free group (p<0.0001) Conclusions Spontaneous colitis development in FF-free fed Muc2-/- mice was observed within 10 days (5–6 weeks old) of weaning whereas Chow or IN fed mice showed minimal to no signs of colitis, respectively. A diet devoid of fibre also led to less mucin staining and a gut microbiome shift that favoured BCFA over SCFA production, suggesting the microbiota of FF-free fed Muc2-/- mice are targeting mucosal proteins concurrent with disease exacerbation. These findings support future studies examining whether consumption of low fibre diets in UC patients worsen disease outcomes whereas diets rich in fibre and/or inulin prove beneficial Funding Agencies CCC, CIHRMSFHR Trainee Award, CH.I.L.D foundation

COVID-19 triggers attacks in HAE patients without worsening disease outcome

COVID-19 triggers attacks in HAE patients without worsening disease outcome

Evaluation of a 5-HT2B receptor agonist in a murine model of amyotrophic lateral sclerosis

Degeneration of brainstem serotonin neurons has been demonstrated in ALS patients and mouse models and was found responsible for the development of spasticity. Consistent with involvement of central serotonin pathways, 5-HT2B receptor (5-HT2BR) was upregulated in microglia of ALS mice. Its deletion worsened disease outcome in the Sod1G86R mouse model and led to microglial degeneration. In ALS patients, a polymorphism in HTR2B gene leading to higher receptor expression in CNS, was associated with increased survival in patients as well as prevention of microglial degeneration. Thus, the aim of our study was to determine the effect of a 5-HT2BR agonist : BW723C86 (BW), in the Sod1G86R mouse model. Despite good pharmacokinetic and pharmacological profiles, BW did not ameliorate disease outcome or motor neuron degeneration in a fast progressing mouse model of ALS despite evidence of modulation of microglial gene expression.

Premature immunosenescence: role of stress-related factors

The ageing of the immune system, immunosenescence, is associated with important remodelling changes in all lymphoid organs and its cells. However, the pace of ageing differs between people with the same chronological age, indicating that it may be thus influenced by extrinsic factors, including the chronic stress. In fact, young adults under chronic psychosocial stress may present immunosenescence features similarly to those found in older adults. Immunosenescence contributes to the onset and clinical course of age-related diseases such as neurodegenerative disorders, cancer, rheumatoid arthritis, cardiovascular and metabolic diseases. This article reviews that premature immunosenescence is observed in adults under chronic psychological stress as well as in healthy adolescents with history of childhood maltreatment. Superimposing stress during ageing is associated with further decline in adaptive immunity, increased inflammaging, accelerated cellular senescence and increased susceptibility to infections. This is of great interest for the COVID-19 pandemic, making the chronically stressed older adults of particular risk for worsen disease outcomes and poor vaccine responses.

In vitro and in vivo efficacy of Metformin against dengue

Dengue is a prevalent mosquito-borne viral infection in the tropical and sub-tropical regions. Its potential to progress into severe, life-threatening disease, has pressed the research community to develop safe, effective and affordable antivirals. Metformin (MET), a first-line antidiabetic drug and indirect AMP-activated protein kinase (AMPK) activator, has recently emerged as a potential anti-DENV therapeutic candidate, based on some experimental evidence supporting anti-DENV activity in vitro and widely reported anti-inflammatory properties. Here, we examined MET in vitro activity against the four DENV serotypes and in two different mammalian cell lines. MET displayed a poor anti-DENV activity in BHK-21 cells with IC50 in the mM range, which was associated with increased p-AMPKα levels, thereby supporting that MET antiviral activity is mediated through AMPK activation. In contrast, MET exerted a pro-DENV activity in Vero cells that did not correlate with increased AMPK activation, suggesting AMPK-independent effects. Treatment with compound 991, a direct AMPK activator, led to reduced viral titers against all four serotypes and across both mammalian cell lines. In vivo, oral administration of MET did not reduce viremia titers in an asymptomatic mouse model, neither did it improve disease severity and progression in a mouse model of severe dengue. Instead, high dose regimen worsened disease outcome as evidenced by increased mortality, higher viremia and hyper-inflammation. Therefore, while AMPK may represent a potential host target, MET does not seem to hold great promise as a pan-serotype anti-dengue drug.

Effects of inflammatory stimuli on responses of macrophages to Mycoplasma bovis infection

Inflammation in the respiratory tract is thought to worsen the disease response to Mycoplasma bovis infection. This study investigated the cells involved in this response with a focus on proteases and cytokines as harmful effector mechanisms. By immunohistochemistry, Mac387-positive macrophages were the main cell type comprising the foci of caseous necrosis in cattle with M. bovis pneumonia. Thus, the study evaluated how priming of different types of macrophages with bacterial lysate (or pro-inflammatory cytokines induced by the bacterial lysate) affected their responses to M. bovis infection. Inducible responses were detected in monocyte-derived macrophages (M1-MDMs and M2-MDMs), whereas pulmonary alveolar macrophages (PAMs) were minimally affected by priming or infection. M. bovis-infected MDMs secreted MMP-12 and SPLA2, and priming with pro-inflammatory cytokines increased the secretion of cathepsin B in response to M. bovis infection. Of these, there were higher concentrations of cathepsin B and SPLA2 in lungs with M. bovis pneumonia compared to healthy lungs, and these are potential mechanisms for macrophage-induced lung damage in M. bovis infection. Priming of MDMs with either bacterial lysate or with pro-inflammatory cytokines caused an enhanced response to M. bovis infection with respect to IL-8 and IL-1β secretion. The findings of this study suggest proteases, lipases and cytokines derived from monocyte-derived macrophages as possible mediators by which prior inflammation in the respiratory tract worsen disease outcomes from M. bovis infection.

Coinfections with Other Respiratory Pathogens among Patients with COVID-19.

ABSTRACTEmerging evidence indicates that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals are at an increased risk for coinfections; therefore, physicians need to be cognizant about excluding other treatable respiratory pathogens. Here, we report coinfection with SARS-CoV-2 and other respiratory pathogens in patients admitted to the coronavirus disease (COVID) care facilities of an Indian tertiary care hospital. From June 2020 through January 2021, we tested 191 patients with SARS-CoV-2 for 33 other respiratory pathogens using an fast track diagnostics respiratory pathogen 33 (FTD-33) assay. Additionally, information regarding other relevant respiratory pathogens was collected by reviewing their laboratory data. Overall, 13 pathogens were identified among patients infected with SARS-CoV-2, and 46.6% (89/191) of patients had coinfection with one or more additional pathogens. Bacterial coinfections (41.4% [79/191]) were frequent, with Staphylococcus aureus being the most common, followed by Klebsiella pneumoniae. Coinfections with SARS-CoV-2 and Pneumocystis jirovecii or Legionella pneumophila were also identified. The viral coinfection rate was 7.3%, with human adenovirus and human rhinovirus being the most common. Five patients in our cohort had positive cultures for Acinetobacter baumannii and K. pneumoniae, and two patients had active Mycobacterium tuberculosis infection. In total, 47.1% (90/191) of patients with coinfections were identified. The higher proportion of patients with coinfections in our cohort supports the systemic use of antibiotics in patients with severe SARS-CoV-2 pneumonia with rapid de-escalation based on respiratory PCR/culture results. The timely and simultaneous identification of coinfections can contribute to improved health of COVID-19 patients and enhanced antibiotic stewardship during the pandemic.IMPORTANCE Coinfections in COVID-19 patients may worsen disease outcomes and need further investigation. We found that a higher proportion of patients with COVID-19 were coinfected with one or more additional pathogens. A better understanding of the prevalence of coinfection with other respiratory pathogens in COVID-19 patients and the profile of pathogens can contribute to effective patient management and antibiotic stewardship during the current pandemic.

Acceleration of TAA-Induced Liver Fibrosis by Stress Exposure Is Associated with Upregulation of Nerve Growth Factor and Glycopattern Deviations.

Liver fibrosis results from many chronic injuries and may often progress to cirrhosis and hepatocellular carcinoma (HCC). In fact, up to 90% of HCC arise in a cirrhotic liver. Conversely, stress is implicated in liver damage, worsening disease outcome. Hence, stress could play a role in disrupting liver homeostasis, a concept that has not been fully explored. Here, in a murine model of TAA-induced liver fibrosis we identified nerve growth factor (NGF) to be a crucial regulator of the stress-induced fibrogenesis signaling pathway as it activates its receptor p75 neurotrophin receptor (p75NTR), increasing liver damage. Additionally, blocking the NGF decreased liver fibrosis whereas treatment with recombinant NGF accelerated the fibrotic process to a similar extent than stress challenge. We further show that the fibrogenesis induced by stress is characterized by specific changes in the hepatoglycocode (increased β1,6GlcNAc-branched complex N-glycans and decreased core 1 O-glycans expression) which are also observed in patients with advanced fibrosis compared to patients with a low level of fibrosis. Our study facilitates an understanding of stress-induced liver injury and identify NGF signaling pathway in early stages of the disease, which contributes to the established fibrogenesis.