Cathelicidin (hCAP-18/LL-37) and beta-defensin 1 (HBD-1) are human antimicrobial peptides (AMPs) with high basal expression levels, which form the first line of host defence against infections over the epithelial surfaces. The antimicrobial functions owe to their direct microbicidal effects as well as the immunomodulatory role. Pathogenic microorganisms have developed multiple modalities including transcriptional repression to combat this arm of the host immune response. The precise mechanisms and the pathogen-derived molecules responsible for transcriptional downregulation remain unknown. Here, we have shown that enteric pathogens suppress LL-37 and HBD-1 expression in the intestinal epithelial cells (IECs) with Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) exerting the most dramatic effects. Cholera toxin (CT) and labile toxin (LT), the major virulence proteins of V. cholerae and ETEC, respectively, are predominantly responsible for these effects, both in vitro and in vivo. CT transcriptionally downregulates the AMPs by activating several intracellular signalling pathways involving protein kinase A (PKA), ERK MAPKinase and Cox-2 downstream of cAMP accumulation and inducible cAMP early repressor (ICER) may mediate this role of CT, at least in part. This is the first report to show transcriptional repression of the AMPs through the activation of cellular signal transduction pathways by well-known virulence proteins of pathogenic microorganisms.

Vero cell cytotoxins and cytotonic enterotoxins produced by E. coli are toxic proteins, which have been implicated in a number of specific diseases in humans and animals. Nomenclature for these toxins is complicated by the existence of different names for the same toxin. The Vero cell cytotoxins are called verotoxins because they are lethal for Vero cells in culture; they are also known as Shiga-like toxins (SLTs) because they are clearly related to Shiga toxin in structure, amino acid sequence, mechanism of action, and biological activity. SLTs belong to two classes. SLT-I is identical with Shiga toxin and is in a class by itself (class I). The other SLTs are closely related to each other and form a second class (class II). Class II SLTs include SLT-II, SLT-IIv, SLT-IIvha, SLT-IIvhb, and SLT-IIva. All SLTs that have been investigated are A-B subunit protein toxins, whose A subunits possess N-glycosidase activity against 28S rRNA and cause inhibition of protein synthesis in eukaryotic cells. These toxins are enterotoxic as well as cytotoxic. SLTs produced in the intestine are absorbed into the blood stream and affect vascular endothelial cells in target organs. They may also have a direct toxic effect on enterocytes. Diseases in which E. coli SLTs have been implicated include diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome in humans and edema disease in pigs. Variation in receptor specificities among SLTs may be the reason for different disease syndromes in different host species. The E. coli enterotoxins belong to three distinct classes: heat-labile enterotoxin (LT), heat-stable enterotoxin type I or type a (STI, STa), and heat-stable enterotoxin type II or type b (STII, STb). There is clear evidence that these cytotonic enterotoxins play an essential role in diarrheal disease. LT is an A-B subunit protein toxin, closely related to cholera toxin. Following binding of LT to receptors in enterocytes the A subunit is internalized. The enzymatically active A subunit transfers ADP-ribose from NAD to a GTP-dependent adenylate cyclase regulatory protein, thereby elevating intracellular levels of adenylate cyclase. The increased levels of cyclic AMP cause stimulation of A kinase and lead to hypersecretion of electrolytes and fluid. STI is a small peptide of 18 or 19 amino acids. It binds to receptors in enterocytes and stimulates particulate guanyl cyclase. Elevated intracellular cyclic GMP stimulates G kinase, resulting in increased Cl- secretion and impaired absorption of Na+Cl-. STII is a peptide toxin whose mechanism of action is unknown.(ABSTRACT TRUNCATED AT 400 WORDS)

There is increasing evidence that inflammation plays an important role in the development of cardiovascular complications in patients with obstructive sleep apnoea (OSA). No previous works have studied levels of soluble tumour necrosis factor-alpha receptor (sTNFR)-1 in patients with OSA. The aims of the present study were to examine serum levels of sTNFR-1 and the effect of nasal continuous positive airway pressure (CPAP) in patients with OSA. A prospective, randomised, placebo-controlled crossover study was performed. In total, 30 consecutive newly diagnosed OSA patients (apnoea/hypopnoea index 43.8+/-27.0 events x h(-1)) and 15 healthy obese patients were selected. Urinary levels of norepinephrine and epinephrine, as well as plasma sTNFR-1, tumour necrosis factor (TNF)-alpha, interleukin (IL)-6 and leukotriene (LT)B(4) levels were obtained at baseline and after 3 months of CPAP or sham CPAP. Nocturnal urinary levels of norepinephrine, epinephrine and sTNFR-1 (1,053+/-269 versus 820+/-166 pg x mL(-1)) were significantly higher in OSA patients. There were no significant differences in plasma levels of IL-6, LTB(4), or TNF-alpha between the two study groups. There were no significant differences in blood pressure, urinary catecholamine levels, or plasma IL-6, LTB(4) and TNF-alpha levels after both treatment modalities. However, after 3 months of effective CPAP usage, sTNFR-1 levels were significantly reduced (1,053+/-269 versus 899+/-254 pg x mL(-1)). Obstructive sleep apnoea patients have higher levels of soluble tumour necrosis factor-alpha receptor 1 than individuals without OSA; soluble tumour necrosis factor-alpha receptor 1 levels are lowered by continuous positive airway pressure therapy. These findings further corroborate a potential role of inflammation in the natural history of obstructive sleep apnoea.

<A<em>b</em>stractText>Inflammation indexes and <em>b</em>ody mass index (BMI) are easily evaluated, predict survival, and are potentially modifia<em>b</em>le. We evaluated the potential association of inflammatory indexes and BMI with the clinical outcome of patients with renal cell carcinoma (RCC) undergoing immune checkpoint inhi<em>b</em>itor therapy.</A<em>b</em>stractText><p><div>(<em>b</em>)EXPERIMENTAL DESIGN</<em>b</em>)</div>A prospective cohort of patients with metastatic RCC treated with nivoluma<em>b</em> enrolled in the Italian Expanded Access Program from July 2015 through April 2016 was examined. Reference measures of inflammation were identified for neutrophil-to-lymphocyte ratio (NLR) &<em>lt</em>;/≥ 3, systemic immune inflammation index (SII) &<em>lt</em>;/≥ 1,375, and platelet-to-lymphocyte ratio (PLR) &<em>lt</em>;/≥ 232. Patients were classified as high BMI (≥25 kg/m²) versus normal BMI (&<em>lt</em>;25 kg/m²).</p><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>Among 313 evalua<em>b</em>le patients, 235 (75.1%) were male, and median age was 65 years (range, 40-84 years), with 105 (33.69%) ≥70 years. In univariate analysis, age, performance status, BMI, SII, NLR, and PLR were a<em>b</em>le to predict outcome. In mu<em>lt</em>ivariate analyses, SII ≥1,375, BMI &<em>lt</em>;25 kg/m², and age ≥70 years independently predicted overall survival [OS; HR = 2.96, 95% confidence interval (CI), 2.05-4.27; HR = 1.59, 95% CI, 1.10-2.30; and HR = 1.65, 95% CI, 1.07-2.55, respectively). A patient with <em>b</em>oth SII ≥1,375 and BMI &<em>lt</em>;25 kg/m² was estimated to have much worse OS (HR, 3.37; 95% CI, 2.29-4.95; <i>P</i> &<em>lt</em>;0.0001) than a patient with neither or only one risk factor. SII changes at 3 months predicted OS (<i>P</i> &<em>lt</em>;0.0001).</p><A<em>b</em>stractText>Normal BMI com<em>b</em>ined with inflammation tripled the risk of death, suggesting that these <em>b</em>iomarkers are critical prognostic factors for OS in patients with RCC treated with nivoluma<em>b</em>.</A<em>b</em>stractText>

B lymphocytes are generally recognized for their potential to mediate humoral immunity by producing different antibody isotypes and being involved in opsonization and complement fixation. Nevertheless, the non-classical, antibody-independent immune potential of B cell subsets has attracted much attention especially in the past decade. These B cells can release a broad variety of cytokines (such as IL-2, IL-4, IL-6, IL-10, IL-17, IFN-α, IFN-γ, TNF-α, TGF-β, LT), and can be classified into distinct subsets depending on the particular cytokine profile, thus emerging the concept of cytokine-producing B cell subsets. Although there is still controversy surrounding the key cell surface markers, intracellular factors and cellular origins of cytokine-producing B cell subsets, accumulating evidence indicates that these B cells are endowed with great potential to regulate both innate and adaptive arms of immune system though releasing cytokines. On the one hand, they promote immune responses through mounting Th1/Th2/Th17 and neutrophil response, inducing DC maturation and formation of lymphoid structures, increasing NK cell and macrophage activation, enhancing development of themselves and sustaining antibody production. On the other hand, they can negatively regulate immune responses by suppressing Th cell responses, inhibiting Tr1 cell and Foxp3(+) Treg differentiation, impairing APC function and pro-inflammatory cytokine release by monocytes, and inducing CD8(+) T cell anergy and CD4(+) T cell apoptosis. Therefore, cytokine-producing B cell subsets have multifunctional functions in health and diseases, playing pathologic as well as protective roles in autoimmunity, infection, allergy, and even malignancy. In this review, we revisit the history of discovering cytokine-producing B cells, describe the identification of cytokine-producing B cell subsets, introduce the origins of cytokine-producing B cell subsets as well as molecular and cellular mechanisms for their differentiation, and summarize the recent progress made toward understanding the unexpectedly complex and potentially opposing roles of cytokine-producing B cells in immunological disorders.

<A<em>b</em>stractText>More effective and safer treatments are needed for antineutrophil cytoplasmic anti<em>b</em>ody (ANCA)-associated vasculitis.</A<em>b</em>stractText><p><div>(<em>b</em>)METHODS</<em>b</em>)</div>We conducted a randomized trial with a 2-<em>b</em>y-2 factorial design to evaluate the use of plasma exchange and two regimens of oral glucocorticoids in patients with severe ANCA-associated vasculitis (defined <em>b</em>y an estimated glomerular fi<em>lt</em>ration rate of &<em>lt</em>;50 ml per minute per 1.73 m² of <em>b</em>ody-surface area or diffuse pulmonary hemorrhage). Patients were randomly assigned to undergo plasma exchange (seven plasma exchanges within 14 days after randomization) or no plasma exchange (control group). Patients were also randomly assigned to follow either a standard-dose regimen or a reduced-dose regimen of oral glucocorticoids. Patients were followed for up to 7 years for the primary composite outcome of death from any cause or end-stage kidney disease (ESKD).</p><A<em>b</em>stractText>Death from any cause or ESKD occurred in 100 of 352 patients (28.4%) in the plasma-exchange group and in 109 of 352 patients (31.0%) in the control group (hazard ratio, 0.86; 95% confidence interval [CI], 0.65 to 1.13; P = 0.27). The resu<em>lt</em>s were similar in su<em>b</em>group analyses and in analyses of secondary outcomes. We also assessed the noninferiority of a reduced-dose regimen of glucocorticoids to a standard-dose regimen, using a noninferiority margin of 11 percentage points. Death from any cause or ESKD occurred in 92 of 330 patients (27.9%) in the reduced-dose group and in 83 of 325 patients (25.5%) in the standard-dose group (a<em>b</em>solute risk difference, 2.3 percentage points; 90% CI, -3.4 to 8.0), which met the criterion for noninferiority. Serious infections at 1 year were less common in the reduced-dose group than in the standard-dose group (incidence rate ratio, 0.69; 95% CI, 0.52 to 0.93), <em>b</em>ut other secondary outcomes were similar in the two groups.</A<em>b</em>stractText><A<em>b</em>stractText>Among patients with severe ANCA-associated vasculitis, the use of plasma exchange did not reduce the incidence of death or ESKD. A reduced-dose regimen of glucocorticoids was noninferior to a standard-dose regimen with respect to death or ESKD. (Funded <em>b</em>y the U.K. National Institute for Hea<em>lt</em>h Research and others; PEXIVAS Current Controlled Trials num<em>b</em>er, ISRCTN07757494; ClinicalTrials.gov num<em>b</em>er, NCT00987389.).</A<em>b</em>stractText>

Severe respiratory syncytial virus (RSV) infection is a major cause of morbidity and mortality in infants &lt;2 years-old. Here we describe that high-fiber diet protects mice from RSV infection. This effect was dependent on intestinal microbiota and production of acetate. Oral administration of acetate mediated interferon-β (IFN-β) response by increasing expression of interferon-stimulated genes in the lung. These effects were associated with reduction of viral load and pulmonary inflammation in RSV-infected mice. Type 1 IFN signaling via the IFN-1 receptor (IFNAR) was essential for acetate antiviral activity in pulmonary epithelial cell lines and for the acetate protective effect in RSV-infected mice. Activation of Gpr43 in pulmonary epithelial cells reduced virus-induced cytotoxicity and promoted antiviral effects through IFN-β response. The effect of acetate on RSV infection was abolished in Gpr43^-/- mice. Our findings reveal antiviral effects of acetate involving IFN-β in lung epithelial cells and engagement of GPR43 and IFNAR.

Background: Lutetium-177 [¹⁷⁷Lu]Lu-PSMA-617 is a radiolabelled small molecule that delivers β radiation to cells expressing prostate-specific membrane antigen (PSMA), with activity and safety in patients with metastatic castration-resistant prostate cancer. We aimed to compare [¹⁷⁷Lu]Lu-PSMA-617 with cabazitaxel in patients with metastatic castration-resistant prostate cancer.

Methods: We did this multicentre, unblinded, randomised phase 2 trial at 11 centres in Australia. We recruited men with metastatic castration-resistant prostate cancer for whom cabazitaxel was considered the next appropriate standard treatment. Participants were required to have adequate renal, haematological, and liver function, and an Eastern Cooperative Oncology Group performance status of 0-2. Previous treatment with androgen receptor-directed therapy was allowed. Men underwent gallium-68 [⁶⁸Ga]Ga-PSMA-11 and 2-flourine-18[¹⁸F]fluoro-2-deoxy-D-glucose (FDG) PET-CT scans. PET eligibility criteria for the trial were PSMA-positive disease, and no sites of metastatic disease with discordant FDG-positive and PSMA-negative findings. Men were randomly assigned (1:1) to [¹⁷⁷Lu]Lu-PSMA-617 (6·0-8·5 GBq intravenously every 6 weeks for up to six cycles) or cabazitaxel (20 mg/m² intravenously every 3 weeks for up to ten cycles). The primary endpoint was prostate-specific antigen (PSA) response defined by a reduction of at least 50% from baseline. This trial is registered with ClinicalTrials.gov, NCT03392428.

Findings: Between Feb 6, 2018, and Sept 3, 2019, we screened 291 men, of whom 200 were eligible on PET imaging. Study treatment was received by 98 (99%) of 99 men randomly assigned to [¹⁷⁷Lu]Lu-PSMA-617 versus 85 (84%) of 101 randomly assigned to cabazitaxel. PSA responses were more frequent among men in the [¹⁷⁷Lu]Lu-PSMA-617 group than in the cabazitaxel group (65 vs 37 PSA responses; 66% vs 37% by intention to treat; difference 29% (95% CI 16-42; p<0·0001; and 66% vs 44% by treatment received; difference 23% [9-37]; p=0·0016). Grade 3-4 adverse events occurred in 32 (33%) of 98 men in the [¹⁷⁷Lu]Lu-PSMA-617 group versus 45 (53%) of 85 men in the cabazitaxel group. No deaths were attributed to [¹⁷⁷Lu]Lu-PSMA-617.

Interpretation: [¹⁷⁷Lu]Lu-PSMA-617 compared with cabazitaxel in men with metastatic castration-resistant prostate cancer led to a higher PSA response and fewer grade 3 or 4 adverse events. [¹⁷⁷Lu]Lu-PSMA-617 is a new effective class of therapy and a potential alternative to cabazitaxel.

Funding: Prostate Cancer Foundation of Australia, Endocyte (a Novartis company), Australian Nuclear Science and Technology Organization, Movember, The Distinguished Gentleman's Ride, It's a Bloke Thing, and CAN4CANCER.

<AbstractText>Non-alcoholic steatohepatitis (NASH) is characterised by hepatic steatosis, inflammation, hepatocellular injury, and progressive liver fibrosis. Resmetirom (MGL-3196) is a liver-directed, orally active, selective thyroid hormone receptor-<em>β</em> agonist designed to improve NASH by increasing hepatic fat metabolism and reducing lipotoxicity. We aimed to assess the safety and efficacy of resmetirom in patients with NASH.</AbstractText><AbstractText>MGL-3196-05 was a 36-week randomised, double-blind, placebo-controlled study at 25 centres in the USA. Adu<em>lt</em>s with biopsy confirmed NASH (fibrosis stages 1-3) and hepatic fat fraction of at least 10% at baseline when assessed by MRI-proton density fat fraction (MRI-PDFF) were eligible. Patients were randomly assigned 2:1 by a computer-based system to receive resmetirom 80 mg or matching placebo, orally once a day. Serial hepatic fat measurements were obtained at weeks 12 and 36, and a second liver biopsy was obtained at week 36. The primary endpoint was relative change in MRI-PDFF assessed hepatic fat compared with placebo at week 12 in patients who had both a baseline and week 12 MRI-PDFF. This trial is registered with ClinicalTrials.gov, number NCT02912260.</AbstractText><AbstractText>348 patients were screened and 84 were randomly assigned to resmetirom and 41 to placebo at 18 sites in the USA. Resmetirom-treated patients (n=78) showed a relative reduction of hepatic fat compared with placebo (n=38) at week 12 (-32·9% resmetirom vs -10·4% placebo; least squares mean difference -22·5%, 95% CI -32·9 to -12·2; p&<em>lt</em>;0·0001) and week 36 (-37·3% resmetirom [n=74] vs -8·5 placebo [n=34]; -28·8%, -42·0 to -15·7; p&<em>lt</em>;0·0001). Adverse events were mostly mild or moderate and were balanced between groups, except for a higher incidence of transient mild diarrhoea and nausea with resmetirom.</AbstractText><AbstractText>Resmetirom treatment resu<em>lt</em>ed in significant reduction in hepatic fat after 12 weeks and 36 weeks of treatment in patients with NASH. Further studies of resmetirom will allow assessment of safety and effectiveness of resmetirom in a larger number of patients with NASH with the possibility of documenting associations between histological effects and changes in non-invasive markers and imaging.</AbstractText><AbstractText>Madrigal Pharmaceuticals.</AbstractText>

B cells are important in the pathogenesis of multiple sclerosis (MS) and some of the effects are not dependent on maturation of B cells into immunoglobulin (Ig) producing plasmablasts and plasma cells. B cells present antigen, activate T cells, and are involved in immunoregulation and cytokine secretion. To determine if B cells from MS patients secrete products that have deleterious effects on glial cells not mediated by Ig, and to compare effects with secretory products of normal controls (NC), we isolated B cells from 7 patients with relapsing remitting MS (RRMS) and 4 NC. B cells were cultured alone or after stimulation with CD40 ligand (CD40L), CD40L+cross-linking of the B cell antigen receptor (xBCR) and CD40L+xBCR+stimulation of toll like receptor 9 (TLR9). Supernatants were harvested and incubated with mixed central nervous system (CNS) neonatal rat glial cells. Supernatants from unstimulated NC B cells induced on average death of 7% (range 0-24%) of differentiated oligodendrocytes (OL); in contrast, supernatants from unstimulated B cells from RRMS patients induced death of 57% (range 35-74%) of OL. Supernatants of stimulated B cells from NC did not increase the minimal OL death whereas stimulation of B cells from RRMS had variable results compared to unstimulated B cells. Supernatants from both NC and RRMS induced microglial enlargement and loss of normal resting bipolar morphology. OL death did not correlate with levels of tumor necrosis alpha (TNF-α), lymphotoxin alpha (LT-α), interleukin 6 (IL-6), IL-10, transforming growth factor beta 1 (TGF-β1) or any combination or ratio of these cytokines. Analysis of 26 supernatants from NC and RRMS patients failed to detect IgM. There were very low levels of IgG in 8 of the 26 supernatants, and no correlation between of OL death and presence or absence of IgG. Sera used in both the B cell and glial cell cultures were heated, which inactivates complement. The effects of B cell supernatants on OL could be direct and/or indirect involving either microglia and/or astrocytes. The identity of the toxic factor(s) is as yet unknown. Thus we have demonstrated that B cells from patients with RRMS but not NC secrete one or more factors toxic to OL. It is possible that such factors produced by peripheral blood B cells when within the CNS could contribute to demyelination in MS patients.

We have determined the sequence of the DNA encoding the A2 (gamma) and B subunits of Vibrio cholerae enterotoxin. The order of the subunits as they would be transcribed is A2-B and the termination codon of the A2 subunit overlaps the initiation codon of the B subunit by four bases. Sequence analysis revealed a region capable of coding for a 21-amino acid leader peptide located at the NH2 terminus of the B subunit. While the nucleotide sequence homology between the cholera enterotoxin subunits and the analogous sequences of the genes for Escherichia coli heat-labile enterotoxin (LT) was 72 and 77% for the A2 and B subunits, respectively, the predicted amino acid sequences of the A2 subunits were less similar. Twenty-nine of 46 (63%) amino acids of the A2 subunits and 98 of 124 (79%) amino acids of the B subunits were identical between cholera enterotoxin and LT. The predicted amino acid sequence of the enterotoxin from a V. cholerae El Tor biotype strain reported here differs from the previously published amino acid sequences of the toxin from a classical biotype of V. cholerae at seven residues. Comparison of the A2 and B amino acid sequences among El Tor and classical biotypes of V. cholerae and E. coli LT demonstrates two regions of highly conserved sequences: 12 and 22 uninterrupted amino acids are the same among the A2 and B subunits, respectively, from the three strains.

Type II heat-labile enterotoxin (LT-II) from Escherichia coli 41 was purified and compared with prototype LT-II encoded by genes from E. coli SA53. Both toxins were oligomeric proteins consisting of polypeptides A (Mr, 28,000) and B (Mr, 11,800). The A polypeptides were cleaved by trypsin into fragments A1 (Mr, 21,000) and A2 (Mr, about 7,000). These two toxins were shown to belong to two different subclasses of LT-II. We propose to designate the prototype toxin LT-IIa and the new variant LT-IIb. The pI of LT-IIb was between 5.2 and 5.6, significantly lower than the pI of 6.8 for LT-IIa, and the behavior of LT-IIb during purification differed significantly from that of LT-IIa. The toxic dose of unnicked LT-IIb in the Y1 adrenal-cell assay was 94 pg, but trypsin-treated, nicked LT-IIb was toxic at about 3 pg. In contrast, the toxic dose of LT-IIa was previously shown to be 0.5 to 1 pg for several preparations that varied from unnicked to partially nicked, and treatment with trypsin was not required for full toxicity. The titer of LT-II antiserum in neutralization tests was 100-fold greater against LT-IIa than against LT-IIb. In immunodiffusion tests, LT-IIa and LT-IIb gave a reaction of partial identity. In a radioimmunobinding assay, the titer of LT-IIa antiserum against homologous LT-IIa was approximately 10-fold greater than against LT-IIb. The cholera-E. coli family of heat-labile enterotoxins has been divided into serogroup I, which includes cholera toxin and the antigenic variants of E. coli heat-labile toxin designated LTh-I and LTp-I, and serogroup II, which includes LT-IIa and LT-IIb. The type I and type II toxins do not cross-react in neutralization or immunodiffusion tests. By using very sensitive radioimmunobinding assays, it was possible to demonstrate common antigenic determinants between the type I and type II toxins. However, the titers of antibodies in hyperimmune sera that recognized these common determinants were very low.

T lymphocytes use several specialized mechanisms to induce apoptotic cell death. The tumor necrosis factor (TNF)-related family of membrane-anchored and secreted ligands represent a major mechanism regulating cell death and cell survival. These ligands also coordinate differentiation of tissue to defend against intracellular pathogens and regulate development of lymphoid tissue. Cellular responses are initiated by a corresponding family of specific receptors that includes two distinct TNFR (TNFR60 and TNFR80), Fas (CD95), CD40, p75NTF, and the recently identified lymphotoxin beta-receptor (LT beta R), among others. The MHC-encoded cytokines, TNF and LT alpha, form homomeric trimers, whereas LT beta assembles into heterotrimers with LT alpha, creating multimeric ligands with distinct receptor specificities. The signal transduction cascade is initiated by transmembrane aggregation (clustering) of receptor cytoplasmic domains induced by binding to their multivalent ligands. The TRAF family of Zn RING/finger proteins bind to TNFR80; CD40 and LT beta R are involved in induction NF kappa B and cell survival. TNFR60 and Fas interact with several distinct cytosolic proteins sharing the "death domain" homology region. TNF binding to TNFR60 activates a serine protein kinase activity and phosphoproteins are recruited to the receptor forming a multicomponent signaling complex. Thus, TNFRs use diverse sets of signaling molecules to initiate and regulate cell death and survival pathways.

Powerful noninvasive imaging technologies enable real-time tracking of pathogen-host interactions in vivo, giving access to previously elusive events. We visualized the interactions between wild-type Bacillus anthracis and its host during a spore infection through bioluminescence imaging coupled with histology. We show that edema toxin plays a central role in virulence in guinea pigs and during inhalational infection in mice. Edema toxin (ET), but not lethal toxin (LT), markedly modified the patterns of bacterial dissemination leading, to apparent direct dissemination to the spleen and provoking apoptosis of lymphoid cells. Each toxin alone provoked particular histological lesions in the spleen. When ET and LT are produced together during infection, a specific temporal pattern of lesion developed, with early lesions typical of LT, followed at a later stage by lesions typical of ET. Our study provides new insights into the complex spatial and temporal effects of B. anthracis toxins in the infected host, suggesting a greater role than previously suspected for ET in anthrax and suggesting that therapeutic targeting of ET contributes to protection.

The type II heat-labile enterotoxins (LT-IIa and LT-IIb) of Escherichia coli have an ABB subunits and different ganglioside receptor specificities. LT-II holotoxins and their nontoxic B subunits display unique properties as immunological adjuvants distinct from those of CT and its B subunits. In contrast to type II holotoxins, the corresponding pentameric B subunits, LT-IIaB and LT-IIbB, stimulated cytokine release in both human and mouse cells dependent upon Toll-like receptor 2 (TLR2). Induction of interleukin-1beta (IL-1beta), IL-6, IL-8, or tumor necrosis factor alpha in human THP-1 cells by LT-IIaB or LT-IIbB was inhibited by anti-TLR2 but not by anti-TLR4 antibody. Furthermore, transient expression of TLR1 and TLR2 in human embryonic kidney 293 cells resulted in activation of a nuclear factor-kappaB-dependent luciferase gene in response to LT-IIaB or LT-IIbB. Moreover, peritoneal macrophages from TLR2-deficient mice failed to respond to LT-IIaB or LT-IIbB, in contrast to wild-type or TLR4-deficient cells. These results demonstrate that besides their established binding to gangliosides, the B subunits of type II enterotoxins also interact with TLR2. Although a ganglioside-nonbinding mutant (T34I) of LT-IIaB effectively induced cytokine release, a phenotypically similar point mutation (T13I) in LT-IIbB abrogated cytokine induction, suggesting a variable requirement for gangliosides as coreceptors in TLR2 agonist activity. TLR2-dependent activation of mononuclear cells by type II enterotoxin B subunits appears to be a novel mechanism whereby these molecules may exert their immunomodulatory and adjuvant activities.

Isolated lymphoid follicles (ILFs) are recently appreciated members of the mucosal immune system. The architecture, composition, and inducible nature of these structures indicates that these structures are tertiary lymphoid structures. The process leading to the formation of tertiary lymphoid structures, lymphoid neogenesis, has been observed in a number of inflammatory and autoimmune conditions. Given this association, there is considerable interest in identifying the factors promoting lymphoid neogenesis, and understanding the steps in this process. Using murine ILF formation as a model, we have examined the roles of different cellular sources of lymphotoxin (LT) and the adaptive immune response in lymphoid neogenesis. In this study, we report that, although other cellular sources of LT may supplant B lymphocytes in the formation of immature ILFs (loosely organized clusters of B lymphocytes), LT-sufficient B lymphocytes are required for the progression of immature ILFs to mature ILFs (organized lymphoid aggregates with a follicle-associated epithelium). ILF formation occurs in the absence of T lymphocytes and Ag-specific B lymphocyte responses, and ILF B lymphocytes express elevated levels of LT in the absence of antigenic stimulation. Consistent with a role for chemokines inducing LT expression in Ag-naive B lymphocytes, and a chemokine-driven positive-feedback loop driving mature ILF formation, mature ILFs express elevated levels of B lymphocyte chemoattractant in the absence of Ag-specific B lymphocyte stimulation. These observations indicate that ILFs contain Ag-naive lymphocytes, and suggest that events occurring within ILFs shape subsequent immune responses mediated by these lymphocytes.

Normal human T cells purified from the peripheral blood and cultured for 6 days in the presence of OKT3 and IL-2 were shown to produce simultaneously TNF and lymphotoxin (LT) mRNA. Both biologically active TNF and LT were detected in supernatants of the cultured T cells using WEHI 164 as target cells. The cytotoxic activities of both cytokines could be inhibited with specific antibodies against TNF or LT. The T cell cultures were free of macrophages, NK cells, or B lymphocytes as assessed by flow cytometric analysis.

BACKGROUND

The meniscus plays critical roles in the knee, contributing to load transmission, shock absorption, and joint stability. Little is known about gene expression in meniscal tears, particularly in relation to injury pattern and patient age and sex. The purpose of this study was to test the hypothesis that gene expression in meniscal tears varies depending on patient age and sex and whether the anterior cruciate ligament (ACL) is also torn.

METHODS

Meniscal tissue from twenty-eight patients with an isolated meniscal tear or a meniscal tear with a concomitant ACL tear was collected at the time of clinically indicated partial meniscectomy. Messenger RNA (mRNA) expression was examined by quantitative real-time polymerase chain reaction for molecular markers of osteoarthritis including proinflammatory cytokines (interleukin [IL]-1α, IL-1β, IL-6, and tumor necrosis factor-alpha [TNFα]), chemokines (IL-8, CCL3, CCL3L1, CXCL1, CXCL3, CXCL6, and CCL20), aggrecanases (ADAMTS-4 [a disintegrin and metalloproteinase with thrombospondin type-4 motifs] and ADAMTS-5), matrix metalloproteinases (MMP-1, MMP-3, MMP-9, and MMP-13), transcription factors (NFκBBBIA [NF-kappa B inhibitor alpha], and IκBA [inhibitor of kappa B alpha]), and matrix components (bone morphogenetic protein [BMP]-2, type-I collagen alpha 1 [Col1a1], Col2a1, and aggrecan).

RESULTS

Expression of IL-1β (p = 0.02), ADAMTS-5 (p = 0.001), MMP-1 (p = 0.007), MMP-9 (p = 0.002), MMP-13 (p = 0.01), and NFκ<em>B</em>2 (p = 0.01) was significantly higher in patients with a meniscal tear who were under the age of forty years than it was in those over the age of forty years. Similarly, the expression of ADAMTS-4 (p = 0.002), ADAMTS-5 (p = 0.02), MMP-1 (p = 0.02), and MMP-13 (p = 0.0002) was higher in patients with a meniscal tear and an ACL tear who were under the age of forty years than it was in those over forty years. In patients with a meniscal tear and an ACL tear, the expression of IL-1β (p = 0.01), TNFα (p = 0.02), MMP-13 (p = 0.004), CCL3 (p = 0.03), and CCL3L1 (p = 0.03) was significantly higher, while that of aggrecan (p = 0.03) was lower, than that in patients with a meniscal tear alone. The only sex-based difference in gene expression was higher levels of CCL3L1 in female patients (p < 0.05) of all ages with combined injuries.

CONCLUSIONS

These findings suggest clinically relevant differences in the response of the knee to meniscal tears on the basis of patient age and sex. Elevated expression levels of arthritis-related markers indicate an increased catabolic response in patients under forty years old. Higher expression of catabolic markers in patients with meniscal and ACL tears suggests this combined injury pattern is more likely to lead to the development of osteoarthritis. Catabolic activity in meniscal tissue may predict patients who are at risk for progression of osteoarthritis following partial meniscectomy.

We have produced a functional heat labile enterotoxin (LT-) B subunit of Escherichia coli in maize. LT-B is a multimeric protein that presents an ideal model for an edible vaccine, displaying stability in the gut and inducing mucosal and systemic immune responses. Transgenic maize was engineered to synthesize the LT-B polypeptides, which assembled into oligomeric structures with affinity for G(M1) gangliosides. We orally immunized BALB/c mice by feeding transgenic maize meal expressing LT-B or non-transgenic maize meal spiked with bacterial LT-B. Both treatments stimulated elevated IgA and IgG antibodies against LT-B and the closely related cholera toxin B subunit (CT-B) in serum, and elevated IgA in fecal pellets. The transgenic maize induced a higher anti-LT-B and anti-CT-B mucosal and serum IgA response compared to the equivalent amount of bacterial LT-B spiked into maize. Following challenge by oral administration of the diarrhea inducing toxins LT and CT, transgenic maize-fed mice displayed reduced fluid accumulation in the gut compared to non-immunized mice. Moreover, the gut to carcass ratio of immunized mice was not significantly different from the PBS (non-toxin) challenged control group. We concluded that maize-synthesized LT-B had features of the native bacterial LT-B such as molecular weight, G(M1) binding ability, and induction of serum and mucosal immunity. We have demonstrated that maize, a major food and feed ingredient, can be efficiently transformed to produce, accumulate, and store a fully assembled and functional candidate vaccine antigen.

(<em>b</em>)Rationale</<em>b</em>): Up to date, the exploration of clinical features in severe COVID-19 patients were mostly from the same center in Wuhan, China. The clinical data in other centers is limited. This study aims to explore the feasi<em>b</em>le parameters which could <em>b</em>e used in clinical practice to predict the prognosis in hospitalized patients with severe coronavirus disease-19 (COVID-19). Methods: In this case-control study, patients with severe COVID-19 in this newly esta<em>b</em>lished isolation center on admission <em>b</em>etween 27 January 2020 to 19 March 2020 were divided to discharge group and death event group. Clinical information was collected and analyzed for the following o<em>b</em>jectives: 1. Comparisons of <em>b</em>asic characteristics <em>b</em>etween two groups; 2. Risk factors for death on admission using logistic regression; 3. Dynamic changes of radiographic and la<em>b</em>oratory parameters <em>b</em>etween two groups in the course. (<em>b</em>)Resu<em>lt</em>s</<em>b</em>): 124 patients with severe COVID-19 on admission were included and divided into discharge group (n=35) and death event group (n=89). Sex, SpO2, <em>b</em>reath rate, diastolic pressure, neutrophil, lymphocyte, C-reactive protein (CRP), procalcitonin (PCT), lactate dehydrogenase (LDH), and D-dimer were significantly correlated with death events identified using <em>b</em>ivariate logistic regression. Further mu<em>lt</em>ivariate logistic regression demonstrated a significant model fitting with C-index of 0.845 (p&<em>lt</em>;0.001), in which SpO2≤89%, lymphocyte≤0.64×10⁹/L, CRP>77.35mg/L, PCT>0.20μg/L, and LDH>481U/L were the independent risk factors with the ORs of 2.959, 4.015, 2.852, 3.554, and 3.185, respectively (p&<em>lt</em>;0.04). In the course, persistently lower lymphocyte with higher levels of CRP, PCT, IL-6, neutrophil, LDH, D-dimer, cardiac troponin I (cTnI), <em>b</em>rain natriuretic peptide (BNP), and increased CD4+/CD8+ T-lymphocyte ratio and were o<em>b</em>served in death events group, while these parameters stayed sta<em>b</em>le or improved in discharge group. (<em>b</em>)Conclusions</<em>b</em>): On admission, the levels of SpO2, lymphocyte, CRP, PCT, and LDH could predict the prognosis of severe COVID-19 patients. Systematic inflammation with induced cardiac dysfunction was likely a primary reason for death events in severe COVID-19 except for acute respiratory distress syndrome.

Keywords: COVID-19; Critical care; Prognosis; Radiography; Risk factors.

The B subunit of the heat labile toxin of enterotoxigenic Escherichia coli (LTB) was used as a model immunogen for production in soybean seed. LTB expression was directed to the endoplasmic reticulum (ER) of seed storage parenchyma cells for sequestration in de novo synthesized inert protein accretions derived from the ER. Pentameric LTB accumulated to 2.4% of the total seed protein at maturity and was stable in desiccated seed. LTB-soybean extracts administered orally to mice induced both systemic IgG and IgA, and mucosal IgA antibody responses, and was particularly efficacious when used in a parenteral prime-oral gavage boost immunization strategy. Sera from immunized mice blocked ligand binding in vitro and immunized mice exhibited partial protection against LT challenge. Moreover, soybean-expressed LTB stimulated the antibody response against a co-administered antigen by 500-fold. These results demonstrate the utility of soybean as an efficient production platform for vaccines that can be used for oral delivery.

Coronavirus disease 2019 (COVID-19) pandemic has affected health care systems worldwide. Severe presentations of COVID-19 such as severe pneumonia and acute respiratory distress syndrome (ARDS) have been associated with the post-viral activation and release of cytokine/chemokines which leads to a "cytokine storm" causing inflammatory response and destruction, mainly affecting the lungs. COVID-19 activation of transcription factor, NF-kappa B (NF-κB) in various cells such as macrophages of lung, liver, kidney, central nervous system, gastrointestinal system and cardiovascular system leads to production of IL-1, IL-2, IL-6, IL-12, TNF-α, LT-α, LT-β, GM-CSF, and various chemokines. The sensitised NF-κB in elderly and in patients with metabolic syndrome makes this set of population susceptible to COVID-19 and their worse complications, including higher mortality. Immunomodulation at the level of NF-κB activation and inhibitors of NF-κB (IκB) degradation along with TNF-α inhibition will potentially result in a reduction in the cytokine storm and alleviate the severity of COVID-19. Inhibition of NF-κB pathway has a potential therapeutic role in alleviating the severe form of COVID-19.

Keywords: COVID-19; Cytokine storm; Immunomodulation; MERS; NF-κb; SARS; Therapeutic role.

(<em>b</em>)Purpose:</<em>b</em>) Coronavirus disease 2019 (COVID-19) is an emerging disease that was first reported in Wuhan city, the capital of Hu<em>b</em>ei province in China, and has su<em>b</em>sequently spread worldwide. Risk factors for mortality have not <em>b</em>een well summarized. Current meta-analysis of retrospective cohort studies was done to summarize availa<em>b</em>le findings on the association <em>b</em>etween age, gender, comor<em>b</em>idities and risk of death from COVID-19 infection.(<em>b</em>)Methods:</<em>b</em>) Online data<em>b</em>ases including We<em>b</em> of Science, Pu<em>b</em>Med, Scopus, Cochrane Li<em>b</em>rary and Google scholar were searched to detect relevant pu<em>b</em>lications up to 1 May 2020, using relevant keywords. To pool data, random-effects model was used. Furthermore, sensitivity analysis and pu<em>b</em>lication <em>b</em>ias test were also done.(<em>b</em>)Resu<em>lt</em>s:</<em>b</em>) In total, 14 studies with 29,909 COVID-19 infected patients and 1445 cases of death were included in the current meta-analysis. Significant associations were found <em>b</em>etween older age (≥65 vs &<em>lt</em>;65 years old) (pooled ORs = 4.59, 95%CIs = 2.61-8.04, <i>p</i> &<em>lt</em>; .001), gender (male vs female) (pooled ORs = 1.50, 95%CIs = 1.06-2.12, <i>p</i> = .021) and risk of death from COVID-19 infection. In addition, hypertension (pooled ORs = 2.70, 95%CIs = 1.40-5.24, <i>p</i> = .003), cardiovascular diseases (CVDs) (pooled ORs = 3.72, 95%CIs = 1.77-7.83, <i>p</i> = .001), dia<em>b</em>etes (pooled ORs = 2.41, 95%CIs = 1.05-5.51, <i>p</i> = .037), chronic o<em>b</em>structive pulmonary disease (COPD) (pooled ORs = 3.53, 95%CIs = 1.79-6.96, <i>p</i> &<em>lt</em>; .001) and cancer (pooled ORs = 3.04, 95%CIs = 1.80-5.14, <i>p</i> &<em>lt</em>; .001), were associated with higher risk of mortality.(<em>b</em>)Conclusions:</<em>b</em>) Older age (≥65 years old), male gender, hypertension, CVDs, dia<em>b</em>etes, COPD and malignancies were associated with greater risk of death from COVID-19 infection. These findings could help clinicians to identify patients with poor prognosis at an early stage.

Keywords: COVID-19; SARS-CoV-2; meta-analysis; mortality; systematic review.

Certain classes of dendritic cells (DCs) meet rare cognate Ag-specific T and B cells inside primary B cell follicles for the development of germinal centers. However, the mechanisms underlying this coordination are still undefined. Cysteine-rich (CR) domain of the mannose receptor (CR-Fc)(+) DCs are a newly discovered subset of DCs that migrate rapidly into the primary lymphoid follicles from marginal zone after immunization. In this work, we uncover the key role of B cells in the establishment of a microenvironment that allows these DCs to be in the B cell area in a lymphotoxin (LT)-dependent fashion. CR-Fc(+) DCs are absent from the spleens of both LTbetaR- and LTalpha-deficient mice, suggesting that signaling by membrane LT is required for the presence of CR-Fc(+) DCs in the spleen. Interestingly, analysis of mutant mice that lack T, B, or NK cells demonstrates that B cell-derived membrane LT is essential for the unique localization of CR-Fc(+) DCs in the spleen. Using bone marrow transfer and ligand-blocking approaches, we provide evidence that B cell-derived LT acts indirectly on CR-Fc(+) DCs through LTbetaR(+) stromal cells. In analogous fashion to certain Ag-activated T and B cells, CR-Fc(+) DCs, expressing CXCR5, localize to primary lymphoid follicles in response to CXC ligand 13 (B lymphocyte chemoattractant). Together, we propose that B cells play a central role in establishing the chemotactic gradient that attracts not only Ag-activated T and B cells but also Ag-carrying CR-Fc(+) DCs. In turn, CR-Fc(+) DCs and T cells home to B cell follicles to interact with B cells in the developing germinal center.