Nanocurcumin combined with insulin alleviates diabetic kidney disease through P38/P53 signaling axis.

Treatments for diabetic kidney disease (DKD) mainly focus on managing hyperglycemia and hypertension, but emerging evidence suggests that inflammation also plays a role in the pathogenesis of DKD. This 10-week study evaluated the efficacy of daily oral nanoparticulate-curcumin (nCUR) together with long-acting insulin (INS) to treat DKD in a rodent model. Diabetic rats were dosed with unformulated CUR alone, nCUR alone or together with INS, or INS alone. The progression of diabetes was reflected by increases in plasma fructosamine, blood urea nitrogen, creatinine, bilirubin, ALP, and decrease in albumin and globulins. These aberrancies were remedied by nCUR+INS or INS but not by CUR or nCUR. Kidney histopathological results revealed additional abnormalities characteristic of DKD, such as basement membrane thickening, tubular atrophy, and podocyte cytoskeletal impairment. nCUR and nCUR+INS mitigated these lesions, while CUR and INS alone were far less effective, if not ineffective. To elucidate how our treatments modulated inflammatory signaling in the liver and kidney, we identified hyperactivation of P38 (MAPK) and P53 with INS and CUR, whereas nCUR and nCUR+INS deactivated both targets. Similarly, the latter interventions led to significant downregulation of renal NLRP3, IL-1ß, NF-ĸB, Casp3, and MAPK8 mRNA, indicating a normalization of inflammasome and apoptotic pathways. Thus, we show therapies that reduce both hyperglycemia and inflammation may offer better management of diabetes and its complications.

Prevalence and Molecular Characteristics of Avian Pathogenic Escherichia coli in "No Antibiotics Ever" Broiler Farms.

Avian pathogenic Escherichia coli (APEC) causes significant economic and welfare concerns to the broiler industry. For several decades, prophylactic supplementation of antimicrobial growth promoters was the primary method to control APEC; however, the recent shift to no antibiotics ever (NAE) production has increased colibacillosis incidence. The objectives of this study were to determine the influence of season, flock age, and sample type on the prevalence and virulence of E. coli and to identify the serogroups and antimicrobial susceptibility of virulent and nonvirulent E. coli in NAE broiler farms. Litter, feces, cloacal swabs, and tracheal swabs were collected from 4 NAE farms during spring and summer seasons, and E. coli was isolated and confirmed by PCR. Confirmed E. coli isolates were tested for 5 APEC-virulence-associated genes (VAGs) using quantitative PCR (qPCR). Further, E. coli isolates with all five VAGs (100 isolates) and E. coli isolates without any VAGs (87 isolates) were screened against 11 antimicrobials through Kirby-Bauer disk diffusion assay, and their serogroups were tested using PCR. Data were analyzed using the GLIMMIX procedure of SAS 9.4, and statistical significance was determined at a P value of ≤0.05. Overall, the prevalence of E. coli was not affected by season, flock age, or sample type. However, the prevalence of all tested VAGs decreased from spring to summer (P ≤ 0.002). The frequency of resistance was highest for tetracycline, and serogroups O8 (31%) and O78 (11%) were most frequent in virulent E. coli. In conclusion, there is a high prevalence of virulent E. coli in NAE farms, especially in the spring season. IMPORTANCE Avian pathogenic Escherichia coli causes one of the most detrimental bacterial diseases to the United States poultry industry, colibacillosis. Colibacillosis leads to decreased performance, early mortality, and subsequent production loss. Previously, colibacillosis was largely mitigated by the use of antimicrobial growth promoters. Due to concerns about antimicrobial resistance, the use of these promoters has been largely removed from the broiler industry. With recent shifts in the poultry industry to NAE broiler production, there is an increase in bacterial disease and mortality. We do not know how this shift to NAE affects APEC prevalence within broiler farms. Therefore, in the current study, we attempted to assess the prevalence and virulence of E. coli within an antibiotic-free broiler environment, assessed antimicrobial susceptibility, and identified the serogroups of virulent and nonvirulent E. coli.

Research Note: Prevalence and molecular characteristics of Clostridium perfringens in "no antibiotics ever" broiler farms.

Clostridium perfringens (C. perfringens) is the etiological agent of necrotic enteritis and gangrenous dermatitis; 2 diseases that cause significant economic and welfare concerns to the broiler industry. Previously, Clostridium-related diseases were managed with the use of antimicrobial growth promoters fed to broilers that improved gut health and performance. The recent shift to no antibiotics ever (NAE) production has increased the incidence of Clostridium-related diseases. The objective of this study was to identify C. perfringens prevalence and toxinotypes in NAE farms. Samples of litter, feces, and cloacal swabs were collected from 4 NAE broiler farms in the summer of 2019, on d 28 and d 56 of one flock cycle. A total of 734 presumptive isolates were obtained from 192 samples collected in the study. Irrespective of the age of flock and sample type, all 192 samples contained at least one colony presumptively identified as C. perfringens on Perfringens agar plate with morphology as a single, round colony with opaque ring and black center. All isolates were further screened using PCR for confirmation, toxinotyping, and identification of virulence-associated genes. Only 9 isolates among the 734 presumptive isolates were confirmed as C. perfringens and all confirmed isolates were toxinotype A with variation in presence of netB, cpb2, and tpeL. More extensive studies are required to assess the prevalence and virulence of C. perfringens in NAE farms.

Transcriptomic analysis of early B-cell development in the chicken embryo.

The chicken bursa of Fabricius is a primary lymphoid tissue important for B-cell development. Our long-term goal is to understand the role of bursal microenvironment in an early B-cell differentiation event initiating repertoire development through immunoglobulin gene conversion in the chick embryo. We hypothesize that early bursal B-cell differentiation is guided by signals through cytokine receptors. Our theory is based on previous evidence for expression of the receptor tyrosine kinase superfamily members and interleukin receptors in unseparated populations of bursal B-cells and bursal tissue. Knowledge of the expressed genes that are responsible for B-cell differentiation is a prerequisite for understanding the bursal microenvironment's function. This project uses transcriptomic analysis to evaluate gene expression across early B-cell development. RNA-seq was performed with total RNA isolated from bursal B-cells at embryonic day (ED) 16 and ED 19 (n = 3). Approximately 90 million high-quality clean reads were obtained from the cDNA libraries. The analysis revealed differentially expressed genes involved in the Jak-STAT pathway, Wnt signaling pathway, MAPK signaling pathway, metabolic pathways including tyrosine metabolism, Toll-like receptor signaling pathway, and cell-adhesion molecules. The genes predicted to encode surface receptors, signal transduction proteins, and transcription factors identified in this study represent gene candidates for controlling B-cell development in response to differentiation factors in the bursal microenvironment.

Germ line variants of human N-methylpurine DNA glycosylase show impaired DNA repair activity and facilitate 1,N6-ethenoadenine-induced mutations.

Human N-methylpurine DNA glycosylase (hMPG) initiates base excision repair of a number of structurally diverse purine bases including 1,N(6)-ethenoadenine, hypoxanthine, and alkylation adducts in DNA. Genetic studies discovered at least eight validated non-synonymous single nucleotide polymorphisms (nsSNPs) of the hMPG gene in human populations that result in specific single amino acid substitutions. In this study, we tested the functional consequences of these nsSNPs of hMPG. Our results showed that two specific arginine residues, Arg-141 and Arg-120, are important for the activity of hMPG as the germ line variants R120C and R141Q had reduced enzymatic activity in vitro as well as in mammalian cells. Expression of these two variants in mammalian cells lacking endogenous MPG also showed an increase in mutations and sensitivity to an alkylating agent compared with the WT hMPG. Real time binding experiments by surface plasmon resonance spectroscopy suggested that these variants have substantial reduction in the equilibrium dissociation constant of binding (KD) of hMPG toward 1,N(6)-ethenoadenine-containing oligonucleotide (ϵA-DNA). Pre-steady-state kinetic studies showed that the substitutions at arginine residues affected the turnover of the enzyme significantly under multiple turnover condition. Surface plasmon resonance spectroscopy further showed that both variants had significantly decreased nonspecific (undamaged) DNA binding. Molecular modeling suggested that R141Q substitution may have resulted in a direct loss of the salt bridge between ϵA-DNA and hMPG, whereas R120C substitution redistributed, at a distance, the interactions among residues in the catalytic pocket. Together our results suggest that individuals carrying R120C and R141Q MPG variants may be at risk for genomic instability and associated diseases as a consequence.

Loss of function of the melanocortin 2 receptor accessory protein 2 is associated with mammalian obesity.

Melanocortin receptor accessory proteins (MRAPs) modulate signaling of melanocortin receptors in vitro. To investigate the physiological role of brain-expressed melanocortin 2 receptor accessory protein 2 (MRAP2), we characterized mice with whole-body and brain-specific targeted deletion of Mrap2, both of which develop severe obesity at a young age. Mrap2 interacts directly with melanocortin 4 receptor (Mc4r), a protein previously implicated in mammalian obesity, and it enhances Mc4r-mediated generation of the second messenger cyclic adenosine monophosphate, suggesting that alterations in Mc4r signaling may be one mechanism underlying the association between Mrap2 disruption and obesity. In a study of humans with severe, early-onset obesity, we found four rare, potentially pathogenic genetic variants in MRAP2, suggesting that the gene may also contribute to body weight regulation in humans.

IL-23-dependent IL-17 drives Th1-cell responses following Mycobacterium bovis BCG vaccination.

The generation of effective type 1 T helper (Th1)-cell responses is required for immunity against intracellular bacteria. However, some intracellular bacteria require interleukin (IL)-17 to drive Th1-cell immunity and subsequent protective host immunity. Here, in a model of Mycobacterium bovis Bacille Calmette-Guerin (BCG) vaccination in mice, we demonstrate that the dependence on IL-17 to drive Th1-cell responses is a host mechanism to overcome bacteria-induced IL-10 inhibitory effects. We show that BCG-induced prostaglandin-E2 (PGE2) promotes the production of IL-10 which limits Th1-cell responses, while simultaneously inducing IL-23 and Th17-cell differentiation. The ability of IL-17 to downregulate IL-10 and induce IL-12 production allows the generation of subsequent Th1-cell responses. Accordingly, BCG-induced Th17-cell responses precede the generation of Th1-cell responses in vivo, whereas the absence of the IL-23 pathway decreases BCG vaccine-induced Th17 and Th1-cell immunity and subsequent vaccine-induced protection upon M. tuberculosis challenge. Importantly, in the absence of IL-10, BCG-induced Th1-cell responses occur in an IL-17-independent manner. These novel data demonstrate a role for the IL-23/IL-17 pathway in driving Th1-cell responses, specifically to overcome IL-10-mediated inhibition and, furthermore, show that in the absence of IL-10, the generation of BCG-induced Th1-cell immunity is IL-17 independent.