Role of interleukin-1β in the regulation of porcine corpora lutea during the late luteal phase of the cycle and during pregnancy

Interleukin-1β (IL-1β) may regulate ovarian physiology. In this study, the influence of IL-1β on secretory activity within the corpora lutea (CL) of cyclic and gravid pigs was determined in vitro during different stages of the CL lifespan, e.g. on Days 10-11, 12-13 and 15-16 of the oestrous cycle and pregnancy. IL-1β (10 ng/ml) increased prostaglandin E2 (PGE2) secretion from CL of the cyclic and gravid pigs during studied days of the oestrous cycle and pregnancy. Increase (P < 0.05) of prostaglandin F2α (PGF2α) in IL-1β-treated CL was demonstrated only on Days 10-11 of the oestrous cycle. More potent stimulatory effect of IL-1β on PGE2 than PGF2α secretion resulted in the enhancement of the PGE2:PGF2α ratio in cyclic and early pregnant CL. IL-1β increased (P < 0.05) progesterone (P4) secretion only in gravid CL and had no effect on oestradiol-17β (E2) release. Expression of cyclooxygenase-2 (COX-2) mRNA was stimulated (P < 0.05) in IL-1β-treated cyclic and gravid CL. Expression of prostaglandin synthase mRNAs in response to IL-1β did not increase. In conclusion, IL-1β modulates PGE2, PGF2α and P4 secretion from porcine CL, depending on luteal stage and the surrounding hormonal milieu. The cytokine may act locally in porcine CL for luteotrophic support throughout the PGE2-mediated synthesis and secretion.     

Effects of sexual maturation and Salmonella infection on the expression of avian β-defensin genes in the chicken testis

Rooster infertility is a major concern in the poultry industry and protection of the male reproductive organs from pathogens is an essential aspect of reproductive physiology. During the last years, research on antimicrobial protection has elucidated the critical role of the antimicrobial peptides avian β-defensins (AvBDs) in the innate immunity in chickens. AvBDs have been reported to be expressed in the hen reproductive organs, providing protection against microbial pathogens including Salmonella Enteritidis (SE). However, mechanisms of antimicrobial protection of rooster reproductive organs and especially the testis, mediated by AvBDs are poorly understood. The aim of this study was to investigate the expression of the complete family of the 14 AvBD genes, in the rooster testis in vivo, to determine whether sexual maturation affects their testicular mRNA abundance and to investigate whether SE infection alters their expression. Expression analysis revealed that 9 members of the AvBD family, namely AvBD1, 2, 4, 5, 6, 9, 10, 12 and 14 were expressed in the testis. Quantitative real-time PCR analysis revealed that the mRNA abundance of three AvBDs was up regulated and of three AvBDs was down regulated with respect to sexual maturation. In addition, SE infection resulted in a significant induction of AvBD4, 10, 12 and 14 in the testis of sexually mature roosters. These findings provide strong evidence to suggest that an AvBD-mediated immune response mechanism exists in the rooster testis providing protection against bacterial pathogens including Salmonella species.     

Temporal changes in the expression of avian β-defensins in the chicken vagina during sexual maturation and Salmonella infection

Avian β-defensins (AvβDs) constitute a family of antimicrobial peptides that are critical to innate immunity in chickens, providing protection against microbial pathogens including Salmonella Enteritidis (SE). As apart from the digestive tract another main route of SE colonization in birds is via infection of the oviduct and specifically of the vagina, the aim of this study was to investigate the expression of the complete family of AvβDs, in the chicken vagina in vivo, to determine whether sexual maturation affects their mRNA abundance and to investigate whether SE infection alters the vaginal AvβDs expression. Expression analysis revealed that 11 members of the AvβD family were expressed in the chicken vagina. Quantitative real-time PCR analysis revealed that the mRNA abundance of five AvβDs was up regulated and of one AvβD was down regulated with respect to sexual maturation. In addition SE infection resulted in a significant induction of AvβD5, 7, 10, 11, 12 and 14 in the vagina of sexually mature birds, and in a significant induction of AvβD5 and 11 in the vagina of aged birds. These findings provide strong evidence to suggest that an AvβD-mediated immune response mechanism exists in the chicken vagina providing protection against bacterial pathogens including Salmonella species.     

Possible roles of myostatin and PGC-1α in the increase of skeletal muscle and transformation of fiber type in cold-exposed chicks: Expression of myostatin and PGC-1α in chicks exposed to cold

This study examined the hypothesis that myostatin and PGC-1α are involved in the increase in skeletal muscle mass and transformation of fiber type in cold-exposed chicks. One-week-old chicks were exposed to acute (24 h) or long-term (8 d) cold at 4 °C or kept warm at 30 °C. Acute cold exposure induced a significant increase in the skeletal muscle weight and the ratio of slow- to fast-fiber specific troponin I expression (sTnI/fTnI), accompanied by a significant decrease in lactate dehydrogenase activity. Expression of myostatin mRNA in the muscle was significantly lower in cold-exposed chicks than in the controls, whereas PGC-1α mRNA expression was significantly enhanced. These changes in the gene expression rapidly returned to the levels of the control chicks after the end of cold exposure, whereas the changes in fiber type and enzymatic activity were not resumed within 24 h after removal of cold exposure. On the other hand, long-term exposure to cold resulted in a remarkable increase in skeletal muscle weight, accompanied by a significant increase in the ratio of sTnI/fTnI and the enzymatic activities of cytochrome oxidase and lactate dehydrogenase. However, the expression level of myostatin mRNA in cold-exposed chicks was not different from that in their age-matched control chicks and that of PGC-1α mRNA was significantly lower than in the controls. These results indicate that myostatin and PGC-1α expression in the skeletal muscle rapidly change in response to acute cold, suggesting the possibility that these two genes could be involved in the increase in muscle mass and transformation of fiber type, respectively, at the initial stage of adaptation in cold-exposed chicks.     

Endometrial response to conceptus-derived estrogen and interleukin-1β at the time of implantation in pigs

The establishment of pregnancy is a complex process that requires a well-coordinated interaction between the implanting conceptus and the maternal uterus. In pigs, the conceptus undergoes dramatic morphological and functional changes at the time of implantation and introduces various factors, including estrogens and cytokines,interleukin-1β2(IL1 B2), interferon-γ(IFNG), and IFN-δ(IFND), into the uterine lumen. In response to ovarian steroid hormones and conceptus-derived factors, the uterine endometrium becomes receptive to the implanting conceptus by changing its expression of cell adhesion molecules, secretory activity, and immune response. Conceptus-derived estrogens act as a signal for maternal recognition of pregnancy by changing the direction of prostaglandin(PG) F2αfrom the uterine vasculature to the uterine lumen. Estrogens also induce the expression of many endometrial genes,including genes related to growth factors, the synthesis and transport of PGs, and immunity. IL1 B2, a pro-inflammatory cytokine, is produced by the elongating conceptus. The direct effect of IL1 B2 on endometrial function is not fully understood. IL1 B activates the expression of endometrial genes, including the genes involved in IL1 B signaling and PG synthesis and transport. In addition, estrogen or IL1 B stimulates endometrial expression of IFN signaling molecules,suggesting that estrogen and IL1 B act cooperatively in priming the endometrial function of conceptus-produced IFNG and IFND that, in turn, modulate endometrial immune response during early pregnancy. This review addresses information about maternal-conceptus interactions with respect to endometrial gene expression in response to conceptus-derived factors, focusing on the roles of estrogen and IL1 B during early pregnancy in pigs.     

Central injection of exogenous IL-1β in the control activities of hypothalamic-pituitary-gonadal axis in anestrous ewes

This study was performed to determine the effect of intracerebroventricular (icv) injection of interleukin (IL)-1β on the gene expression, translation and release of gonadotropin-releasing hormone (GnRH) and the GnRH receptor (GnRHR) gene expression in the hypothalamus of anestrous ewes. In the anterior pituitary gland (AP), the expression of genes encoding: GnRHR, β subunits of luteinizing hormone (LH) and folliculotropic hormone (FSH) was determined as well as the effect of IL-1β on pituitary gonadotropins release. The relative mRNA level was determined by real-time PCR, GnRH concentration in the cerebrospinal fluid (CSF) was assayed by ELISA and the plasma concentration of LH and FSH were determined by radioimmunoassay. Our results showed that icv injection of IL-1β (10 or 50 μg/animal) decreased the GnRH mRNA level in the pre-optic area (POA) (35% and 40% respectively; p ≤ 0.01) and median eminence (ME) (75% and 70% respectively; p ≤ 0.01) and GnRHR gene expression in ME (55% and 50% respectively; p ≤ 0.01). A significant decrease in GnRHR mRNA level in the AP in the group treated with the 50 μg (60%; p ≤ 0.01) but not with the 10 μg dose was observed. The centrally administrated IL-1β lowered also GnRH concentration in the CSF (60%; p ≤ 0.01) and reduced the intensity of GnRH translation in the POA (p ≤ 0.01). It was not found any effect of icv IL-1β injection upon the release of LH and FSH. However, the central injection of IL-1β strongly decreased the LHβ mRNA level (41% and 50%; p ≤ 0.01; respectively) and FSHβ mRNA in the case of the 50 μg dose (49%; p ≤ 0.01) in the pituitary of anestrous ewes. These results demonstrate that the central IL-1β is an important modulator of the GnRH biosynthesis and release during immune/inflammatory challenge.     

Dominant expression of interleukin-8 vs interleukin-1β and tumour necrosis factor alpha in lungs of lambs experimentally infected with Mannheimia haemolytica

Abstract

AIMS: To quantify the number of cells infected with Mannheimia haemolytica and expressing interleukin (IL)-1β, tumour necrosis factor alpha (TNFα) and IL-8 using immunohistochemistry, and to measure the immunoreactivity of cytokines in pulmonary tissue extracts using ELISA, in the lung of lambs experimentally infected with M. haemolytica, and to compare the patterns of expression of cytokines in airways at different times post-infection (p.i.).

METHODS: Twenty 3-month-old lambs of both sexes were randomly assigned to two groups, viz infected (n=15), and uninfected controls (n=5). Each lamb in the infected group was inoculated with 1.5 x 10⁹ cfu M. haemolytica in 5 mL sterile nutrient broth, control lambs were inoculated with 5 mL sterile nutrient broth and clinical signs were monitored. Infected and control animals were killed at 1, 3, 5, 7, and 15 days p.i. Histopathology and immunohistochemistry were conducted to determine the number of immunolabelled cells in pneumonic lungs, and study the pattern of expression of IL-1β, TNFα and IL-8 in lung extracts using ELISA.

RESULTS: Lesions in bronchi and bronchioles ranged from epithelial desquamation to bronchiolitis obliterans and necrosis. The alveoli had areas of seroproteinaceous fluid, fibrin and bacterial aggregates that evolved to foci of pyogranulomatous inflammation with clustered inflammatory cells, referred to as ‘oat cells’. M. haemolytica antigen was observed in the cytoplasm of inflammatory cells. Labelling of IL-1β, TNFα and IL-8 was observed in bronchial and bronchiolar epithelial cells, alveolar exudate, and in interstitial inflammatory infiltrate, with increased expression on 1 and 3 days p.i. for IL-1β and TNFα, and 1, 3, and 5 days p.i. for IL-8. In lung tissue extracts, peak concentrations of IL-1β (55 (SD 5) ng/mL), TNFα (92 (SD 6) pg/mL) and IL-8 (8 [SD 2] μg/mL) occurred at 3 days p.i.

CONCLUSIONS: The results of this study suggested that the inflammatory cytokines IL-1β, TNFα and IL-8 may play an important role in enhancing the biological response to M. haemolytica, and contribute to the development of lesions in the lung in pulmonary pasteurellosis in sheep. Given that the expression of IL-8 in lung was much greater than that of IL-1β and TNFα, anti-cytokine agents directed at this mediator could be useful in the prevention and treatment of this disease.     

Genetics of canine diabetes mellitus: Are the diabetes susceptibility genes identified in humans involved in breed susceptibility to diabetes mellitus in dogs?

Diabetes mellitus is a common endocrinopathy in companion animals, characterised by hyperglycaemia, glycosuria and weight loss, resulting from an absolute or relative deficiency in the pancreatic hormone insulin. There are breed differences in susceptibility to diabetes mellitus in dogs, with the Samoyed breed being overrepresented, while Boxers are relatively absent in the UK population of diabetic dogs, suggesting that genetic factors play an important role in determining susceptibility to the disease. A number of genes, linked with susceptibility to diabetes mellitus in humans, are associated with an increased risk of diabetes mellitus in dogs, some of which appear to be relatively breed-specific. Diabetes mellitus in dogs has been associated with major histocompatibility complex (MHC) class II genes (dog leucocyte antigen; DLA), with similar haplotypes and genotypes being identified in the most susceptible breeds. A region containing a variable number of tandem repeats (VNTR) and several polymorphisms have been identified in the canine insulin gene, with some alleles associated with susceptibility or resistance to diabetes mellitus in a breed-specific manner. Polymorphisms in the canine CTLA4 promoter and in other immune response genes are associated with susceptibility to diabetes mellitus in a number of pedigree breeds. Genome wide association studies are currently underway that should shed further light on the genetic factors responsible for the breed profile seen in the diabetic dog population.     

Breed-related expression patterns of Ki67, γH2AX,and p21 during ageing in the canine liver

Veterinary Research Communications - Cellular senescence is a molecular hallmark of ageing that is associated with multiple pathologies, and DNA damage marker γH2AX, together with cell cycle...     

Validation of the Postprandial Interleukin-1β Response in Horses Using Equine-Specific Antibodies

Interleukin (IL)-1β is a commonly studied proinflammatory cytokine, with relevance to arthritis, obesity, aging, and other inflammatory diseases of horses. Evaluating protein concentrations in plasma is a useful measurement for research in these areas of equine health. The objective of this research was to validate a commercially available enzyme-linked immunosorbent assay (ELISA) for equine IL1β and to compare concentrations with those previously published using an ELISA that is no longer available. The ELISA was assessed for linear parallelism and recovery using plasma from four healthy Standardbred mares. The assay was found to have linear parallelism for samples diluted 1:2, when the detection antibody concentration was 3 μg/mL. The average recovery of spiked IL1β was 98.9%. To compare concentrations, plasma was collected from six geldings at −0.5, 1, 2, and 4 hours after consumption of a meal high in starch and sugar (1.2 g/kg bodyweight). Consumption of 1.2 g of starch and sugar per kg of BW increased plasma IL1β concentrations 1 hour after feeding (P = .053). In conclusion, the commercially available ELISA is validated, with modifications, for use in equine plasma, and it detects a rise in plasma IL1β concentrations at 1 hour after meal consumption, a finding that is similar to previously reported data.     

Prevalence of Flavobacterium psychrophilum bacterial cells in farmed rainbow trout: Characterization of metallothionein A and interleukin1-β genes as markers overexpressed in spleen and kidney of diseased fish

The aim of the present study was to assess the prevalence of the flavobacteria within farmed trout and to quantify their bacterial burden. A total of 61 fish were sampled from seven farms, and were distributed in two groups: (1) visibly diseased fish suffering from the rainbow trout fry syndrome or the bacterial cold water disease caused by the bacteria Flavobacterium psychrophilum and (2) normally appearing fish. F. psychrophilum cells were titered by qPCR, targeting a specific area of the 16S rRNA gene in skin, muscle, gills, liver, spleen and kidney from all fish. The pathogen was detected in these organs whatever the health status, with titers ranging from 10⁴ to 6 × 10⁷ bacteria/g of tissue in normally appearing fish, thus showing they were bacterial carriers. Two organs allowed differentiation between diseased and normally appearing fish: spleen and kidney, with titers ranging from 10⁶ to 10⁷ bacteria/g of tissue in normally appearing fish vs 10¹¹ to 10¹² bacteria/g of tissue in diseased fish. No relationship was found between immunoglobulin M-like titer in plasma and health status. Gene expression analysis in fish organs revealed two genes that were markers of the bacterial infection: mt-a and il-1β genes encoding the metallothionein A and the interleukin1-β, respectively. These genes were both over-expressed in gills, liver, spleen and kidney of diseased fish. Four genes encoding immunity markers were down-regulated in spleen (a key organ implicated in immunity) of diseased fish: tgf-β, cd8-α, mhc2-β and igt, suggesting a weakened immune system in diseased fish.     

Negative regulation of interleukin 1β expression in response to DnaK from Pseudomonas aeruginosa via the PI3K/PDK1/FoxO1 pathways

Interleukin (IL)-1β is crucial for a wide range of inflammatory responses. Previously, we reported that IL-1β is produced in response to Pseudomonas aeruginosa-derived DnaK via NF-κB and JNK pathways; however, the signaling pathways that counter the process to maintain IL-1β homeostasis are unknown. Here, we show that DnaK-mediated expression of IL1β is increased markedly in macrophages upon blockade of PI3K/PDK1. This was verified by measuring released IL-1β protein. The negative effect of PI3K on IL-1β production was dependent on suppression of both NF-κB and JNK activation. Intriguingly, PDK1 (an underlying mediator of PI3K) acted as an upstream regulator for the activation of NF-κB, but downregulated JNK activation. Furthermore, production of IL-1β and activation of JNK were triggered by inhibition of phosphorylated FoxO1; phosphorylation of FoxO1 was controlled by PDK1 signaling in response to DnaK. Thus, IL-1β production is modulated by P. aeruginosa-derived DnaK via cross-talk between JNK and PI3K/PDK1/FoxO1 pathways.     

Regulation of matrix metalloproteinase-9 protein expression by 1α,25-(OH)2D3 during osteoclast differentiation

To investigate 1α,25-(OH)₂D₃ regulation of matrix metalloproteinase-9 (MMP-9) protein expression during osteoclast formation and differentiation, receptor activator of nuclear factor κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) were administered to induce the differentiation of RAW264.7 cells into osteoclasts. The cells were incubated with different concentrations of 1α,25-(OH)₂D₃ during culturing, and cell proliferation was measured using the methylthiazol tetrazolium method. Osteoclast formation was confirmed using tartrate-resistant acid phosphatase (TRAP) staining and assessing bone lacunar resorption. MMP-9 protein expression levels were measured with Western blotting. We showed that 1α,25-(OH)₂D₃ inhibited RAW264.7 cell proliferation induced by RANKL and M-CSF, increased the numbers of TRAP-positive osteoclasts and their nuclei, enhanced osteoclast bone resorption, and promoted MMP-9 protein expression in a concentration-dependent manner. These findings indicate that 1α,25-(OH)₂D₃ administered at a physiological relevant concentration promoted osteoclast formation and could regulate osteoclast bone metabolism by increasing MMP-9 protein expression during osteoclast differentiation.