Protected biofactors and antioxidants reduce the negative consequences of virus and cold challenge by modulating immunometabolism via changes in the interleukin-6 receptor signaling cascade in the liver.

Protected biofactors and antioxidants (PBA), and protected biofactors and antioxidants with protected organic acids and essential oils (PBA+POAEO) have been shown to have benefits in stressed or challenged birds. Here, we describe the immunometabolic changes observed in the liver of Ross 308 broilers during feed supplementation and brief physiological stress. These studied additives contain protected essential oils, organic acids, and vitamins which may have protective effects on the liver. Thus, we aimed to determine the signaling changes induced by these supplements and the resultant immunometabolic effects in the liver. All birds received a 2X dose of live bronchitis vaccine at d 0 and a 48-h cold challenge by reducing the temperature from 30 to 32°C, to 20 to 23°C on d 3 to 5. Control birds were fed a standard diet without supplementation. Liver samples were collected to evaluate the effects of these treatments on cytokine gene expression and protein phosphorylation via kinome peptide array. ANOVA was used for statistical analysis of the gene expression data (significance at a p-value of 0.05), and PIIKA2 was used for statistical evaluation and comparative analysis of the kinome peptide array data. At d 15, the kinome peptide array analysis and gene expression data showed stimulation of the interleukin 6 receptor (IL-6R) signal transduction for host protection via heightened immune response while inducing immune modulation and reducing inflammation in both supplement treated groups. Significant changes were observed via IL-6R signaling in the metabolic profiles of both groups compared to control and no significant differences when compared to each other. In the liver, these 2 feed additives induced immunometabolic changes predominantly via the IL-6 receptor family signaling cascade. Differences between the 2 treated groups were predominantly in the metabolic pathways, centered around the mTOR pathway and the proteins AMPK, mTOR and S6K, with a more anabolic phenotype following the addition of essential oils.

Novel model for chronic intestinal inflammation in chickens: (2) Immunologic mechanism behind the inflammatory response.

Intestinal inflammation in poultry is a complex response that involves immune and intestinal cells which is still not fully understood. Thus, to better understand the mechanisms that drive the chronic intestinal inflammation in fowl we conducted an experiment applying a previously established nutritional model of low-grade chronic intestinal inflammation to evaluate cytokine and chemokine profiles in the chicken intestine. For this, we placed 90 one-day chickens into two treatments: (1) a control group (CNT) fed a corn-soybean diet, and (2) a group fed a diet high in non-starch polysaccharides (NSP). At days 14, 22, 28 and 36 of age, 6 birds from each treatment were euthanized, jejunal and ileal samples were collected for histological examination and cytokine measurements. The cytokines interferon-alpha (IFN-α), IFN-γ, interleukin-16 (IL-16), IL-10, IL-21, IL-6, macrophage-colony stimulating factor (M-CSF), chemokine C-C motif ligand 20 (CCL20), CCL4, CCL5 and vascular endothelial growth factor (VEGF) were quantified in the intestinal tissue. Histologically, both jejunum and ileum of broilers fed NSP diet showed marked infiltration of mononuclear immune cells into the villi. Further, these birds exhibited a significant (P < 0.05) increase in CCL20 concentration in the jejunum at 14d, but a dramatic reduction of M-CSF at 14 and 21d. Later at 28d and 36d, birds fed the NSP diet exhibited increased IL-16 concentration in the jejunum. Since M-CSF is a monocyte stimulatory cytokine and CCL20 a chemokine of T-cells, the reduced M-CSF and increased production of CCL20 may indicate the involvement of the adaptive immune response, specifically driven by T-cells, occurring around the third week of age in the NSP model. Lastly, as a result of the mononuclear cell infiltration and activation of T-cells, IL-16, a pro-inflammatory T-cell cytokine, increased. Therefore, the current work indicates the importance of adaptive immune cells, especially T-cells, in the chronic intestinal inflammation in broiler chicken.

Protected biofactors and antioxidants reduce the negative consequences of virus and cold challenge while enhancing performance by modulating immunometabolism through cytoskeletal and immune signaling in the jejunum.

The aim of this study was to evaluate the effectiveness and mechanism of action of 2 feed additives in reducing the impacts of virus and temperature stressors. We determined the effects of protected biofactors and antioxidants (P(BF+AOx)), and protected biofactors and antioxidants with protected organic acids and essential oils (P(BF+AOx)+P(OA+EO)) on the immune and metabolic health of Ross 308 broiler chickens. These biofactors and antioxidants were derived from vitamins, and Aspergillus niger, Aspergillus oryzae and Bacillus subtilis fermentation extracts. All Ross 308 chickens were exposed to a double-dose of live bronchitis vaccine at d 0 and environmentally challenged by reducing the temperature from 32°C to 20°C at d 3 for 48 h. Control birds were fed without feed additives in the diet. Performance data and jejunum samples were collected to evaluate the effects of these treatments on growth, cytokine expression, and protein phosphorylation via kinome peptide array. ANOVA was used for statistical analysis of the performance and gene expression data (p-value of 0.05), and PIIKA2 was used for statistical evaluation and comparison of the kinome peptide array data. The P(BF plus;AOx) and P(BF+AOx)+P(OA+EO) treatments significantly increased bird weight gain and decreased feed conversion. The kinome peptide array data analysis showed increased activity of cytoskeletal, cell growth and proliferation proteins, and metabolic signaling in the jejunum of P(BF+AOx)+P(OA+EO) treated chickens. There was a significant decrease in IL-6 gene expression in the jejunum of P(BF+AOx)+P(OA+EO) samples compared to control at d 15. P(BF+AOx)+P(OA+EO) treatments in the jejunum showed strong immunomodulatory effects, perhaps to control inflammation. P(BF+AOx)+P(OA+EO) improves gut health via growth and metabolic signaling in the jejunum while inducing stronger immunomodulation.

A protected complex of biofactors and antioxidants improved growth performance and modulated the immunometabolic phenotype of broiler chickens undergoing early life stress.

We evaluated the supplementation of a protected complex of biofactors and antioxidants [P(BF+AOx)] on growth performance, antioxidant activity, expression of immune-related genes, and immunometabolic phenotype of broilers submitted to early life stressors. The treatments were a nutritionally complete basal diet supplemented or not with P(BF+AOx) (Jefo Nutrition Inc., Saint-Hyacinthe, QC, Canada) from 1 to 14 d of age. 720 one-day old male Ross 308 chickens were placed into pens of 30 birds (12 replicates/treatment). Birds were double-vaccinated against infectious bronchitis (IB; MILDVAC-Ma5T) at the hatchery and submitted, on d 3, to an acute reduction on environmental temperature (from 32° C to 20°C) for 48 h. Feed intake (FI), body weight gain (BWG), and feed conversion ratio (FCR) were calculated weekly. On d 7 and 15, samples were collected for expression of immune-related genes and kinome array analysis, and serum to evaluate the antioxidant status. Data were analyzed by ANOVA using SAS (SAS 9.4). From d 1 to 21 and d 1 to 28, the dietary supplementation of P(BF+AOx) significantly increased BWG (P < 0.05) by 3.6 and 3.8%, respectively, and improved FCR (P < 0.05) by 1.2 and 1.8%, respectively. From d 1 to 35, dietary supplementation enhanced BWG (P = 0.03) by 4%. Serum glutathione reductase activity on d 15 was higher in birds fed diets supplemented with P(BF+AOx) compared to the control diet-fed birds (P = 0.04). Dietary supplementation reduced the expression of IL-1ß (P = 0.03) in the lungs on d 7. On d 15, dietary supplementation increased the expression of IL-6 (P = 0.02) and IL-10 (P = 0.03) in the liver. It was observed that, via decreased phosphorylation, catalase was activated in the jejunum and liver, and the phosphorylation of immunoregulatory or proinflammatory proteins was decreased. Other important cellular signaling pathways were also changed in the liver and jejunum due to the supplementation. The supplementation of P(BF+AOx) improves growth performance by promoting a general anti-inflammatory and antioxidant response in chickens undergoing early life stress.

Selection for pro-inflammatory mediators produces chickens more resistant to Campylobacter jejuni.

Campylobacter spp. are the second leading cause of bacterial-induced foodborne illnesses with an estimated economic burden of nearly $2B USD per year. Most human illness associated with campylobacteriosis is due to infection by C. jejuni and chickens are recognized as a reservoir that could lead to foodborne illness in humans resulting from handling or consuming raw or undercooked chicken. We recently developed a novel breeding strategy based on identification and selection of chickens with an inherently high and low phenotype of pro-inflammatory mediators including IL-6, CXCLi2, and CCLi2, hereafter referred to as the high and low lines, respectively. We have shown the high line chickens are more resistant to the foodborne and poultry pathogens Salmonella enterica serovar Enteritidis, Eimeria tenella, and Clostridium perfringens-induced necrotic enteritis compared to the low line. The objective of this study was to determine whether the same trend of enhanced resistance in the high line birds was observed for C. jejuni. Birds were challenged at 2 d of age by oral gavage (0.5 mL) with 5 × 106 colony forming units (cfu) of C. jejuni/mL, necropsied 4 d post challenge, and cecal content collected to determine if there was a difference in C. jejuni resistance between the high and low line chickens. There were fewer (P = 0.01) chickens from the high line (28/40 = 71.8%) that were colonized by C. jejuni compared to the low line (37/39 = 94.9%). The amount of C. jejuni recovered from the ceca of infected birds was quantified; however, no differences were observed (P = 0.10). Since the high line birds were also more resistant to C. jejuni, it provides additional validation of selection based on pro-inflammatory mediators producing a line of chickens with increased natural resistance against diverse foodborne and poultry pathogens. The poultry industry is moving towards reduced therapeutics and, as such, our breeding strategy would be a viable method to incorporate into traditional poultry breeding programs.

Outcomes of emergent embolisation of ruptured hepatocellular carcinoma in a western population.

AIM: To evaluate the authors' experience with interventional radiological management of tumour rupture in hepatocellular carcinoma (HCC) in a Western population. MATERIALS AND METHODS: A retrospective review was performed of all consecutive patients treated at a single institution with transcatheter embolisation for ruptured HCC between 2000 and 2013. Patient age, sex, aetiology of liver disease, degree of underlying liver dysfunction, and clinical presentation were assessed. Embolisation was performed in a selective fashion when possible. Success, complications, and survival were assessed. RESULTS: Twenty-three patients were treated with embolisation for ruptured HCC. Of these patients, nine, nine, and five patients were Child-Pugh Class A, B, and C respectively. Embolisation was successful in 22 patients; one patient remained haemodynamically unstable and transfusion dependent despite embolisation. No major complications occurred. Median survival time was 260 days and the 30 day and 1 year survival rates were 83% and 45%, respectively. Child-Pugh class B or C (p = 0.04), Model for End-Stage Liver Disease score greater than 10 (p = 0.04), lobar embolisation (p = 0.04), and presence of portal vein thrombosis (p = 0.01) were significantly associated with worse prognosis. CONCLUSION: Transcatheter embolisation is effective at controlling haemorrhage in patients with ruptured HCC. Although major procedural complications are low, embolisation should proceed with an understanding of poor prognosis in patients with decompensated liver disease. Superselective embolisation is associated with improved prognosis and should be performed when feasible.

Effects of avian triggering receptor expressed on myeloid cells (TREM-A1) activation on heterophil functional activities.

A class of innate receptors called the triggering receptors expressed on myeloid cells (TREM) has been discovered and shown to be involved in innate inflammatory responses. The TREM family has been found in the chicken genome and consists of one activating gene (TREM-A1) and two inhibitory genes (TREM-B1 and TREM-B2). However, to date, there have been no reports on the effects of activating the TREM molecules on the functional activity of the primary avian polymorphonuclear cell, the heterophil. To characterize the activation of avian heterophils, we evaluated the effect of receptor ligation on heterophil effector functions. A specific agonistic antibody (Ab) was generated against the peptide sequence of chicken TREM-A1 38-51aa (YNPRQQRWREKSWC). To study TREM-A1 mediated activation, purified peripheral blood heterophils were incubated with various concentrations of the anti-TREM-A1 Ab or control Ab against an irrelevant antigen. Activation via TREM-A1 induces a significant increase in phagocytosis of Salmonella enteritidis, a rapid degranulation, and a dramatic up-regulation in gene expression of the pro-inflammatory cytokine, IL-6, and the inflammatory chemokine, CXCLi2. However, we found no direct TREM-A1 stimulation of the heterophil oxidative burst. Like mammalian TREM, avian TREM-A1 ligation synergizes with the activation of Toll-like receptor-4 (TLR4) ligand, LPS. In addition, the synergistic activity of LPS and TREM-A1 resulted in a significantly (p⩽0.05) increased production of an oxidative burst. Taken together, these results suggest, unlike in mammalian neutrophils, TREM-A1 engagement activates a differential functional activation of avian heterophils, but like mammalian neutrophils, acts in synergy with TLR agonists. These results provide evidence of the function of TREM-A1 in heterophil biology and avian innate immunity.

Caecal transcriptome analysis of colonized and non-colonized chickens within two genetic lines that differ in caecal colonization by Campylobacter jejuni.

Campylobacter jejuni is one of the most common causes of human bacterial enteritis worldwide. The molecular mechanisms of the host responses of chickens to C. jejuni colonization are not well understood. We have previously found differences in C. jejuni colonization at 7-days post-inoculation (pi) between two genetic broiler lines. However, within each line, not all birds were colonized by C. jejuni (27.5% colonized in line A, and 70% in line B). Therefore, the objective of the present experiments was to further define the differences in host gene expression between colonized and non-colonized chickens within each genetic line. RNA isolated from ceca of colonized and non-colonized birds within each line was applied to a chicken 44K Agilent microarray for the pair comparison. There were differences in the mechanisms of host resistant to C. jejuni colonization between line A and line B. Ten times more differentially expressed genes were observed between colonized and non-colonized chickens within line B than those within line A. Our study supports the fact that the MAPK pathway is important in host response to C. jejuni colonization in line B, but not in line A. The data indicate that inhibition of small GTPase-mediated signal transduction could enhance the resistance of chickens to C. jejuni colonization and that the tumour necrosis factor receptor superfamily genes play important roles in determining C. jejuni non-colonization in broilers.

Flagellin and lipopolysaccharide stimulate the MEK-ERK signaling pathway in chicken heterophils through differential activation of the small GTPases, Ras and Rap1.

The TLR agonists, flagellin (FLG) and lipopolysaccharide (LPS) stimulate functional activation and cytokine gene expression via the extracellular signal regulated kinase 1/2 (ERK1/2) MAP kinase cascade. However, the upstream mechanisms of these signaling events remain unknown. In mammals, the small GTP-binding protein Ras mediates ERK1/2 activation through activation of downstream effectors Raf-1-MEK1/2-ERK1/2 in response to a variety of stimuli. It is not clear whether this classic Ras cascade plays a role in TLR signaling in avian cells. In the present study, we investigated the role of Ras in FLG- and LPS-mediated signaling in ERK activation in chicken heterophils. Treatment of heterophils with LPS caused a rapid (within 5min) activation of Ras-GTP. The role of Ras activation in LPS-induced stimulation of ERK1/2 was corroborated when the specific Ras inhibitor, FTI-277, inhibited ERK1/2 activation. The classic Ras-mediated pathway of ERK1/2 activation by LPS was confirmed when the specific Raf-1 inhibitor, GW 5074, and the MEK1/2 inhibitor, U0126, both reduced ERK activation by 51-60%. Of more interest was that treatment of the heterophils with FLG did not activate Ras-GTP. Likewise, neither FTI-277 nor GW 5074 had any effect on FLG-mediated activation of ERK1/2. Another small GTPase, Rap1, has been shown to play a role in mammalian neutrophil function. Using a Rap1-GTP pull-down assay, we found that FLG stimulation, but not LPS, of avian heterophils induced a rapid and transient Rap1 activation. Rap1 has been shown to activate the ERK1/2 via a different Raf family member B-Raf whose downstream effector is MEK1/2. We show here that FLG stimulation of heterophils induces the phosphorylation of Rap1. The FLG induction of the Rap1-->B-Raf-->MEK1/2-->ERK1/2 cascade was confirmed by the reduction of ERK1/2 activation by the specific Rap1 inhibitor (GGTI-298) and U0126. The results demonstrate that for the first time that the small GTPase Ras family is involved in TLR signaling of avian heterophils with the TLR agonists LPS (Ras) and FLG (Rap1) inducing differential signaling cascades to activate the downstream ERK MAP kinase.

Cytokines and prevention of infectious diseases in poultry: a review.

Cytokines are soluble, low molecular weight polypeptides and glycopeptides produced by a broad range of cell types of haematopoietic and nonhaematopoietic origin that have suppressive or enhansive effects on cellular proliferation, differentiation, activation, and motility. Like hormones, cytokines mediate cellular responses through autocrine, paracrine, and endocrine activities. Both inflammatory responses and specific immune responses to invasive microbes, which were evolved to protect the host from pathogens, are controlled by a complex network of cytokines. As regulators of the initiation and maintenance of host defenses, cytokines ultimately determine the type of response generated and the effector mechanisms generated to mediate resistance. As effector molecules, cytokines are produced transiently and locally to control the amplitude and duration of the response. Likewise, excessive or insufficient production of a cytokine or cytokines may contribute significantly to the pathophysiology of disease. Therefore, cytokines play pivotal but paradoxical roles in both in the regulation of inflammation and immunity. The use of exogenous cytokines against infectious agents in poultry medicine has centered on: (a) their use as adjuvants for vaccines, (b) their direct effects on inducing protection against infections and/or the undesired effects of immune responses elicited by pathogens, and (c) their ability to stimulate the ontogeny and activation of neonatal host defenses. This monograph will review what has been reported about the use of cytokines in poultry in these three main areas.

Ontogeny of the phagocytic and bactericidal activities of turkey heterophils and their potentiation by Salmonella enteritidis-immune lymphokines.

Heterophils, the functional equivalent to the mammalian neutrophil, are important mediators of natural resistance against invasive pathogens in poultry. Young poultry are susceptible to pathogens, such as Salmonella enteritidis, during the first week post-hatch. No studies have evaluated the ontogeny of heterophil function in turkeys during the first few weeks post-hatch. Previous studies from our laboratory have shown day-old poults were protected against S. enteritidis organ invasion following immunoprophylactic administration of chicken S. enteritidis immune lymphokines. Therefore, the objective in the present study was to characterize the development of phagocytosis and bacterial killing by turkey heterophils during the first 3 weeks of life and to compare the effect of immune lymphokines on the development of heterophil phagocytosis and killing during the first 3 weeks post-hatch. Both functional phagocytosis and killing activities were age-dependent events. During the first 1-7 days post-hatch, little functional activity was demonstrated which apparently is associated with susceptibility. Optimal heterophil phagocytosis and killing activities were reached 14-21 days post-hatch. Administration immune lymphokines significantly potentiated phagocytosis (P < 0.01) and killing (P < 0.001) by turkey heterophils. In fact, immune lymphokine administration to 1-7-day-old poults augmented phagocytosis and killing activities of heterophils equivalent to levels found in functionally mature 14-21-day-old poults. These results demonstrate the ontogeny of the functional activity of the turkey heterophil is an age-related phenomenon, with inefficient phagocytosis and killing during the first week post-hatch. Prophylactic administration of immune lymphokines significantly potentiated the functional activity of the heterophil poults during the first 3 weeks of life. Most importantly the administration of immune lymphokines enhanced the functional activity of heterophils from 1-7-day-old poults to levels comparable to that of an immunologically mature bird.

Neutralization of G-CSF inhibits ILK-induced heterophil influx: granulocyte-colony stimulating factor mediates the Salmonella enteritidis-immune lymphokine potentiation of the acute avian inflammatory response.

Hematopoietic colony stimulating factors (CSF) regulate the growth and development of phagocytic cell progenitors and also augment functional activation of phagocytes. Granulocyte-CSF (G-CSF) is the CSF that acts specifically upon granulocyte progenitor cells and mature granulocytes. We have shown that lymphokines (ILK) from T cells of birds immunized against Salmonella enteritidis (SE) induce a granulocytic (PMN) inflammatory response in chicks challenged with SE. This inflammatory response was characterized by: (a) a dramatic emigration of granulocytic cells from the bone marrow into the peripheral blood, (b) an enhancement of the biological functions of the circulating PMNs, and (c) a directed influx of these activated PMNs to the site of bacterial invasion. In the current study, we determined the presence of G-CSF in ILK by Western blot analysis using a goat polyclonal antihuman G-CSF antibody (Ab). Using this Ab, we then evaluated the role of G-CSF in the ILK-induced protective inflammatory response in chickens against SE. Pretreatment of ILK with the Ab totally abolished the colony-stimulating activity of the ILK. Furthermore, Ab treatment of ILK resulted in: (a) an elimination of the ILK-induced peripheral blood heterophilia with a dramatic inhibition of ILK-mediated protection against SE organ invasion and (b) an elimination of accumulation of inflammatory PMNs in the peritoneum with subsequent decrease in the survival rate of chicks challenged i.p. with SE. Taken together these studies demonstrate for the first time the contribution of G-CSF to avian PMN activation and the immunoprophylaxis of SE infection by ILK in neonatal chickens.

Characterisation of colony-stimulating activity in the avian T cell-derived factor, Salmonella enteritidis-immune lymphokine.

This investigation was designed to characterise the specific cytokine activity from the conditioned medium of concanavalin A-stimulated avian T cells derived from Salmonella enteritidis-immune chickens, S enteritidis-immune lymphokine (ILK). Studies were designed to determine first, whether colony-stimulating activity was present in ILK, second, the type(s) of colonies from the bone marrow that were supported in vitro by the potential colony-stimulating factors in ILK and, third, whether colony-stimulating activity was present in serum from chicks treated with ILK and challenged with S enteritidis, and to use physicochemical treatment as a means of identifying the potential colony-stimulating factor(s) in ILK. Both ILK alone and serum from chicks treated with ILK and challenged with S enteritidis caused significant increases in the number of colony-forming units (CFU) from the bone marrow in vitro. After 10 days of incubation, ILK alone supported the in vitro growth of granulocytic bone marrow colonies. The colony-stimulating activity from serum derived from chicks treated with ILK and challenged with S enteritidis peaked two hours after the challenge. When ILK was either heated at 100 degrees C or treated with trypsin or acid and then injected into chicks, all the chicks responded with significant increases in circulating polymorphonuclear leucocytes (PMNs). However, when assayed for in vitro colony-stimulating activity, only trypsinisation destroyed the activity in ILK. The results indicate that a colony-stimulating factor which preferentially supported the growth of granulocytic bone marrow colonies was present in ILK and that the factor was stable to heat and acid but sensitive to trypsin.

Eimeria tenella or Eimeria adenoeides: induction of morphological changes and increased resistance to Salmonella enteritidis infection in Leghorn chicks.

The effect of either low doses of the chicken coccidium, Eimeria tenella (ET) or high doses of the turkey coccidium, Eimeria adenoeides (EA) on Salmonella enteritidis (SE) organ invasion, as well as histological and morphometric changes in the ceca of Leghorn chicks, was investigated. In these studies, chicks were inoculated orally with either saline solution (control) or one of three doses of sporulated oocysts of either ET or EA at 1 d of age. Five days later, 10(4) cfu of SE were administered per os to all chickens. Chicks were killed and organs cultured for SE 6 d following bacterial challenge. A clear relationship of decreased SE organ invasion was observed by increasing the dose of sporulated oocysts of either ET or EA. Furthermore, the increased resistance to organ infectivity in both experiments was matched with an increase in lamina propria thickness based on morphometric analysis (P < .05), probably due to a marked infiltration of inflammatory cells in the cecal mucosa. In both trials, a significant correlation was found (r = -.98 for ET; r = -.99 for EA) between the rate of bacterial organ infectivity and lamina propria thickness of the cecum. These data indicate that the increased resistance to SE organ invasion following subclinical coccidial infections were associated with morphological changes in the ceca of Leghorn chicks.

In vitro interleukin-1 and tumor necrosis factor-alpha production by macrophages from chickens infected with either Eimeria maxima or Eimeria tenella.

We measured the in vitro production of interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF- ) by macrophages from chickens during and immediately following an infection with one of two different species of Eimeria, E. maxima and E. tenella. Quantitatively, the amounts of IL-1 produced during each infection were nearly identical regardless of the oocyst dose of each parasite. TNF production followed a biphasic pattern of increased production with the first peak associated with the pathogenesis of disease and the second peak associated with the development of protective immunity. These experiments together with others we have reported imply cells taken from chickens infected with coccidia have a greater capacity to produce cytokines upon stimulation in vitro than cells from non-infected birds. The production of significantly greater amounts of TNF during the days 3-6 after inoculation correlates with the appearance of the most characteristic local and systemic pathophysiological changes in the host induced by the coccidia. The excessive release of TNF in response to a heavy coccidial infection may account for many of the pathological features observed with avian coccidiosis probably through the release of other mediators.

Recombinant interferon-gamma inhibits cell invasion by Eimeria tenella.

Treatment of primary chicken kidney (CK) cultures with supernatants from concanavalin A-treated Eimeria tenella-immune avian splenic T cells significantly inhibited the invasion of the cells by sporozoites of E. tenella. Biochemical evaluation of this T-cell factor is consistent with the idea that this anti-Eimeria activity in the avian lymphokines is due to interferon-gamma (IFN-gamma). We then studied the effect of recombinant bovine and human IFN-gamma on the invasion of sporozoites of E. tenella in cultures of bovine and human cell lines, respectively. The initial infection of either the Madin-Darby bovine kidney cell line (MDBK) or the human larynx epidermoid carcinoma cell line (HEp-2) by the obligate intracellular protozoan parasite E. tenella was inhibited following a 24-h pretreatment with recombinant bovine or human IFN-gamma, respectively. The IFN-gamma-mediated inhibition was dose-dependent for both cell lines. Incubation of sporozoites of E. tenella with IFN-gamma alone before infection had no detectable inhibitory effect on cell invasion of either cell line by the parasite. In addition, the establishment of the antiinfective cellular state by IFN-gamma required at least a 16- to 24-h preincubation for significant inhibitory effects. These results are suggestive of a direct effect of the IFN-gamma on the host cells and not on the parasite themselves. Taken together these studies provide evidence for a role for IFN-gamma as a nonspecific mediator of host protection against coccidiosis.

Gonadal function in patients with galactosaemia.

Gonadal function was followed in 26 females and 12 males with galactosaemia due to deficiency of the enzyme galactose-1-phosphate (Gal-1-P) uridyl transferase over a 4 year period. Gonadal function was normal in males, but all females except two had evidence of acquired ovarian failure. Twelve females with ovarian failure documented at the beginning of this study continued to have either primary or secondary amenorrhoea on follow-up. Five of six patients, who previously had normal gonadal function developed either hypergonadotrophic hypogonadism or an abnormal response to gonadotrophin-releasing hormone (LRH) indicative of acquired ovarian damage. Seven of eight female patients, 1-12 years of age, who were evaluated for the first time had an exaggerated release of gonadotrophins during LRH stimulation tests diagnostic of gonadal insufficiency. The pathogenesis of ovarian failure remains unknown, but it appears likely that galactose or Gal-1-P is toxic to the ovary. The source of galactose metabolites, which may begin to accumulate prenatally and continue to damage the gonad in the postnatal period, is likely to be derived from the diet and from the endogenous synthesis of Gal-1-P from glucose via a variety of metabolic pathways. The testis appears to be relatively resistant to the effects of abnormal galactose metabolism.