Natural Immunoglobulin Research Articles

Gene expression profiling in the lungs of pigs with different susceptibilities to Glässer's disease

BackgroundHaemophilus parasuis is the causative agent of Glässer's disease in pigs. Currently, little is known about the molecular mechanisms that contribute to disease susceptibility. This study used a porcine oligonucleotide microarray to identify genes that were differentially expressed (DE) in the lungs of colostrum-deprived animals previously characterized as being either 'Fully Resistant' (FR) or 'Susceptible' to infection by H. parasuis in a bacterial challenge experiment.ResultsGene expression profiles of 'FR' and 'Susceptible' animals were obtained by the identification of genes that were differentially expressed between each of these groups and mock-inoculated 'Control' animals. At 24 hours post-inoculation, a total of 21 and 58 DE genes were identified in 'FR' and 'Susceptible' animals respectively. At 72 hours, the numbers of genes were 20 and 347 respectively. 'FR' animals at 24 hours exhibited an increased expression of genes encoding extracellular matrix and TGF-β signalling components, possibly indicative of tissue repair following the successful early resolution of infection. The gene expression profile of 'FR' animals at 72 hours supported the hypothesis that higher levels of antibacterial activity were responsible for the 'FR' phenotype, possibly due to an increase in natural immunoglobulin A and decrease in signalling by the immunoregulatory transcription factor peroxisome proliferator-activated receptor gamma (PPAR-γ). The expression profile of 'Susceptible' animals at both time-points was characterized by an imbalance in signalling between pro and anti-inflammatory cytokines and an increased expression of genes involved in biological processes associated with inflammation. These include the pro-inflammatory cytokine genes resistin (RETN) and interleukin 1-beta (IL1B). At 72 hours, a reduction in the expression of genes involved in antigen presentation by both MHC class I and II molecules was observed, which could have contributed to the inability of 'Susceptible' animals to clear infection.ConclusionsThis study is the first to have identified discrete sets of DE genes in pigs of differing susceptibility to H. parasuis infection. Consequently, several candidate genes and pathways for disease resistance or susceptibility phenotypes have been identified. In addition, the findings have shed light on the molecular pathology associated with Glässer's disease.

Viral Antigens Trigger Granzyme B Secretion by Human CD5+ B1 Cells in the Presence of Interleukin 21.

Abstract 4729CD5+ B1 cells are thought to play a key role in innate immune responses by producing and secreting natural immunoglobulins, which bind to a wide range of bacterial, viral antigens as well as auto-antigens. Here we demonstrate that B1 cells from human cord blood are able to express the pro-apoptotic serine protease granzyme B (GrB) in response to viruses including tick-borne encephalitis virus, rabies virus, and hepatitis B virus. Using pharmacological tools we reveal that this response depends on B cell receptor stimulation, toll-like receptor 9 engagement and interleukin 21 (IL-21), a cytokine primarily found in the acute phase of viral infections. While up to 35% of freshly isolated B1 cells directly respond with GrB expression to stimulation with inactivated viruses in the presence of IL-21, similar, though lower reactions in CD5- B2 cells are only found after, but not before vaccination of the donor against the respective virus. Of note, GrB-expressing B1 cells feature a homogeneous CD19+CD5+GrB+CD20+CD27-CD38-IgD- phenotype and B1 cell-derived GrB is secreted in an enzymatically active state, reaching levels comparable to those secreted by activated cytotoxic cells. GrB induction requires activation of similar signaling pathways as in CTL and NK cells including members of the JAK/STAT pathway. Our findings suggest GrB secretion by B1 cells represents a novel innate immune response mechanism. GrB-secreting B1 cells may play a role in early anti-viral immune defense, and may contribute to elevated serum GrB levels found in various viral diseases. Further studies will elucidate whether CD5+ B1 cells, possibly in co-operation with NK cells, exhibit cytotoxicity towards virus-infected as well as tumor cells. Disclosures:No relevant conflicts of interest to declare.

ATTENUATION OF THE EFFECTS OF RAT HEMORRHAGIC SHOCK WITH A REPERFUSION INJURY-INHIBITING AGENT SPECIFIC TO MICE

Death after hemorrhagic shock (HS) may be caused by a generalized reperfusion injury, particularly noticeable in the gut. A period of tissue ischemia followed by reinstitution of perfusion produces severe inflammation that can be blocked in mice by preventing the binding of a pathogenic natural immunoglobulin M (IgM) of defined specificity to antigens in reperfused tissue by using a soluble peptide analogue of the IgM tissue target. We hypothesize that this agent can improve end points of rat HS: death, intestinal injury, and lung injury. Male Sprague-Dawley rats were anesthetized; 50% of calculated blood volume was removed for 120 min, shed blood, then returned; and animals were sacrificed at 72 h. One group of rats received i.v. analogue ([N2] 300 microg) with the return of shed blood. Small intestine and lung were evaluated by histological examination and immunohistochemistry. Lung edema was assessed by Evans blue extravasation and histological examination. I.v. N2 decreased experimental mortality from 62% to 12% (P < 0.05). Associated with this was diminution of gut injury score from 57.8% +/- 5.5% to 19.5% +/- 2.5% (P < 0.05), lung injury from 21.4 +/- 1.5 to 14.8 +/- 1.3 polymorphonuclear leucocytes per high-power field x400 (P < 0.05), and Evans blue extravasation index from 0.61 +/- 0.14 to 0.18 +/- 0.06 (P < 0.05). As well, the deposition of IgM and C3 that is seen in intestinal villi from HS was not present in N2-treated rats. The N2 peptide agent that blocks reperfusion injury in mice prevents death from rat HS, as well as attenuates gut reperfusion injury and its remote target injuries. These data suggest that death from HS is caused by reperfusion injury, and that an agent derived from mice is effective in rats when given in real therapeutic time.

Octamer binding protein 2 (Oct2) regulates PD-L2 gene expression in B-1 cells through lineage-specific activity of a unique, intronic promoter.

PD-L2 expression extends beyond macrophages/dendritic cells to B-1 B cells, a distinct B cell lineage that is responsible for natural immunoglobulin and which is repertoire skewed toward autoreactive specificities. PD-L2 expression is constitutive in B-1 cells, whereas it is inducible in other cell types, suggesting that PD-L2 is regulated differently in the former versus the latter, and this proved to be the case, both in transcription and promotion. B-1 cells express a PD-L2 transcript that lacks exon1, in contrast to macrophages/dendritic cells for which exon1 is included, reflecting a unique start site upstream of exon2. PD-L2 transcription in B-1 cells is regulated by a novel intronic promoter located between exon1 and exon2. This intronic promoter binds Oct1 and Oct2, and although these transcription factors are present in all B cells, Oct2 binding is found in vivo only in B-1 cells and not PD-L2 negative B-2 cells. Moreover, the proximal promoter upstream of exon1 that is active in macrophages is inactive in B-1 cells. Thus, PD-L2 expression is regulated by two different promoters that function in a lineage-specific fashion, with the B-1-specific promoter being constitutively active as a result of Oct1 and Oct2 binding.

Evolution of major histocompatibility complex G and C and natural killer receptors in primates

Major histocompatibility complex (MHC)-G and -C molecules bear ligands to natural killer immunoglobulin receptors (KIR). MHC-G evolution in primates shows some anomalies. In New World monkeys MHC-G molecules show a high polymorphism and most likely are classical antigen presenters; they also cluster closer to MHC-E in a relatedness dendrogram. Their genes lack intron 2 deletion, which is typical of all other primates in regard to MHC-G. Medium-sized Eurasian-African monkeys ( Cercopithecinae) show stop codons in exon 3: only MHC-G isoforms without exon 3 are possible. Big apes such as the orangutan, gorilla, and chimpanzee as well as human beings show limited HLA-G polymorphism. HLA-C has not been found in medium-size Eurasian-African monkeys, but we have found MHC-C DNA sequences in more evolutionary ancient New World monkeys. Taking into account that the KIR inhibitory receptors signal is dominated by MHC-C in human beings, this suggests that both MHC-C molecules and their ligands within natural killer lymphocyte KIR also exist in the most evolutionary ancient apes (New World monkeys were present on Earth before 40 million years ago), as KIR receptors also appeared before 130 million years ago in evolution. Indeed, KIR receptor genes have recently been found in a New World monkey.

Restoration of skeletal muscle ischemia-reperfusion injury in humanized immunodeficient mice

Ischemia and reperfusion (I/R) of tissue provokes an inflammatory process that is highly dependent on circulating natural immunoglobulin M (IgM) and the complement cascade. In mice, a single IgM specificity produced by peritoneal B cells can initiate reperfusion injury. It is unknown whether humans express natural IgM with a similar specificity. It is also unknown whether pathogenic IgM is produced solely from peritoneal B cells or can also be made by circulating B cells. Immunodeficient mice lacking endogenous immunoglobulin were used. Mice were reconstituted with 0.9% normal saline, human serum, or xenografted human peripheral blood lymphocytes (PBLs) and then subjected to tourniquet-induced hindlimb I/R. Serum human IgM and immunoglobulin G (IgG) were measured by enzyme-linked immunosorbent (ELISA) assay. Skeletal muscle was harvested for injury assessment by histology and for immunohistochemistry. Immunodeficient mice were protected from skeletal muscle injury after hindlimb I/R. Transfer of human serum restored skeletal muscle damage. Rag2/gammaR-/- mice that were engrafted with human PBL (huPBL-SCID) had high levels of human IgM. huPBL-SCID mice developed significantly more skeletal muscle injury than control saline-treated mice (P < or = .01) and human serum-reconstituted Rag2/gammaR-/- mice (P < or = 0.01). Sham-treated huPBL-SCID mice had no muscle injury, demonstrating that human lymphocyte engraftment did not cause injury in the absence of ischemia. Deposition of human IgM was observed on injured but not sham-injured muscle. Human serum can initiate murine skeletal muscle I/R injury. Circulating human PBL may be a source of pathogenic IgM. The huPBL-SCID mouse may be a useful model to define the specificity of pathogenic human IgM and to test therapeutics for I/R injury.

Modern Technologies for Creating Synthetic Antibodies for Clinical Application

The modular structure and versatility of antibodies enables one to modify natural immunoglobulins in different ways for various clinical applications. Rational design and molecular engineering make it possible to directionally modify the molecular size, affinity, specificity, and immunogenicity and effector functions of an antibody, as well as to combine them with other functional agents. This review focuses on up-to-date methods of antibody engineering for diagnosing and treating various diseases, particularly on new technologies meant to refine the effector functions of therapeutic antibodies.

Natural Antibody to Conserved Targets ofHaemophilus influenzaeLimits Colonization of the Murine Nasopharynx

Nasopharyngeal colonization represents the initial interaction between Haemophilus influenzae and its human host. Factors that influence bacterial carriage likely affect transmission and incidence of infection. Therefore, we investigated host factors involved in limiting H. influenzae colonization in BALB/c mice, as colonization can be established in this genetic background. Unlike what is observed in the C57BL/6 background, initial colonization of BALB/c mice was mainly limited by adaptive immune components. This effect on colonization did not require either CD4- or CD8-positive T cells. Instead, initial colonization by genetically diverse strains was limited by preexisting natural antibody with a lesser contribution of complement activity and the presence of neutrophils. Natural serum immunoglobulin from mice was able to bind to the bacterial surface and exhibited complement-dependent bactericidal activity against these genetically diverse H. influenzae strains. Moreover, natural immunoglobulin G (IgG) recognizing these strains was detected at the nasopharyngeal mucosal surface. This antibody-mediated effect required exposure to the normal mouse microbial flora, since mice raised under germfree (GF) conditions showed increased levels of H. influenzae colonization that were not limited by adaptive immunity. In addition, serum IgG from GF mice exhibited less surface binding to H. influenzae, suggesting that natural antibody, induced through prior exposure to the microbial flora, mediated the observed reduction in initial colonization. The broad effect of natural IgG against genetically diverse isolates suggests the presence of conserved species-wide protective targets of antibody.

Natural Human Immunoglobulins in Cancer Immunotherapy

The natural or innate immunity is the first-line defense against transformed cells. It guarantees the recognition and removal of malignant cells at an early stage and makes manifest cancers an exceptional event. Natural antibodies, which are predominantly IgM molecules, play a major role in these defense mechanisms and they have some typical features in common. They are coded by specific germline families and equipped mainly with lambda-chains, in contrast to the majority of circulating antibodies. The targets that are recognized by these antibodies are not newly synthesized proteins, but instead post-translationally modified carbohydrate structures on membrane-bound glycoproteins and glycolipids. Another typical feature of these natural IgM antibodies is their ability to induce apoptosis in vitro and in vivo in a death domain-independent manner. These results show that natural IgM antibodies represent a huge reservoir of therapeutic antibodies.

Ischemia-mediated aggregation of the actin cytoskeleton is one of the major initial events resulting in ischemia-reperfusion injury

Ischemia-reperfusion (IR) injury represents a major clinical challenge, which contributes to morbidity and mortality during surgery. The critical role of natural immunoglobulin M (IgM) and complement in tissue injury has been demonstrated. However, cellular mechanisms that result in the deposition of natural IgM and the activation of complement are still unclear. In this report, using a murine intestinal IR injury model, we demonstrated that the beta-actin protein in the small intestine was cleaved and actin filaments in the columnar epithelial cells were aggregated after a transient disruption during 30 min of ischemia. Ischemia also led to deposition of natural IgM and complement 3 (C3). A low dose of cytochalasin D, a depolymerization reagent of the actin cytoskeleton, attenuated this deposition and also attenuated intestinal tissue injury in a dose-dependent manner. In contrast, high doses of cytochalasin D failed to worsen the injury. These data indicate that ischemia-mediated aggregation of the actin cytoskeleton, rather than its disruption, results directly in the deposition of natural IgM and C3. We conclude that ischemia-mediated aggregation of the actin cytoskeleton leads to the deposition of natural IgM and the activation of complement, as well as tissue injury.

Clearance of Adenovirus by Kupffer Cells Is Mediated by Scavenger Receptors, Natural Antibodies, and Complement

Kupffer cells (KCs) rapidly remove intravenously injected adenovirus (Ad) vectors from the circulation. A better understanding of the mechanisms involved could suggest strategies to improve Ad gene delivery by suppressing or evading KC uptake. We recently showed that clearance of Ad type 5 vectors by KCs does not involve the interaction of Ad with the well-established Ad receptors, namely, integrins or the coxsackievirus and Ad receptor (J. S. Smith, Z. Xu, J. Tian, S. C. Stevenson, and A. P. Byrnes, Hum. Gene Ther. 19:547-554, 2008). In the current study, we systematically quantified the contributions of various receptors and plasma proteins to the clearance of Ad by KCs. We found that scavenger receptors are a predominant mechanism for the clearance of Ad by KCs. In addition, we found that Ad is opsonized by natural immunoglobulin M antibodies and complement and that these opsonins play a contributory role in the clearance of Ad by KCs. We also examined additional mechanisms that have been postulated to be involved in the clearance of Ad, including the binding of Ad to platelets and vitamin K-dependent coagulation factors, but we found that neither of these were required for the clearance of Ad by KCs.

Genetic interactions of KIR and G1M immunoglobulin allotypes differ in obese from non-obese individuals with type 2 diabetes

We analyzed the natural killer cell immunoglobulin-like receptor (KIR) genes and immunoglobulin allotypes in the development of type 2 diabetes (T2D) based on body mass index (BMI) measurements (obese vs. non-obese) in Puerto Rican Americans. Genetic interactions between the KIR haplotype A homozygotes (HAH) and its fraction containing two inhibitory receptors 2DL3 and 2DL1 and the activating receptor 2DS4 with immunoglobulin allotypes were studied. We found a significant association between the HAH and T2D ( p = 0.002; OR = 7.97) and its interaction with the immunoglobulin allotype z: GM f/f (−) ( p = <0.0001; OR, not determined) only in non-obese individuals. This association were due to the interactions between the 2DL3/2DL3, 2DL1/2DL1, and 2DS4 fragment with GM f/f (−) in T2D patients ( p = 0.0017; OR = 3.45). Analysis based on BMI demonstrated associations in both obese ( p = 0.037; OR = 2.43; 95% CI = 0.97–6.31) and non-obese individuals ( p = <0.0001; OR = 8.38; 95% CI = 2.49–29.31). By contrast, the interaction of the GM allotype f/f (−) with the HAH fragment was associated with T2D only in non-obese individuals ( p = <0.0001; OR = 18.2; 95% CI = 3.71–113.4). As expected, interaction of both HAH and its fragment with HLA-C group's ligands were significant. We used informative short tandem repeats (STRs) that distinguish major populations to determine genetic admixture and found that there was no genetic stratification in our cohort. Our findings are consistent with the possibility of an autoimmune and/or innateimmune component in the pathogenesis of T2D: NK receptors with chronic inflammation in obese and genetic interactions with G1M allotype in T2D non-obese possibly mediating autoimmunity.

Inhibition of HIV‐1 transmission in trans from dendritic cells to CD4+ T lymphocytes by natural antibodies to the CRD domain of DC‐SIGN purified from breast milk and intravenous immunoglobulins

The present study demonstrates that human breast milk and normal human polyclonal immunoglobulins purified from plasma [intravenous immunoglobulins (IVIg)] contain functional natural immunoglobulin A (IgA) and IgG antibodies directed against the carbohydrate recognition domain (CRD) domain of the dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) molecule, which is involved in the binding of human immunodeficiency virus (HIV)-1 to dendritic cells (DCs). Antibodies to DC-SIGN CRD were affinity-purified on a matrix to which a synthetic peptide corresponding to the N-terminal CRD domain (amino-acid 342-amino-acid 371) had been coupled. The affinity-purified antibodies bound to the DC-SIGN peptide and to the native DC-SIGN molecule expressed by HeLa DC-SIGN+ cells and immature monocyte-derived dendritic cells (iMDDCs), in a specific and dose-dependent manner. At an optimal dose of 200 microg/ml, natural antibodies to DC-SIGN CRD peptide purified from breast milk and IVIg stained 25 and 20% of HeLa DC-SIGN+ cells and 32 and 12% of iMDDCs, respectively. Anti-DC-SIGN CRD peptide antibodies inhibited the attachment of virus to HeLa DC-SIGN by up to 78% and the attachment to iMDDCs by only 20%. Both breast milk- and IVIg-derived natural antibodies to the CRD peptide inhibited 60% of the transmission in trans of HIV-1(JRCSF), an R5-tropic strain, from iMDDCs to CD4+ T lymphocytes. Taken together, these observations suggest that the attachment of HIV to DCs and transmission in trans to autologous CD4+ T lymphocytes occur through two independent mechanisms. Our data support a role of natural antibodies to DC-SIGN in the modulation of postnatal HIV transmission through breast-feeding and in the natural host defence against HIV-1 in infected individuals.

Medical management of Acute Radiation Syndromes:Immunotherapy by Antiradiation Immunoglobulin G .

Immunoglobulins is an important part of Acquired Immunity and participate in such important immunological processes as recognition, regulation and elimination of foreign antigens. At the present time, Intravenous Immunoglobulins are used for an efficient therapy for immune deficiency syndromes, thrombocytopenias, inflammatory reactions, modulation of autoimmunity and a wide range of hematologic disorders. Traditionally, the treatment of Acute Radiation Syndromes (ARS) includes supportive therapy, cytokine therapy, blood component transfusions and stem cell transplantation. However, results of treatment of ARS remain limited and in cases of severe radiation injury insufficient. Studies of therapy effects of Anti-radiation Immunoglobulin G in vivo have established that specific antibodies to Radiation Toxins of SRD group can be important, effective part of medical management of ARS and can play a significant role in neutralization of radiation induced toxicity. Multiple-organ failure at Acute Radiation Syndromes is a major cause of mortality after high doses of gamma irradiation. Radiation Toxins of Specific Radiation Deteminant group(neuro-toxic, neuro-vascular-toxic, entero-toxic, hemato-toxic) play an important role in development of Acute Radiation Syndromes and development of multi-organ involvement and multi-organ failure. Radiation Toxins possess high toxic properties. Radiation Neurotoxin isolated from lymphatic system of irradiated animals (with a clinical picture of cerebral radiation syndrome) and injected to healthy animals in toxic doses 0.03 mg/kg, 0.5 mg/kg, 10.0 mg/kg, 15.0 mg/kg had initiated development of acute failure of blood circulation and breathing ventilation. Death of laboratory animals had occurred within 5 min-3 days after injection of toxic doses of Radiation Neuro-Toxin and depended on a concentration and a type of active substance of Radiation Toxins. Hyper-immunization of non-irradiated animals by non-toxic doses of Radiation Toxins were provided. The immunoglobulin fraction of pooled hyperimmune anti-radiation plasma was separated. Immunoglobulines to Radiation Toxins were used for a treatment of Acute Radiation Syndromes and the efficacy of this bio-pharmaceutical agent was initially evaluated. Therapeutic application of Specific Anti-radiation Immunoglobulin had significantly diminished mortality rate at Acute Radiation Syndromes and was much more effective compare with natural immunoglobulins preparations and irradiated forms of natural irradiated immunoglobulins.

C-reactive protein and natural IgM antibodies are activators of complement in a rat model of intestinal ischemia and reperfusion

The role of C-reactive protein (CRP), natural immunoglobulin M (IgM), and natural IgM against phosphorylcholine (anti-Pc IgM) was investigated in relation with complement activation in a rat model of intestinal ischemia and reperfusion (II/R). The effect of C1-esterase inhibitor (C1-Inh) on this complement activation along with other inflammatory mediators was also studied. Rats were subjected to 1 h of superior mesenteric artery occlusion and 3 h of reperfusion. Intravenous administration of vehicle (human albumin) or C1-Inh (200 U/kg) was performed before (n = 8) or after ischemia (n = 8). II/R increased levels of C4b/c, CRP, IgM, anti-Pc IgM, and myeloperoxidase activity in the intestinal homogenates and induced vascular leakage. A good correlation was observed in the intestinal homogenates between C4b/c and CRP levels. Clear depositions of C3, CRP, and IgM in intestinal tissue were demonstrated after II/R, and a strong correlation of both CRP and IgM with complement was observed. C1-Inh administered before ischemia reduced the complement activation response after II/R, as reflected by decreased levels of C4b/c in conjunction with reduced anti-Pc IgM in the intestinal homogenates. C1-Inh also decreased leakage of albumin when administered before ischemia. C1-Inh after ischemia reduced C4b/c levels and myeloperoxidase activity in the homogenates. CRP and IgM depositions correlated well with local complement activation, which suggests a role of these molecules in complement activation. Furthermore, C1-Inh inhibited potentially II/R injury either administered before or after ischemia, by attenuating complement activation induced by CRP and/or natural IgM antibodies.

Natural antibody and complement mediate neutralization of influenza virus in the absence of prior immunity.

Early control of virus replication by the innate immune response is essential to allow time for the generation of a more effective adaptive immune response. As an important component of innate immunity, complement has been shown to be necessary for protection against numerous microbial infections. This study was undertaken to investigate the role of complement in neutralizing influenza virus. Results demonstrated that the classical pathway of complement mediated serum neutralization of influenza virus. Although nonimmune serum neutralized influenza virus, the mechanism of virus neutralization (VN) required antibody, as sera from RAG1-deficient mice lacked VN activity; moreover, purified natural immunoglobulin M (IgM) restored VN activity to antibody-deficient sera. The mechanism of VN by natural IgM and complement was associated with virion aggregation and coating of the viral hemagglutinin receptor; however, viral lysis did not significantly contribute to VN. Additionally, reconstitution of RAG1-deficient mice with natural IgM resulted in delayed morbidity during influenza virus infection. Collectively, these results provide evidence that natural IgM and the early components of the classical pathway of complement work in concert to neutralize influenza virus and that this interaction may have a significant impact on the course of influenza viral pneumonia.

Immune responses of piglets to weaning stress: Impacts of photoperiod1

An acute stress response can be provoked by abrupt social, nutritional, and environmental changes associated with weaning, and this may disrupt homeostasis and thus compromise well-being. Manipulating environmental factors, such as photoperiod, might provide a simple way to reduce the physiological consequences that piglets experience due to weaning stress. The objective of this study was to evaluate the impacts of photoperiod manipulation across various weaning ages on leukocyte populations, lymphocyte proliferation, natural killer cytotoxicity (NK), chemotaxis, phagocytosis, and immunoglobulin G, cortisol, and BW of piglets during the nursery phase. Sixty-eight crossbred piglets were obtained from sows kept on a short-day (8 h of light/d) photoperiod from d 90 of gestation until weaning. Piglets were weaned at 14, 21, or 28 d of age and kept on a short or long (16 h of light/d) photoperiod until 10 wk of age. Piglet BW and blood samples were collected at weaning and at 6, 8, and 10 wk of age. Pigs weaned at 28 d had reduced neutrophil counts (P < 0.001), phagocytosis (P < 0.001), and lymphocyte proliferation (P < 0.05) at weaning compared with those weaned at 14 and 21 d. Pigs weaned at 21 d tended to have lower (P = 0.08) lymphocyte counts than did pigs weaned at 14 or 28 d. Pigs weaned at 14 d had reduced (P < 0.01) NK relative to those weaned at 21 or 28 d. Photoperiod also influenced pig BW and immune status. Generally, those pigs on the long-day photoperiod and weaned at 28 d were heavier (P < 0.001) than their counterparts weaned at 14 or 21 d. At 6 wk of age, NK was greater (P = 0.002) in pigs kept on a long day and weaned at 14 or 21 d than in pigs weaned at 28 d. Phagocytosis was less (P = 0.005) at 6 wk of age, but was greater at 8 wk, in piglets kept on the long day and weaned at 28 d than in long-day pigs weaned at 14 or 21 d. These results suggest that photoperiod differentially influences immune responses in piglets weaned at different ages and indicate an inverse relationship between growth and immune status. Here, weaning at 28 d and a long-day photoperiod was the treatment combination that was most physiologically beneficial to piglets, whereas a 14-d weaning and short-day photoperiod was least physiologically beneficial.

Nutritional modulation of exercise-induced immunodepression in athletes: a systematic review and meta-analysis

Heavy exercise induces marked immunodepression that is multifactorial in origin. Nutrition can modulate normal immune function. To assess the efficacy of nutritional supplements in exercise-induced immunodepression in athletes. Systematic review. Randomised and/or controlled trials of athletes undertaking nutritional supplements to minimise the immunodepression after exercise were retrieved. The primary outcome measure was incidence of upper respiratory tract (URT) illness symptoms after exercise, and secondary outcomes included cortisol, cell counts, plasma cytokine concentration, cell proliferative response, oxidative burst, natural killer cell activity and immunoglobulins. When data were available for a pooled estimate of the effect of intervention, meta-analyses were conducted for direct comparisons. Forty-five studies were included (1603 subjects). The studies were heterogeneous in terms of exercise interventions, selection of athletes, settings and outcomes. The overall methodological quality of most of the trials was poor. Twenty studies addressed carbohydrate supplementation, eight glutamine, 13 vitamin C and four others interventions. Three trials assessed the effect of intervention on prevention of URT infections. The pooled rate ratio for URT infections after vitamin C supplementation against placebo was 0.49 (0.34-0.71). Carbohydrate supplementation attenuated the increase in cortisol and neutrophils after exercise; vitamin C attenuated the decrease in lymphocytes after exercise. No other interventions had significant or consistent effect on any of the studied outcomes. Although the prevention of URT infections by vitamin C was supported by two trials, further studies are needed. The available evidence failed to support a role for other nutritional supplements in preventing exercise-induced immune suppression. Larger trials with clinically relevant and uniform end points are necessary to clarify the role of these nutritional interventions.

A study of the xenoantigenicity of neonatal porcine islet‐like cell clusters (NPCC) and the efficiency of adenovirus‐mediated DAF (CD55) expression

The pig pancreas is considered to be the most suitable source of islets for xenotransplantation in patients with type I diabetes. The objective of this study was to assess the antigenicity of neonatal porcine islet-like cell clusters (NPCC), including the Galalpha1-3Galbeta1-4GlcNAc-R (alpha-Gal) and Hanganutziu-Deicher (H-D) antigens, and the pathway involved in human complement activation. The efficiency of expression of human decay-accelerating factor (DAF: CD55) on NPCC by adenoviral transduction was also examined, and the functional capacity of DAF was also estimated. The deposition of human natural antibodies, immunoglobulin (Ig)G and IgM, and the expression of alpha-Gal and H-D antigens on NPCC were investigated by FACS analysis. The downregulation in the antigenicity to human natural antibodies, including the alpha-Gal and H-D antigens on NPCC by treatment with tunicamycin, PDMP and neuraminidase were also examined. In addition, complement-mediated islet lysis was examined using factor D-deficient and C1-deficient sera. An adenovirus encoding DAF under the control of the cytomegalovirus promoter, Ad.pCMV-DAF, was then constructed, and used for transducing NPCC. The amelioration of complement-dependent cytotoxicity of the NPCC by the transduced DAF was assessed as an in vitro hyperacute rejection model of a pig to human xenograft. The NPCC clearly expressed the alpha-Gal epitope, and the human natural antibodies, IgG and IgM, and the anti-H-D antibody also reacted with the NPCC. Treatment of NPCC with tunicamycin led to a drastic reduction in the extent of deposition of IgG, indicating the importance of N-linked sugars on the islets, presumably related to alpha-Gal expression on N-linked sugars. Neuraminidase treatment indicated the presence of, not only the H-D antigen, but also other sialic acid antigens which reacted with the human natural antibody, especially IgG. The complement deposition of factor B on NPCC was clear, and the alternative pathway-mediated NPCC killing accounted for approximately 30% of that by the total complement pathway. On the other hand, approximately 90% of the NPCC could be transduced to express DAF by the adenovector, Ad.pCMV-DAF. The expressed DAF showed an approximately 50-62% suppression in complement-dependent NPCC lysis. The origin of the antigenicity of NPCC is mainly N-linked sugars including alpha-Gal and sialic acid antigens, and NPCC expressed the transduced molecule in high efficiency by the adenovector.

Herpes Simplex Virus Type 1 and 2 Glycoprotein C Prevents Complement-Mediated Neutralization Induced by Natural Immunoglobulin M Antibody

Glycoprotein C (gC) of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) binds complement component C3b and protects virus from complement-mediated neutralization. Differences in complement interacting domains exist between gC of HSV-1 (gC1) and HSV-2 (gC2), since the amino terminus of gC1 blocks complement C5 from binding to C3b, while gC2 fails to interfere with this activity. We previously reported that neutralization of HSV-1 gC-null virus by HSV antibody-negative human serum requires activation of C5 but not of downstream components of the classical complement pathway. In this report, we evaluated whether activation of C5 is sufficient to neutralize HSV-2 gC-null virus, or whether formation of the membrane attack complex by C6 to C9 is required for neutralization. We found that activation of the classical complement pathway up to C5 was sufficient to neutralize HSV-2 gC-null virus by HSV antibody-negative human serum. We evaluated the mechanisms by which complement activation occurred in seronegative human serum. Interestingly, natural immunoglobulin M antibodies bound to virus, which triggered activation of C1q and the classical complement pathway. HSV antibody-negative sera obtained from four individuals differed over an approximately 10-fold range in their potency for complement-mediated virus neutralization. These findings indicate that humans differ in the ability of their innate immune systems to neutralize HSV-1 or HSV-2 gC-null virus and that a critical function of gC1 and gC2 is to prevent C5 activation.