Yael Harari

Reconstruction of immune-mediated ion secretion in gut mucosa of the athymic rat

Summary A defect in an immune‐mediated, physiological reaction in the small intestine of congenitally immunosuppressed rats has been identified and corrected by recouping the immunological trigger in the mucosa with epithelial effector cells. Antigenic challenge of jejunum from euthymic rats sensitized by infection with Trichinelta spiralis evoked an anaphylactic response in vitro that was characterized by an elevation in transmural short‐circuit current (Isc) and caused by net Cl‐ secretion. In contrast, jejunum from previously infected athymic counterparts failed to respond electrophysiotogically to antigenic stimulation. Passive immunization of athymic rats with anti‐Trichinella serum restored jejunal responsiveness to antigen. Jejunal unresponsiveness to anligcnic challenge in the athymic rat compared with the euthymic rat was not due to a difference in the duration of exposure to the sensitizing antigens, mast cell numbers, inflammation‐related hislological changes nor to reduced responsiveness of epithelium to Cl‐ sccretagogues. Because the transduction of the antigenic signal into epithelial ion transport changes in euthymic rais requires IgE, mucosal mast cells, mast cell‐derived mediators and enteric nerves, the results support the conclusion that an antibody deficiency is the only functional defect preventing the antigen‐induced change in mucosal ion transport in the athymic host.

Intestinal anaphylaxis in chickens: epithelial ion secretion as a determinant and potential component of functional immunity.

Immunity to secondary protozoan infections in chickens is accompanied by rapid onset of intestinal permeability to serum proteins, an event in mammals associated with local anaphylaxis. The permeability changes in the chicken intestine are hypothesized to be mediated by mast cell-derived paracrine factors. In a test of this hypothesis, we demonstrated, using an electrophysiological correlate of intestinal anaphylaxis (antigen-induced Cl(-) secretion), that the response of the chicken intestine to antigenic stimulation is consistent with type I hypersensitivity reactions. Day-old, single-comb-white-Leghorn chickens were sensitized to bovine serum albumin (BSA). At 3 weeks of age ileal segments were mounted in Ussing-type chambers. Serosal challenge with BSA elevated the transmural short circuit current (DeltaIsc) within 1min and was maximally expressed (DeltaIsc=50-60microA/cm(2)) within 2-3 min. The magnitude of the DeltaIsc was directly related to the concentration of antigen (10-200 microg antigen/ml), was only expressed in immunized chickens, and was blocked by the mucosal application of a Cl(-) channel blocker. Data obtained in the present investigation identify epithelial ion secretion as a potential mechanism of functional immunity in the mucosal immune system of the chicken small intestine.

Sialic acid deficiency in lectin-resistant intestinal brush border membranes from rats following the intestinal phase of trichinellosis

Maximal binding (Bmax) of the lectin, wheat germ agglutinin, by small intestinal brush border membrane is significantly reduced in rats infected with Trichinella spiralis. Wheat germ agglutinin specificity is for N-acetylglucosamine and sialic acid. Whereas total hexosamine and N-acetylglucosaminidase-labile N-acetylglucosamine were comparable in membranes from uninfected as compared with infected rats, the total sialic acid content and neuraminidase-released sialic acid were significantly higher in BBM from uninfected hosts. N-Acetylglucosaminidase treatment of membranes reduced Bmax for wheat germ agglutinin in both hosts. Neuraminidase treatment reduced Bmax in uninfected hosts, but tended to increase it in infected rats. Membranes from uninfected rats incorporated more N-acetylglucosamine from UDP-N-[14C]acetylglucosamine into oligosaccharide-lipid than did membranes from infected hosts. However, lipid and protein fractions were labeled at the same rate in both sets of membranes. Sialic acid was incorporated into protein at a slightly faster rate in brush border membrane from uninfected rats, indicative of a higher level of sialotransferase activity. These results suggest that the reduction in Bmax for wheat germ agglutinin in gut epithelial cell membranes from infected rats is related to a reduced level of sialic acid available for lectin binding as well as specific interactions between N-acetylglucosamine and sialic acid.

Biliary and gut function following shock

The aim of this study was to characterize the alterations in gallbladder and intestinal function after hemorrhagic shock and blood reperfusion in opossums. Animals were subjected to a shock of 30 mm Hg of arterial blood pressure for 60 minutes and resuscitated with blood reinfusion. Gallbladder epithelial ion transport, gallbladder motility in vitro and in vivo, gastrointestinal motility, and flora of the stomach and small bowel were studied 2 and 24 hours after shock. Changes at 2 hours included decreased gallbladder contractility in vitro and decreased emptying in vivo, loss of coordination with intestinal motor activity, decrease in frequency of intestinal electrical slow waves, and reduced duration of the intestinal migrating motor complex cycle. By 24 hours, gallbladder epithelial permeability was increased and in vitro contractility remained reduced but the in vivo functions showed partial recovery. Gastrointestinal flora was not affected by these changes. These data demonstrate that hemorrhagic shock and reperfusion affect digestive motility. The early timing of the alterations observed and the partial recovery 24 hours post shock suggest an ischemia-hypoxia mechanism of injury.

Fish oil supplementation does not impair the gut immune response to Trichinella spiralis infection in rats.

BACKGROUND Fish oil has been recommended as a source of omega-3 fatty acids for preterm infants and for therapy of some inflammatory diseases. METHODS Because fish oil supplementation could downregulate the hosts immune response, we studied the gut inflammatory response to an enteric infection in 72 rats assigned to three dietary groups with differing fatty acid profile: 1) fish oil, rich in eicosapentaenoic and docosahexaenoic acid; 2) olive oil, containing 71% monounsaturated fat; and 3) rat chow, containing 57% saturated fat. One half (n = 36) of the rats were infected with Trichinella spiralis larvae; the other half served as controls. The inflammatory response to initial infection (study 1), and type I hypersensitivity response to a subsequent parasite-derived antigenic challenge (study 2) were assessed. Jejunal inflammatory cell infiltrate, mean villus height, disaccharidase levels, changes in short-circuit current in response to glucose absorption, and chloride secretagogues (study 1) were measured 9 days after infection. Short-circuit current changes induced by chloride secretion were measured when the proximal jejunum was challenged with T. spiralis-derived antigen 40 days after infection (study 2). RESULTS In study 1, jejunal tissue from infected animals had more eosinophilic infiltrate, lower disaccharidase levels, and less glucose absorptive and chloride secretory capacity than tissue from noninfected animals. In study 2, the jejunum of infected animals showed an antigen-induced chloride secretory response, whereas no response was obtained from jejunal tissue from noninfected animals. Type of diet did not affect the response in either study. CONCLUSION Under the conditions of this experiment, fish oil supplementation did not interfere with the local intestinal inflammatory response after T. spiralis infection.

Phosphatidylethanolamine methylation in intestinal brush border membranes from rats resistant to Trichinella spiralis

Methylation of phospholipids is proposed as a mechanism to explain changes in properties of intestinal brush border membrane that coincide with development of immunity to the intraepithelial parasite, Trichinella spiralis. Methylation was measured by the incorporation of the [3H]methyl group from S-adenosyl-L-[3H]methyl methionine into phospholipids. At least two enzymatic components were detected that converted phosphatidylethanolamine to phosphatidylcholine. The first, designated methyltransferase I, catalyzed the formation of phosphatidylmonomethylethanolamine from phosphatidylethanolamine and had a low Km for S-adenosyl-L-methyl-methionine (5 microM). The second, designated methyltransferase II, which catalyzed the methylation of phosphatidylmonomethylethanolamine to phosphatidyldimethylethanolamine and phosphatidyldimethylethanolamine to phosphatidylcholine, had a high Km for S-adenosyl-L-methyl methionine (167 microM). Both enzymes had two pH optima, were most active at 37 degrees C and were Mg2+ dependent. A decrease in methylation activity was present in brush border membranes from rats immunized against T. spiralis. Although the synthesis of phosphatidylcholine was not significantly altered there was a substantial decrease in the formation of phosphatidylmonomethylethanolamine and phosphatidyldimethylethanolamine as compared with nonimmunized rats. Since phospholipid composition influences membrane fluidity and cell function, it is proposed that altered methylation activity may influence the characteristics of brush border membrane in the immune host.

EVALUATION OF A POSSIBLE FUNCTIONAL RELATIONSHIP BETWEEN CHEMICAL STRUCTURE OF INTESTINAL BRUSH BORDER AND IMMUNITY TO TRICHINELLA SPIRALIS IN THE RAT

Primary exposure to Trichinella spiralis in the rat, while immunizing against reinfection, induces changes in the carbohydrate structure of intestinal brush border membranes. Immunity is expressed in heightened resistance to mucosal invasion by L1 larvae, and the change in structure is evident in reduced membrane binding of the lectin, wheat germ agglutinin. The possibility that altered membrane composition is a requisite for expression of immunity was hypothesized and this was evaluated by correlating the maximum, specific binding of wheat germ agglutinin by isolated brush border membranes with (1) the expression of immunity acquired passively through serum transfer, and (2) the loss of immunity acquired from serial infections terminated in the intestinal phase. The hypothesis was further evaluated by determining whether the change in membrane structure represents a stimulus-specific response. We observed that (1) passively acquired immunity was not associated with a reduction in lectin binding and (2) short-term exposure to the intestinal stages of T. spiralis led to a reduction in lectin binding that was detectable at a time when rats were incapable of resisting reinfection. The change in lectin binding associated with trichinosis also accompanied infection with Nippostrongylus brasiliensis. Results uniformly support the conclusion that immunity to T. spiralis is independent of brush border membrane changes reflected in reduced binding of wheat germ agglutinin.

Immunological sensitization of opossum gallbladder by naturally acquired stomach roundworm infection

1. The opossum gallbladder develops an immediate type hypersensitivity as a result of chronic infection with the gastric nematode, Physaloptera turgida. 2. Hypersensitization is evident from a change in net transepithelial ion transport in response to parasite-derived antigen. 3. The antigen-induced change in ion transport involves mediation by histamine and possibly prostaglandins and is independent of intrinsic neural modulation. 4. The functions of the gallbladder epithelium are influenced by reactions of local immune elements that are part of the common mucosal immune system.