Effect of visfatin on the structure and immune levels in the small intestine of LPS-induced rats / Efecto de la visfatina sobre la estructura y los niveles de inmunidad en el intestino delgado de ratas inducidas por lipopolisacáridos

This study investigated the effects of visfatin on the structure and the immunity levels in the small intestine of LPS-induced rats. Forty Wistar male and female SPF rats were randomly and equally divided into four groups: the saline (control), vistfatin, lipopolysaccharide (LPS), and visfatin+LPS co-stimulated. The functions of visfatin in the intestinal mucosal immunity were investigated by examining the variation of tissue structure, inflammation and immunity-related proteins in the intestine of immunologically stressed rats using HE staining, ELISA, immunohistochemistry and Western Blot. The results showed that, when compared with the control group, the visfatin-treated group showed a decrease in the intestinal villus height and width, and a significant increase in the levels of IL-6 and TNF-ð as well as Immunoglobulin A (IgA) positive cells. Additionally, when compared with the LPS-treated group, the visfatin+LPS co-stimulated group showed a decrease in the villus height and width as well as the levels of IL-6 and TNF-ð, and an increase in IgA levels, implying a shrinking response to LPS injection. All the results suggest that, under normal physiological conditions, visfatin disturbs the body's homeostasis and causes intestinal villus atrophy by increasing IgA expression. While under immune response conditions, LPS acts as an exogenous antigen to promote visfatin against LPS-induced inflammation by decreasing the expression of IgA. Under immune stress conditions, visfatin as an exogenous stimulus promotes the immune response by regulating the protein levels of IL-6, TNF-ð and IgA.

Este estudio investigó los efectos de la visfatina sobre la estructura y los niveles de inmunidad en el intestino delgado de ratas inducidas por lipopolisacáridos (LPS). Cuarenta ratas Wistar se dividieron aleatoriamente e igualmente en cuatro grupos: solución salina (control), vistafin, LPS y visfatina + LPS co-estimuladas. Las funciones de la visfatina en la inmunidad de la mucosa intestinal se investigaron mediante el examen de variación de la estructura del tejido, la inflamación y las proteínas relacionadas con la inmunidad en el intestino de ratas estresadas inmunológicamente; usando tinción HE, ELISA, inmunohistoquímica y Western Blot. Los resultados mostraron que, en comparación con el grupo control, el grupo tratado con visfatina presentó una disminución en la altura y ancho de las vellosidades intestinales, y un aumento significativo en los niveles de IL-6 y TNF-ð, así como inmunoglobulina A (IgA células positivas). Además, al comparar este grupo con el grupo tratado con LPS- el grupo visfatina + LPS co-estimulado mostró una disminución en la altura y ancho de las vellosidades, así como en los niveles de IL-6 y TNF-ð, y un aumento en los niveles de IgA, lo que implica reducción de una respuesta a la inyección LPS. Todos los resultados sugieren que, en condiciones fisiológicas normales, la visfatina perturba la homeostasis del cuerpo y provoca la atrofia de las vellosidades intestinales mediante el aumento de la expresión de IgA. Mientras que bajo condiciones de la respuesta inmune, LPS actúa como un antígeno exógeno para promover visfatina contra la inflamación inducida por LPS por la disminución de la expresión de IgA. En condiciones de estrés inmunológico, la visfatina como estímulo exógeno promueve la respuesta inmune mediante la regulación de los niveles de proteína de IL-6, TNF-ð e IgA.

Modulation of protective immunity against herpes simplex virus via mucosal genetic co-transfer of DNA vaccine with beta2-adrenergic agonist

Cholera toxin, which has been frequently used as mucosal adjuvant, leads to an irreversible activation of adenylyl cyclase, thereby accumulating cAMP in target cells. Here, it was assumed that beta2-adrenergic agonist salbutamol may have modulatory functions of immunity induced by DNA vaccine, since beta2-adrenergic agonists induce a temporary cAMP accumulation. To test this assumption, the present study evaluated the modulatory functions of salbutamol co-administered with DNA vaccine expressing gB of herpes simplex virus (HSV) via intranasal (i.n.) route. We found that the i.n. co-administration of salbutamol enhanced gB-specific IgG and IgA responses in both systemic and mucosal tissues, but optimal dosages of co-administered salbutamol were required to induce maximal immune responses. Moreover, the mucosal co-delivery of salbutamol with HSV DNA vaccine induced Th2-biased immunity against HSV antigen, as evidenced by IgG isotypes and Th1/Th2-type cytokine production. The enhanced immune responses caused by co-administration of salbutamol provided effective and rapid responses to HSV mucosal challenge, thereby conferring prolonged survival and reduced inflammation against viral infection. Therefore, these results suggest that salbutamol may be an attractive adjuvant for mucosal genetic transfer of DNA vaccine.

Adjuvant requirement for successful immunization with recombinant derivatives of Plasmodium vivax merozoite surface protein-1 delivered via the intranasal route

Recently, we generated two bacterial recombinant proteins expressing 89 amino acids of the C-terminal domain of the Plasmodium vivax merozoite surface protein-1 and the hexa-histidine tag (His6MSP1(19)). One of these recombinant proteins contained also the amino acid sequence of the universal pan allelic T-cell epitope (His6MSP1(19)-PADRE). In the present study, we evaluated the immunogenic properties of these antigens when administered via the intra-nasal route in the presence of distinct adjuvant formulations. We found that C57BL/6 mice immunized with either recombinant proteins in the presence of the adjuvants cholera toxin (CT) or the Escherichia coli heat labile toxin (LT) developed high and long lasting titers of specific serum antibodies. The induced immune responses reached maximum levels after three immunizing doses with a prevailing IgG1 subclass response. In contrast, mice immunized by intranasal route with His6MSP1(19)-PADRE in the presence of the synthetic oligonucleotides adjuvant CpG ODN 1826 developed lower antibody titers but when combined to CT, CpG addition resulted in enhanced IgG responses characterized by lower IgG1 levels. Considering the limitations of antigens formulations that can be used in humans, mucosal adjuvants can be a reliable alternative for the development of new strategies of immunization using recombinant proteins of P. vivax.

Spontaneous inflammatory pelvic disease in adult non-castrated female rats treated with estrogen

The adaptive immune response of the genital tract is under the control of sexual steroids; however, the influence of sex hormones on innate immune mechanisms of the genital mucosa are only beginning to be understood. We found that long-term estrogen treatment increases the risk for inflammatory pelvic diseases in adult non-castrated female rats. Female rats (110 g to 130 g) received estrogen (10 rats; 17-beta estradiol, 50 mg pellet; 10 rats: subcutaneous weekly injection of estradiol valerate 0.166 mg/kg). Ten rats received a pellet of 17-beta estradiol and were treated with amoxicillin, 50 mg/kg after the 90th day of exposure to estrogen. Three control groups of ten rats were also used. The estrogen-treated rats developed an inflammatory pelvic disease, with abscess formation after the third month of hormonal treatment. All the surviving animals were killed after six months of hormonal exposure. Among 15 survivors of the two groups that received estrogen 13 animals presented tuboovarian abscesses. Among eight survivors of the group treated with amoxicillin, six had tuboovarian abscesses. None of the 30 control rats presented macro or microscopic signs of inflammatory disease in the uterus, tubes or ovaries. We conclude that estrogen impairs the defense mechanisms of the genital tract of non-castrated female rats, enhancing bacterial growth in the vagina and ascending infection to the uterus, tubes and ovaries.

Iron chelator inducesMIP-3alpha/CCL20 in human intestinal epithelial cells: implication for triggeringmucosal adaptive immunity

A previous report by this laboratory demonstrated that bacterial iron chelator (siderophore) triggers inflammatory signals, including the production of CXC chemokine IL-8, in human intestinal epithelial cells (IECs). Microarray-based gene expression profiling revealed that iron chelator also induces macrophage inflammatory protein 3 alpha (MIP-3alpha)/ CC chemokine-ligand 20 (CCL20). As CCL20 is chemotactic for the cells involved in host adaptive immunity, this suggests that iron chelator may stimulate IECs to have the capacity to link mucosal innate and adaptive immunity. The basal medium from iron chelator deferoxamine (DFO)-treated HT-29 monolayers was as chemotactic as recombinant human CCL20 at equivalent concentrations to attract CCR6+ cells. The increase of CCL20 protein secretion appeared to correspond to that of CCL20 mRNA levels, as determined by real-time quantitative RT-PCR. The efficacy of DFO at inducing CCL20 mRNA was also observed in human PBMCs and in THP-1 cells, but not in human umbilical vein endothelial cells. Interestingly, unlike other proinflammatory cytokines, such as TNF-alpha and IL-1beta, a time-dependent experiment revealed that DFO slowly induces CCL20, suggesting a novel mechanism of action. A pharmacologic study also revealed that multiple signaling pathways are differentially involved in CCL20 production by DFO, while some of those pathways are not involved in TNF-alpha-induced CCL20 production. Collectively, these results demonstrate that, in addition to some bacterial products known to induce host adaptive immune responses, direct chelation of host iron by infected bacteria may also contribute to the initiation of host adaptive immunity in the intestinal mucosa.