Effects On Mouse CD4~+T Cells By Methionine Enkephalin Alone Or In Combination With Interleukin-2 Or Interferon-γ

IntroductionMethionine enkephalin (MENK), the endogenous neuropeptide, is derivatized by prehormone and proenkephalin which is producted by adrenal,is suggested to be involved in the regulatory loop between immune and neuroendocrine systems.It is proposed that this molecule be classified as an important cytokine or opioid growth factor. Thus MENK appears to be an important immunomodulatory signaling molecule to exert regulatory actions concerned with the expressing of receptors. The classical opioid growth factor receptor(OGFr) containedμ(mu),δ(delta),κ(kappa).Immune widespread MENK cell-surface receptor, MENK with the body's opioid receptors, regulate the immune system function. Studies have shown that MENK alone and in combination with Interleukin-2 or Interferon-y could increase the function of immune,but the mechanism was not clear.This investigation measured the effects on the opioid growth factor receptors (OGFr) on CD4+T cells by methionine enkephalin(MENK) alone and in combination with interleukin-2 or interferon-γin vivo and vitro,included the number of CD4+T cells,the expression of opioid growth factor receptors and cytokine levels of supernatants which to explore the immunological effects of MENK,the findings may provide meaningful theoretical and experimental basis for the clinical application.MaterialsAnimalMale C57BL/6 mice 6-8 weeks (weighing 18-20g) were purchased from the Shanghai SIPPR/BK Experimental Animal Co,Ltd.Methods 1. Primary cell culture,treatment and Magnetic bead enriched CD4+T cells.2. Flow cytometry detected the number of CD4+T cells with different treatment.3. RT-PCR analysis for expressions of three OGFr mRNA from spleen CD4+T Cells of mice.4. ELISA detected the levels of IL-2 and IFN-y in CD4+T cells.Results1. Flow cytometryThe number of CD4+T cells in all experimental groups in vivo and in vitro, compared with the control group, were significantly increased (P<0.05). MENK+IL-2 group compared with MENK group,as well as MENK+IFN-γgroup compared with MENK group,the number of CD4+T cells were significantly increased (P＜0.05) in vivo, however, the comparison between groups was no significant increase in vitro (P> 0.05).2. RT-PCR confirmationThe expression of three kinds opioid receptorμ,δ,κmRNA were detected in spleen CD4+T cells of mice, and each group compared with control group,μandδOGFr were significantly (P<0.05),KOGFr was no significant increase (P> 0.05) in vivo and in vitro. MENK+IL-2 group compared with MENK group,as well as MENK+ IFN-y group compared with MENK group, the expression ofμ,δOGFr mRNA were significantly (P＜0.05) in vivo. However,both of them were no significant increase (P> 0.05) in vitro.3. ELISAIL-2, IFN-y levels of CD4+T cells supernatants:in vivo, the treated group compared with the control and the comparison group were significantly (P＜0.05); while in vitro, MENK combined IL-2 or IFN-y group compared with MENK group and IFN-y or IL-2 group were no significant (P> 0.05).ConclutionMENK alone or combined with IL-2 or IFN-y could increase the expression ofμ,δopioid growth factor receptors and the levels of cytokines in culture supernatants significantly in vivo;while MENK combined with IL-2 or IFN-y had no significant effect in vitro.