Selenium Supplementation Alleviates Autoimmune Thyroiditis by Regulating Expression of Th1/Th2 Cytokines

Selenium （Se） is an essential trace element. Autoimmune thyroid diseases （AITD） are destructive inflammatory or anti-receptor autoimmune diseases characterized by reactivity to self-thyroid antigens. However, the effects of Se on the cytokines in AITD are still unclear. So we researched the role of Selenium （Se） and Thl/Th2 cytokine productions in the pathogenesis of autoimmune thyroid diseases （AITD）.

Local Th1/Th2 Cytokine Expression in Experimental Murine Vaginal Candidiasis

In order to investigate the expression of Th1 and Th2 cytokines in the vaginal candidiasis caused by Candida, the fungal vaginitis model was established in female ICR mice by intravaginal inoculation of suspension of C. albicans after the animals were pretreated with estradiol. Reverse transcriptase-polymerase chain reaction （RT-PCR） was used to detect the expression of IL-2, IL-4, IL-10 and TGF-β1 in the vagina in the mice of different groups at different time points after the beginning of the experiment. The average expression level of IL-2 mRNA in group D （estrogen-treated mice） was significantly higher than that in groups H （estrogen-untreated mice） and I （control group） on the day 2. The average expression level of IL-4 mRNA in group D was significantly higher than that in groups I and H on the day 5. The average expression level of IL-10 mRNA in group D was significantly higher than that in groups H and I from day 7 to 11. The average expression level of TGF-β1 mRNA in group D was significantly higher than that in groups H and I at all time points. It was concludes that the high-level expression of IL-2 mRNA during early infection was associated with clearance of mucosal C. albicans, and the high-level expression of IL-10 mRNA during late stage of the infection was related to susceptibility to infection. TGF-β1 may play a predominant role when the virtual absence of changes in other Th-type cytokines during infection.

A Novel LNP-Based <i>Chlamydia</i>Subunit Vaccine Formulation That Induces Th1 Responses without Upregulating IL-17 Provides Equivalent Protection in Mice as Formulations That Induced IL-17 and Th1 Cytokines

We evaluated novel Chlamydial vaccines, consisting of major outer membrane protein (MOMP) alone or in combination with polymorphic membrane proteins D (PmpD) and G (PmpG) using a C57BL/6 mouse model. Native MOMP (nMOMP) isolated from C. muridarum elementary bodies (EBs) and recombinant PmpD and PmpG proteins were adjuvanted with Monophosphoryl lipid A (MPLA), with either lipid nanoparticles (LNPs) or the cationic lipid dimethyldioctadecylammonium bromide (DDA). Antibody titers to C. muridarum nMOMP, and EBs were evaluated by ELISA, and T-cell responses were analyzed by intracellular cytokine staining (ICS). Protection from challenge was determined by qPCR. Vaccine immunized mice showed significantly higher antibody titers to nMOMP (P < 0.001) and C. muridarum EBs (P < 0.001), when compared to the adjuvant alone group. Antibody titers in vaccine groups with Monophosphoryl lipid A (MPLA) + LNP were higher as compared to the MPLA + DDA group (P < 0.001) except for (Cm nMOMP + PmpG + PmpD p73 + PmpD p82 + MPLA + DDA) vs (Cm nMOMP + PmpG + PmpD p73 + PmpD p82 + MPLA + LNP) for both C. muridarum EBs and PmpG. ICS analysis showed more robust CD4 + T-cell responses (IFN-γ/IL-2/TNF-a) in the DDA and LNP groups compared to the adjuvant alone group. The DDA + MPLA gave robust Th17 responses in comparison to MPLA and LNP group. Mice immunized with Chlamydia antigens also showed protection from C. muridarum challenge, by reduction in bacterial shedding for all groups (P < 0.003) compared to shedding from the adjuvant control. Both vaccine formulations generated robust immunological responses, and both were protective by reducing bacterial shedding after challenge. This data indicates equal protection can be achieved without the induction of Th17 responses.

Effect of short- and long-term immunization of recombinant disorganized muscle protein-1(rDIM-1) against human filarial parasite Brugia malayi in rodents

Objective: To evaluate the effect of short-term and long-term immunization of recombinant disorganized muscle protein-1(r DIM-1) in rodents against human filarial parasite Brugia malayi.Methods: Recombinant Brugia malayi DIM-1(rDIM-1 bm) protein was cloned, expressed and purified using a Ni-NTA affinity column. Mastomys coucha were immunized with rDIM-1 bm in three immunization schedules: short-term(3-dose of rDIM-1 bm), and long-term(booster doses till 3-and 6-week) and subsequently challenged with infective third-stage larvae of filarial parasite Brugia malayi(L3). Microfilaraemia was monitored in L3 exposed groups on day 90 post larval inoculation(p.l.i.) and continued till day 205 p.l.i. On day 205 p.l.i. all the infected animals were killed and total worm burden was estimated. Cellular proliferative response, macrophage activity, nitric oxide(NO) release, specific IgG and its subtypes, IgE, IgA and Th1(IFN-γ, TNF-ααand IL-2) and Th2(IL-4, IL-5, IL-6, IL-10 and IL-13) cytokine release were determined. Results: Of the 3 different immunization schedules, shortterm immunization(3-dose schedule) showed better reduction in microfilarial burden(36%-63%) in the peripheral circulation, adult worm load(52%), whereas long-term immunization(3-and 6-week schedule) exerted less effect on peripheral microfilariae count(9%-58%), and adult worm burden(9%-12.5%). Short-term immunization resulted in upregulation of cellular proliferation, macrophages activity, NO release, specific IgG, IgG1, IgG2 a, Ig G2 b, IgE and IgA levels and both Th1(IFN-γ, TNF-α and IL-2) and Th2(IL-4, IL-5, IL-6, IL-10 and IL-13) cytokine release whereas long-term immunization(3-and 6-week schedule) exerted less effect on parasite burden and showed mixed immunological responses. None of the rDIM-1 bm administration schedules induced any pathology in lymphoid tissues, or alteration in mast cell number and granularity. Conclusions: The short-term immunization with rDIM-1 bm(3-dose schedule) induces robust immune responses and protects the host from filarial parasite infection.

Effects of combined immune therapy on survival and Th1/Th2 cytokine balance in rat orthotopic liver transplantation

Background The induction of immune tolerance and suppression of allograft rejection has become the focus in the study of liver transplantation. The effect of immune therapy with anti-CD40L mAb alone or in combination with cyclosporine A （CsA） on the recipient survival and Th1/Th2 cytokine profile was studied to elucidate its immunological mechanism and role in rat orthotopic liver transplantation.Methods The model of rat orthotopic liver transplantation was established by modified Kamada＇s technique. Recipients were divided into group A （control group）： SD→SD; group B （group of rejection）： SD→Wistar without any treatment; group C： SD→Wistar with CsA monotherapy from day 1 to day 5; and group D： SD→Wistar with CsA from day 1 to day 5 and anti-CD40L mAb on day 0 and day 2. The survival of the recipients in all groups was observed and ELISA technique was used to detect the level of cytokines in peripheral blood on post-transplant day 7.Results The survival period of recipients in groups A （〉60 days） and D （〉60 days） was significantly longer than that in group B （13.8±2.4 days）. The serum levels of interleukin 2 （IL-2） and interferon y in group B were significantly higher than those in other groups; the level of tumor necrosis factor a was higher but not statistically significant. In contrast, the serum levels of IL-4 and IL-10 in group D were elevated more significantly than those in group B （P〈0.05）.Conclusions Combined immune therapy can prolong the survival of al log rafts. Increased expression of Th2 cytokines, which is closely related to the induction of tolerance and suppression of rejection, is beneficial to the long-term survival of recipients and allografts.

Effect of Chinese Herbal Treatment on Th1- and Th2-Type Cytokines,Progesterone andβ-Human Chorionic Gonadotropin in Early Pregnant Women of Threatened Abortion

Objective： To study the clinical effects of Prescription Zhuyun-Ⅲ （助孕3号方, ZYⅢ） on early pregnant women diagnosed as threatened abortion, and its mechanism in immunity and endocrine by determining serum Th1- and Th2-type cytokines, progesterone, and β-human chorionic gonadotropin （β-HCG）. Methods： The treatment group comprised 30 early pregnant women diagnosed as threatened abortion of deficiency syndrome of Pi （脾）, Shen （肾）, or both. The control group consisted of 20 normal early pregnant women of similar gestational age. Patients in the treatment group were administered with ZYⅢ for 4 weeks. Peripheral blood samples were collected pre- and post-treatment from both the treatment and the control groups. Serum Thl-type cytokine [interleukin-2 （IL-2）] and Th2-type cytokine [interleukin-10 （IL-10）] were determined by flow cytometry, and serum progesterone and 13-HCG were determined by ELISA. Results： （1） The treatment was effective in 26 and ineffective in 4 patients of the treatment group. Therefore, the cure percentage was 86.67%. （2） In the treatment group before the treatment, IL-2 was significantly higher, IL-10 tended to be less, and the Th1/Th2 balance shifted toward Thl compared with those in the control group. （3） After the treatment, IL-2 was decreased, IL-10 was increased, and IL-2/IL-10 was decreased. Both progesterone and β-HCG were increased. Changes of progesterone were positively correlated with changes of IL-10, whereas changes of β-HCG were negatively correlated with changes of IL-2. Conclusions： Our study suggests that ZYⅢ has an evident function of protecting the fetus, and one of its mechanisms is inhibiting the secretion of Thl cytokines, promoting the secretion of Th2 cytokines, and recovering the pathological shift of the Th1/Th2 balance. The other possible mechanism is increasing serum progesterone and β-HCG concentrations. Moreover, there are some correlations between the above two effects.

Pleural fluid from tuberculous pleurisy inhibits the functions of T cells and the differentiation of Th1 cells via immunosuppressive factors

Immunosuppressive mediators in tuberculosis pleurisy(pleural fluid(PF))are associated with the course of disease,but they remain poorly defined.To study the local immune status of patients with tuberculosis pleurisy,we examined the effect of PF on the functions of T cells and the differentiation of Th1 cells.PF could inhibit the ability of T cells to produce cytokines.However,tumor-necrosis factor(TNF)-a derived from non-T cells was not impaired.Further analysis indicated that cell activation and cell cycle progression were also suppressed.Moreover,PF could inhibit Th1 cell differentiation.Importantly,we found that inhibitors of indoleamine 2,3-dioxygenase(IDO)and adenosine and neutralizing antibodies against IL-10 and transforming growth factor(TGF)-b could reverse cytokine production,suggesting that IDO,adenosine,IL-10 and Transforming growth factor–b1 in PF might take part in impairing T-cell functions.Taken together,our data demonstrate for the first time that several immunopathological factors participate in the downregulation of T-cell functions in local PF.

Immunostimulatory role of rBmHSP60 from filarial parasite Brugia malayi

Objective:To evaluate the immunostimulatory potential of crossreactive molecule heat shock protein 60(HSP60)of filarial parasite Brugia malayi and Leishmania donovani.Methods:HSP60 of Brugia malayi(BmHSP60)was amplified using gene-specific primer,cloned in p Tri Ex4 vector,expressed in BL21-DE3 cells,and recombinant HSP60(rHSP60)of~65 k Da was purified by affinity chromatography using Ni-NTA column.The recombinant protein was desalted by the dialysis membrane,and the presence of endotoxin level was determined by Limulus amebocyte lysate assay.The recombinant protein was tested for cell proliferation,nitric oxide release,expression of Th1 and Th2 cytokines,and transcription factors(STATs)in vitro using murine macrophage cell line(J774 A.1).Results:Higher cell proliferation indicated that BmHSP60 had immunostimulatory potential.rBmHSP60 exposure upregulated the expression of iNOS,STAT1,STAT4,Th1 cytokines(IFN-γ,TNF-α,IL-12),and nitric oxide release.In addition,no remarkable change was observed in the expression of IL-6,IL-10,and STAT3 in macrophage cell line J774 A.1.The ELISA analysis showed the levels of IFN-γ,TNF-α,and IL-12 were upregulated while IL-10 level was downregulated,revealing that BmHSP60 triggered a Th1 immune response.Conclusions:Our study demonstrates that rBmHSP60 has immunogenic properties which effectively enhances the Th1 type immune responses,and can be used as an immunoprophylactic agent against leishmaniasis.Furthermore,in vivo studies are in progress to determine the protective role of rBmHSP60 against Leishmania donovani infection in a mouse model.

Antisense-Induced Blockade of GATA-3 Expression Could Inhibit Th2 Excursion of Tumor Cells in vitro and in vivo

Previous studies have shown that tumor cells predominantly express Th2 type cytokines and transcription factors. GATA-3, as a Th2-specific transcription factor, plays a central role in positive-regulating Th2 development. So whether the expression of GATA-3 in tumor cells has any effect on tumor development is a question of interest. In the present study, we inhibited the expression of GATA-3 in tumor cells through antisense RNA blockade technique, and observed its effects on tumor in vitro and in vivo. Our results showed that antisense GATA-3 treatment could inhibit the expression of TNF-α and Th2 cytokines in tumor cells, and antisense-induced blockade of GATA-3 could also depress tumor growth in tumor-bearing mice. We suggest that the ratio of T-bet/GATA-3 can be evaluated as a more important marker of the status of Thl/Th2 type. And our results might provide some evidence about the molecular regulatory mechanisms in tumor cell development. Cellular ＆ Molecular Immunology. 2005 ;2（3 ）： 189-196.