Inhibitory effects of methamphetamine on mast cell activation and cytokine/chemokine production stimulated by lipopolysaccharide in C57BL/6J mice.

Previous studies have demonstrated that methamphetamine (MA) influences host immunity; however, the effect of MA on lipopolysaccharide (LPS)-induced immune responses remains unknown. Mast cells (MCs) are considered to serve an important role in the innate and acquired immune response, but it remains unknown whether MA modulates MC activation and LPS-stimulated cytokine production. The present study aimed to investigate the effect of MA on LPS-induced MC activation and the production of MC-derived cytokines in mice. Markers for MC activation, including cluster of differentiation 117 and the type I high affinity immunoglobulin E receptor, were assessed in mouse intestines. Levels of MC-derived cytokines in the lungs and thymus were also examined. The results demonstrated that cytokines were produced in the bone marrow-derived mast cells (BMMCs) of mice. The present study demonstrated that MA suppressed the LPS-mediated MC activation in mouse intestines. MA also altered the release of MC cytokines in the lung and thymus following LPS stimulation. In addition, LPS-stimulated cytokines were decreased in the BMMCs of mice following treatment with MA. The present study demonstrated that MA may regulate LPS-stimulated MC activation and cytokine production.

Human cathelicidin LL-37 enhance the antibiofilm effect of EGCG on Streptococcus mutans.

BACKGROUND: Streptococcus mutans forms biofilms as a resistance mechanism against antimicrobial agents in the human oral cavity. We recently showed that human cathelicidin LL-37 exhibits inhibitory effects on biofilm formation of S. mutans through interaction with lipoteichoic acid (LTA), but without antibacterial or biofilm dispersal abilities. (-)-Epigallocatechin gallate (EGCG) is the most abundant constituent of tea catechins that has the greatest anti-infective potential to inhibit the growth of various microorganisms and biofilm formation. Therefore, in this study, we evaluated whether LL-37 interacts with EGCG to enhance the antibiofilm effect of EGCG on S. mutans biofilm formation. METHODS: Clinical S. mutans strains (n = 10) isolated from children's saliva were tested in a biofilm formation assay. The antibiofilm effect of EGCG with and without LL-37 was analyzed by the minimum biofilm eradication concentration assay and confirmed using field emission-scanning electron microscopy. In addition, the interaction among EGCG, LL-37, and LTA of S. mutans was determined using quartz crystal microbalance analysis. RESULTS: EGCG killed 100 % of planktonic S. mutans within 5 h, inhibited biofilm formation within 24 h, and reduced bacteria cells in preformed biofilms within 3 h at a concentration of 0.2 mg/mL. However, EGCG did not appear to interact with LTA. LL-37 effectively enhanced the bactericidal activity of EGCG against biofilm formation and preformed biofilms as determined by quantitative crystal violet staining and field emission-scanning electron microscopy. In addition, quartz crystal microbalance analysis revealed that LL-37 interacted with EGCG and promoted binding between EGCG and LTA of S. mutans. CONCLUSIONS: We show that LL-37 enhances the antibiofilm effect of EGCG on S. mutans. This finding provides new knowledge for dental treatment by using LL-37 as a potential antibiofilm compound.

The effects of D3R on TLR4 signaling involved in the regulation of METH-mediated mast cells activation.

Accumulating studies have revealed that the dopamine D3 receptor (D3R) plays an important role in methamphetamine (METH) addiction. However, the action of D3R on METH-mediated immune response and the underlying mechanism remain unclear. Mast cells (MCs) are currently identified as effector cells in many processes of immune responses, and MC activation is induced by various stimuli such as lipopolysaccharide (LPS). Moreover, CD117 and FcεRI are known as MC markers due to their specific expression in MCs. To investigate the effects of D3R on METH-mediated alteration of LPS-induced MCs activation and the underlying mechanism, in this study, we examined the expression of CD117 and FcεRI in the intestines of wild-type (D3R(+/+)) and D3R-deficient (D3R(-/-)) mice. We also measured the production of MC-derived cytokines, including TNF-α, IL-6, IL-4, IL-13 and CCL-5, in the bone marrow-derived mast cells (BMMCs) of WT and D3R(-/-) mice. Furthermore, we explored the effects of D3R on METH-mediated TLR4 and downstream MAPK and NF-κB signaling induced by LPS in mouse BMMCs. We found that METH suppressed MC activation induced by LPS in the intestines of D3R(+/)mice. In contrast, LPS-induced MC activation was less affected by METH in D3R(-/-) mice. Furthermore, METH altered LPS-induced cytokine production in BMMCs of D3R(+/+) mice but not D3R(-/-) mice. D3R was also involved in METH-mediated modulation of LPS-induced expression of TLR4 and downstream MAPK and NF-κB signaling molecules in mouse BMMCs. Taken together, our findings demonstrate that the effect of D3R on TLR4 signaling may be implicated in the regulation of METH-mediated MCs activation induced by LPS.

The dopamine D3 receptor regulates the effects of methamphetamine on LPS-induced cytokine production in murine mast cells.

Previous studies have demonstrated that methamphetamine (METH) alter inflammatory and anti-inflammatory cytokine production in the periphery. However, the effect of METH on lipopolysaccharide (LPS)-induced immune responses and its underlying mechanism of action remains unclear. The dopamine D3 receptor (D3R) plays an important role in METH addiction, indicating that the D3R may regulate METH-mediated immune responses. In this study, we examined the effect of METH on mast cell released cytokines in the lungs and thymi of mice stimulated by LPS, and on LPS-induced murine bone marrow-derived mast cells (BMMCs). Moreover, we used D3R-deficient mice to investigate the effect of this receptor on LPS-stimulated mast cell released cytokine production after METH treatment in the lungs and thymi. The effects of a D3R agonist and antagonist on LPS-induced cytokine production after METH treatment in murine BMMCs were also evaluated. METH suppressed LPS-induced cytokine production in the lungs and thymi of wild-type (WT) mice and BMMCs. However, METH did not alter LPS-induced cytokine production in the lungs and thymi of D3R-deficient mice. When BMMCs were treated with the D3R receptor antagonist, NGB2904 hydrochloride (NGB-2904), METH did not alter LPS-induced cytokine production. However, treatment with the D3R agonist, 7-hydroxy-(di-n-propylamino) tetralin (7-OH-DPAT), significantly enhanced the effects of METH on LPS-induced cytokine production. Our results suggest that METH regulates mast cell released cytokines production in an LPS-induced mouse model via the D3R.

Evaluation of protective effect of multi-epitope DNA vaccine encoding six antigen segments of Toxoplasma gondii in mice.

To investigate the vaccine potential of multi-epitope vaccines against toxoplasmosis, a multi-epitope DNA vaccine, eukaryotic plasmid pcDNA3.1/T-ME expressing six antigen segments (SAG1(238-256), SAG1(281-320), GRA1(170-193), GRA4(331-345), GRA4(229-245), and GRA2(171-185)) of Toxoplasma gondii was constructed. We investigated the efficacy of pcDNA3.1/T-ME with or without co-administration of a CpG-oligodeoxynucleotide (CpG-ODN) as an adjuvant to protect mice (BALB/c and C57BL/6) against toxoplasmosis. High survival rates were observed in mice immunized with pcDNA3.1/T-ME when challenged with T. gondii RH strain. Lymphocyte proliferation assays, cytokine, and antibody determinations show that mice immunized with pcDNA3.1/T-ME produced stronger humoral and Th1-type cellular immune responses compared to untreated mice or those immunized with empty plasmids. However, co-immunization with CpG-ODN resulted in impaired immune responses. Our data demonstrates that multi-epitope DNA vaccination is a potential strategy for the control of toxoplasmosis and paves the way for further investigations into producing a multi-epitope anti-T. gondii DNA vaccine.

The detection of sLFA-3 in plasma of patients with hemorrhagic fever with renal syndrome.

Hemorrhagic fever with renal syndrome (HFRS) is an acute viral disease characterized by endothelial dysfunction. The cellular immune response, especially the virus-specific CD8+ T lymphocytes, is known to attack vascular endothelial cells (VEC) and to contribute to the diffuse damage and penetrability increasing of VEC. Lymphocyte function associated antigen 3 (LFA-3) is expressed on T lymphocytes and VEC, which is contributed to the activation of T lymphocytes. The expression of LFA-3 on the activated T lymphocytes and VEC is highly increased, which can exfoliate into plasma to increase the level of soluble LFA-3 (sLFA-3) in plasma. So the change of sLFA-3 levels is correlated with the activation of T lymphocytes. In this study we detected the levels of sLFA-3 in plasma of patients with HFRS. We examined the levels of sLFA-3 in plasma samples collected from 53 HFRS patients by double antibody sandwich ELISA. We found variable, but persistently elevated levels of sLFA-3 throughout the various phases and types of the HFRS disease, which suggest that sLFA-3 levels have correlation with disease stages. Moreover, elevated sLFA-3 levels are closely correlated to the severity of HFRS and the degree of kidney damage.

Levels of soluble vascular cell adhesion molecule-1 and soluble intercellular adhesion molecule-2 in plasma of patients with hemorrhagic fever with renal syndrome, and significance of the changes in level.

Hemorrhagic fever with renal syndrome (HFRS) is an acute viral disease characterized by endothelial dysfunction. Vascular cell adhesion molecule (VCAM)-1 and intercellular adhesion molecule (ICAM)-2 provide costimulatory signals for the activation of T lymphocytes; these adhesion molecules play key roles in leukocyte adherence and propagation of inflammatory responses. They may be involved in the immunologic response that leads to vascular endothelial cell (VEC) and kidney damage of HFRS patients, and increased levels of soluble (s)VCAM-1 and sICAM-2 in plasma may indicate the severity of HFRS. We examined the presence of sVCAM-1 and sICAM-2 in 52 plasma samples collected from 52 patients. We tested these plasma samples for sVCAM-1 and sICAM-2 by double-antibody sandwich ELISA. We found variable, but persistently elevated, levels of sVCAM-1 and sICAM-2 throughout the various phases and types of the disease, which suggested sVCAM-1 may play an important role in the immunopathological lesions of HFRS and is closely correlated to the severity of HFRS and the degree of kidney damage. sICAM-2 may be associated with the hyperfunctioning of the cellular immune response.

Effective induction of type 1 cytotoxic T cell responses in mice with DNA vaccine encoding two hepatitis C virus cytotoxic T lymphocyte epitopes.

The aims of this study were to explain whether a multiple cytotoxic T lymphocyte (CTL) epitope-based anti-hepatitis C virus (HCV) DNA vaccine can induce specific CTL responses to each HCV CTL epitope independently and long-term CD8(+) T cell memory responses, and to determine the cytokine secretion pattern and subtype of epitope-specific cytotoxic T cells. A multi-CTL epitope gene, which consists of two epitopes of HCV (H-2(d)-restricted HCV core(133142) and E1(315322)), was cloned into the eukaryotic expression vector pcDNA3.1. BALB/c mice (H-2(d) restricted) were vaccinated intramuscularly with this multi-CTL epitope-based DNA vaccine. The epitope-specific CTLs against target cells (P815,H-2(d) restricted) pulsed with various CTL epitope peptides were detected by lactate dehydrogenase release assay, and the precursor frequency of epitope-specific CTLs was determined by limiting dilution analysis. Cytokines (interleukin [IL]-2, IL-4, and interferon-) in culture supernatants were determined by enzyme-linked immunosorbent assay. The multi-CTL epitope-based DNA vaccine directed against two HCV CTL epitopes could induce specific CTL responses to each of the two CTL epitopes independently and long-term CD8(+) T cell memory responses. The epitope-specific cytotoxic T cells produced helper T cell type 1 cytokines. This work demonstrated that multiepitope DNA vaccination is a potential strategy to control HCV infection.