Novel T-cell epitopes on Schistosoma japonicum SjP40 protein and their preventive effect on allergic asthma in mice.

Allergic asthma is a chronic inflammatory disease mediated by Th2 cell immune responses. Currently, immunotherapies based on immune deviation are attractive, preventive, and therapeutic strategies for asthma. Many studies have shown that intracellular bacterial infections such as mycobacteria and their components can suppress asthmatic reactions by enhancing Th1 responses, while helminth infections and their proteins can inhibit allergic asthma via immune regulation. However, some helminth proteins such as SmP40, the major egg antigen of Schistosoma mansoni, are found as Th1 type antigens. Using a panel of overlapping peptides, we identified T-cell epitopes on SjP40 protein of Schistosoma japonicum, which can induce Th1 cytokine and inhibit the production of Th2 cytokines and airway inflammation in a mouse model of allergic asthma. These results reveal a novel form of immune protective mechanism, which may play an important role in the modulating effect of helminth infection on allergic asthmatic reactions.

Phospholipase Cε deficiency delays the early stage of cutaneous wound healing and attenuates scar formation in mice.

This study aimed to investigate the role of phospholipase Cε (PLCε) in the skin wound healing process. PLCε, an effect factor of Ras/Rap small G protein, plays a crucial role in skin inflammation by regulating inflammatory cytokines. Inflammatory responses are closely associated with wound healing. Full-thickness skin wounds were made in the PLCε knockout (KO) and wild-type (WT) mice, and the healing process was analyzed. The macroscopic wound closure rate declined in the PLCε KO mice on days 3, 4, and 5 after wounding, following the decreased expression of interleukin (IL)-6, chemokine (C-X-C motif) ligand (Cxcl)-1, Cxcl-2, and chemokine (C-C motif) ligand (Ccl) 20. The proliferation rate of epidermal keratinocytes was not affected by PLCε, but silencing of PLCε resulted in the delayed migration of keratinocytes. Moreover, the scars were found to be much smaller in the PLCε KO mice than in the WT mice. The mRNA expression of Ccl20, collagen (Col) 6a1, and Col17a1 decreased in the PLCε KO mice. These results were in agreement with a previous hypothesis that PLCε might delay the early stage of cutaneous wound healing by inhibiting the migration of keratinocytes, and decrease the expression of Col6a1, Col17a1, and Ccl20 by inhibiting the inflammatory response to reduce scar formation. This study shed light on a novel role of PLCε in wound healing and provided new therapeutic approaches to target PLCε for diminishing scar formation after injury.

Function of AURKA protein kinase in the formation of vasculogenic mimicry in triple-negative breast cancer stem cells.

Tumor cell vasculogenic mimicry (VM), a newly defined pattern of tumor blood supply, signifies the functional plasticity of aggressive cancer cells forming vascular networks. VM and cancer stem cells (CSCs) have been shown to be associated with tumor growth, local invasion, and distant metastasis. In our previous study, CSCs in triple-negative breast cancer were potential to participate in VM formation. In this study, breast CSCs were isolated from the triple-negative breast cancer cell line MDA-MB-231 by using mammosphere culture. Western blotting and reverse transcription polymerase chain reaction showed that mammosphere cells displayed an increased expression of AURKA protein kinase and stem cell marker c-myc and sox2. The VM formation by mammosphere cells was inhibited by AURKA knockdown or the addition of AURKA inhibitor MLN8237. In the meantime, MLN8237 induced the increased E-cadherin and decreased c-myc, sox2, and ß-catenin expressions. The function of AURKA in VM formation was further confirmed using a xenograft-murine model. The results suggested that AURKA protein kinase is involved in VM formation of CSCs and may become a new treatment target in suppressing VM and metastasis of breast cancer.

[PCR in evaluating the effect of allicin and its combination with SMZco on murine toxoplamosis].

OBJECTIVE: To evaluate the effect of allicin alone or combined with SMZco on murine toxoplasmosis by a specific, rapid, and sensitive PCR technique. METHODS: 147 mice were infected with 2 x 10(4) tachyzoites intraperitoneally and divided into 5 groups at random. Each group was divided into two sub-groups except an untreated group. One subgroup was used to get samples for PCR test and the other for observing the survival duration. The therapeutic grouping was as follows: group A, a combination of allicin and SMZco administered orally for 7 days and continued by allicin alone till 21 days; group B, the combination administered for 14 days and continued with allicin till 21 days; group C, allicin alone for 21 days; group D, SMZco alone for 7 days; group E, untreated control. The dosage was: SMZco 400 mg/(kg.d) and allicin 35 mg/(kg.d). PCR test was used to detect the parasites in samples of liver and blood from infected mice at 5, 10, 15, 20, 25, 30, 40 and 50 days after infection. RESULTS: Parasites were eliminated in the blood because no signal was seen in all the blood samples except for samples from group C at day 5 after infection. From day 10 after infection until the end of the experiment, no amplification of DNA was seen in all the samples. As for liver samples, signals were clear at day 5 post infection. From day 10 post infection till the day 50 post-infection, parasites were still detected, but the PCR products decreased significantly than that of day 5 post-infection. Result showed that a combination of SMZco with allicin provided a significant protection. SMZco alone was also effective, but allicin alone was not. CONCLUSION: When SMZco is used in combination with allicin, a much higher efficacy is received in the treatment of acute murine toxoplasmosis.

Doxycycline as an inhibitor of the epithelial-to-mesenchymal transition and vasculogenic mimicry in hepatocellular carcinoma.

This study was conducted to examine the effects of doxycycline on the survival time and proliferation of hepatocellular carcinoma (HCC) in vivo and on the biologic functions of HCC in vitro. This study was also designed to evaluate the effects of doxycycline on epithelial-to-mesenchymal transition (EMT)- and vasculogenic mimicry (VM)-related protein expression and on matrix metalloproteinase (MMP) and DNA methyltransferase (DNMT) activity in vitro. Human MHCC97H cells were injected into BALB/c mice, which were divided into treatment and control groups. Doxycycline treatment prolonged the mouse survival time and partly suppressed the growth of engrafted HCC tumor cells, with an inhibition rate of 43.39%. Higher amounts of VM and endothelium-dependent vessels were found in the control group than the treatment group. IHC indicated that epithelial (E)-cadherin expression was increased in the doxycycline-treated mice compared with the control group. In in vitro experiments, doxycycline promoted HCC cell adhesion but inhibited HCC cell viability, proliferation, migration, and invasion. Western blot analysis, semiquantitative RT-PCR, qRT-PCR, and immunofluorescence demonstrated that doxycycline inhibited the degradation of the epithelial marker E-cadherin and downregulated the expression levels of EMT promoters, the mesenchymal marker vimentin, and the VM-associated marker vascular endothelial (VE)-cadherin. Furthermore, the activities of MMPs and DNMTs were examined in different groups via gelatin zymography and a DNMT activity assay kit. A methylation-specific PCR was performed to assess the promoter methylation of CDH1 (the gene encoding E-cadherin). Doxycycline prolonged the mouse survival time by inhibiting EMT progression and VM formation.

The PH domain adaptor protein Bam32/DAPP1 functions in mast cells to restrain FcɛRI-induced calcium flux and granule release.

Mast cell activation triggered by IgE binding to its high affinity receptor FcɛRI is highly dependent on signaling via phosphoinositde 3-kinases (PI3K). The phosphoinositide phosphatase SHIP controls mast cell activation by regulating accumulation of D3 phosphoinositide second messengers generated by PI3K. The PH domain adaptor protein Bam32/DAPP1 binds specifically to the D3 phosphoinositides PI(3,4,5)P3 and PI(3,4)P2 (the substrate and product of SHIP respectively). In B cells, Bam32 is phosphorylated by Src family kinases including Lyn, and is required for antigen receptor-induced activation; however the function of Bam32 in mast cells is unknown. Here we report that Bam32 is expressed in mast cells, is recruited to the plasma membrane upon stimulation and functions in FcɛRI signaling. Examination of bone marrow-derived mast cells (BMMC) isolated from Bam32-deficient mice revealed enhanced FcɛRI-induced degranulation and IL-6 production, indicating that Bam32 may function to restrain signaling via FcɛRI. These enhanced degranulation responses were PI3K-dependent, as indicated by blockade with PI3K inhibitors wortmannin or IC87114. While Bam32-deficient BMMC showed reduced FcɛRI-induced activation of mitogen-activated protein kinases ERK and JNK, FcɛRI-induced calcium flux and phosphorylation of PLCγ1 and Akt were increased. Bam32-deficient BMMC showed significantly reduced phosphorylation of Lyn and SHIP, indicating reduced activity of inhibitory signaling pathways. Together our results identify Bam32 as a novel regulator of mast cell activation, potentially functioning in membrane-proximal integration of positive and negative signaling pathways.