[Mannose-capped lipoarabinomannan (ManLAM) binding TLR2 activates mast cells to release exosomes and induces M2 polarization of macrophages].

Objective To explore the mechanism of mannose-capped lipoarabinomannan (ManLAM) lipopolysaccharide of mycobacterium tuberculosis (MTB) inducing exosomes of mast cells to recruit macrophages and influence macrophage polarization for immune evasion. Methods H37Rv MTB was cultured and ManLAM extracted and identified. The extracted ManLAM was used to treat mast cells, and the exosomes secreted by mast cells were collected. The protein components of the exosomes were analyzed and identified by proteomic analysis and Western blotting. The collected exosomes were co-cultured with macrophages differentiated from THP-1 cells. The chemotaxis of exosomes released by mast cells on macrophages was detected by TranswellTM assay. The macrophage polarization induced by ManLAM was detected by flow cytometry and real-time quantitative PCR. Results ManLAM was successfully extracted and purified, and the ratio of mannose to arabinose was about 12.6:9.4 identified by gas chromatography. ManLAM bound to Toll-like receptor 2 of mast cells, the exosomes produced by those mast cells had a high levels of CCL2, IL-4, and IL-13. ManLAM-induced mast cell exosomes recruited macrophages and promoted high expression of M2 polarization of macrophages molecular markers: arginase-1, IL-10, and FIZZ-1. Conclusions ManLAM stimulates mast cells to secrete exosomes and indirectly induces M2 polarization of macrophages, which makes MTB evade immune clearance.

[HpaA promotes gastric mucosa injury via stimulating IL-21 secretion of T cells].

Objective To demonstrate HpaA can intensify the inflammatory response and gastric mucosa injury by IL-21 from induced T cell. Methods Biopsy specimens were taken from gastric mucosa of 56 patients with H.pylori infection before and after H.pylori radical elimination by endoscope. The levels of IL-21, matrix metalloproteinase-2 (MMP2) and MMP9 from the biopsy were detected by reverse transcription PCR and Western blot analysis. Meanwhile, the recombinant HpaA was cloned, expressed and purified to stimulate the magnetic cell sorting CD3+ T cells from healthy donors' peripheral blood mononuclear cells (PBMCs), and the level of IL-21 in the supernatant fluid was detected by ELISA. Thereafter, AGS cells were cultured and Western blot analysis was performed to detect the levels of MMP2 and MMP9 in the AGS cells with human IL-21 and anti-IL-21 antibody treatment for 24 hours. Results The protein levels of IL-21 and MMP2 and MMP9 in gastric mucosa infected with H. pylori was significantly higher than that in gastric mucosa after radical treatment of H. pylori. Meanwhile, the recombinant HpaA promoted IL-21 secretion by induced CD3+T cells in vitro. IL-21 stimulated the expression of MMP2 and MMP9 in AGS cells. When IL-21 was blocked by the antibody, the levels of MMP2 and MMP9 in AGS cells decreased significantly. Conclusion HpaA plays a significant role in the gastric mucosa injury caused by H.pylori infection through IL-21 from induced T cells.

[Screening and characterization of nucleic acid aptamers specifically binding to adhesin HpaA from Helicobacter pylori].

Objective To screen aptamers that specifically bind to adhesin HpaA from Helicobacter pylori (H. pylori) by systematic evolution of ligands by exponential enrichment (SELEX) and identify the binding properties of aptamers. Methods The prokaryotic expression recombinant plasmid pET28a/HpaA was constructed and the HpaA protein was expressed and purified with IPTG. With the recombinant HpaA protein as target, we screened aptamers with high affinity and specificity binding force by SELEX. The binding force between aptamers and H. pylori in vitro and the performance of aptamers in H. pylori detection from the biopsy of gastric mucosa were examined using the aptamers we had screened. Results We extracted genome from H. pylori ATCC26695 strains and amplified 699 bp HpaA gene using PCR. The recombinant plasmid pET28a/HpaA was constructed successfully. The recombinant HpaA was expressed and purified up to 98% as target for aptamer screening. The six highest affinity aptamers were obtained and named HA1 to HA6 through 10-round positive screening and five-round negative screening by SELEX. The full-length aptamer HA6 and the core sequence of HA6 showed highest affinity and specificity in H. pylori detection in vitro. In view of this, the FAM-labelled aptamer HA6 was used to detect H. pylori in gastric mucosa from 166 patients. The aptamer HA6 showed a higher detection rate (94.58%) than URT (87.95%) in the same batch of clinical samples. Conclusion The aptamers that specifically bind to HpaA may be applied for the detection of H. pylori in gastric mucosa as a novel method.

GSK1838705A, an insulin-like growth factor-1 receptor/insulin receptor inhibitor, induces apoptosis and reduces viability of docetaxel-resistant prostate cancer cells both in vitro and in vivo.

Prostate cancer is the leading malignancy and the second most common cause of cancer-related death in men. Despite high cure rates with surgery and/or radiation, 30%-40% of patients eventually develop advanced cancer. Docetaxel is one of the most effective and well established chemotherapeutic agents for prostate cancer. However, docetaxel resistance often develops within months. Combination therapies have been proposed to improve the therapeutic efficacy of docetaxel in prostate cancer, and there is an urgent need to identify agents that are effective for treatment of the disease, especially docetaxel-resistant prostate cancer. In this work, we investigated the activity of GSK1838705A, a potent insulin-like growth factor-1 receptor (IGF1R)/insulin receptor (IR) inhibitor, in prostate cancer, especially docetaxel-resistant prostate cancer. We found that GSK1838705A could effectively reduce the viability of both docetaxel-sensitive and docetaxel-resistant prostate cancer cells. GSK1838705A induced marked apoptosis in docetaxel-resistant cells, and also dramatically inhibited migration of these cells. Further, GSK1838705A significantly inhibited phosphorylation of IGF1R/IR. Importantly, GSK1838705A significantly suppressed docetaxel-resistant PC-3R tumor growth in vivo. This is the first study of GSK1838705A in prostate cancer. Our results indicate that GSK1838705A is a promising compound for the treatment of prostate cancer, especially for those who develop resistance to docetaxel, and might shed new light on treatment for prostate cancer.