The effects of Particulate matter (PМ2.5) pollutants on cancer cells in in vitro model / Монголын Анагаах Ухаан

Introduction@#Air pollution has become one of the major problems in socio-economic and health issues in Mongolia. Among the various hazards of particulate matter (PM) pollutants, microorganisms in PM2.5 and PM10 are thought to be responsible for various allergies and for the spread of respiratory diseases. Recent studies have shown that PM2.5 particles can cause chronic heart failure, heart arrhythmias, and strokes, as well as lung damage, cirrhosis, inflammation, cancer, cardiovascular disease, and metabolic disorders. Furthermore, some studies have concluded that PM2.5 particles in the environment are a risk factor for gastrointestinal, liver, colon, and lung cancer as well as it affects the growth and metastasis of various cancer cells caused by other factors. In our country, the health effects of air pollution and the relationship between the pathogenesis of cancer research are scarce. Therefore, the study of the effects of PM2.5 particles on cancer cell proliferation, migration (metastasis) can provide a significant role for cancer treatment, diagnosis, and prevention.@*Purpose@#Determining the effects of PM2.5 particles on cancer cell proliferation, migration (metastasis) in in-vitro@*Material and Methods@#A human liver cancer cell line (HepG2), human gastric cancer cell line (AGS) were obtained from the central scientific research laboratory in the Institute of medical sciences. HepG2, AGS cells were seeded at a concentration of 1*105 cells/mL in a culture flask and cultured in RPMI-1640 medium supplemented with 10% FBS, 1% antibiotic mix (penicillin, streptomycin) in a humidified atmosphere of 5% CO2 at 37 °C. The cytotoxic effect of PM 2.5 in AGS, HepG2 cells were evaluated by MTT, CCK8 assays. AGS, HepG2 cells were incubated in 96 well plates for 24h then treated with different concentrations (0, 5, 10, 25, 50 and 100 μg ) of Bayankhoshuu, Buhiin urguu, and Zaisan samples for 24h, respectively.@*Results@#Concentrations of 10, 25, and 50 μg/ml of samples collected from the Bukhiin urguu and Zaisan in March increased HepG2 cell growth, while doses of 25, 50 μg/ml of samples collected from Bayankhoshuu in March and December increased HepG2 cell growth. Therefore, concentrations of 25 and 50 μg/ml of samples collected from Bayankhoshuu in March increased AGS cell growth, while concentrations of 25, 100 and μg/ml of samples collected in December increased AGS cell growth. However, no cytotoxic effect was observed in the sample collected from Zaisan in March, whereas the PM2.5 sample enhanced AGS cell growth in dose dependent manner in December.(p <0.05) @*Conclusion@#High levels of heavy metals were detected in samples collected in December from Bayankhoshuu, Bukhiin urguu and Zaisan of Ulaanbaatar. Concentration of 25 μg/ml of samples collected from the Bukhiin urguu and Zaisan in March increased HepG2 cell growth. Concentrations of 25 μg/ml of PM2.5 collected from three regions around Ulaanbaatar increased HepG2 and AGS cell migration.

Involvement of Vitamin D in Immune system / Монголын Анагаах Ухаан

@#Research of function of vitamin D on immune system has been studying since the study revealed that vitamin D receptor is expressed on the surface of the immune cells. 1,2-dihydroxyvitamin D3 [1,25(OH)2D], physiologically active form, can be generated through hydroxylation of 25-hydroxyvitamin D3 [25(OH)D], inactive form of vitamin D, in a liver, connecting with specific VDR make biological action. Vitamin D make different biological actions depends on connecting with different immunological cells. Some studies indicated that Vitamin D plays pivotal role in antibacterial innate immune responses through regulating reaction of the main cells as macrophages and dendritic cells. Moreover, calcitriol, the active form of vitamin D, is connected with VDRE, modulates the innate immune response through directly inducing expression of catelicithin and β-defensin as antimicrobial peptides, reducing secretion of IL-1b, IL-6, TNF-a, RANKL, COX-2 as proinflammatory cytokines and increasing production of IL-10, an anti-inflammatory cytokine. Vitamin D plays in proliferation and differentiation of T and B cells and regulates the activities of over 500 genes. Vitamin D differently impacts on per se stages of T cells’ proliferation. Vitamin D indirectly mitigates the differentiation from immature B cells to plasma B cells while it directly impacts on regulation of overloaded production of antibodies in plasma B cells. In conclusion, vitamin D modulates the innate- and adaptive immune response through regulation on activation of APCells, proliferation and differentiation of immune cells, secretion of some antibacterial peptides.

Study on influence of the CpG DNA on activation of IFN-γ signaling transduction regulatory proteins / Монголын Анагаах Ухаан

Introduction@#When human body encounters external pathogens primary/innate immunity cells are activated by recognizing them and secondary/adaptive immunity is activated consecutively. In our previous study, we revealed that there is a synergistic action between TLR9 and IFN-γ signaling in the endothelial cells. @*Purpose@#To determine the role of negative and positive regulator proteins on the IFN-γ/TLR9 signaling pathway. @*Methods@#In this study, murine endothelial cell (END-D) culture was used. END-D cells pre-treated with TLR9 ligand CpG DNA and then stimulated with IFN-γ. The negative (SHP-2, SOCS1, PIAS1) and positive (p38) regulator protein expression was detected by Western blotting. @*Results and Conclusion@#Treatment by TLR9 ligand CpG DNA and IFN-γ increased positive regulator p38 phosphorylation in 0.5 hour. CpG DNA inhibited IFN-γ negative regulator PIAS1 protein expression in 6 hour and SOCS1 and SHP-2 expression could not affect in 4 hour.

Role of negative regulators on the TLR7 ligand/IFN-γ signaling in the endothelial cells / Эрүүл мэндийн лаборатори

Introduction@#Toll like receptors (TLRs) are a class of proteins that key role in the innate immune system. The SOCS1 and SHP2 proteins are negative-feed loop inhibitors of signaling of JAK/STAT and TLRs pathways.@*Purpose@#To determine negative regulator protein activation which is activated through TLR7 ligand/IFN-γ signal transduction in endothelial cells. @*Methods@#We used mouse aortic linear endothelial cell (END-D); protein expressio was detected by western blotting @*Results@#We analyzed a time dependent stimulation effects of negative regulator proteins stimulated by TLR7 ligand/IFN-γ in endothelial cell cultures. Imiquimod of 10 μg/ml treatment of 1 hr was followed by 100 ng/ml IFN-γ stimulation for 1-8hr to analysis of negative regulator SOCS1 and SHP2 protein expression. </br> In untreated cells, there was low activations of negative regulator SOCS1 and SHP2 proteins. IFN-γ stimulation alone had increased SOCS1 and SHP2 protein expressions, also imiquimod treatment highly <i>elevated</i> SOCS1 and SHP2 expressions. However imiquimod and IFN-γ doubled treatment have decreased activation of negative regulator SOCS1 and SHP2 proteins. These findings suggest SOCS1 and SHP2 proteins are inhibitors in the TLR7 ligand/IFN-γ signaling. @*Conclusion@#Negative regulators, SOCS1 and SHP2 strongly suppressed activations of TLR7 ligand/IFN-γ signaling

The effect of TLR9 ligand on IFN-ү signaling / Эрүүл мэндийн лаборатори

Introduction@#The aim of this research project is to elucidate the crosstalk of innate and adaptive immune reactions against the DNA containing bacteria.@*@#This study held in the Core laboratory, Science Technology Center, Mongolian National University of Medical Sciences (MNUMS). Murine aortal endothelial cells, END-D cultured and the cell viability checked by MTT assay. In addition, the NO production, protein and gene expression studied by Griess Reagent assay, R.T-PCR and immunoblotting, respectively. @*Results@#0.1µM, 1µM and 10µM of TLR9 ligand exhibited no cytotoxic action against the cells by MTT assay. IFN-ү alone induced NO production in END-D cells. In the other hand, TLR9 ligand at 0.1µM, 1µM and 10µM up-regulated IFN-ү induced NO production in dose dependent manner. RTPCR results exhibit that TLR9 ligand up regulates iNOS mRNA. Immunoblotting analysis showed the enhanced iNOS protein expression and phosphorylation of STAT1 in cells pre-treated with TLR9 ligand.@*Discussion@#We have demonstrated CpG DNA, TLR9 ligand, up-regulates IFN-ү induced NO via enhanced IFN-ү signaling. The result of Western Blotting and RT-PCR support the up-regulation of NO. CpG DNA can be used as agent against virus and bacteria. Further research need to be conducted. @*Conclusion@#TLR9 ligand, CpG DNA up-regulates IFN-ү induced NO production in time and dose dependent manner. TLR9 ligand augments the expression of iNOS mRNA and STAT1 phosphorylation in response to IFN-ү.

EFFECT OF TLR7 LIGAND ON SIGNAL TRANSDUCTION OF INTERFERON GAMMA / Шинэ санаа Шинэ нээлт

@#BACKGROUND: Toll like receptors (TLRs) are a class of proteins that key role in the innate immune system. TLR7 is expressed on monocytes, macrophages and dendritic cells, T cell, B cell and eosinophiles. TLR7, originally identified as recognizing imidaquinoline, loxibrine, broprimine and ssRNA, ssRNA viruses such as vesicular stomatitis virus, influenza A virus and human immunodefiency virus. It is known that virus ssRNA affects signaling molecule of IFN-y. Objective: To determine gene and protein activation of IFN-y signal transduction by TLR7 ligand in the endothelial cells. MATERIAL: In study we used mouse aortic linear endothelial cell which is cultured (END-D) in 5% heat- inactivated fetal calf serum (FCS), medium (DMEM) containing antibiotic mix(penicillin G, streptomycin, amphotericin B) at 37°C (5% CO2). Endothelial cells treated with synthetic IFN-γ and imiquimodligands, then the NO (nitric oxide) concentration in the supernatant is determined by Griess reagent. Endothelial cells are cultured in 6 well cell culture plate and in each well 2*104cells are expected to be grown for 24 hours of culture. Then, the cells are treated with synthetic IFN-γ and имиквимод ligand for 6 hours and the NO signaling gene activation iNOS mRNA expression which is induced by IFN-γ is determined by RT-qPCR. Endothelial cells are cultured in 12 well cell culture plate and in each well 2*104 cells are expected to be grown for 18 hours of culture. Then, the cells are treated with synthetic IFN-γ and imiquimodligands for 24 hours and the NO signaling protein iNOS expression which is induced by IFN-γ is determined by western blotting. The experiment was conducted as representation mean of at least three test results. The difference between statistical probabilities is determined by the “Students” t test. The p<0.01 value is assumed to be statistically different. RESULTS: TLR7 ligand imiquimodaugmented interferon gamma induced nitric oxide production TLR7 ligand imiquimodincreased interferon gamma induced iNOS mRNA gene expression. TLR7 ilgand imiquimodup-regulated interferon gamma induced iNOS protein expression. CONCLUSIONS: TLR7 ligand imiquimod augments IFN-γ signaling in the endothelial cells. This synergistic effect has revealed in the levels of gene and protein expression.