Background: According to the World Health Organization’s 2022 Global Oral Health Status Report, approximately 3.5 billion people worldwide - nearly half of the global population - suffer from some form of oral disease, among which dental caries is the most prevalent, affecting close to 2 billion people in permanent teeth (global average prevalence of 29%) and 514 million people in primary teeth (global average prevalence of 43%). In Mongolia, the prevalence of dental caries also remains high: a 2017 survey of 1,754 individuals from six provinces and five districts found caries prevalence of 82.9% among children under 5 years of age, 80.9% among those under 12, 97.8% among the population aged 35-44, and 99.1% among those aged 65-74. A 2010 study among Mongolian preschool and school-age children found a caries prevalence of 87.8-90%, with S. mutans identified alone in 75% of culture-positive plaque samples and together with S. sobrinus in the remaining 25%. Given the persistently high global burden of dental caries, preventing the disease rather than merely treating it - for example through “replacement therapy,” in which a less cariogenic strain of S. mutans is used to colonize the oral cavity - requires a reliable rat model in which standard and clinical strains of S. mutans can reproducibly induce dental caries, so that the efficacy of such preventive strategies can be properly evaluated. Untreated dental caries is also estimated to impose an annual global economic burden of approximately US$387 billion in direct costs and US$323 billion in indirect costs, underscoring the growing importance of research into new preventive approaches. Aim: Development of a dental caries model caused by Streptococcus mutans infection in rat teeth. Materials and Methods: This study used an experimental research design. It was conducted from March 30 to April 14, 2021, at the Core Laboratory and the Education Laboratory of the Department of Microbiology and Infection Prevention Control of the Mongolian National University of Medical Sciences (MNUMS). Ethical approval was obtained from the Medical Ethics Review Committee under the Ministry of Health (meeting No. 02, March 18, 2019). Five 8-week-old Wistar rats were divided into four experimental groups - differing by bacterial strain (S. mutans ATCC 25175 standard strain versus a clinical isolate) and by drinking-water composition (with or without sucrose) - plus one negative control group, with one rat assigned to each group. A bacterial suspension was applied to the rats’ molar teeth with a sterile cotton swab three times daily at two-hour intervals for 28 consecutive days. Rats were lightly sedated with midazolam during application, and the suspension was applied during the daytime, when salivary flow is naturally reduced in rodents, in order to promote bacterial colonization of the tooth surface. Result: On day 28, microscopic examination of each rat’s maxillary and mandibular teeth revealed carious spots and cavities in all groups except the sucrose-free negative control. Cavities were deeper in the groups that received sucrose-supplemented drinking water, whereas the sucrose-free groups showed comparatively milder lesions, indicating that bacterial infection alone was sufficient to induce caries. Conclusion Infection of 8-week-old Wistar rats with either the standard or a clinical strain of S. mutans for 28 days successfully induced dental caries, and sucrose intake increased the severity of the lesions. This baseline model can be reproduced within a relatively short time frame and at low cost, and may be applied in future studies evaluating the efficacy of replacement-therapy strains or probiotic products against dental caries.

Background: Peripheral artery occlusion is the final stage of complications in peripheral arterial disease, which has a serious negative impact on the quality of life of patients. A study conducted in Germany found that 66% of the over 40,000 patients diagnosed with peripheral artery occlusion required limb amputation within four years of diagnosis. The primary treatment goals are to improve limb blood flow, alleviate pain, accelerate tissue regeneration, and prevent cardiovascular complications. Recent studies emphasize that microsurgical techniques may accelerate tissue regeneration and improve treatment outcomes. Aim: This study aimed to evaluate and compare the effects of end-to-end (ETE) and end-to-side (ETS) microvascular anastomosis in a rat model of ischemia, focusing on tissue regeneration. Materials and Methods: Thirty Wistar rats were divided into two groups (control group 10 rats, ischemic group 20 rats) at the Bio-Medical Research Institute of the Mongolian National University of Medical Sciences. Ischemia was induced in the experimental group, and ETE and ETS anastomoses were performed using donor arteries, which were preserved for three days. Preoperative and 21-day postoperative body weight, artery size, and blood flow were measured and analyzed to determine the results. Results: In the control group, the average body weight was 227.86±21.11 grams before surgery, which increased by 12.38 grams to 240.24±25.77 grams post-surgery (p<0.01). In the ischemic group, the rats that underwent ETE anastomosis had an average weight of 239.67±26.96 grams before surgery, which decreased to 234.3±27.20 grams immediately after surgery, but increased to 250.67±27.09 grams after 21 days (p<0.05). The ETS group showed an increase from 235.18±22.39 grams to 249.62±21.12 grams after 21 days (p<0.01). The surgical duration for the control group was 24.51±2.29 minutes, while the ischemic group required 39.75±0.89 minutes for ETE and 46.65±8.12 minutes for ETS anastomoses (p=0.001). Intraoperative blood loss in the control group was 0.27±0.12 ml, whereas the ischemic group showed blood loss of 0.58±0.13 ml for ETE and 1.12±0.61 ml for ETS, with some instances reaching 2.35 ml (p<0.01). Conclusions 1. The body weight of the control rats increased to 240.24±25.77 grams after 21 days, while the ischemic group showed an increase to 250.67±27.09 grams, indicating improved tissue regeneration and successful microsurgical anastomosis. 2. Although PCNA and KGF protein expression levels decreased, tissue regeneration was robust, as evidenced by the continued regeneration observed 21 days after the microsurgery, indicating successful microvascular surgery. Differences in the response to ischemic changes were observed across tissues, with increased expression of KGF and PCNA in the skin and muscle tissues, further supporting the effectiveness of the regenerative process.

Introduction: Valproic acid (VPA) has been used in the treatment of seizures and bipolar disorders. In the present study, we examined how VPA affected PI3K-Akt pathway in response to LPS by using mouse RAW 264.7 macrophage cells. Material and methods: Mouse RAW 264.7 macrophage-like cells cultured and the cell viability checked by MTT and TUNEL assay. In addition, protein expression and protein interaction were detected by immune blotting and immune precipitation, respectively. TLR4 expression on cell surface studied by FACS analysis. Results: The MTT and TUNEL assays demonstrated no significant difference between VPA at 2 mM treated and untreated control cells. VPA attenuated LPS-induced phosphorylation of phosphatidylinositol 3-kinase (PI3K) and Akt, but not nuclear factor (NF)-κB and mitogen activated protein kinases (MAPKs). There was no significant difference in the TLR4 expression on the cell surface between cells treated with or without VPA. VPA inhibited LPS-induced PI3K/Akt signal transduction in a dose dependent manner. Conclusion VPA at 2mM exhibits nontoxic effect in the RAW 264.7 cells. VPA down regulates LPS-induced phosphorylation of Akt via inhibition of PI3K activation.

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.
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 elevated 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

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.

BACKGROUND Bovine colostrums is the milk secreted by cows during the first few days after parturition. It contains many essential nutrients and bioactive components, including growth factors, immunoglobulins, lactoperoxidase, lactoferrin and cytokines ets. Lactoferrin has been reported for its multifunctional properties such as antifungal, antibacterial, antiviral antioxidant and anticancer activities. The aims of this study focused on the isolation and purification of lactoferrin from Mongolian bovine colostrums. Lactoferrin purified using HiTrap DEAE an ion exchange chromatography. Lactoferrin purification efficiency was about 60.5%. The single band of purified lactoferrin has been observed in SDS-PAGE electrophoresis. METHODS Bovine colostrum was collected at a cow farm in the Darkhan province of Mongolia. At first the cream was separated by centrifugation (10000 xg 20 min at 4oC). In order to separate the whey, the samples were precipitated with 1mol/l to pH 4.6 and centrifuged at 10000 g 20 min again. The samples of whey were stored at -18oC to the analysis. Lactoferrin was purified by HiTrap DEAE an ion exchange chromatography using 0.005 M phosphate buffer (pH 7.7) and linear gradient NaCl from 0.25M, 0.5M, 1M. During chromatography, protein in the eluents was monitored by ultraviolet absorbation at 280 nm with the instrument. Purity test done by using polyacrylamide gel electrophoresis under denaturated condition (SDS-PAGE) method by Laemmli (1970). For HPLC determination of the lactoferrin by Shimadzu Nexera X2 HPLC system with UV/ VIS detector were used. Detection was carried out at the wavelength 280 nm. Separation was performed on a chromatographic column Protein R C18 ,2.2 x 150 mm, 5 μm particle size. Linear gradient and flow rate 0.2 ml/min were used. Mobile phase a consisted of water / acetonitrile/ trifluoroacetic acid ( 95:5:0.1). The column temperature was set at 40oC and injection volume was 10 μl. Data were collected and evaluated by software Lab Solution. An external standard method for quantification analytes was used. RESULTS Purified lactoferrin in the present study had a good concentration and purification efficiency was about 60.5 %. Protein fraction from 1M NaCl gradient delivers sharp and clean peak to HPLC chromatogram that fits intensity and retention time of standard bovine lactoferrin. Ammount of lactoferrin in bovine colostrums was 0.6 mg/ml and it`s molecular weight 80 kDa as a standard sample. The retention time of lactoferrin fraction which is purified by SDS-PAGE gel electrophoresis. The peak of fraction same compared to the standard lactoferrin 5.8 minutes by HPLC analysis. CONCLUSION Ion exchange chromatography shows reliable and easy isolation of lactoferrin from Mongol bovine colostrum.

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.

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.

Background: In the era of science and technology /development, it have been using a laboratory animals to confirm theory and evidences of biology, medical and veterenary sciences. The value of laboratory animals sciences are still a very important. Because, the studies of laboratory animals are used for instead of human being and biosafety evaluation, and so on. Goal: To describe the current sitution analysis of general usage of laboratory animals in the Mongolian institutes, universities and biotechnological factories. Materials and Method: In this study, we used a questionnaire that is covering belongs to the quantity counting numbers of laboratory animals, information of colonies and breeding, euthanizing methods, infectious disease in the laboratory animals, experimental types, special food and bedding. Results: Each year were used more than 10000 animal of 9 species. Mouse, rat, rabbit and guine pig are commonly used in laboratory experiments in Mongolia. Laboratory animals imported from Russia and China, mainly. These animals uses to study toxicity and virulences, lethal dose of newly revealed substances, drug testing and diagnosis. We have conducted some comparison of laboratory animal used with the same situation of other countries. Conclusions
1. Study results are indicating that the usage of laboratory animals on biological studies among the Mongolian scientists is too low as compaired to other countries.
2. It has no standarzided technical, legal provisions technical legal documents such as SOPs and other douments, but also specialized HRs.
3. Therefore, it has been raised following requirements which are to expand national- and international cooperation, to be a official member in the international association of laboratory animal sciences, to share experiences and reports between the specialists in the Asian- and international level.