PCS-16 Determaining Surgical Method by Meniscectomy Induction on Garut sheep (Ovis aries) for early stage of Osteoarthritis

Osteoarthritis (OA) is the most common joint disease that cause of pain and disability by various factors such as advanced age, obesity, trauma, and arthritis disease. These factors affect by degeneration of the cartilage surface, leading to loss of matrix include proteoglycan osteophyte formation, subcondral and synovial membrane affected. In the healthy joint, meniscus, articular cartilage, subchondral bone, and synovial membrane provide support to the joint. The meniscus is an important load bearing structure and has nutritive as well as lubricating properties in the knee joint as well (Little et al. 2010).Animal models are research materials that can be used in studying potential pathogenesis and therapy in various diseases in humans. Sheep are commonly large animal model of OA because of the availability, ease of handling, and have a similarities with humans in size and structure of joint. In the development of science, sheep can be used as an animal model in studying the pathogenesis of diseases in human orthopedics studies such as joints, ligaments, and bones. Garut sheep is an Indonesian germplasm indigenous that has the structure, density, and size of joint anatomy that are similar in human joints rather than other small animals. This is the basis of the utilization of Garut sheep as an animal model in human orthopaedic. (Little et al. 2010; Gregory et al. 2012).The aim of this study was to identify and analyze the determining surgical method by meniscectomy induction on Garut sheep with 8 weeks post meniscectomy observation for early stage of OA

Optimization of Activation Methods for Mouse Oocytes Using Calcium-free CZB Medium, SrCl2, and Cytochalasin B in Vitro

Embryonic stem cells can be obtained by generating an embryo through fertilization; however, an embryo can also be generated asexually through parthenogenesis. This procedure will overcome the ethical issues regarding the use of embryos initially generated for reproductive purposes. The aim of this study was to obtain an optimized oocyte activation method through parthenogenesis by using mice oocytes as a model. Ten mM SrCl2 and 5 µg/ml Cytochalasin B (CB) in calcium-free Chatot Ziomek Bavister (CZB) were used as a medium for an in vitro activation of mouse oocytes. Treatment combinations for the oocyte activation methods were (A) activation in CZB & SrCl2 (prepared in stock) for two hours and in CZB & CB for four hours; (B) activation in CZB & SrCl2 (fresh medium) for two hours and in CZB & CB for four hours; and (C) activation in CZB & CB & SrCl2 (fresh medium) for six hours. The results show that the activation rate of  mouse oocytes  with  method C has  been  the best among all the protocols. This optimized protocol clearly provides a new insight in the generation of embryos for further use, particularly for producing embryonic stem cells

Induction of Matrix Metalloproteinases in Chondrocytes by Interleukin IL-1β as an Osteoarthritis Model

Osteoarthritis (OA) is a chronic disease of the joints and bones due to trauma or other joint-related diseases (secondary). Synovial inflammation commonly causes disturbance in joint homeostasis, which is associated with OA. Enzymes such as aggrecanase and metalloproteinase generate cartilage damage, mediated by tumor necrosis factor (TNF-α) and interleukin (IL)-1. Pro-inflammatory cytokines, including TNF-α, IL-1β, and IL-6, are responsible for regulation of the extracellular matrix, cartilage degradation, and apoptosis of chondrocytes. This study aimed to observe the cell viability and expression level of matrix metalloproteinases (MMP-1 and MMP-3) and tissue inhibitor metalloproteinases (TIMP-1 and TIMP-2) in human chondrocyte cells (CHON-002) induced by IL-1β. CHON-002 was induced with IL-1β (0.1, 1 and 10 ng/mL) as an OA model. The viability of the cells was measured with a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxyme-thoxyphenyl)-2-(4-sulfophenyl)-2H-tetra zolium (MTS) assay, while expression of MMP-1, MMP-3, TIMP-1, and TIMP-2, was evaluated by RT-PCR. The viability of IL-1β-induced CHON-002 (CHON-002- IL-1β) cells at day 1 and 5 showed that treatment with up to 10 ng/mL of IL-1β was not toxic. Expression of TIMP-1 and TIMP-2 in CHON-002-IL-1β was lower compared to control, while that of MMP-1 and MMP-3 was higher compared to control. These results indicate that CHON-002 treated with 10 ng/mL IL-1β has expression patterns consistent with chondrocyte damage, so the CHON-002-IL-1β system may serve as a model for MMP induction in OA

Effects of Conditioned Medium of Co-Culture IL-2 Induced NK Cells and Human Whartonâs Jelly Mesenchymal Stem Cells (hWJMSCs) on Apoptotic Gene Expression in a Breast Cancer Cell Line (MCF-7)

Breast cancer (BC) is the most prevalent type of cancer among women and one of the major causes of cancer mortality in women. Metastasis in breast cancer (BC) occurs due to immunosurveillance deficiency, including impairment of natural killer (NK) cell maturation. Conditioned medium (CM) from human Wharton's jelly mesenchymal stem cells (hWJMSC-CM) is known to possess anticancer activity. The CM of co-culture of human recombinant IL-2 treated NK cells and hWJMSCs is expected to boost anticancer activity toward BC cells which can be analyzed from the effect of CM towards secretion of effector molecules and expression of BC cell apoptosis-related genes, and cytotoxic granules in human recombinant IL-2 treated NK (IL-2 NK) and hWJMSCs (IL-2 hWJMSCs). TNF-α, IFN-γ, perforin, granzyme were measured by ELISA, while the inhibition of cell proliferation was measured by MTS assay and BC cell apoptosis by flow cytometry and apoptotic gene expression by RTPCR. CM from co-cultured hWJMSCs and IL-2 NK cells inhibited NK and BC cell proliferation, increased expression of Bax and p53 and decreased the number of Bcl-2 in BC cells. In conclusion, CM of co-culture IL-2 treated NK cells and hWJMSCs induce apoptosis in BC cells as indicated by increased Bax and p53 expression and decreased Bcl-2 expression.

Effects of insulin-like growth factor-induced Wharton jelly mesenchymal stem cells toward chondrogenesis in an osteoarthritis model

Objective(s): This study aimed to determine the collagen type II (COL2) and SOX9 expression in interleukin growth factor (IGF-1)-induced Wharton’s Jelly mesenchymal stem cells (WJMSCs) and the level of chondrogenic markers in co-culture IGF1-WJMSCs and IL1β-CHON002 as osteoarthritis (OA) cells model. Materials and Methods: WJMSCs were induced with IGF1 (75, 150, and 300 ng/ml) to enhance their chondrogenesis capability. The gene expression of SOX9 and COL2 was evaluated with quantitative RT-PCR. Furthermore, IGF1-WJMSCs were co-cultured with IL1β-CHON002 cells in varied ratios (1:2, 1:1, 2:1). Chondrogenic markers ADAMTS1, ADAMTS5, MMP3, MMP1, and RANKL were measured with ELISA. Results: The IGF1-WJMSCs had an increased expression of COL2 and SOX9. ADAMTS1, ADAMTS5, MMP1, MMP3, and RANKL levels were decreased in the co-culture IGF1-WJMSCs and IL1β-CHON002. Conclusion: The IGF1-induced WJMSCs were capable to enhance chondrogenesis, indicated by increased expression of SOX9 and COL2 and decreased expression of ADAMTS1, ADAMTS5, MMP3, MMP1, and RANKL. These findings can be further used in the osteoarthritis treatment

EFFECT OF FLAVONOIDS ON OXIDATIVE STRESS, APOPTOSIS, AND CELL MARKERS OF PERIPHERAL BLOOD-DERIVED ENDOTHELIAL PROGENITOR CELLS: AN IN VITRO STUDY

Objective: Circulating EPCs (endothelial progenitor cells) play a role in neovascularization and vascular repair. Oxidative stress impairs endothelial progenitor. Flavonoid is a phytochemical compound for antioxidant activity. Flavonoid effects toward oxidative stress, apoptosis, and expression of the cell markers on EPCs are not fully understood. This study was aimed to elucidate the effects of quercetin, kaempferol, and myricetin toward oxidative stress, apoptosis, and cell markers of peripheral blood-derived-EPCs. Methods: EPCs (endothelial progenitor cells) were isolated from peripheral blood mononuclear cells (PBMNCs) using cultivation under EPCs spesific media. Oxidative stress in EPCs was induced by H2O2 and then treated by quercetin, kaempferol, and myricetin. Cytotoxicity was measured by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, while intracellular reactive oxygen species (ROS), apoptosis and characterization of cells, which expressed CD133 and KDR, was measured using flow cytometry. Results: Quercetin, kaempferol, and myricetin at concentration 12.50 µmol/l were not toxic on EPCs as the cells viability were 96.11±4.03%, 95.42±7.75%, and 94.22±9.49%, respectively. Flavonoids decreased intracellular ROS level in EPCs (quercetin: 14.38±1.47%, kaempferol: 20.21±6.25%, and myricetin: 13.88±4.02%) compared to EPCs treated with H2O2 (30.70%±1.04). Percetage of EPCs apoptosis was not significantly different among each treatment. Immunophenotyping showed the increasing of CD133 and KDR expression in EPCs treated with flavonoids. Conclusion: Quercetin, kaempferol, and myricetin were safe for EPCs, decreased ROS levels, and increased CD133 and KDR expression. However, the flavonoids did not significantly affect EPCs apoptosis

Kinetics Study of Yttrium Leaching from Zircon Tailings Using Sulfuric Acid

From the analysis of zircon tailings using X-Ray Fluorescence (XRF), Yttrium is a rare earth element (REE) with the highest concentration compared to other REEs. The purpose of this study is to determine the best kinetic model for describing how sulfuric acid extracts Yttrium from zircon tailings. Leaching temperatures of 200, 250, and 300 °C were used to determine the kinetics. Samples were obtained at 0, 20, 40, 60, 80, 100, and 120 min for each temperature. This study discovered that the chemical reaction model's kinetics are the most closely related to those of the leaching process. The evaluation of the model utilizing the coefficient of determination (R2) on the relationship between each model and time lends support to this conclusion. The activation energy (Ea) of the leaching process is determined by the Arrhenius plot between ln k and 1/T. In the Yttrium leaching procedure, the Ea value is 14.42 kJ/mol. The chemical reaction model was in charge of the leaching process, according to the Ea value. The premise of the chemical reaction model is that chemical reactions regulate the rate of the reaction

Direct and Indirect Effect of TNFα and IFNγ Toward Apoptosis in Breast Cancer Cells

Background: Breast cancer (BC) is the leading cause of death cancer in women. Cancer therapies using TNFα and IFNγ have been recently developed by direct effects and activation of immune responses. This study was performed to evaluate the effects of TNFα and IFNγ directly, and TNFα and IFNγ secreted by Conditioned Medium-human Wharton’s Jelly Mesenchymal Stem Cells (CM-hWJMSCs) toward apoptosis of BC cells (MCF7).Materials and Methods: BC cells were induced by TNFα and IFNγ in 175 and 350ng/mL, respectively. CM-hWJMSCs were produced by co-culture hWJMSCs and NK cells that secreted TNFα, IFNγ, perforin (Prf1), granzyme B (GzmB) for treating BC cells. The BC cells were treated with CM-hWJMSCs in 50%. The expression of apoptotic genes Bax, p53, and the antiapoptotic gene Bcl-2 were determined using RT-PCR.Results: TNFα and IFNγ at concentration of 350 ng/mL induced higher Bax expression compared to 175 ng/mL. TNFα and IFNγ 350 ng/mL, 175 ng/mL induced p53 expression, whilst TNFα and IFNγ at 350 ng/mL decreased Bcl-2 expression. Perf1, GzmB, TNFα and IFNγ-containing CM-hWJMSCs induced significantly apoptosis percentage, induced Bax expression, but did not effect p53, Bcl-2 expression.Conclusion: TNFα and IFNγ directly induce Bax, p53, decrease Bcl-2 gene expression. The Prf1, GzmB, TNFα, IFNγ-containing CM-hWJMSCs induce apoptosis and Bax expression.Keywords: breast cancer, Wharton’s Jelly mesenchymal stem cells, TNFα, IFN

Effect of interleukins (IL-2, IL-15, IL-18) on receptors activation and cytotoxic activity of natural killer cells in breast cancer cell

Introduction: Breast cancer is one of the leading cause of cancer deaths in women. Metastasis in BC is caused by immunosurveillance deficiency, such NK cell maturation, low NK activity and decreasing cytotoxicity. This study was performed to improve activating receptors and cytotoxicity of NK cells using interleukins (ILs). Methods: Human recombinant IL-2, -15, and -18 were used to induce NK cells. We measured the activating and inhibiting receptors, proliferation activity of NK cells, and the cytotoxicity of NK cells on BC cells (MCF7). The effects of ILs were tested on the NK cell receptors CD314, CD158a and CD107a with flowcytometry, proliferation at various incubation times with 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy methoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and concentrations of TNF-\u3b1 and IFN-\u3b3 by NK cells with ELISA. Results: ILs increased NK cell receptor levels (CD314, CD158a, and CD107a) at 24 hours of incubation. ILs increased NK cell viability, which increased with longer incubation. Moreover, ILs-induced NK cells inhibited proliferation in MCF7 cells, as well as increased TNF-\u3b1, IFN-\u3b3, PRF1 and GzmB secretion. Conclusion: IL-2, IL-15, and IL-18 improved activating receptors and proliferation of NK cells. IL-induced NK cells increased TNF-\u3b1, IFN-\u3b3, PRF1 and GzmB secretion and cytotoxic activity on BC cells. High NK cell numbers increased BC cell growth inhibition