The Indonesian Biomedical Journal (Prodia Education and Research Institute)
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microRNA-1 Induces Transdifferentiation of Peripheral Blood CD34+ Cells into Cardiomyocytes-like Cells
Get PDFBACKGROUND: Transdifferentiation is a method to provide cells sources for cellular cardiomyoplasty. CD34+ cells are potential cells sources because these cells can differentiate into cardiomyocytes through several mechanisms. MicroRNA (miR-1) is known to have the ability to inhibit the expression of histone deacetylase 4 (HDAC4). HDAC4 is a gene that essentially contributes in cardiomyocytes differentiation. However, the study reporting an evidence that miR-1 can induce transdifferentiation of CD34+ peripheral blood cells into mature cardiomyocytes is limited.METHODS: CD34+ cells were taken from peripheral blood and isolated using a magnetic-activated cell sorting (MACS) method in vitro. Mature mimics of miR-1 were transfected into isolated CD34+ cells and then incubated for 48 hours for quantification of HDAC4 mRNA using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). On the fifth day after miR-1 transfection, cardiomyocyte-like cells were identified based on their morphology and cardiac troponin expression using immunocytochemistry.RESULTS: Transfection of miR-1 in CD34+ isolated cells decreased HDAC4 gene expression by -0.54 fold at second day and caused a significant increase in percentage of cardiac troponin positive cells (median: 31.34; p<0.05) at fifth-day post-transfection. The efficiency of transdifferentiation was 32%. The miR-1 transfection had a significant negative relationship with HDAC4 gene expression (B=-1.000; p=0.001). HDAC4 gene expression had a negative and significant relationship with the percentage of cardiac troponin-positive cells (B=-0.701; p=0.001).CONCLUSION: This study suggests that miR-1 can induce transdifferentiation of peripheral blood CD34+ cells into cardiomyocytes-like cells by decreasing HDAC4 gene expression.KEYWORDS: transdifferentiation, microRNA-1, CD34, cardiomyocyte, HDAC
Anti-Osteoporosis Potencies of Zingiber officinale Rosc. Rhizome Water Extract and DFA III Produced from Dahlia spp. L.: in vivo and in vitro Studies
Get PDFBACKGROUND: Zingiber officinale Rosc. is estrogenic and thus can be developed as an anti-osteoporosis. Difructose anhydride III (DFA III), possesses anti-osteoporosis potencies. This study aimed to investigate the anti-osteoporosis activity of ginger rhizome water extract (GE) and DFA III from dahlia tubers in ovariectomized (OVX) rat models and to determine their anti-osteoclastogenic effect in vitro.METHODS: This study was conducted using 25 female rats. Blood sampling was carried out at the beginning and end of treatments. Femur bones were isolated after daily 14-day treatments, measured for density, and processed for histological staining. RAW 264.7 cells were induced by osteoclast differentiation factor. A cell viability assay was employed to determine the cytotoxicity of DFA III and GE. The inhibition of osteoclastogenesis was investigated by tartrate-resistant acid phosphatase staining.RESULTS: All groups showed no difference in body weight elevation and serum lipid profiles. The GE and DFA III caused no effect on bone density. However, the GE or DFA III groups showed higher osteoblast numbers compared with the control groups. A significantly less osteoclast was found in the GE+DFA III group. The GE and DFA III showed no toxicity on RAW 264.7 cells. GE showed strong inhibitory effects on the post stimulation osteoclastogenesis model. The combination of GE and DFA III was synergistic in reducing the osteoclastogenesis confluency in RAW 264.7 cells.CONCLUSION: The data support our hypothesis that GE and DFA III can decrease the risk of osteoporosis by osteoclastogenesis inhibition.KEYWORDS: Dahlia spp., estrogenic, ginger, osteoclast, osteoporosis, ovariectomy, RAW 264.7 cel
Single or Divided Administration of Cisplatin Can Induce Inflammation and Oxidative Stress in Male Sprague-Dawley Rats
Get PDFBACKGROUND: Cisplatin is one of the most potent chemotherapy drugs to treat various types of cancer, however the use of cisplatin has some the adverse effect, such as the increase of oxidative stress and inflammation by malondialdehyde (MDA) and nuclear factor kappa B (NF-kB) activation. Since the dosing of cisplatin is critical, we observed the effect of single and multiple doses of cisplatin injection on rats’ inflammation and oxidative stress level.METHODS: Total of 27 male Sprague-Dawley rats were divided into 9 sub-groups, each consisted of 3 rats. The baseline sub-group received no treatments; Group 1 (sub-group 1.1, 1.2, 1.3, and 1.4) were administered one single dose of 5 mg/kg BW/intravenously (i.v) of cisplatin; and Group 2 (sub-group 2.1, 2.2, 2.3, and 2.4) were given 0.2 mg/kg BW/i.v of cisplatin twice a week for two months. Rats were observed for their body weight, NF-kB, and MDA level based on the assigned group. RESULTS: Body weight loss was observed in the 1st week after treatment for Group 1, and 7th week for Group 2. Group 1 and Group 2 showed increasing level of NF-kB and MDA since the 1st observation, which was the 1st week and 5th week, respectively. NF-kB and MDA and levels were also significantly increasing in both groups for every week of observation (p<0.05).CONCLUSION: Cisplatin injection either in single or divided dose can induce inflammation and oxidative stress thus decrease the body weight. However, dividing cisplatin in smaller dose can delay the inflammation effect on subjects.KEYWORDS: cisplatin dose, MDA, NF-kB, body weight, inflammation, oxidative stress
Dioscorea alata L. Tubers Improve Diabetes through Anti-hyperglycemia, Anti-inflammation, Ameliorate Insulin Resistance and Mitochondrial Dysfunction
Get PDFBACKGROUND: Dioscorea alata L. tubers (DA) are suspected to prevent diabetes mellitus (DM) because they have a low glycemic index. However, only a few reports about the anti-diabetic effect of DA were reported up to date. This study aims to analyze the effect of DA consumption on several diabetic biomarkers through in vitro, in vivo, and in silico analysis.METHODS: In vitro experiments were conducted by observing the anti-inflammatory activity of DA extract, steroidal saponins (SDA) isolated from DA, and diosgenin in lymphocyte cell cultures. The tumor necrosis factor (TNF)-α and interferon (IFN)-γ percentages were analyzed by flow cytometry. In vivo study involved 24 healthy adolescents that were given a boiled DA 10 hours post-prandial. The blood sugar levels were measured at 0, 30, 60, and 120 min after treatment. Furthermore, the in silico study was carried out by analyzing the active compounds and predicting the biological activity, the target proteins, and interactions of target proteins with compounds contained in DA .RESULTS: DA extract, SDA isolated from DA, and diosgenin at 50 µg/mL significantly reduced pro-inflammatory cytokines TNF-α and IFN-γ in lymphocyte cell culture. The blood glucose levels in the DA group were lower at 30 and 60 min after treatment. Based on the in silico study, the anti-diabetic activity of DA was speculated to be attributed to the mechanisms of anti-hyperglycemia, prevention of mitochondrial dysfunction, anti-inflammation, and treated insulin resistance. Several proteins included in the DM pathway became the protein target of compounds contained in DA.CONCLUSION: DA potentially have an anti-diabetic activity through several mechanisms.KEYWORDS: hyperglycemia, inflammation, insulin resistance, ya
Immunomodulatory Activity of Agarwood Aquilaria malaccensis Lamk. Leaf Extracts on Staphylococcus aureus-infected Macrophages in vitro
Get PDFBACKGROUND: Aquilaria malaccensis has been consumed as herbal medicine, and in vitro study showed that the leaf extract possesses high antioxidant activities. A brief preliminary study indicated that A. malaccensis showed a promising immunomodulatory activity when evaluated using latex beads. This current study aimed to evaluate the immunomodulatory activity of A. malaccensis leaf extract on the macrophage, which was challenged with pathogenic bacteria Staphylococcus aureus.METHODS: Bioactivity was determined by evaluating the phagocytic capacity of macrophages isolated from Mus musculus against S. aureus. First, the cytotoxicity of extracts on macrophages was evaluated using MTT assays, and the IC50 value was used to determine the dose of immunomodulatory assays. The highest toxicity was observed on chloroform extract with an IC50 value of 111.4 µg/mL. Therefore, the treatment was 100 and 50 µg/mL. Two parameters, including the phagocytic activity and phagocytic capacity of macrophages infected with S. aureus, were used to evaluate immunomodulatory activity. The analysis of variance was done at p<0.05 to determine the significant difference among treatments.RESULTS: Chloroform and ethanol extracts at a 50 µg/mL concentration showed the best results with the phagocytic activity of 82.33%±9.61% and 80.33±1.53%. The ethyl acetate showed lower phagocytic activities of 70.67±1.53. All extracts significantly increased phagocytic activity and phagocytic capacity, and the results differed significantly between negative and positive controls. Thin-layer chromatography indicated that the extract contained terpenoid, flavonoid, phenolic, and tannin.CONCLUSION: A. malaccensis leaf extracts showed immunomodulatory activity. Both chloroform and ethanol extracts showed comparable activity, while the ethyl acetate extract was lower. The extracts contained diverse bioactive compounds that may support activating macrophage cells for immunomodulatory activity.KEYWORDS: Aquilaria malaccensis, immunomodulator, phagocytosis, macrophages, Staphylococcus aureu
Probiotic Lactobacillus acidophilus FNCC 0051 Improves Pancreatic Histopathology in Streptozotocin-induced Type-1 Diabetes Mellitus Rats
Get PDFBACKGROUND: Intestinal microbial dysbiosis and its metabolites can affect the immune activity of intestinal mucosal cells, causing insulitis and pancreatic β-cell death. Probiotic Lactobacillus acidophilus plays an important role in reducing inflammatory cytokines, hence improves oxidative stress that affects pancreatic β-cell apoptosis. Current study examined the feature of pancreatic histopathology affected by the administration of probiotic L. acidophilus in rats with type-1 diabetes mellitus (DM) induced by streptozotocin (STZ).METHODS: Twelve rats were induced by STZ at double dose of 50 mg/kgBB before administered with probiotic L. acidophilus at a dose of 1.5x10 8 or 1.5x10 9 CFU/mL/day, while other 4 rats were used as control. After 21 days of the L. acidophilus treatment, the average of fasting blood glucose (FBG) levels of rats were measured, then the pancreatic histopathology was assessed to evaluate the degree of insulitis in islet of Langerhans.RESULTS: The induction of STZ had been succeeded to increase blood glucose levels, which indicate DM condition. The highest FBG level after 21 days of treatment was found in DM group with glucose level of 512±81.51 mg/dL. The administration of probiotic L. acidophilus during 21 days treatment at both dose 1.5x10 8 and 1.5x10 9 CFU/mL/day significantly improved pancreatic histopathology (p=0.04 and p=0.034, respectively), with significant decrease on insulitis scores compared to DM group.CONCLUSION: The administration of L. acidophilus at both dose of 1.5x10 8 and 1.5x10 9 CFU/mL/day for 21 days can improve pancreatic histopathology of type-1 DM rats induced by STZ, therefore probiotic L. acidophilus may be potential as supplementation treatment for type-1 DM.KEYWORDS: Lactobacillus acidophilus, pancreatic histopathology, streptozotocin, type-1 diabetes mellitu
Increased Platelet-derived Microparticles Counts is Correlated with Elevated Blood LDL Cholesterol in Acute Myocardial Infarction
Get PDFBACKGROUND: Platelet-derived microparticles (PDMPs) and low-density lipoprotein (LDL) cholesterol are contributing factors to acute myocardial infarction (AMI). However, the association between LDL cholesterol and PDMPs in AMI has not fully discovered. This study assessed the correlation between these two parameters in patients diagnosed with AMI.METHODS: This was an observational cross-sectional study involving 95 subjects with AMI. The blood measurement of PDMPs counts and LDL cholesterol levels were conducted concomitantly within 24 hours of admission. PDMPs count was analyzed by flow-cytometry method, meanwhile the LDL cholesterol was measured with enzymatic and colorimetric methods. For further analysis, subjects were further divided into LDL cholesterol level ≥130 mg/dL and <130 mg/dL. A statistical test was conducted for a correlative and comparative analyses.RESULTS: A correlative analysis to assess the association between PDMPs counts and LDL cholesterol level depicted a low but significant positive correlation (r=0.231, p=0.024). Furthermore, mean PDMPs counts was significantly higher in subjects with LDL cholesterol level ≥130 mg/dL compared to LDL cholesterol level <130 mg/dL (12,499.59 (95% CI: 8,507.44-16,491.74) counts/μL vs. 9,267.23 (95% CI: 4,445.45-14,089.01) counts/μL; p=0.039).CONCLUSION: There was a significant correlation between PDMPs counts and LDL cholesterol levels in AMI. A significantly increased PDMPs counts were found in subjects with LDL cholesterol level ≥130 mg/dL. Therefore, it is recommended to measure PDMPs in patients with high LDL cholesterol levels as both might be significant AMI biomarkers.KEYWORDS: acute myocardial infarction, LDL-cholesterol, platelet microparticles, platelet activatio
Caffeic Acid Induces Apoptosis in MG-63 Osteosarcoma Cells via Protein Kinase C Delta (PKCδ) Translocation and Mitochondrial Membrane Potential Reduction
Get PDFBACKGROUND: Caffeic acid has been reported to activate caspases in MG-63 osteosarcoma cells, which can lead to apoptosis via both extrinsic and intrinsic apoptotic pathways. Translocation of protein kinase C delta (PKCδ), which reduces mitochondrial membrane potential (ΔΨm), is involved in apoptosis. The role of PKCδ translocation and ΔΨm alteration in caffeic acid-induced MG-63 cell apoptosis are largely unknown. Present study investigated the effect of caffeic acid on PKCδ translocation and ΔΨm in MG-63 cells.METHODS: MG-63 cells were cultured and starved, followed by pretreatment with or without Z-VAD-FMK and treatment with or without 10 μg/mL caffeic acid. MG-63 cells were collected, lysed, and processed to obtain cytosolic and mitochondrial fractions. Each fraction was subjected to immunoblotting analysis by using anti-PKCδ antibody. Mitochondrial membrane potential (ΔΨm) was measured using flow cytometry.RESULTS: Cytosolic PKCδ levels were higher than mitochondrial PKCδ levels in untreated and 1 h caffeic acid treatment groups. Inversely, cytosolic PKCδ levels were lower than the mitochondrial PKCδ levels after 6 and 12 h caffeic acid treatment. By Z-VAD-FMK pretreatment, cytosolic PKCδ levels were higher than mitochondrial PKCδ after 6 and 12 h caffeic acid treatment. After 6 h treatment with caffeic acid, ΔΨm was slightly shifted. More shifting occurred in MG-63 cells treated with caffeic acid for 12 h. The ΔΨm shifting was inhibited by Z-VAD-FMK pretreatment.CONCLUSION: Caffeic acid could trigger apoptosis of MG-63 osteosarcoma cells by inducing PKCδ translocation to mitochondria and reducing ΔΨm, which might cause MMP.KEYWORDS: caffeic acid, MG-63, osteosarcoma, PKCδ, mitochondrial membrane potential, mitochondrial membrane permeabilization, Z-VAD-FM
The Role of Klotho G395A Gene Polymorphism in Atherosclerotic Cardiovascular Disease and Mortality Risk Scores in Non-dialysis Chronic Kidney Disease
Get PDFBACKGROUND: Cardiovascular disease (CVD) is the leading cause of death in patients with chronic kidney disease (CKD). Klotho expression was reduced in patients with CKD, leading to vascular calcification, endothelial dysfunction, and atherosclerosis. We investigated the role of the klotho G395A gene polymorphism and plasma klotho level in the ten-year risk of atherosclerotic cardiovascular disease (ASCVD) and CVD mortality in CKD patients.METHODS: We used the PCR-CTPP assay method to genotype klotho G395A single nucleotide polymorphism (SNP) in 72 non-dialysis CKD patients. The klotho level was determined using the enzyme-linked immunoassay (ELISA) method. Path analysis was used to determine the relationship between the klotho G395A SNP, plasma klotho level, ASCVD risk score, and CVD mortality risk score.RESULTS: Our results showed that the GA genotype had lower plasma klotho levels than the GG genotype (path coefficient=-0.185, p=0.000). There was a significant negative correlation between plasma klotho level and the ASCVD risk score (r=-0.243, p=0.040), but no significant correlation was found between plasma klotho level and the CVD mortality risk score (r=-0.145, p=0.225). Path analysis showed that plasma klotho level had a significant negative direct effect on ASCVD risk score (path coefficient=-0.272, p=0.000) and an indirect effect on CVD mortality risk score (path coefficient=0.187, p=0.005).CONCLUSION: Klotho G395A SNP might reduce lower plasma klotho levels, which increased ASCVD and CVD mortality risk scores in non-dialysis CKD patients. However, other risk factors such as age, CKD stages, hypertension, and smoking should be taken into consideration. Therefore, large-scale genetic association studies with adjusted variables could be conducted in various ethnic groups for a more robust result.KEYWORDS: klotho, single nucleotide polymorphism, cardiovascular disease, chronic kidney diseas
Caffeic Acid Induces Intrinsic Apoptotic Pathway in MG-63 Osteosarcoma Cells Through Bid Truncation and Cytochrome c Release
Get PDFBACKGROUND: Caffeic acid has been reported to induce apoptosis in MG-63 osteosarcoma cells via caspases activation. However, apoptotic pathway that is involved in the caffeic acid-induced apoptosis is still unclear. Present study aimed to investigate the role of cytochrome c (Cyt c) release and BH3-interacting death (Bid) activation in caffeic acid-induced apoptosis in MG-63 osteosarcoma cells.METHODS: MG-63 cells were cultured, pretreated with/without Z-VAD FMK and treated with/without 10 μg/mL caffeic acid. Treated MG-63 cells were then lysed, homogenized, and processed further to prepare cell lysate and mitochondrial fraction. Immunoblotting method was used to measure the amount of Bid and truncated Bid (t-Bid) as well as mitochondrial and cytosolic Cyt c.RESULTS: The amount of Bid and mitochondrial Cyt c in MG-63 cells decreased in a time-dependent manner, while the amount of t-Bid and cytosolic Cyt c increased in a time-dependent manner. By pretreatment of 100 μM Z-VAD-FMK for 2 h, the amount of Bid and mitochondrial Cyt c was significantly higher, while the amount of t-Bid and cytosolic Cyt c was significantly lower after caffeic acid treatment for 6 and 12 h compared to MG-63 cells that were not pretreated.CONCLUSION: Caffeic acid could induce Cyt c release through the activation of Bid in MG-63 osteosarcoma cells.KEYWORDS: caffeic acid, osteosarcoma, MG-63 cells, Bid, t-Bid, cytochrome c, Z-VAD-FM