The Indonesian Biomedical Journal

349-57

Authors: Marwa Magdy Hamido, Nashwah Ismail Zaki, Sawsan Ahmed Nasr, Wael Mohamed El-Sayed

BACKGROUND: The interaction of carbamazepine (CBZ) and acetaminophen (APAP) could result in hepatic failure and mortality. This study was conducted to analyzed the potential of rosemary ethanol extract (REE) or coenzyme Q10 (CoQ10) to alleviate the interactions between CBZ and APAP. METHODS: Fourty-eight adult male rats were treated differently based on the assigned groups. Oxidative stress parameters, including malondialdehyde (MDA), glutathione (GSH), glutathione peroxidase (GPx), superoxide dismutase (SOD), catalase, and glutathione S-transferase (GST), and the expression levels of CYP3A4, CYP2E1, IL-6, TNF-α, and IL-1B in the liver were estimated. In addition, the histopathology of liver was examined and the plasma clearance rate of CBZ and APAP was estimated. RESULTS: Combination of CBZ and APAP significantly elevated alanine aminotransferase (ALT) activity and hepatic MDA, and reduced the activities of GPx, GST, and GSH level in liver. The gene expression of CYP3A4 and CYP2E1 was upregulated by CBZ and CoQ10, respectively. The expression of IL-6 has decreased in the groups treated with CBZ alone or in combination with APAP. TNF-α expression was significantly downregulated in the groups treated with CBZ, APAP, REE, CoQ10, or combination CBZ and APAP. The liver from CBZ and APAP combination group showed centrolobular degeneration and necrosis. REE and CoQ10 were able to alleviate most of these detrimental effects. The combined administration of CBZ and APAP extended the plasma clearance time of APAP and CBZ from 6 to 24 and from 9 to 24 hours, respectively. CONCLUSION: REE and CoQ10 alleviated the detrimental effects of the combination of CBZ and APAP through enhancing the cellular antioxidant milieu, induction of metabolizing enzymes, reduction of the plasma half-life of APAP and CBZ preventing their accumulation and potential interaction.

365-75

Authors: Sri Nabawiyati Nurul Makiyah, Masaki Kita, Ika Setyawati, Sri Tasminatun

BACKGROUND: 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.

382-92

Authors: Soraya Rahmanisa, Ekavianty Prajatelistia, Indra Wibowo, Anggraini Barlian

BACKGROUND: Biosilica derived from Indonesian marine sponge Melophlus sarasinorum and Xestospongia testudinaria is one of the biomaterials that can be developed together with synthetic polymer as a composite. Poly E-caprolactone (PCL) used as a composite role as an osteoconductive material together with biosilica and also tailored the slow rate of degradation in the body. This study aimed to create a biocompatible biosilica-based scaffold and supports osteogenic differentiation of human Wharton's Jelly mesenchymal stem cell (hWJ-MSCs). METHODS: Biosilica was extracted from M. sarasinorum and X. testudinaria with the acid digestion method. Scaffold was prepared using the salt leaching method. The composite scaffolds were made from seven different biosilica extract and PCL. All of the scaffolds were tested for the cell morphology, Fourier-transform infrared spectroscopy (FTIR), immunocytochemistry, and cytotoxicity. RESULTS: Composite scaffolds of 50% M. sarasinorum and X. testudinaria increased the cell viability and supported the cell growth within 14 days, whereas the osteogenic differentiation can be seen by the presence of collagen type 1 in day 12 based on immunocytochemistry result. CONCLUSION: The biosilica scaffolds from PCL+50% M. sarasinorum and PCL+50% X. testudinaria were promising 3D scaffolds for potential application in bone tissue engineering. In conclusion, this study shows evidence for the osteogenic differentiation of hWJ-MSC, which might be developed for bone tissue engineering.

401-9

Authors: Banun Kusumawardani, Larissa Tania, Ari Satia Nugraha

BACKGROUND: Cervical cancer has a high rate of morbidity and mortality in women with cancer. Recent studies have found that tobacco (Nicotiana tabacum L.) is a potential source of anti-cancer agents. Hence, this study was conducted to determine the potential of Kasturi tobacco leaf resinoids as apoptotic agents against cervical cell malignancies, since it has not been fully elucidated before. METHODS: The phytochemical diversity of Kasturi tobacco resinoids was generated by gas chromatography-mass spectrometry (GC-MS) analysis followed by spectral similarity to National Institute of Standards and Technology (NIST) database. Cytotoxicity and proliferative activity of HeLa cells treated with Kasturi tobacco resinoids at various concentrations were evaluated by MTT assay. The expression of Caspase-3, cyclooxygenase-2 (COX-2) and heat shock protein 90 (HSP-90) in HeLa cells was analyzed by immunocytochemistry. Next, the migration ability of HeLa cells was observed by the scratch method. RESULTS: Kasturi tobacco resin contains 4,8,13-cyclotetradecatriene-1,3-diol, 1,5,9-trim with α-2,7,11-cembratriene-4,6-diol (α-CBD) structure in the form of a diterpenoid compound with the chemical formula C20H34O2 and a molecular weight of 306 Da. Kasturi tobacco resinoid with IC50 value of 2500 μg/mL inhibited proliferative activity during 72 hours. At a concentration of 1¼ IC50 and incubation for 48 hours, Caspase-3 expression increased by 74.1%, while COX-2 and HSP-90 expression decreased by 28.3% and 26.1%, respectively. HeLa cell migration was inhibited by Kasturi tobacco resinoid at 24 hours incubation. CONCLUSION: Kasturi tobacco resinoids with a concentration of 1¼ IC50 have potential as cervical anti-cancer agents by increasing Caspase-3 expression and decreasing COX-2 and HSP-90 expression within 48 hours.

416-20

Authors: Ferry Sandra, Dewi Ranggaini, Laifa Annisa Hendamin, Nurrani Mustika Dewi, Melanie Sadono Djamil

BACKGROUND: Formation of tumour mass is one symptom of osteosarcoma development. Caffeic acid has been known to provide effective treatment but has less side effect for some cancer therapy. Studies reported that caffeic acid might promote apoptosis in MG-63 osteosarcoma cells, however, the effect of caffeic acid treatment in preventing tumour mass formation has not been well elucidated, especially in MG-63 cells-induced nude mice in vivo. METHODS: MG-63 cells were pre-treated with 0, 1, or 10 µg/mL caffeic acid, and 6 hours after pre-treatment, MG-63 cells were injected into subcutaneous space of mice to induce osteosarcoma. Another model was also created by subcutaneously injecting MG-63 cells to the back of mice, and after 48 days, the visible tumour mass was injected intra-tumour with 0 or 10 µg/mL caffeic acid every 7 days for 6 times. After 90 days, mice were anaesthetised, and the nodule pictures were taken for observation and measurement. RESULTS: In pre-treated MG-63 cells-induced mice, volumes of the mass decreased in reverse with the dose of caffeic acid given. Ten µg/mL caffeic acid pre-treatment was able to significantly lower the mass volume compared to the untreated (p<0.05). Meanwhile, the intra-tumour treatment of 10 µg/mL caffeic acid, even though not significant, was able to inhibit tumour mass formation. CONCLUSION: Results of caffeic acid pre-treatment and caffeic acid treatment in tumour mass of mice show that caffeic acid is able to inhibit the MG-63 cells formation. This suggests that caffeic acid can be a potential anti-cancer agent.

429-35

Authors: Jonathan Alvin Nugraha Halim, Endang Sri Lestari, Sigit Adi Prasetyo, Muflihatul Muniroh, Agung Aji Prasetyo

BACKGROUND: Cholestasis caused by obstruction of the common bile duct and may developed gut-derived sepsis due to reactive oxygen species (ROS) accumulation. Ursodeoxycholic acid (UDCA) and glutathione are widely known for their antioxidant properties, that might be beneficial against ROS. However, the effects of UDCA-glutathione combination against ROS have not been well elucidated in previous studies. Thus, this study was conducted to evaluate tumor necrosis factor (TNF)-α level and height of terminal ileal mucosal villus after UDCA-glutathione administration in cholestasis rat model. METHODS: Twenty-eight male Sprague Dawley rats were randomly grouped into four treatment groups, each group consisted of seven rats that had previously undergone bile duct ligation. Three groups received treatment of UDCA-glutathione combination on stratified dose, while the other one only received UDCA. Each treatment was given for 21 days. Ileal samples were collected from the rats and stained with mouse anti TNF-a antibody and hematoxylin-eosin (HE). Immunohistochemistry and histopathological examination were done using microscope and then calculated with ImageJ. RESULTS: The combination of UDCA and glutathione treatment decreased the TNF-α expression (p<0.05) compared to UDCA only group, particularly in group that received 20 mg UDCA and 15 mg glutathione supplementation (p<0.05) and group that received 30 mg UDCA and 20 mg glutathione supplementation (p<0.05). The height of the mucosa villous was higher in the UDCA-glutathione combination groups for all the three dosage variations given (p<0.05) compared to UDCA only group. CONCLUSION: UDCA-glutathione combination downregulates TNF-α expression and improves ileum mucosal villus height in cholestasis.

243-51

Authors: Raghda Abdulsamad Abdualhay, Adnan Jassim Mohammed Al-Fartosy

BACKGROUND: Gestational diabetes mellitus (GDM) has a strong relationship with an increased risk of maternal and perinatal complications. However, in Basrah, Iraq, studies regarding GDM are still limited. In current study, we aimed to investigate the association between insulin resistance and some clinical predictors of GDM among pregnant women in 1st and 3rd trimesters of gestation. METHODS: This case-control study was conducted on 44 pregnant women with GDM and 45 without GDM aged 20 to 40 years who applied for GDM screening during the first (9-13 week) and third trimester (24-28 week) of pregnancy. Demographics, blood glucose, HbA1c, insulin, homeostatic model assessment for insulin resistance (HOMA-IR), spexin, nesfatin-1, orexin-A, vaspin and lipid profile levels were compared between groups. RESULTS: Subjects with GDM showed a higher level of glucose, insulin HOMA-IR, HbA1c, spexin, vaspin in the first and third trimesters of pregnancy (p<0.01) compared to the healthy subjects. Meanwhile in the first and third trimester, subjects with GDM showed significantly lower level of nesfatin-1 and orexin-A compare to the control. In third trimester, oral glucose tolerance test (OGTT) outcomes for fasting glucose at 1 hour, 2 hours, and 3 hours after glucose load were significantly higher (p<0.01). According to the area under the receiver operating characteristics (ROC) curve (AUC) findings, HOMA-IR, spexin, and vaspin may be more effective predictors biomarkers for GDM in pregnant subjects, while orexin-A and nesfatin-1 were ineffective. CONCLUSION: The correlation of insulin resistance and adipokines in the first and thrid trimester was not significantly different, which may cast new light on the possible role as an etiological cause of GDM and might be a better monitoring parameter in women with GDM.

261-8

Authors: Kelvin Supriami, Ira Puspitawati, Dyah Samti Mayasari, Anggoro Budi Hartopo

BACKGROUND: 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.

276-81

Authors: Ferry Sandra, Muhammad Ihsan Rizal, Nurrani Mustika Dewi, Toshio Kukita

BACKGROUND: Increase in inflammatory cytokine levels promotes pathological osteoclast differentiation. Caffeic acid has anti-inflammatory properties and can inhibit osteoclast bone resorption. In vitro studies have reported the ability of caffeic acid in inhibiting osteoclastogenesis pathways, however the in vivo study is rarely conducted. The aim of this study is to examine the role of caffeic acid in reducing inflammation and inhibiting osteoclastogenesis in Adjuvant-Induced Arthritis (AIA) rats. METHODS: Rats were injected with Freund’s Complete Adjuvant (CFA) and mineral oil. One day after injection, various concentration (0, 5, 25, 125 mg) of caffeic acid were given gastro-intestinally. Swelling degree in rats’ ankle joints was determined by measuring height and width of each ankle joint. Bone loss level was examined with soft X-ray, and then bone density was calculated. To examine osteoclastogenesis, ankle joints were stained with Tartrate-Resistant Acid Phosphatase (TRAP) and evaluated microscopically. RESULTS: Ankle joints of AIA rats had severe swelling before treated, yet the swelling was reduced based on concentration-dependent after receiving caffeic acid. Severe bone loss in AIA rats’ ankle joints were also observed, however the treatment of 125 mg caffeic acid showed remarkable inhibition effect toward rats’ bone loss. Osteoclastogenesis in AIA rats’ ankle joints were higher than the normal ones, as indicated with high TRAP-positive Multinucleated Cells (MNCs). But low number of TRAP-positive MNCs was observed in ankle joint of AIA rats that received 125 mg caffeic acid. CONCLUSION: Administration of caffeic acid can reduce the degree of swallowing, inhibit bone loss, and inhibit osteoclastogenesis in ankle joint of arthritis-induced rats.

289-93

Authors: I Ketut Edy Sudiarta, Monicha Zalzabilla Aldinasyah, Cindy Jennilyn Candra, Supriyono Supriyono, Annisa Ullya Rasyida

BACKGROUND: Premature Rupture of Membranes (PROM) is significantly linked to the infections-related maternal deaths. In the inflammatory process, the influencing stressor will stimulate the activation of Nuclear Factor Kappa B (NF-κB) and Receptor of Advanced Gyclation End product (RAGE). Yet up to date, the expression of NF-κB and RAGE in pregnant women with PROM are still rarely studied. Therefore, this study aimed to observe the differences of NF-κB and RAGE expression from PROM and non-PROM subjects. METHODS: This was a cross-sectional study involving 20 PROM subjects and 20 non-PROM subjects with infections and complications. Samples from the fetal membrane tissue of subjects were obtained and put into paraffin block preparation for the determination of NF-κB and RAGE expression. The detection of NF-κB and RAGE expression was conducted using immunohistochemical staining and observed under an upright light microscope. The expressions were later calculated using ImageJ software. RESULTS: Both NF-κB and RAGE expression were found to be higher in PROM subjects compare to the non-PROM subjects. The median of NF-κB in PROM and non-PROM subjects were 32.47±1.22 and 5.59±1.09, respectively (p=0.000). While the median of RAGE in PROM subjects was 53.58±3.46, and in non-PROM subjects was 11.64±2.49 (p=0.013). CONCLUSION: There is significant difference between NF-κB and RAGE expression in fetal membranes of PROM and non-PROM subjects. Therefore, the increased of NF-κB and RAGE expression can be used as a potential marker to detect complication of PROM.

303-8

Authors: Nataniel Tandirogang, Silvia Anitasari, Enos Tangke Arung, Swandari Paramita, Yung Kang Shen

BACKGROUND: Indonesia's tropical forest is home to around 80% of the world's medicinal plants. One of these is Zingiber purpureum, which have traditionally been used to treat joint discomfort, the common cold, and jaundice. The rhizomes of this plant have been suggested to possess antibacterial action in the treatment of infections. In this study, Z. purpureum was screened for antibacterial activity against 13 bacteria (Gram-positive and Gram-negative). METHODS: Z. purpureum rhizomes were obtained and the distillated extracts were made to generate essential oil. The minimum inhibitory concentration (MIC) and Kirby Bauer disk diffusion methods were used to determine the antibacterial activity. RESULTS: All bacteria activity were inhibited by the essential oil of Z. purpureum at concentrations ranging from 2.5 vol% to 10 vol%. However, several bacterias (Escherichia coli, Pseudomonas aeruginosa, and Enterobacter cloacae) were inhibited at the lowest concentration (0.63 vol %), with the inhibition zones ranging from 6.7 mm to 8.0 mm. Meanwhile, the widest inhibition zone (13.3 mm) was reported on E. cloacae at 10 vol% concentration. CONCLUSION: A 10 vol% Z. purpureum rhizome extract inhibits Gram-positive and Gram-negative bacteria, particularly those that are resistant to a variety of antibiotics.

316-22

Authors: Valentinus Besin, Abdul Hafid Bajamal, I Ketut Sudiana, Mangestuti Agil, Jusak Nugraha, Mohammad Hasan Machfoed, Paulus Budiono Notopuro, Naesilla Naesilla

BACKGROUND: Interleukin (IL)-10 is involved in Wallerian degeneration after peripheral nerve crush injury. Oral thymoquinone was previously observed to decrease local immunoglobulin-G (IgG) in a crush-injured rat model. No study has evaluated the pathway of various thymoquinone dosages on local IgG and IL-10 in this injury. METHODS: This experimental study used 126 Rattus norvegicus Wistar rats that were divided into 18 groups: six groups received a placebo, the other six groups received thymoquinone at 100 mg/kg/day and the last six groups received thymoquinone at 250 mg/kg/day, respectively. Rats were sacrificed at 12, 18, 24, 5x24, 6x24, and 7x24 hours. Matrix metalloproteinase-9 (MMP-9), IL-10, and local IgG levels were assessed by Enzyme-Linked Immunosorbent Assay (ELISA). The nuclear factor KappaB (NF-κB) expressions on Schwann cells were examined by flow cytometry. Path analysis was performed using SmartPLS. RESULTS: The path analysis showed that 100mg/kg/day of thymoquinone significantly decreased NF-κB expression. However, NF-κB did not affect local MMP-9, and MMP-9 had no significant relationship with local IL-10 and IgG. Thymoquinone 250 mg/kg/day also significantly inhibited NF-kB expression, decreased local MMP-9, and, in turn, decreased local IL-10 and IgG. CONCLUSION: Administration of oral thymoquinone 250 mg/kg/day decreases local IgG and IL-10 levels via suppressing NF-κB expression and MMP-9 levels.

139-47

Authors: Nor Raihan Mohammad Shabani, Che Muhammad Khairul Hisyam Ismail, Chiuan Herng Leow, Munirah Mokhtar, Kirnpal Kaur Banga Singh, Chiuan Yee Leow

BACKGROUND: Shigella is a Gram-negative rod-shaped intracellular bacterial pathogen that causes bacterial dysentery or shigellosis among children under five years old. Antibiotics have been less effective in treating shigellosis due to the multi-drug resistance of Shigella. Therefore, an effective vaccine is urgently needed to prevent this disease. The present study aims to determine the peptides presented by major histocompatibility complex (MHC) class II molecules of Shigella-infected macrophages using mass spectrometry-based immunopeptidomics approaches. The MHC class II-associated peptides derived from Shigella-infected macrophages are candidates for developing subunit-based Shigella vaccine. METHODS: THP-1-derived macrophages were infected with Shigella flexneri 2a at the multiplicity of infection equal to 10. The lysate was immunoprecipitated and analyzed by liquid chromatography–tandem mass spectrometry (LC-MS/MS). The sequences retrieved were analyzed using bioinformatics tools. RESULTS: The Shigella-infected THP-1-derived macrophages contained sample peptides from source proteins of almost all subcellular localizations. Eight peptides from S. flexneri 2a-infected macrophages were predicted to be localized at the outer membrane proteins (OMPs) of S. flexneri 2a by the PSORTb server. Two of the OMP-associated peptides were predicted as antigenic, non-allergenic, and non-toxic by respective bioinformatics tools. CONCLUSION: The findings in this study showed two selected OMPs have great potential for vaccine development against shigellosis.

156-63

Authors: Hiqmah Yusi Yana, Lisna Hidayati, Nastiti Wijayanti, Tri Rini Nuringtyas

BACKGROUND: 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.

172-9

Authors: Ferdian Ramadhan, Jamaluddin Jamaluddin, Ismi Zuhria, Luki Indriaswati, Evelyn Komaratih

BACKGROUND: Pterygium is an ocular surface disease that often occurs in tropical countries with a high recurrence rate. Matrix metalloproteinase-3 (MMP-3) play a key role in the inflammatory process of pterygium. This study aims to investigate the ability of curcumin and fibrin glue (FG) in suppressing the expression of MMP-3, and whether can be expected as adjuvant therapy to reduce pterygium recurrence. METHODS: Human pterygium fibroblasts (HPF) obtained from primary cultured of pterygium were treated with no treatment, curcumin, mitomycin-C (MMC), and FG. MMP-3 expression was analyzed using immunocytochemistry and the intensity measurement was done using ImageJ software. Cell migration was measured by scratching and stratification of fibroblast culture after cell confluence, and assessed for 48 hours. RESULTS: The expression of MMP-3 were lower in the HPF treated with 100 mol/mL curcumin, 200 mol/mL, and FG (2205.84±86.1 pg/mL, 1002.51±25.22 pg/mL, 1131.55±17.71 pg/mL, respectively) in comparison with untreated HPF (4703.49±108.9 pg/mL). The expression of MMP-3 were significantly different between groups (p<0.001). Cell migration of HPF after scratching with curcumin intervention at 200 mol/mL decrease from 178.67±2.85 (24 hours) to 88.83±1.48 (48 hours). Meanwhile the migration in FG group also decrease from 180.4±2.56 (24 hours) to 72.45±1.25 (48 hours). CONCLUSION: Curcumin and FG able to reduce the expression of MMP-3 and inhibit the migration of HPF cells.

186-92

Authors: Erlina Sih Mahanani, Ikhsan Nur Arifin, Arya Nur Ihsan, Yusrina Lukitasari, Ferry Sandra

BACKGROUND: Parkia speciosa seeds have been reported to have an anticancer property due to the presence of various antioxidant compounds. Since the potential uses of P. speciosa for the tongue cancer has not been clearly disclosed, we conducted a study to investigate anticancer properties of P. speciosa seed ethanol extract (PSSEE) as well as its effect on cardiac cells. METHODS: Tongue cancer rat model were treated with/without Doxorubicin and various concentrations of PSSEE. After treatment, tongue and heart samples were collected and processed further for histological examinations. Tongue epithelium thickness and damaged heart tissues was observed by HE staining, while tongue cancer cell proliferation was assessed by Ki-67 immunohistochemistry. Analyses were performed under an upright light microscope to measure tongue epithelium thickness, state of cancer cell proliferation, and degree of heart tissue damage. RESULTS: Addition of 400 mg/kg body weight (BW) PSSEE to 4.6 mg/kg BW Doxorubicin reduced the average tongue epithelial thickness and Ki-67+ cells number. Upon addition of PSSEE to Doxorubicin, the damage of heart tissue was reduced in a concentration dependent manner. Among all groups, the group of tongue cancer treated with 4.6 mg/kg BW Doxorubicin and 400 mg/kg BW PSSEE had the lowest percentage as well as the lowest degree of heart tissue damage. CONCLUSION: Since addition of PSSEE to Doxorubicin reduced epithelial thickness, number of Ki-67+ cells and heart tissue damage, PSSEE could be a potential co-chemotherapeutic agent for Doxorubicin toward tongue cancer.

199-205

Authors: Bertoka Fajar Surya Perwira Negara, Jae Suk Choi

BACKGROUND: Bifidobacterium is a genus of lactic acid bacteria that lives in the large intestine of humans and animals. The health benefits of this genus are well established; however, the anti-inflammatory activity of this genus, specifically Bifidobacterium lactis, has not been well defined. Therefore, in this study, we evaluated anti-inflammatory activity of B. lactis hydrolysates using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. METHODS: RAW 264.7 cells were cultured using Dulbecco’s Modified Eagle’s Medium in 5 % CO2 incubator at 37 ℃. One µg/mL of LPS was used to stimulate RAW 264.7 cells. Nitric oxide (NO) production, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) were measured to evaluate anti-inflammatory activity of B. lactis hydrolysates. The cytotoxicity of the inhibitor was also measured in present study through 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. RESULTS: The results showed that B. lactis hydrolysates at 25–200 μg/mL inhibited NO production. In concentration-dependent manner, B. lactis hydrolysate showed inhibition of iNOS expression. However, no inhibition on COX-2 expression was observed. The MTS assay of the B. lactis hydrolysates showed no side effects on the cell viability at all concentrations. CONCLUSION: The current study revealed that B. lactis hydrolysates possess specific anti-inflammatory effects by inhibiting iNOS expression without cytotoxicity and therefore could potentially be developed as a new iNOS inhibitor.

211-7

Authors: Ni Ketut Susilawati, I Wayan Putu Sutirta Yasa, Wayan Suardana, Sri Maliawan, I Made Jawi, Achmad Chusnu Romdhoni, Hamsu Kadriyan, I Wayan Gede Artawan Eka Putra, I Gusti Kamasan Nyoman Arijana

BACKGROUND:Purple sweet potato tuber ethanol extract (PSPTEE) has been known to have benefits in various disease, including to prevent nasopharyngeal carcinoma (NPC). However, there has been no research on PSPTEE in preventing the proliferation of nasopharyngeal mucosal epithelial cells induced by formaldehyde through the oxidative stress mechanisms and inflammatory process. Studies have proven the effect of PSP on various cancer cells, but the carcinogenesis process of the nasopharyngeal mucosal epithelium is still limited. Hence, this study aimed to demonstrate the mechanism for preventing histopathology changes with PSPTEE due to formaldehyde exposure. METHODS: Thirty-two formaldehyde-induced Wistar rats were treated with or without 1g/kgBW/day PSPTEE for 16 weeks. Malondialdehyde (MDA) level were examined with spectrophotometry method, while tumor necrosis factor (TNF)-α and p53 with enzyme linked immunosorbent assay (ELISA) method. The histopathology appearance of the nasopharyngeal epithelium was observed with hematoxylin-eosin (HE) staining. RESULTS: MDA and TNF-α levels in control and treatment group were 22.89±2.84 μM; 9.83±0.89 μM and 84.18±11.58 ng/L; 73.92±10.59 ng/L, respectively, and they showed significant difference. Meanwhile, the level of p53 showed no significant difference. Histopathology appearance showed a significant difference. Path analysis for MDA, TNF-α and p53 levels contributed 28.7% to histopathology appearance. MDA have an effect on significant direct effect on TNF-α. TNF-α has a significant direct effect to histopathological appearance. Indirect influence MDA on histopatological appearance were smaller than the direct effect. TNF-α has the greatest influence on histopatological appearance. CONCLUSION: Formaldehyde induces the histopathology appearance in nasopharyngeal epithelium. This study also demonstrates the mechanism to prevent the nasopharynx epithelial histopathology appearance by administration of PSPTEE.

11-28

Authors: Anna Meiliana, Nurrani Mustika Dewi, Andi Wijaya

BACKGROUND: Aging is an unavoidable part of life, defined by a gradual loss in tissue and organ function and an increasing chance of death. Current studies of aging connected the genetic and epigenetic changes to cause this process. CONTENT: When the aging-related epigenetic alterations is accumulated, it may result in irregulated gene expression, metabolic instability, stem cell senescence and exhaustion, and imbalance of tissue homeostasis, which all accelerate the aging process. Altered epigenetic gene regulatory mechanisms such as DNA methylation, histone modification and chromatin remodeling, and non-coding RNAs can induce aging process, thus manipulating these processes give a chance for the success of age-delaying interventions. SUMMARY: Given updated tools and technologies to investigate the epigenetic regulation affecting aging processes, new therapeutic strategies to delay this process can be developed to increase longevity and improve quality of life.

36-44

Authors: Mykyta Valilshchykov, Volodymyr Babalyan, Тetiana Markina, Marina Kumetchko, Liudmyla Boiko, Sergey Romaev

BACKGROUND: In patients with a fracture of the proximal femur and concomitant arterial hypertension, there is a disturbance of the reparative processes of bone tissue. This research aimed to study the regulation of the reparative processes of fractures of the proximal femur with intramedullary osteosynthesis during the correction of concomitant hypertension, which was examined based on some markers using the rat model. METHODS: The study involved healthy Wistar rats and spontaneously hypertensive rats (SHR). The subjects were then grouped into healthy rats without exposure (1.1) SHR without exposure (2.1), healthy rats with modeled fractures of the proximal femur (1.2), SHR with modeled fractures of the proximal femur (2.2), SHR underwent hypertension correction with enalapril in subgroups without fracture (2.3) and SHR underwent hypertension correction with enalapril in subgroups with fracture (2.4). The levels of interleukin (IL)-6, tumor necrosis factor alpha (TNF-a), IL-10, amino-terminal propeptide procollagen type III (PIIINP), glucose, uric acid, creatinine, urea, cholesterol, and albumin were determined in the blood serum of the animals. Femur preparations were examined after the removal of intramedullary fixation. RESULTS: Serum IL-6 level of animal in group 2.4 (2.297±0.361 pg/mL) were reduced compared to the corresponding indicators of rats in group 2.3 (4.054±0.491 pg/mL, p<0.05). Serum glucose and urea levels of animal in group 2.4 (3.951±0.156 mmol/L, 6.552±0.426 mmol/L, respectively) were significantly reduced in comparison with the group 2.3 (6.384±0.890 mmol/L, 10.369±0.888 mmol/L, respectively). The histological results indicated a positive effect of the drug enalapril on the healing of fractures of the proximal femur in animals with hypertension. CONCLUSION: Correction of arterial hypertension with enalapril in fractures of the proximal femur improves the reparative processes of bone tissue.

52-8

Authors: Dewi Wulandari, Alida Roswita Harahap, Suhendro Suhendro, R. Tedjo Sasmono, Aryati Aryati, Herdiman Theodorus Pohan, Iris Rengganis, Saptawati Bardosono

BACKGROUND: Dengue infection remains a major public health problem in Indonesia. Severe dengue associated with plasma leakage require hospitalization and potentially life threatening. However, the mechanism remains unclear, and the occurrence is unpredictable. The role of anti-endothelial antibody is predicted play an important role in the pathogenesis of plasma leakage, as severe dengue is more prevalent in secondary infection or post vaccinated individuals. METHODS: Serum samples from 127 single Dengue Virus (DENV) serotype infected subjects were obtained in day 2 of fever onset. Subjects were divided into plasma leakage and non-plasma leakage based on World Health Organization (WHO) criteria. Anti-endothelial antibody in patient sera were detected using western blot of Human Umbilical Vein Endothelial Cells (HUVEC). To confirm cross-reactivity, the sera was preabsorb with mix-DENV lysate. RESULTS: Three prominent bands were identified on western blot strips that inhibited by pre-absorption with DENV lysate. Plasma leakage patient expressed significantly more antibodies, with 51.7% of plasma leakage patients expressed at least two bands out of those three, compared to 18.5% of non-plasma leakage. CONCLUSION: Antibodies found in sera of dengue patients with plasma leakage cross-reacted with DENV proteins and endothelial proteins 37 kDa, 75 kDa, 120 kDa, and therefore may be involved in the pathogenesis of plasma leakage. Proteomic identification of those protein targets is needed and may be useful for vaccine studies and further development of predictor marker for plasma leakage in dengue.

66-73

Authors: Todung Silalahi, Idrus Alwi, Frans Suyatna, Katarina Dewi Sartika, Christopher Surya Suwita

BACKGROUND: Percutaneous coronary intervention (PCI) is the most common intervention for coronary artery disease (CAD) with very low complications. High oxidative stress post-PCI is associated with further atherosclerosis progression. Curcumin, extracted from a specific type of herbs, exhibits anti-oxidant properties by acting as hydrogen and electron donor for superoxide radicals. The aim of this study is to determine the effect of curcumin’s antioxidant properties in reducing oxidative stress of post-PCI in stable CAD. METHODS: This study was a double-blind parallel randomized controlled trial among 50 stable CAD patients undergoing PCI in Cipto Mangunkusumo General Hospital and Jakarta Heart Center. The subjects received either 45 mg/day curcumin or placebo 7 days pre-PCI until 48 hours post-PCI. Reduced oxidative stress markers (decreased MDA or increased GSH) were measured in 3 phases (7 days pre-PCI, 24 hours post-PCI, 48 hours post-PCI). RESULTS: Curcumin group showed increased MDA from baseline to 24 hours (Δ1=0.01 vs. 0.03; p=0.3) and decreased MDA from baseline to 48 hours (Δ2=-0.06 vs. 0.03; p=0.9). While, curcumin group showed decreased GSH from baseline to 24 hours (Δ1=-49.7% vs. 12.2%; p=0.4) and from baseline to 48 hours (Δ2=-19.09% vs. 11.4%; p=0.6). However, no significant changes were found in malondialdehide (MDA) and glutathione (GSH) level after the intervention. CONCLUSION: The 45 mg/day curcumin supplementation from 7 days pre-PCI until 48 hours post-PCI had no significant antioxidant effect in stable CAD post-PCI.

84-90

Authors: Kusmardi Kusmardi, Bagas Ariffandi, Nadar Sukri Lubis, Tri Wahyuni Lestari, Putri Reno Intan, Alfred Pakpahan

BACKGROUND: Malaria remains a global health concern and an endemic disease in Indonesia. Sambiloto (Andrographis paniculata) and spirulina (Spirulina platensis) are two potential antimalarial agents which act as antioxidants and antiinflammatories that can suppress morbidities during chronic inflammation in malaria, such as proliferation in the colon. This study aims to investigate the effects of A. paniculata extract and S. platensis administration on Ki-67 expression in medial colon epithelial cells of Plasmodium berghei-infected mice measured by H-score. METHODS: Thirty P. berghei-infected male Swiss-Webster mice were divided into five groups: negative controls (carboxymethyl cellulose/CMC); positive controls (dihydroartemisinin-piperaquine/DHP); A. paniculata extract alone (AP); A. paniculata extract in combination with S. platensis extract (AP+ES); and with S. platensis powder (AP+PS). All mice were infected with P. berghei on day 0. The treatment for each group were given 3 days before infection (D-3) until the day of infection (D0) for 28 days after infection. Colon tissues were processed with immunohistochemistry to detect Ki-67. RESULTS: A difference in Ki-67 expression was observed among the groups (p<0.01). The mean H-score for the CMC control group is 135.503±6.723. The lowest level of Ki-67 expression was observed in the AP+PS group (H-score= 110.941±7.079). AP group did not show a significant difference from the CMC group (p=0.514) and neither did the AP+ES group (p=0.234). CONCLUSION: In conclusion, administration of A. paniculata extract and S. platensis powder lowers Ki-67 expression in medial colon epithelial cells of P. berghei-infected mice.

98-103

Authors: Mabruratussania Maherdika, Indranila Kustarini Samsuria, Meita Hendrianingtyas, Nyoman Suci Widyastiti, Muji Rahayu

BACKGROUND: Inflammation occurs in chronic diseases such as type 2 diabetes mellitus (DM). Insulin resistance and inflammation in type 2 DM with obesity can increase interleukin (IL)-6, causing an increase in hepcidin synthesis in the liver. Increased inflammation can exacerbate the course of type 2 DM. This study aims to prove that there are differences in the levels of hepcidin and IL-6 between obese and non-obese type 2 DM. METHODS: This cross-sectional study was conducted on 61 patients with type 2 DM, consist of 22 male and 39 female with an age of more than 40 years. Type 2 DM subjects were obtained from a doctor's diagnosis and were divided into obese and non-obese groups based on body mass index (BMI). Hepcidin and IL-6 levels were examined using the Enzyme-Linked Immunosorbent Assay (ELISA) principle. The data were analyzed using an independent t-test and Mann-Whitney test. RESULTS: The mean level of hepcidin in the obese with type 2 DM group was 25.32±11.54 ng/mL, and non-obese was 11.94±5.31 ng/mL. The median level of IL-6 in the obese with type 2 DM group was 11.9 (5-61) pg/mL, and non-obese 4.8 (1.5-9.8) pg/mL. There was a significant difference in hepcidin and IL-6 levels between the obese and non-obese groups (p=0.000). CONCLUSION: Hepcidin and IL-6 levels in the obese group with type 2 DM were higher than non-obese group.