Improvement of renal functions in mice with septic acute kidney injury using secretome of mesenchymal stem cells.

Background: A potentially fatal complication of sepsis is septic acute kidney injury. Stem cell therapy is a potential new method of treating sepsis and has been applied to treat some human diseases. Objectives: This study investigated the effects of secretome-MSCs on NGAL, CRP, NF-κB, and MMP-9 proteins, and histopathology in mice with septic AKI. Methods: A post-test-only group design was conducted in 30 Balb/C male mice, which were randomly assigned to five groups: the control group was intraperitoneally injected with 0.5 ml of 0.9 % NaCl, the septic AKI, and the treatment groups (T1, T2, and T3) were intraperitoneally injected with 0.5 ml of 0.9 % NaCl and 0.3 mg/kg BW LPS single dose for three days. Three-day treatments of 150, 300, and 600 µl secretome-MSCs were administered intraperitoneally into the treatment groups. Furthermore, kidney and blood samples were collected for biochemical and histopathological analyses. Results: The T1, T2, and T3 groups had lower expression of NF-κB and MMP-9 and significantly lower CRP and NGAL levels than that of septic AKI group. T1 (1.21 ± 0.19), T2 (0.75 ± 0.22), and T3 (0.38 ± 0.14) groups demonstrated lower average scores for inflammation, necrosis, hemorrhage, and degeneration compared to septic AKI group (2.17 ± 0.13). Conclusions: Administration of 600 µl/20 g BW secretome-MSCs suppresses NF-κB and MMP-9 expression and reduces CRP and NGAL levels. Meanwhile, the 150 and 300 µl/20 g BW doses also indicated a greater improvement in renal tissue damage of mice with septic AKI.

Dose-response effect of L-citrulline on skeletal muscle damage after acute eccentric exercise: an in vivo study in mice.

Background: Eccentric exercise may trigger mechanical stress, resulting in muscle damage that may decrease athletic performance. L-citrulline potentially prevents skeletal muscle damage after acute eccentric exercise. This study aimed to assess the dose-response effect of L-citrulline as a preventive therapy for skeletal muscle damage in mice after acute eccentric exercise. Methods: This is a controlled laboratory in vivo study with a post-test-only design. Male mice (BALB/c, n = 25) were randomized into the following groups: a normal control (C1) (n = 5); a negative control (C2) with downhill running and placebo intervention (n = 5); treatment groups: T1 (n = 5), T2 (n = 5), and T3 (n = 5), were subjected to downhill running and 250, 500, and 1,000 mg/kg of L-citrulline, respectively, for seven days. Blood plasma was used to determine the levels of TNNI2 and gastrocnemius muscle tissue NOX2, IL-6, and caspase 3 using ELISA. NF-κB and HSP-70 expressions were determined by immunohistochemistry. Results: Skeletal muscle damage (plasma TNNI2 levels) in mice after eccentric exercise was lower after 250 and 500 mg/kg of L-citrulline. Further, changes in oxidative stress markers, NOX2, were reduced after a 1,000 mg/kg dose. However, a lower level of change has been observed in levels of cellular response markers (NF-κB, HSP-70, IL-6, and caspase 3) after administration of L-citrulline doses of 250, 500, and 1,000 mg/kg. Conclusion: L-citrulline may prevent skeletal muscle damage in mice after acute eccentric exercise through antioxidant effects as well as inflammatory and apoptotic pathways. In relation to dose-related effects, it was found that L-citrulline doses of 250, 500, and 1,000 mg/kg significantly influenced the expression of NF-κB and HSP-70, as well as the levels of IL-6 and caspase 3. Meanwhile, only doses of 250 and 500 mg/kg had an impact on TNNI2 levels, and the 1,000 mg/kg dose affected NOX2 levels.

Anemia in female adolescents at Karanganyar regency: A cross-sectional study associated with polymorphism of duodenal cytochrome B gene and daily consumptions of fruits and vegetables.

BACKGROUND: The prevalence of anemia in female adolescents increases every year. A duodenal Cytochrome B (CYBRD1) enzyme is involved in the regulation of iron metabolism. G797A gene polymorphism of the CYBRD1 reduces nonheme iron transport into the enterocytes, which was mediated by the divalent metal transporter 1 protein. Daily consumption of fruits and vegetables has been recommended for the prevention of non- communicable diseases, including anemia. AIM: This study aimed to analyze the association of CYBRD1 polymorphism and daily consumption of fruits and vegetables with anemia in female adolescents in the Karanganyar regency. METHODS: This cross-sectional study recruited 233 female students in 6 senior and vocational high schools in Karanganyar regency, which were selected using purposive sampling. Data on fruit and vegetable consumption were collected using Semi-Quantitative Food Frequency Questionnaire (SQ-FFQ). Polymorphism of the G797A CYBRD1 gene was determined using the amplification refractory mutation system polymerase chain reaction. All collected data were analyzed using chi-square and multiple logistic regression tests with p < 0.05. RESULTS: Anemia was found in 13.73% of female students. Inadequate intakes of fruits and vegetables were found in 63.09 and 51.07% of female students, respectively. The AA genotype (OR = 5.779; 95%CI: 0.974-34.289; p = 0.053), inadequate fruit consumption (OR = 1.497; 95%CI: 0.603-3.718; p = 0.133), and inadequate vegetable consumption (OR = 11.99; 95%CI: 3.457-41.586; p < 0.001) increased higher risk of anemia, compared with their counterparts. CONCLUSIONS: G797A CYBRD1 gene polymorphism and daily consumption of vegetables increase the risk of anemia but not for daily consumption of fruits in female adolescents in the Karanganyar regency.

The acute effects of nonstructural-1 protein dengue virus type 2 on wet liver weight, zonulin expression and serum zonulin.

Background and Objectives: Intestinal leakage commonly occurs in severe dengue infection with zonulin as a biomarker. The aim of this study was to determine the effects of NS1 on liver weight, zonulin expression and serum zonulin levels. Materials and Methods: This laboratory experiment used 18 ddY mice, which were randomly divided into control (C), PBS (T1), and PBS + NS1 (T2) groups. Mice in the T1 and T2 groups were intravenously injected with 500 µl PBS only and 50 µg NS1 respectively. Mice blood samples were collected before and after three-day treatment for measurement of zonulin level. The fresh liver was weighted directly and were then used for immunostaining. Results: The C group had lower wet liver weight compared to the T groups (p=0.001). Increased expression of liver zonulin was found in the T2 group, significant different from the C (p=0.014) and T1 groups (p=0.020). After treatment, serum zonulin levels in the T1 group was higher than that of the T1 group before treatment (p=0.035) but not in control (p=0.753) and T2 groups (p=0.869). Conclusion: Administration of 50 µg NS 1 increases wet liver weight and zonulin expression in hepatocytes, but did not increase serum zonulin levels in ddY mice.

Association between carotid intima media thickness and acute ischemic stroke at an Indonesian tertiary referral hospital.

Objectives: A high prevalence of tobacco smoking contributes to a high incidence of acute ischemic stroke (AIS) in Indonesia. Large-artery atherosclerosis is known to be a significant cause of AIS. The present study was aimed at evaluating the association between AIS and atherosclerosis on the basis of carotid intima-media thickness (CIMT) measurements in a tertiary care hospital in Indonesia. Methods: A total of 79 patients with AIS (case study group) and 79 individuals without AIS (control group) were included. Chi-squared tests and odds ratios were used to compare the groups and determine associations. We also considered factors such as age, body mass index (BMI), sex, type-2 diabetes mellitus (T2DM), hypertension, smoking status, dyslipidemia, socioeconomic status, and educational level in the statistical analyses. A p-value <0.05 was considered statistically significant. Results: Stratification of atherosclerosis into case study and control groups with respect to all study variables indicated a significant relationship (p > 0.05) between atherosclerosis and all variables except low socioeconomic status (p = 0.265) and low educational level (p = 0.180). Regression analysis demonstrated that a BMI ≥25 kg/m2, compared with a normal BMI, was associated with a 2.139-fold higher risk of atherosclerosis. Conclusions: AIS was associated with atherosclerosis, on the basis of CIMT measurements, according to age, BMI, sex, T2DM, hypertension, smoking status, dyslipidemia, socioeconomic status, and education level in the Indonesian population.

Alkaloid fraction of Mirabilis jalapa Linn. flowers has low cytotoxicity and increases iron absorption through Erythropoietin-Matriptase-2-Hepcidin pathway in iron deficiency Hepatocarcinoma cell model.

In this study, we investigated the effects of an alkaloid fraction of Mirabilis jalapa L. flowers in terms of cytotoxicity, Erythropoietin (EPO), hepcidin, and Matriptase-2 (MT-2) expression levels in iron deficiency Hepatocarcinoma (HepG2) cell model. The iron deficiency HepG2 cell model was generated by induction with Deferoxamine (DFO) and was then treated with standard therapy Ferric Ammonium Citrate (FAC) and different alkaloid fraction doses. Subsequently, the type II transmembrane serine proteases (TTSPs) activity and MT-2 expression were measured using a fluorometer and immunocytochemistry methods, while the EPO and hepcidin levels and total iron were examined using an ELISA kit and a colorimetric assay, respectively. The data were then analyzed using ANOVA with a significance level of 95 %. According to the UV-vis Spectrophotometry and HPLC results, the alkaloid fraction of M. jalapa flowers had 6.17- and 4-times higher Betaxanthin levels, respectively, compared to the ethanol extract of M. jalapa flower. Furthermore, LC-MS/MS analysis showed that the most dominant compound is Indicaxanthin. The ethanol extract and alkaloid fraction of M. jalapa flowers were not cytotoxic (IC50 > 30 ppm). Furthermore, the alkaloid fraction containing Indicaxanthin, Miraxanthin-V, and Boeravinone F is capable of increasing EPO levels, membrane and soluble TTSPs activity and MT-2 expression, decreasing hepcidin levels, and increasing intracellular iron levels in iron deficiency HepG2 cell model. In conclusion, the obtained alkaloid fraction of M. jalapa flowers has low cytotoxicity and the later increases iron absorption via EPO-MT2-hepcidin pathway in iron deficiency HepG2 cell model.

Modulation effects of Etlingera elatior ethanol extract as anti-inflammatory on chronic kidney disease in mice with hypertension and diabetes.

The incidence of diabetes increased significantly around the world in accordance with lifestyle and change in eating behaviour. Streptozotocin-Nicotinamide (STZ-NA) is capable of inducing Diabetes Mellitus type 2 in experimental animals for insulin resistance. In this research, we inspect the therapeutic potential of Etlingera elatior ethanol extract (EEEE) on diabetes associated with diabetic nephropathy and hypertension complications in mice models. Diabetes and hypertension are induced in mice using STZ 45 mg/kgBB and NA 110 mg/kgBB, followed by unilateral ureter ligation (UUO) for 4 weeks after a week of STZ-NA induction. The EEEE solution was given in the last 4 weeks with doses of 200, 400, 600, and 800 mg/kgBB. The results of this study prove the effect of vanillic acid on improving systolic blood pressure, plasma creatinine, plasma glucose, albuminuria and reducing the inflammatory marker high sensitivity C-reactive protein (hs-CRP). Histopathology of kidney is under investigation for being part of diabetes hypertension pathology. Treatment using EEEE 600 and 800 mg/kgBB for 4 weeks in experimental mice results in the decrease of plasma glucose, systolic blood pressure, plasma creatinine, albuminuria, and hs-CRP, including the restoration of kidney histology significantly compared to 200 and 400 mg/kgBB doses. This result concludes that EEEE offers modulation effects on diabetes hypertension control by reducing blood glucose rate, blood pressure rate, kidney defect, and inflammation markers.

Daily iron intake and BMI for age but not for daily tannin intake increase hemoglobin levels in young females at Karanganyar Regency, Central Java, Indonesia.

Abstract.

Phytochemical evaluation of Plumbago zeylanica roots from Indonesia and assessment of its plumbagin concentration.

INTRODUCTION: Plumbagozeylanica grows widely in many tropical countries. In Indonesia, this plant, known as Daun Encok, has some beneficial effects on human health.

Brazilin Isolated from Caesalpina Sappan Wood Induces Intrinsic Apoptosis on A549 Cancer Cell Line by Increasing p53, caspase-9, and caspase-3.

OBJECTIVE: Lung cancer is the leading cause of death among cancer patients. The majority of lung cancer is the Non-Small Lung Carcinoma (NSLC). This study evaluated the potency of brazilin isolated from Caesalpinia sappan wood to induce apoptosis on non-small lung carcinoma cell line, A549, by examining the expression of p53, caspase-9, and caspase-3. METHODS: Brazilin was isolated from Caesalpinia sappan wood following a guided assay and it was determined by using Brazilin®SIGMA as standard. The activity of brazilin on the growth of A549 cell line was analysed by MTT assay and the apoptosis was evaluated by flowcytometer following Annexin V (FITC) and PI staining. The expression of p53, caspase-9, and caspase-3 was examined by immunocytochemistry. RESULT: The IC50 of brazilin on A549 cell line was 43µg/mL. Cell treatment with 20 µg/mL and 40 µg/mL of brazilin significantly increased early apoptosis (p<0.001). Cell treatment with 40 µg/mL  of Brazilin significantly increased late apoptosis (p<0.001). Brazilin significantly increased the expression of p53, Caspase-9, and caspase-3 (p<0.001). CONCLUSION: This study showed evidence of the activity of brazilin to induce intrinsic apoptosis on a NSLC cell line A549.

Effects of Vegetables Consumption Before Carbohydrates on Blood Glucose and GLP-1 Levels Among Diabetic Patients in Indonesia.

Background: Type 2 Diabetes Mellitus (T2DM) is the prominent public health issue. Pharmacotherapy and diet modification should be integrated into T2DM management. Aims: To investigate the effects of vegetables consumption before carbohydrates on blood glucose and GLP-1 levels in T2DM patients. Methods: A non-randomized quasi experimental study was conducted to recruit T2DM patients who attended at the Gatot Soebroto Central Army Hospital, Jakarta, Indonesia from April to May 2016. The Lemeshow's formula was used to determine sample size. A total of 12 non-diabetic and 24 diabetic patients were participated in our study. Glucose levels were measured using a routine hexokinase method while serum GLP-1 levels were determined using the ELISA. The student t-test was used to compare two groups with parametric data. The significant difference was at P < 0.05. Results: Our data showed that T2DM patients who consumed vegetables before carbohydrates, had relatively stable glucose levels at 0, 60 and 120 mins (164.25 ± 86.89 vs 183.5 ± 55.96 vs 167.83 ± 65.53, P = 0.163) and stay lowered within the normal range compared to T2DM patients who consumed vegetables after carbohydrates (165.08 ± 67.89 vs 241.92 ± 68.03 vs 204.92 ± 81.76, P = 0.022). Additionally, GLP-1 levels remained stable after 60 and 120 min at day 1 (P = 0.816) and day 3 (P = 0.955). Conclusions: Vegetables consumption before carbohydrate is a promising and simple method of diabetes diet for maintaining blood glucose and GLP-1 levels and preventing from vascular complication.

Immunomodulation of tahneeq method in IL-12 and CD8+ T-Lymphocyte, an in-vivo study in neonatal rats.

Stimulation of the neonatal immune system is quite important for the proliferation and differentiation of antigen-presenting cells (APCs) and T cells. Tahneeq is a traditional method to manually rub the palatal mucosa of newborn babies with premasticated Ajwa palm dates. The present study was to investigate the tahneeq effects on IL-12 expression of dendritic cells (DCs) and blood T lymphocytes expressing CD8+ in neonatal Wistar rats. The number of 90 healthy neonatal Wistar rats have randomly divided into three groups: control group received breastmilk only, treatment group (T1) receiving breast milk + mild-scratched intensity of tahneeq, and T2 group received breastmilk + strong-scratched intensity of tahneeq on the palatal and gingival mucosa immediately after birth.  Seven neonatal Wistar rats in all groups were then sacrificed in three hours after birth and days 1, 5, 7, 13, and 30 treatment. IL-12 expression in the palatal and gingival mucosa was determined using immunohistochemical staining, and blood CD8+ T-lymphocytes were quantified using a flow cytometer. One way ANOVA was used to analyze the percentage of IL-12 and CD8+ T-lymphocytes among neonatal Wistar rat groups.  The T1 and T2 newborn rat groups had significantly higher IL-12 expression than the control group (p<0.001). The increased IL-12 expression in T2 groups significantly increased (p<0.001) compared to the IL-12 expression in the T1 and control groups. The percentage of  CD8+ T lymphocytes in all neonatal rat groups increased on three hours after birth and day 30 treatment but remained constant on days 5 and 7 treatment and decreased on day 13 treatment. At 5, 13, and 30th days treatment,  the percentage of CD8+ T lymphocytes in T1 and T2 neonatal rat groups was significantly higher (p<0.05) than that in the control group. In conclusion, the impact on systemic CD8+ T cells did not influence by the depth of the scratch. Both mild and strong tahneeq increased the systemic CD8+ T-lymphocytes in neonatal Wistar rats. The roles of anti-inflammatory cytokines and Treg cells should be further investigated to unravel those different results for the development of mucosal immunity in neonates.

Metformin Modulates Cyclin D1 and P53 Expression to Inhibit Cell Proliferation and to Induce Apoptosis in Cervical Cancer Cell Lines.

Background: Cervical cancer is one of the most prevalent gynecological cancers worldwide and contributes in high mortality of Indonesian women. The efficacy of chemotherapy as a standart therapy for cervical cancer decreases because it frequenly rises adverse effects. Recent studies have found that metformin has a potential anticancer effect mostly through reduction of cyclin expression and activation of Activated Adenosine Monophosphate Kinase (AMPK). This study aimed to investigate the effect of metfomin on expression of cyclin D1 and p53 and apoptosis in HeLa cancer cell line. Methods: HeLa cells were treated with various doses of metformin and doxorubicin as a positive control. Cytotoxic effect of metformin was determined using the MTT assay. Immunocytochemistry was used to assess cyclin D1 and p53 expression and apoptosis levels of treated HeLa cells were analyzed using flowcytometry. Data of cyclin D1 expression was statistically analyzed using the Kruskal-Wallis test followed by the Tamhane test, whilst ANOVA and Tukey post Hoc tests were used to analyze data of p53 and apoptosis level. The significant value was p< 0.05. Results: Metformin was able to inhibit proliferation of HeLa cells with IC50 60 mM. HeLa cells treated with 60 and 120 mM metformin had lower cyclin D1 expression than HeLa cells treated without metformin and reached a significant difference (p= 0.001). Moreover, 30 mM or higher doses of metformin increase significantly p53 expression (p< 0.001). Induction of apoptosis was observed in HeLa cells treated with all doses of metformin and reached statistically difference (p= 0.04 and p < 0.001). Conclusion: Metformin can modulate cyclin D1 and p53 expression in HeLa cancer cell line, leading to inhibition of cell proliferation and induction of apoptosis. Other cyclin family members, CDK inhibitors and AMPK signaling should be further investigated in order to know mechanism of metformin action.

Identifying natural substrates for dipeptidyl peptidases 8 and 9 using terminal amine isotopic labeling of substrates (TAILS) reveals in vivo roles in cellular homeostasis and energy metabolism.

Dipeptidyl peptidases (DP) 8 and 9 are homologous, cytoplasmic N-terminal post-proline-cleaving enzymes that are anti-targets for the development of DP4 (DPPIV/CD26) inhibitors for treating type II diabetes. To date, DP8 and DP9 have been implicated in immune responses and cancer biology, but their pathophysiological functions and substrate repertoire remain unknown. This study utilizes terminal amine isotopic labeling of substrates (TAILS), an N-terminal positional proteomic approach, for the discovery of in vivo DP8 and DP9 substrates. In vivo roles for DP8 and DP9 in cellular metabolism and homeostasis were revealed via the identification of more than 29 candidate natural substrates and pathways affected by DP8/DP9 overexpression. Cleavage of 14 substrates was investigated in vitro; 9/14 substrates for both DP8 and DP9 were confirmed by MALDI-TOF MS, including two of high confidence, calreticulin and adenylate kinase 2. Adenylate kinase 2 plays key roles in cellular energy and nucleotide homeostasis. These results demonstrate remarkable in vivo substrate overlap between DP8/DP9, suggesting compensatory roles for these enzymes. This work provides the first global investigation into DP8 and DP9 substrates, providing a number of leads for future investigations into the biological roles and significance of DP8 and DP9 in human health and disease.