Diah Savitri Ernawati

Hypoxic preconditioning effect on stromal cells derived factor-1 and C-X-C chemokine receptor type 4 expression in Wistar rat's (Rattus norvegicus) bone marrow mesenchymal stem cells (in vitro study)

Aim: To examine the effect of hypoxic preconditions on the ability of bone marrow stem cells culture mediated expression C-X-C chemokine receptor type 4 (CXCR4) and stromal cells derived factor-1 (SDF-1) in vitro. Materials and Methods: Bone marrow mesenchymal stem cells (BMSCs) were derived from 12 femurs of 200 g Wistar male rats. The animals were euthanized before BMSCs isolation. BMSCs were divided into two groups, control group: Normoxic condition 21% O2 and treatment group: Hypoxic condition 1% O2. The characterization of BMSCs was analyzed using flow cytometry by cluster differentiation 34 and cluster differentiation 105. The expression of CXCR4 and SDF-1 measured using immunocytochemistry immunofluorescence label after 48-h incubation in a low-tension oxygen chamber with an internal atmosphere consisting of 95% N2, 5% CO2, and 1% O2. All data were subjected to a normality test and then analyzed using t-test statistic (p<0.05). Results: The characterization of bone marrow stem cells showed positive cluster differentiation 34 and cluster differentiation 105. A hypoxic precondition (1% O2) in culture increases CXCR4 (p=0.000) and SDF-1 expression than normoxic conditions (p=0.000) (p<0.05). Conclusion: Hypoxic preconditioning with 1% O2 increase CXCR4 and SDF1 expression.

Effects of topical application of propolis extract on fibroblast growth factor-2 and fibroblast expression in the traumatic ulcers of diabetic Rattus norvegicus

Background: A traumatic ulcer caused by diabetes mellitus (DM) is a lesion caused by an increase in advanced glycosylation end products (AGEs), which takes a long time to heal. AGEs cause angiogenesis, vasculogenesis and a decrease in leukocytes. Fibroblast proliferation and the number of glycosaminoglycans decline, thereby inhibiting the formation of granulation tissue, collagen deposition and platelet derivatives growth factor. The application of topical propolis extract gel to ulcers has an anti-inflammatory function, triggers angiogenesis and accelerates wound healing. Aims: This study sought to establish whether the topical application of propolis extract gel can increase the expression of fibroblast growth factor-2 (FGF-2) and fibroblasts in the healing process of traumatic ulceration in diabetic Wistar rats (Rattus norvegicus). Methods: This was a genuinely experimental research design featuring posttest-only control groups. The simple random sampling technique involved 24 male DM Wistar rats with traumatic ulcers on the labial mucosa of the lower lip. The samples were divided into two groups: a control group whose members were administered hydroxypropyl methylcellulose gel 5% and a treatment group to which propolis extract gel was applied. The expression of FGF-2 and fibroblasts was observed on days 3, 5, 7 and 9 by means of histology and immunohistochemistry (hypothalamic-pituitary-adrenal) with Ab-Mo FGF-2. Results: The topical application of propolis extract gel increased the expression of FGF-2 and fibroblasts in the treatment group on days 5 and 7. There was a correlation between the increased expression of FGF-2 and the number of fibroblasts (P < 0.05). Conclusion: The topical application of propolis extract gel increases the expression of FGF-2 and fibroblasts within the traumatic ulcer healing process in diabetic R. norvegicus.

Osteogenic potential of gingival stromal progenitor cells cultured in platelet rich fibrin is predicted by core-binding factor subunit-α1/Sox9 expression ratio ( in vitro )

Background: Alveolar bone defect regeneration has long been problematic in the field of dentistry. Gingival stromal progenitor cells (GSPCs) offer a promising solution for alveolar bone regeneration. In order to optimally differentiate and proliferate progenitor cells, growth factors (GFs) are required. Platelet rich fibrin (PRF) has many GFs and can be easily manufactured. Core-binding factor subunit-α1 (CBF-α1) constitutes a well-known osteogenic differentiation transcription factor in SPCs. Sox9, as a chondrogenic transcription factor, interacts and inhibits CBF-α1, but its precise role in direct in vitro osteogenesis remains unknown. GSPCs cultured in vitro in PRF to optimally stimulate osteogenic differentiation has been largely overlooked. The aim of this study was to analyze GSPCs cultured in PRF osteogenic differentiation predicted by CBF-α1/Sox9. Methods: This study used a true experimental with post-test only control group design and random sampling. GPSCs isolated from the lower gingiva of four healthy, 250-gram, 1-month old, male Wistar rats ( Rattus Novergicus) were cultured for two weeks, passaged every 4-5 days. GSPCs in passage 3-5 were cultured in five M24 plates (N=108; n=6/group) for Day 7, Day 14, and Day 21 in three different mediums (control negative group: αModified Eagle Medium; control positive group: High Glucose-Dulbecco’s Modified Eagle Medium (DMEM-HG) + osteogenic medium; Treatment group: DMEM-HG + osteogenic medium + PRF). CBF-α1 and Sox9 were examined with ICC monoclonal antibody. A one-way ANOVA continued with Tukey HSD test (p<0.05) based on Kolmogorov–Smirnov and Levenes tests (p>0.05) was performed. Results: The treatment group showed the highest CBF-α1/Sox9 ratio (16.00±3.000/14.33±2.517) on Day 7, while the lowest CBF-α1/Sox9 ratio (3.33±1.528/3.67±1.155) occurred in the control negative group on Day 21, with significant difference between the groups (p<0.05). Conclusion: GSPCs cultured in PRF had potential osteogenic differentiation ability predicted by the CBF-α1/sox9 ratio.

Inhibitory effects of siwak (Salvadora persica L.) extract on the growth of enterococcus faecalis planktonics and biofilms by in vitro.

Background: Enterococcus faecalis (E. faecalis) is one of the most persistent gram positive bacteria in root canal, resulting in secondary infection after endodontic treatment. E. faecalis pathogenicity is caused by overgrowth of E. faecalis planktonics and biofilms. E. faecalis planktonics produce lipoteichoid acid (LTA) as a virulence factor that can defend their permeability cell. On the other hand, E. faecalis biofilms produce protease, such as Esp (enterococcal surface protein), GelE (gelatinase), and SprE (serin protease), that have quorum-sensing mechanism as an adhesion factor to form extracellular polysaccharide substance (EPS) and increase the growth of the biofilms themselves. Siwak (Salvadora persica L.) has active components, namely benzylisothio-cyanate, trimethylamine, and salvadorine that can inhibit the growth of E. faecalis planktonics and biofilms. Purpose: This study aimed to measure inhibitory effects of siwak extract on the growth of E. faecalis planktonics and biofilms. Method: This research was an antimicrobial research on the culture of E.faecalis incubated in a TSB medium. Siwak extract was diluted into different concentrations, namely 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, and 100%. The extract then was placed into the E. faecalis’s colony and planted into Trypticase Soy Agar medium. After incubated for 24 hours at 37°C, the colony would be measured and compared with the control (+) and control (-). As an antibiofilm research, this research used biofilm microtitter assay method to form E. faecalis biofilms incubated in a well-plate medium containing TSB and 0.1 % glucose. Siwak extract then was diluted into the same range concentration as in first method, and placed into the colony of E. faecalis to form biofilms. The biofilms were measured and compared to the control (+) given siwak extract and the control (-) given 0.1% chlorhexidine. After the incubation, they were washed three times, and staining process was conducted using Chrystal violet. The optical density then was measured by ELISA Reader 595 nm. Result: Siwak extract could inhibit the growth of E. faecalis planktonics at the concentration of 35% as a minimum inhibitory concentration as well as the growth of E. faecalis biofilms at the concentration of 45% as a minimum biofilm inhibitory concentration. Conclusion: Siwak extract has an inhibitory effect, particularly at a concentration of 35% on the growth of E. faecalis planktonics and at the concentration of 45% on the growth of E. faecalis biofilms.