THE EFFECT AND MECHANISM OF ACTION OF PRETREATMENT WITH ETHANOL EXTRACT OF PEPEROMIA PELLUCIDA (L.) KUNTH HERB TO ACCELERATE ALVEOLAR BONE HEALING POST TOOTH EXTRACTION IN WISTAR RAT MODEL

• Main Author: Irsyad, Widya
• Format: Dissertations
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/74786
• Institution: Institut Teknologi Bandung

Alveolar bone resorption, which is more active than its formation up to 6 weeks to 8 months after tooth extraction, causes functional and aesthetic problems when making dentures to replace extracted teeth. Alveolar resorption after a tooth extraction is also detrimental in orthodontics because the alveolar bone is the medium for tooth movement. The limitations of commercial drug preparations to address this problem and the tremendous potential of Indonesia’s biological wealth and biodiversity encourage research to continue to develop. Peperomia pellucida is one of the potential plants, but its mechanism of action still needs to be studied further. This study began with extracting the leaves and stems of P. pellucida with 50% ethanol solvent using the reflux method until an ethanol extract yield of 26.23% w/w was obtained. The results of the phytochemical examination showed that the extract contained compounds of flavonoids, saponins, alkaloids, polyphenols, tannins, quinones, and steroids/triterpene. Examination of the extracted content using the Liquid Chromatography-Mass Spectrometer Quadropole Time of Flight (LC-MS QTOF) method detected the presence of compounds known to modulate bone, among which the main ones are luteolin, genistein, cyanidin-3-glucoside, nuciferin, kaempherol. The in vivo effectiveness tests of the extracts were conducted, and data were obtained on the effects of the extracts on osteoblast cells, osteoblast cell products, and growth factors that affect osteoblasts and osteoclasts. Furthermore, histology and serology studies were conducted. Osteoblast products in the form of bone trabeculae formed by collagen fibers and hydroxyapatite crystals were the first data obtained using Micro-Computed Tomography (µ-CT). Results showed that Bone Volume Fraction (BV/TV), Trabecular Thickness (Tb.Th), Trabecular Number (Tb.N) variables increased (p<0.05), while Trabecular Separation (Tb.Sp) decreased although not significantly. This combination indicates that the extract positively affects trabecular bone formation in the tooth socket. Histology study with hematoxylin eosin staining showed that the extract caused an increase in the number of osteoblasts (p<0.05). This result confirmed the previous finding of an increase in the number of osteoblasts that produce trabecular bone-forming hydroxyapatite crystals. The more significant number of blood vessels and fewer Polymorphonuclear neutrophilic (PMNs) in the extract group also corroborated the extract's efficacy on bone healing. Osteoclasts were more abundant in the extract group. Fibroblasts in the extract and control groups were not significantly different. Serological analysis using the Enzyme-Linked Immunosorbent Assay (ELISA) method was performed to study the effect of the extract on the expression of wingless-related integration site 7b (wnt7b), one of the growth factors that affect osteoblast differentiation and proliferation. Serum wnt7b levels of the extract group were higher although not statistically significant. Although the difference was insignificant, the histology results showed significantly more osteoblast cells in the extract group. The levels of soluble receptor activator of nuclear factor-?b ligand (rankl), carboxylated osteocalcin, and bone sialoprotein were lower in the extract group, although insignificant. X-ray fluorescence (XRF) assays were performed to examine the effect of the extract on the femur bone away from the affected area. The calcium and phosphate contents of femur bones between the extract and control groups were compared. The results showed no significant difference between the two groups indicating that the extract only acted on the induction area. In vitro assay was conducted to study the effects of extracts, hexane, ethyl acetate, and water fractions of the P. pellucida plant on the proliferation ability of MC3T3 cells. Treatment with the extract and some fractions significantly led to an increase in cell metabolic activity, an indicator of MC3T3 cell proliferation (p<0,05). Increasing the dose of ethyl acetate fraction caused a decrease in MC3T3 proliferation ability. The mineralization activity of MC3T3 cells was evaluated using alizarin (C14H8O4) staining, and the results showed that the hexane fraction and ethyl acetate fraction had the ability to increase the mineralization activity of MC3T3 cells (p<0.01). The information obtained from this study is that various chemical compounds contained in P. pellucida extract make this plant has the potential to accelerate alveolar bone healing after tooth extraction. The ability of P. pellucida extract to increase the expression of wnt7b as a growth factor causes the promotion of osteoblast cell proliferation that will form alveolar bone. P. pellucida extract was also shown to suppress bone destruction activity by osteoclast cells. The extract, hexane fraction, ethyl acetate fraction, and water fraction have the activity of accelerating osteoblast proliferation, and only hexane and ethyl acetate fractions have the ability to increase the mineralization activity of osteoblast cells. P. pellucida extract is expected to be added to bone graft preparations that usually only contain scaffolds.