Cobalt Chloride as a Hypoxia Mimicking Agent Induced HIF-1α and mTOR Expressions of Human Umbilical Cord Mesenchymal Stem Cells

ABSTRACT Objective: To assess the effects of cobalt chloride (CoCl2) as a hypoxia mimicking agent on human umbilical cord mesenchymal stem cells (hUCMSCs) expression of HIF-1α and mTOR for use in regenerative dentistry. Material and Methods: Human umbilical cord mesenchymal stem cells were isolated and then cultured. The characteristics of stemness were screened and confirmed by flow cytometry. The experiment was conducted on hypoxia (H) and normoxia (N) groups. Each group was divided and incubated into 24-, 48-, and 72-hours observations. Hypoxic treatment was performed using 100 µM CoCl2 on 5th passage cells in a conventional incubator (37°C; 5CO2). Then, immunofluorescence of HIF-1α and mTOR was done. Data was analyzed statistically using One-way ANOVA and Tukey's HSD. Results: Significant differences were found between normoxic and hypoxic groups on HIF-1α (p=0.015) and mTOR (p=0.000) expressions. The highest HIF-1α expression was found at 48 hours in the hypoxia group, while for mTOR at 24 hours in the hypoxia group. Conclusion: Hypoxia using cobalt chloride was able to increase human umbilical cord mesenchymal stem cells expression of HIF-1α and mTOR.

Human umbilical cord mesenchymal stem cells induction in peri-implantitis Rattus norvegicus accelerates and enhances osteogenesis activity and implant osseointegration.

Peri-implantitis additional treatment generally aims to repair damaged tissue through a regenerative approach. Human umbilical cord mesenchymal stem cells (hUCMSCs) produce a high osteogenic effect and are capable of modulating the immune system by suppressing inflammatory response, modulating bone resorption, and inducing endogenous osteogenesis. AIM: This study was intended to discover the effect of hUCMSCs on an implant osseointegration process in peri-implantitis rat subjects as assessed by several markers including interleukin-10 (IL-10), transforming growth factor-ß (TGF-ß), receptor activator of nuclear factor kappa- ß ligand (RANKL), bone morphogenic protein (BMP-2), osterix (Osx), and osteoprotegerin (OPG). MATERIAL AND METHODS: The research design implemented during this study represented a true experimental design incorporating the use of Rattus norvegicus (Wistar strain) as subjects. RESULTS: Data analysed by means of a Brown Forsythe test indicated differences between the increase in BMP-2 expression (p < 0.000) and Osx expression (p < 0.001) and between RANKL expression (p < 0.001, Tukey HSD) and OPG expression (p < 0.000, Games Howell). CONCLUSION: According to the findings of this research, hUCMSCs induction is successful in accelerating and enhancing osteogenic activity and implant osseointegration in peri-implantitis rat subjects.

Human umbilical cord mesenchymal stem cells accelerate and increase implant osseointegration in diabetic rats.

OBJECTIVE: This study was conducted to assess the effect of hUCMSCs injection on the osseointegration of dental implant in diabetic rats via Runt-related Transcription Factor 2 (Runx2), Osterix (Osx), osteoblasts, and Bone Implant Contact (BIC). METHODOLOGY: The research design was a true experimental design using Rattus norvegicus Wistar strain. Rattus norvegicus were injected with streptozotocin to induce experimental diabetes mellitus. The right femur was drilled and loaded with titanium implant. Approximately 1 mm from proximal and distal implant site were injected with hUCMSCs. The control group was given only gelatin solvent injection. After 2 and 4 weeks of observation, the rats were sacrificed for further examination around implant site using immunohistochemistry staining (RUNX2 and Osterix expression), hematoxylin eosin staining, and bone implant contact area. Data analysis was done using ANOVA test. RESULTS: Data indicated a significant difference in Runx2 expression (p<0.001), osteoblasts (p<0.009), BIC value (p<0.000), and Osterix expression (p<0.002). In vivo injection of hUCMSCs successfully increased Runx2, osteoblasts, and BIC value significantly, while decreased Osterix expression, indicating an acceleration of the bone maturation process. CONCLUSION: The results proved hUCMSCs to accelerate and enhance implant osseointegration in diabetic rat models.

Bone Remodeling in Mandible of Wistar Rats with Diabetes Mellitus and Osteoporosis.

OBJECTIVES: This study aimed to determine some of bone molecular expressions and its possible bone remodeling pathway between diabetes mellitus (DM) and osteoporosis model in the mandibular bone of Wistar rats. MATERIALS AND METHODS: Twenty-seven female Wistar rats were divided randomly into control and treatment groups. Treatment groups were injected with streptozotocin intraperitoneally to induce DM (P1) and underwent bilateral ovariectomy to generate osteoporosis (P2). All groups were terminated after 12 weeks. Immunohistochemical and hematoxylin-eosin staining were performed to determine the expression of Runt-related transcription factor 2 (RUNX2), Osterix, vascular endothelial growth factor (VEGF), receptor activator of nuclear factor κB ligand (RANKL), osteoprotegerin (OPG), tartrate-resistant acid phosphatase (TRAP), and observed the osteoblast and osteoclast. Statistical analysis was performed using one-way analysis of variance. RESULTS: The lowest mean of RUNX2 and VEGF expression was found in the P2 group. The lowest mean of Osterix expression was found in the P1 group. Both P1 and P2 groups of osteoblast/osteoclast ratio were decreased. There were no significant differences in the expression of TRAP between all groups; however, increased expression of RANKL/OPG ratio was only found in the P2 group. CONCLUSION: DM and osteoporosis induce changes in the bone remodeling pathway which are represented by a decrease in osteoblast biomarkers and an increase in osteoclast biomarkers.

Human umbilical cord mesenchymal stem cells accelerate and increase implant osseointegration in diabetic rats

Abstract Objective This study was conducted to assess the effect of hUCMSCs injection on the osseointegration of dental implant in diabetic rats via Runt-related Transcription Factor 2 (Runx2), Osterix (Osx), osteoblasts, and Bone Implant Contact (BIC). Methodology The research design was a true experimental design using Rattus norvegicus Wistar strain. Rattus norvegicus were injected with streptozotocin to induce experimental diabetes mellitus. The right femur was drilled and loaded with titanium implant. Approximately 1 mm from proximal and distal implant site were injected with hUCMSCs. The control group was given only gelatin solvent injection. After 2 and 4 weeks of observation, the rats were sacrificed for further examination around implant site using immunohistochemistry staining (RUNX2 and Osterix expression), hematoxylin eosin staining, and bone implant contact area. Data analysis was done using ANOVA test. Results Data indicated a significant difference in Runx2 expression (p<0.001), osteoblasts (p<0.009), BIC value (p<0.000), and Osterix expression (p<0.002). In vivo injection of hUCMSCs successfully increased Runx2, osteoblasts, and BIC value significantly, while decreased Osterix expression, indicating an acceleration of the bone maturation process. Conclusion The results proved hUCMSCs to accelerate and enhance implant osseointegration in diabetic rat models.

Calcium Hydroxide Increases Human Umbilical Cord Mesenchymal Stem Cells Expressions of Apoptotic Protease-Activating Factor-1, Caspase-3 and Caspase-9.

PURPOSE: Calcium hydroxide is a gold standard dental material generally used for pulpal and periapical therapy including regenerative endodontic procedures because of its positive properties. However, evaluation about this material on stem cells is limited. Human umbilical cord mesenchymal stem cells (HUCMSCs) are potential to be used in regenerative therapy. Regenerative therapy needs a sustainable cell supply to maintain its regenerative capacity. The aim of this study was to ascertain the apoptosis result of calcium hydroxide on HUCMSCs through the expression of apoptotic protease-activating factor-1 (APAF-1), caspase-3, and caspase-9. MATERIALS AND METHODS: This study used a thawed frozen stock of passage 5 HUCMSCs, grown in minimum essential medium (MEM) alpha containing calcium hydroxide at concentration of 0.1 microgram/mL for 1, 3 and 7 days. Polyclonal antibody with fluorescence isothiocyanate (FITC) label was used to evaluate the expressions. APAF-1, caspase-3, and caspase-9 expressions were recorded and compared on every observation day using fluorescence microscope. Analysis of variance was performed to analyze the significance among the results of treatment groups. The results were concluded significant if p<0.05. RESULTS: The addition of calcium hydroxide in MEM alpha medium increases HUCMSCs expression of APAF-1, caspase-3 and caspase-9 significantly, compared to the control group without calcium hydroxide (p<0.05) in all the times. Day 1 showed the lowest increase followed by higher expressions on day 3 and day 7. CONCLUSION: HUCMSCs express increased APAF-1, caspase-3 and caspase-9 after in-vitro calcium hydroxide exposure. This should be considered when using calcium hydroxide on HUCMSCs for regenerative procedures with regard to other positive properties.

Human Umbilical Cord Mesenchymal Stem Cell-induced Osterix, Bone Morphogenetic Protein-2, and Tartrate-resistant Acid Phosphatase Expression in Osteoporotic Mandibular Bone.

OBJECTIVES: The aim of this study was to prove that human umbilical cord mesenchymal stem cell (hUCMSC) therapy conducted according to the mandibular osteoporotic model will increase Osterix (Osx) and bone morphogenetic protein-2 (BMP-2) expression, while reducing tartrate-resistant acid phosphatase (TRAP) expression. PKH26 labeling proves that mandibular bone regeneration is produced by hUCMSCs induction. MATERIALS AND METHODS: This study incorporated a true posttest only control group design. Twenty-five female Wistar rats were randomly divided into five groups consisting of the sham surgery (N) group, osteoporotic groups injected with gelatin for 4 weeks (G4) and 8 weeks (G8), and osteoporotic groups injected with hUCMSC-gelatin for 4weeks (SC4) and 8 weeks (SC8). All subjects were provided for BMP-2, Osx, and TRAP on immunohistochemistry examination and PKH-26 labeling. STATISTICAL ANALYSIS: All data were analyzed using ANOVA and Tukey HSD tests with p < 0.05 being considered as statistically significant. RESULTS: Compared with other groups, the highest level of BMP-2 and Osx occurred in the sham surgery (N) and osteoporotic groups injected with hUCMSCs-gelatin (SC), while the lowest level of TRAP was found in SC4. During 4- and 8-week observation periods, the PKH 26 appeared green (fluorescent). CONCLUSIONS: hUCMSC demonstrates high-osteogenic activity and increased osteoporotic mandibular bone regeneration, as shown by increased expression of Osx and BMP-2 and decreased TRAP expression. From the labeling, PKH-26 proved that viable hUCMSCs in gelatin solvent can be present in the mandibular bone and be capable of promoting osteogenic differentiation and increasing mineralization and bone formation in the osteoporotic mandibular bone.

Human Umbilical Cord Mesenchymal Stem Cells Over Platelet Rich Fibrin Scaffold for Mandibular Cartilage Defects Regenerative Medicine

ABSTRACT Objective: To evaluate the regeneration of mandibular cartilage defect after implantation of human umbilical cord mesenchymal stem cells (hUCMSC) over platelet rich fibrin (PRF) as scaffold. Material and Methods: 20 male Wistar rats were randomly divided into four experimental groups consisting of: a control group featuring untreated mandibular defects (C), experimental groups whose mandibular defects were implanted with hUCMSC (E1), mandibular defects implanted with PRF (E2), mandibular defects implanted with hUCMSC and PRF scaffold (E3). The subjects were sacrificed after six weeks of observation for immunohistochemical examination in order to evaluate the expression of Ki67, Sox9, FGF 18, type 2 collagen, and aggrecan, in addition to histology examination to evaluate chondrocyte number and cartilage thickness. Data was analyzed with univariate analysis (ANOVA). Results: The implantation of hUCMSC and PRF scaffold proved capable of regenerating mandibular cartilage defect through the expression of FGF 18, Sox9, Ki67, chondrosis counts, type 2 collagen, aggrecan, and cartilage thickness. The regeneration were significantly higher in group E3. Conclusion: Human umbilical cord mesenchymal stem cells in platelet rich fibrin scaffold proved capable of regenerating mandibular cartilage defect.

Inflammatory and Immunogenic Response of the Tissue after Application of Freeze-Dried Hydroxyapatite Gypsum Puger Scaffold Compared to Freeze-dried Hydroxyapatite Bovine Scaffold.

BACKGROUND: Inflammation is a mechanism or reaction of the natural immune system to defend from external hazards. All foreign objects that enter the body will trigger an immune response in the form of antibodies. In Indonesia, the prevalence of diseases that involve the inflammatory process in the body is high. Freeze-dried hydroxyapatite gypsum puger (HAGP) scaffold is a gypsum powder which is currently under development as a bone replacement material. Freeze-dried hydroxyapatite bovine (HAB) scaffold is a bone substitute material available on the market. OBJECTIVE: To analyze the inflammatory and immunogenic responses in the tissue after application of freeze-dried HAGP scaffold compared to freeze-dried HAB scaffold through mediators of tumor necrosis factor alpha (TNF-α) and immunoglobulin G (IgG) in rats. MATERIALS AND METHODS: This study used Wistar rats. HAGP group and HAB group were applied subcutaneously, settled for 7 and 14 days, then the levels of TNF-α and IgG were measured using enzyme-linked immunosorbent assay. Statistical analysis was done using nonparametric test with the Kruskal-Wallis test. RESULTS: TNF-α levels at day 7 in the HAGP group were nearly equal to the control group, while those in the HAB group were higher. Statistically, the significance was P = 0.184 (P > 0.05). At the 14th day, the level of IgG on the HAGP and HAB groups the level was higher than the control group, statistically it was found P= 0.127. CONCLUSION: freeze-dried HAGP scaffold compared to freeze-dried HAB scaffold did not cause inflammatory and immunogenic response on rats through mediators of TNF-α and IgG.

Lipopolysaccharides' Cytotoxicity on Human Umbilical Cord Mesenchymal Stem Cells

Abstract Objective: To show the cytotoxicity of Porphyromonas gingivalis lipopolysaccharide (LPS) on human umbilical cord mesenchymal stem cells (HUCMSCs) to better understand the characteristics for its application in regenerative procedures under periodontopathogen LPS influence. Material and Methods: Ultrapure Porphyromonas gingivalis LPS was used in this study. This research used a frozen stock HUCMSCs, previously confirmed by flow cytometry. The biological characteristics, such as cell morphology, proliferation, and protein expression, were screened. To check the cytotoxicity, HUCMSCs were cultured and divided into two groups, the control group and LPS group with various concentrations from 25 to 0.39 µg/mL. MTT assay was done and the cells were observed and counted. The significance level was set at 5%. Results: The percentage of living HUCMSCs on LPS group were not significantly different among concentrations (p>0.05) from 25 to 0.39 µg/mL, even though there were slight mean decrease between groups, but they were not significant. The duration of 24 hours of exposure of LPS does not significantly lower HUCMSCs viability. Conclusion: LPS does not affect the viability of HUCMSCs. The lower the concentration of LPS, the higher the viability of HUCMSCs.

Cytotoxicity of Calcium Hydroxide on Human Umbilical Cord Mesenchymal Stem Cells

Abstract Objective: To examine the cytotoxicity of calcium hydroxide on human umbilical cord mesenchymal stem cells (HUCMSC) to understand the characteristics for use in regenerative dentistry procedures especially regenerative endodontics. Material and Methods: HUCMSC was isolated, cultured, and confirmed by flow cytometry. The biological characteristics, such as cell morphology, proliferation, and protein expression, were screened. To check the cytotoxicity, HUCMSC was cultured and divided into two groups, the control group (cultured in minimum essential medium (MEM) alpha) and calcium hydroxide group (cultured in MEM alpha and calcium hydroxide). Methyl-thiazole-tetrazolium (MTT) assay was done on different concentrations of calcium hydroxide (0.39 to 25 µg/mL) and the cells were observed and counted. One-way ANOVA test was used with a significance level set at 5%. Results: Flow cytometric analysis confirmed positive of CD73, CD90, CD105, negative of CD45 and CD34. A significant difference was found between the concentration of 6.25 and 3.125 µg/mL (p=0.004). There was no significant difference among 6.25, 12.5 and 25 µg/mL concentrations. There was also no significant difference among 0.39, 0.78, 1.56, and 3.125 µg/mL concentrations. Conclusion: Even though calcium hydroxide is a medicament of choice in clinical endodontics, it decreases the viability of HUCMSC. The lower the concentration of calcium hydroxide, the higher the viability of HUCMSC.

COX-2 induces T cell accumulation and IFN-γ production during the development of chromium allergy.

Chromium (Cr) is commonly added into various metal alloys to improve some mechanical properties such as corrosion resistance, strength, and workability. However, Cr is also known to be a metal allergen for some individuals. Metal allergy is a T cell-mediated disease with symptoms of inflammation and swelling that involve inflammatory cytokines and prostaglandins. Hence, suppressing these inflammation paths by using COX-2 inhibitor might be useful in treating Cr allergy. In this study, mice were used with Cr-induced allergy challenge model. The mice were injected with celecoxib once per day for 7 days one hour after the challenge. Footpad samples were stained with haematoxylin and eosin (H&E), and lymphocytes were isolated from popliteal lymph nodes for the flow cytometric analysis. The results show that both prostaglandin E2 (PGE2), a known mediator of inflammation, and cyclooxygenases (COX)-2 have important roles in the development of Cr allergy. Further, COX-2 inhibitor, celecoxib, was effective in relieving swelling and inflammation in Cr-allergic mice concordant with suppression of IFN-γ production by CD8+ T cells and T cell accumulation in the lymph nodes. Therefore, the inhibition of COX-2 may be a therapeutic target for Cr allergy, and additional molecules in the PGE2 signalling pathway may also be an effective therapeutic target for the treatment of metal allergy.

Phenotype Characteristics and Osteogenic Differentiation Potential of Human Mesenchymal Stem Cells Derived from Amnion Membrane (HAMSCs) and Umbilical Cord (HUC-MSCs).

INTRODUCTION: Human amnion membrane mesenchymal stem cells (hAMSCs) and human umbilical cord mesenchymal stem cells (hUC-MSCs) are potential, non invasive sources of stem cells used for bone tissue engineering. Phenotyping characterization is an extremely important consideration in the choice of the appropriate passage in order to maximize its osteogenic differentiation potential. AIM: To explore phenotype characteristics and compare osteogenic differentiation potential of hAMSCs and hUC-MSCs. METHOD: Isolation and culture were performed on hAMSCs and hUC-MSCs from a healthy woman in her 38th weeks of pregnancy. CD90, CD105 and CD73 phenotype characterization was done in passage 4-7. An osteogenic differentiation examination of hAMSCs and hUC-MSCs with Alizarin red staining and RUNX2 expression was performed in the passage that had appropriate expressions of phenotype characteristics. RESULTS: The expression of CD90 hUC-MSCs was higher than that of hAMSCs in all passages. CD105 hUC-MSCs was higher in passage 4-6, while CD105 hAMSCs was equal to that of hUC-MSCs in passage 7. CD73 hUC-MSCs was higher than hAMSCs in passage 4 and 5, while in passage 6 and 7 hAMSCs was higher than hUC-MSCs. There was a decrease in the number of CD90, CD105 and CD73 on hAMSCs and hUC-MSCs in passage 5, then determined as appropriate passage. Alizarin red staining examination showed calcium deposition and revealed no significant difference, but RUNX2 expression of hUC-MSCs was significantly higher than that for hAMSCs. CONCLUSION: Both hAMSCs and hUC-MSCs had phenotype characteristics of mesenchymal stem cell and showed ostegenic differentiation potential.

Human Umbilical Cord Mesenchymal Stem-Cell Therapy to Increase the Density of Osteoporotic Mandibular Bone.

OBJECTIVE: The aim of this study is to evaluate the feasibility of human umbilical cord mesenchymal stem-cell (hUCMSC) therapy in increasing osteoporotic mandibular bone density in a rat model by determining changes in alkaline phosphatase (ALP), osteocalcin, type 1 collagen, and trabecular bone area after treatment. MATERIALS AND METHODS: This research adopted an experimental posttest-only control group design. Thirty female Wistar rats were randomly divided into six groups, namely, a control group with rats postsham surgery (T1), osteoporotic model postovariectomy rats (T2), postovariectomy rats 4 weeks after gelatin injection (T3), postovariectomy rats 8 weeks after gelatin injection (T4), postovariectomy rats 4 weeks after hUCMSC injection (T5), and postovariectomy rats 8 weeks after hUCMSC injection (T6). The rats were all sacrificed for histological and immunohistochemical examinations of ALP, osteocalcin, type 1 collagen, and trabecular bone area. RESULTS: Increased expression of ALP, type 1 collagen, and osteocalcin, as well as increased trabecular bone area, was observed in the treatment groups compared with that in the osteoporotic groups. CONCLUSION: hUCMSCs produce significant osteogenic effects and increase osteoporotic mandibular bone density in the animal model. Increases in bone density are demonstrated by the higher levels of ALP, osteocalcin, and type 1 collagen, as well as increases in the trabecular bone area.

A potential therapy of human umbilical cord mesenchymal stem cells for bone regeneration on osteoporotic mandibular bone.

OBJECTIVE: The aim of this study is to prove that human umbilical cord mesenchymal stem cell (hUCMSC) therapy on mandibular osteoporotic model is able to increase transforming growth factor-beta-1 (TGF)-ß1 expression, Runx2, and osteoblasts. MATERIALS AND METHODS: This research is true experimental posttest control group design. Thirty female Wistar rats were divided into 6 groups randomly, which consisted of sham surgery for control (T1), ovariectomy as osteoporotic group (T2), osteoporotic group injected with gelatine for 4 weeks (T3), 8 weeks (T4) injected with hUCMSC-gelatine for 4 weeks (T5) and 8 weeks (T6). All mice were presented for immunohistochemistry examination for TGF-ß1, Runx2, and histology for osteoblasts. RESULTS: The lowest level of osteoblast was osteoporotic group injected with gelatine in 4 weeks compared to other groups. There were increases of TGF-ß1, Runx2, and osteoblasts from osteoporotic group compared to osteoporotic post-hUCMSC-gelatine injection group. CONCLUSION: The hUCMSC has a high osteogenic effect and increases the osteoporotic mandibular bone regeneration on the animal model that is showed by the increase of the level of TGF-ß1, Runx2, and osteoblasts.

The Effect of Combination Spirulina-chitosan on Angiogenesis, Osteoclast, and Osteoblast Cells in Socket Models of Hyperglycemic Rattus norvegicus.

BACKGROUND: Prolongation of the inflammatory process in hyperglycemic interferes with bone formation, inhibits the healing process, and triggers bone resorption. A combination of spirulina and chitosan in the tooth socket of Rattus norvegicus is expected to promote the bone remodeling process. This study aimed to evaluate the effect of spirulina and chitosan on angiogenesis, osteoclast, and osteoblast cell in tooth socket models of type 1 diabetes. MATERIALS AND METHODS: A laboratory-based experiment involving 36 R. norvegicus, divided into three groups (nondiabetes mellitus (DM), uncontrolled DM, and controlled DM) and further divided into six subgroups. The controlled groups (K1, K2, and K3) were induced with 3% carboxymethyl cellulose Na, while the treated groups were induced with 12% spirulina and 20% chitosan. On the 14th day, the mandibles of the rats were removed. The capillary lumen, osteoblasts, and osteoclast cells were counted by hypothalamic-pituitary-adrenal examination and the results analyzed by means of Shapiro-Wilk, Levene's, one-way ANOVA, and post hoc Tukey's honestly significant difference test. RESULTS: There was a significant increment in the number of capillary lumen, osteoblast cells, and a decrease in osteoclasts in all three treated groups (P1, P2, and P3). CONCLUSIONS: A combination of spirulina and chitosan can effectively promote the healing process in postextraction sockets of type 1 DM R. norvegicus.

The potential of mangosteen (Garcinia mangostana) peel extract, combined with demineralized freeze-dried bovine bone xenograft, to reduce ridge resorption and alveolar bone regeneration in preserving the tooth extraction socket.

BACKGROUND: Following the extraction of a tooth, bone resorption can cause significant problems for a subsequent denture implant and restorative dentistry. Thus, the tooth extraction socket needs to be maintained to reduce the chance of any alveolar ridge bone resorption. OBJECTIVE: The objective of this study is to determine whether the administration of mangosteen peel extracts (MPEs), combined with demineralized freeze-dried bovine bone xenograft (DFBBX) materials for tooth extraction socket preservation, could potentially reduce inflammation by decreased the expression of nuclear factor κß (NfKb) and receptor activator of nuclear factor-κß ligand (RANKL), to inhibit alveolar bone resorption, and increased of bone morphogenetic protein-2 (BMP2) expressions to accelerate alveolar bone regeneration. MATERIALS AND METHODS: This study consists of several stages. First, a dosage of MPE combined with graft materials was applied to a preserved tooth extraction socket of a Cavia cobaya. Second, the C. cobaya was examined using immune histochemical expression of NfKb, RANKL, BMP2, as well as histology of osteoblasts and osteoclasts. The research was statistically analyzed, using an analysis of variance test and Tukey honest significant difference test. RESULTS: The results of this research were that it was determined that MPEs combined with graft materials on a preserved tooth extraction socket can reduce NfKb, RANK, and osteoclasts also increase of BMP2 and osteoblast. CONCLUSION: The induction of MPEs and DFBBX is effective in reducing inflammation, lowering osteoclasts, decreasing alveolar bone resorption, and also increasing BMP2 expression and alveolar bone regeneration.