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Los procedimientos endodónticos regenerativos (REPs) representan una evolución significativa en el campo de la endodoncia, buscando no sólo tratar la infección o lesión presente en el diente, sino tam-bién promover la regeneración de los tejidos denta-rios afectados. El presente caso clínico muestra un incisivo lateral superior izquierdo con apexogénesis incompleta y diagnóstico de absceso alveolar crónico reagudizado en una paciente de 22 años, en el que se aplicó un procedimiento de endodoncia regenerativa (REPs). La estrategia terapéutica elegida se basó en los principios de ingeniería tisular, incorporando la novedosa aplicación de la membrana amniótica hu-mana liofilizada esterilizada como andamio bioactivo intraconducto. Las evaluaciones clínicas, radiográ-ficas y tomográficas a corto, mediano y largo plazo revelaron el éxito de la terapia. La resolución exitosa mostró en los controles a la pieza dentaria asintomá-tica, con una notable remisión de la patología apical, aumento de la longitud radicular y disminución del calibre apical. Se ha podido destacar la eficacia de los REPs, con una exitosa aplicabilidad de la membra-na amniótica como andamio innovador (AU)
Regenerative endodontic procedures (REPs) represent a significant evolution in the field of endodontics, aiming not only to address the infection or injury within the tooth, but also to promote the regeneration of the affected dental tissues. In this clinical case, an upper left lateral incisor with incomplete apexogenesis and diagnosis of acute exacerbation of a chronic periapical lesion in a 22-year-old patient is presented. A regenerative endodontic procedure (REPs) was applied. The chosen therapeutic strategy was based on tissue engineering principles, incorporating the innovative use of sterilized lyophilized human amniotic membrane as an intraconduct bioactive scaffold. Clinical, radiographic, and tomographic assessments at short, medium, and long-term follow-up revealed the success of the therapy. Successful resolution demonstrated an asymptomatic tooth in the follow-up, with a notable resolution of apical pathology, increased root length, and decreased apical caliber. The effectiveness of REPs has been highlighted, demonstrating the successful applicability of amniotic membrane as an innovative scaffold (AU)
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Humanos , Feminino , Adulto , Células-Tronco/fisiologia , Alicerces Teciduais , Argentina , Faculdades de Odontologia , Papila Dentária , Liofilização/métodosRESUMO
Abstract Fetal bovine serum (FBS) is the most used supplement in culture media; however, it may interfere with in vitro assays via effects on cell proliferation and cytokine production. The ideal FBS concentration for assays using apical papilla cells (APCs) remains unknown. Therefore, this study aimed to evaluate the effects of FBS on APC activation, cell viability/proliferation, and cytokine production. Methodology Human APCs were cultured, plated, and maintained in media containing increasing concentrations of FBS for 24 h, 48 h, 72 h, 7 days, and 14 days in the presence of Lipopolysaccharide (LPS - 1 µg/mL). At each time point, the cells were subjected to the MTT assay. The cytokines transforming growth factor (TGF)-β1, osteoprotegerin (OPG), and interleukin (IL)-6, along with the chemokine CCL2, were quantified using the enzyme-linked immunosorbent assay at the 24-h time-point. Statistical analysis was performed using two-way analysis of variance (ANOVA) followed by Tukey's post-hoc test (p<0.05). Results In general, APCs exhibited increasing metabolic activity in an FBS concentration-dependent fashion, regardless of the presence of LPS. In contrast, FBS interfered with the production of all the cytokines evaluated in this study, affecting the response induced by the presence of LPS. Conclusion FBS increased APC metabolism in a concentration-dependent manner and differentially affected the production of TGF-β1, OPG, IL-6, and CCL2 by APCs in vitro.
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Introduction: Mesenchymal stem cells (MSCs) are the adult stem cells with potential to differentiate into various tissues. Like in other tissues, MSCs also reside in dental pulp after toot development and help in repair and regeneration by differentiating into odontoblasts. Dental Pulp Stem Cells (DPSCs) and Stem cells from Apical Papilla (SCAP) are the type of MSCs from dental papilla and apical papilla respectively. Aim: The aim of this paper is to highlight the characteristics of DPSCs and SCAP. Method: Information was obtained and compiled from published literature and electronic database search engine from PubMed and Google Scholar. Results: In spite of both DPSCs and SCAP having similar cell population origin they possess some different characteristics. Conclusion: The Dental stem cells with different characteristics of similar origin can be utilized in the stem cell based tissue engineering.
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A 15-year-old patient reported persistent pain in the left mandibular second premolar (#35) following a traumatic bite 3 months ago. Clinical examination revealed a fractured central cusp suggestive of dens evaginatus. Intraoral periapical radiograph revealed an immature permanent tooth with a periapical radiolucency. A diagnosis of pulp necrosis with symptomatic apical periodontitis was made. The tooth was treated according to the revised guidelines of regenerative endodontic procedure by the American Association of Endodontics. The follow-up evaluation revealed a complete resolution of periapical pathology. A detached radiopaque tissue was appreciated at the 12-month follow-up. It resembled a broken root tip at the 24-month follow-up. Both the main root body and disjointed root tip developed independently. A cone-beam computed tomography evaluation at the 36-month follow-up confirmed the segmented development of the apical root tip.
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Objective@#To explore the effects of red LEDs on the proliferation and osteogenic differentiation of human stem cells from apical papilla (hSCAPs).@*Methods@#hSCAPs were obtained by isolation, culture and flow cytometry in vitro and irradiated with 1, 3, 5, and 7 J/cm2 red LEDs. The proliferation of hSCAPs was detected using a CCK-8 assay. The osteogenic differentiation of hSCAPs was evaluated using alkaline phosphatase (ALP) staining, ALP activity assay and Alizarin red quantitative detection. The effect of 5 J/cm2 red LEDs on the expression levels of the ALP, Runx2, OCN, OPN and BSP genes and proteins was detected by RT-PCR and western blot, respectively.@*Results@# Red LEDs at 1, 3, 5, and 7 J/cm2 promoted the proliferation of hSCAPs (P < 0.05). The effects of red LEDs with different light energies on the proliferation of hSCAPs were different at different time points (P < 0.05). On the 7th and 14th days after irradiation, red LEDs promoted the osteogenic differentiation of hSCAPs, and the effect of 5 J/cm2 red LEDs was the most obvious under osteogenic induction culture conditions (P<0.05). Red LEDs (5 J/cm2) promoted the expression of the ALP, Runx2, OCN, OPN and BSP genes and proteins (P < 0.05).@*Conclusion @#Red LEDs promoted the proliferation and osteogenic differentiation of hSCAPs.
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BACKGROUND: Stem cells from the apical papilla (SCAP) play important roles in the formation and development of dental roots. However, the immune-modulating capacity of SCAP has not been fully elucidated. OBJECTIVE: To test the therapeutic effects of transplantation of SCAP on dextran sulfate sodium-induced experimental colitis. METHODS: Twenty-four C57/BL6 mice were equally divided into four groups (normal control, positive control, SCAP treatment group, and FasL-knockdown SCAP group), and latter three groups of mice were induced to acute experimental colitis by 3% dextran sulfate sodium in drinking water. At day 3 after modeling, model mice were treated with PBS, human SCAP (2×106 cells), and FasL-knockdown SCAP via intraperitoneal injection, respectively. Inflammation was evaluated by measuring body mass and length of the colon, detecting levels of interleukin 1β, interleukin 6 and tumor necrosis factor α, as well as histological analyses at day 10 after modeling. Levels of Tregs in mesenteric lymph nodes in mice were detected using flow cytometric analysis. RESULTS AND CONCLUSION: SCAP transplantation could ameliorate the inflammation in dextran sulfate sodium-induced colitis mice, and body mass loss and symptoms were significantly improved. Pathological score and the levels of three inflammatory cytokines in the colon tissue decreased significantly. Flow cytometric analysis revealed an increased level of Tregs in mesenteric lymph nodes. Knocking down of FasL gene in SCAP abrogated the therapeutic effects of SCAP in ameliorating dextran sulphate sodium-induced colitis. Therefore, Fas-FasL pathway played an important role in the underlying mechanism of the immune-modulating capacity of SCAP.
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OBJECTIVE@#To investigate the effects of autophagy on osteogenic differentiation of stem cells from the apical papilla (SCAPs) in the presence of tumor necrosis factor- (TNF-) stimulation .@*METHODS@#SCAPs treated with TNF- (0, 5, and 10 ng/mL) with or without 5 mmol/L 3-MA were examined for the expression of autophagy marker LC3-Ⅱ using Western blotting. The cells were transfected with GFP-LC3 plasmid and fluorescence microscopy was used for quantitative analysis of intracellular GFP-LC3; AO staining was used to detect the acidic vesicles in the cells. The cell viability was assessed with CCK-8 assays and the cell apoptosis rate was analyzed using flow cytometry. The cells treated with TNF- or with TNF- and 3-MA were cultured in osteogenic differentiation medium for 3 to 14 days, and real- time PCR was used to detect the mRNA expressions of osteogenesis-related genes (ALP, BSP, and OCN) for evaluating the cell differentiation.@*RESULTS@#TNF- induced activation of autophagy in cultured SCAPs. Pharmacological inhibition of TNF--induced autophagy by 3-MA significantly decreased the cell viability and increased the apoptosis rate of SCAPs ( < 0.05). Compared with the cells treated with TNF- alone, the cells treated with both TNF- and 3-MA exhibited decreased expressions of the ALP and BSP mRNA on days 3, 7 and 14 during osteogenic induction ( < 0.05) and decreased expression of OCN mRNA on days 3 and 7 during the induction ( < 0.05).@*CONCLUSIONS@#Autophagy may play an important role during the osteogenic differentiation of SCAPs in the presence of TNF- stimulation.
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Humanos , Autofagia , Fisiologia , Diferenciação Celular , Fisiologia , Sobrevivência Celular , Células Cultivadas , Papila Dentária , Biologia Celular , Proteínas de Fluorescência Verde , Osteogênese , Fisiologia , Células-Tronco , Fisiologia , Transfecção , Fator de Necrose Tumoral alfa , FarmacologiaRESUMO
Objective: To obtain and identify the exosomes derived from human stem cells from the apical papilla (hSCAPs). Methods: hSCAPs were cultured by modified tissue adherence method and the phenotypes were analyzed with stem cell surface markers CD105, CD45, CD44, CD31, CD34 and CD29. The capability of multi-differentiation in hSCAPs was identified by osteogenic and adipogenic differentiation in vitro. Exosomes were isolated from hSCAPs culture supernatants using gradient centrifugation methods. The size of vesicle was assessed by nanoparticle size analyzer. The morphology of exosomes was observed by transmission electronic microscope (TEM), and the expression of exosome molecular markers CD81, CD9, CD63 and TSG101 was analyzed by Western blotting. Results: hSCAPs were positive for the mesenchyme stem cell markers, including CD105, CD44 and CD29 and negative for the hematopoietic markers CD45, CD31 and CD34. hSCAPs could differentiate into osteoblasts and adipocytes. hSCAPs secreted microvesicles which exhibited round vesicle structure with an intact membrane observed by the TEM. The results of nanoparticle size analyzer measurement showed that the diameters of vesicles were ranged from 30 to 100 nm, which were consistent with the results by TEM. Microvesicles could express the molecular markers for exosomes, i.e. CD81, CD9, CD63 and TSG101. Conclusion: The microvesicles were successfully isolated from hSCAPs and identified as exosomes.
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Objective@#To investigate the effects of leptin on the proliferation of stem cells from human stem cells from the apical papilla (hSCAPs) and the expression of osteogenic/dentinogenic genes in vitro to provide an experimental basis for the sustainable development of young permanent teeth. @* Methods @#The tissue block method was used to isolate and culture hSCAPs from the apical papilla of the immature third permanent molar. The expression of leptin and OBRb in hSCAPs was detected using immunocytofluorescence staining, western blotting and reverse transcription polymerase chain reaction (RT-PCR) analysis. The hSCAPs was treated with 0.1 μg/mL of leptin (0.1 μg/mL group) or 1.5 μg/mL of leptin (1.5 μg/mL group) at different time points. The control group was treated with alpha-MEM medium. Cell proliferation was measured using the CCK8 assay and cell cycle analysis. QRT-PCR was used to detect the expression of related osteoblast/odontogenic genes for alkaline phosphatase (ALP), dentin matrix protein -1 (DMP-1), dentin sialophosphoprotein (DSPP), and osteocalcin (OCN) mRNA. The differences between the treatment groups and the control group were analyzed statistically using one-way ANOVA followed by Bonferroni analysis.@*Results@#The expression of both leptin and OBRb were found in hSCAPs. Compared with the control group, the cell proliferation capacity and S phase cells in the treatment groups were higher than those in the control group, with the 1.5 μg /mL group displaying higher levels than 0.1 μg /mL group, and the treated hSCAPs demonstrated a higher proliferation rate and a higher expression of ALP, DSPP, and DMP-1 from day 3 to day 7, with the 1.5 μg /mL group displaying higher levels than 0.1 μg /mL group , and the difference was statistically significant (P < 0.05), at day 7. The treated hSCAPs demonstrated a lower expression of ALP, DSPP, and DMP-1. Compared with the control group, the treated hSCAPs demonstrated a higher expression of OCN from day 7 to day 14, with significantly higher expression in the 1.5 μg /mL group compared to the 0.1 μg /mL group.@*Conclusion@#Leptin may promote cell proliferation and upregulate the expression of relative osteogenic/dentinogenic genes.
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Objective @#To investigate the effect of different concentrations of MTA on the proliferation and differentiation of stem cells from the apical papilla (SCAP) and the potential of the SCAP to differentiate into odontoblasts.@*Methods@#SCAP were cultured in different concentrations of mineral trioxide aggregate（MTA）. MTA experimental group with concentration of 0.01 mg/mL, 0.02 mg/mL, 0.1 mg/mL, 0.2 mg/mL, 1 mg/mL, 2 mg/mL, 10 mg/mL and 20 mg/mL were prepared. The number of cells at 1 day, 3 days, 5 days and 7 days were measured via a CCK-8 assay to observe the effect of MTA on SCAP proliferation. Real-time PCR was used to detect the gene expression changes. Cells cultured in alpha MEM culture containing 15% FBS without MTA were set as the control group.@*Results @#When cultured for 1 d, statistically significant differences in the promotion of in vitro proliferation of SCAP were not observed between each MTA experimental group and the control group (P>0.05). When cultured for 3 d, 5 d and 7 d, the 0.01 mg/mL MTA group presented obvious promotion of SCAP proliferation compared with the control group (P<0.05), whereas the 0.02 mg/mL, 0.1 mg/mL, 0.2 mg/mL, 1 mg/mL groups did not presented differences with the control group (P>0.05). The in vitro proliferation of the 2 mg/mL, 10 mg/mL and 20 mg/mL groups was lower than that of the control group (P<0.05). Real-time PCR detection showed that the expression levels of DSPP (t=-11.12, P < 0.05) and Runx2 (t=-10.62, P < 0.05) in the experimental group treated with 0.01 mg/mL MTA for 7 days were higher than those in the control group. @*Conclusion @#The 0.01 mg/mL concentration of MTA significantly promotes the proliferation of SCAP and shows the best ability to induce osteogenic and odontoblast differentiation in the SCAP, whereas high concentrations of MTA inhibited the proliferation of SCAP.
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Objective @#This study aimed to investigate the effect of cAMP-responsive-element-binding protein (CREB) overexpression on the differentiation of human stem cells from the apical papilla (hSCAPs), stimulated by transforming growth factor- beta (TGF-β1). @*Methods@#Cells were isolated from human immature third molars via enzymatic digestion. Four experimental groups were set up: ①a control group, receiving normal mineralization inducer (α-MEM, 10% FBS, 10 mmol/L β-sodium glycerophosphate, 50 μg/mL vitamin C, 10 nmol/L dexamethasone); ② a TGF-β1 group, receiving normal mineralization inducer and 5 μg/mL TGF-β1; ③ a TGF-β1+LV-empty group, receiving normal mineralization inducer and the transfected empty virus vector with 5 μg/mL TGF-β1; and ④ a TGF-β1+ov-CREB group, receiving normal mineralization inducer and the transfected CREB-overexpressing viral vector, with 5 μg/mL TGF-β1. The transfected cells were cultured in odontogenic medium in the presence or absence of TGF-β1 for 2 weeks. Alizarin red staining was used to detect mineralized nodules, and the mRNA expression of the mineralization genes runt-related transcription factor 2 (RUNX2), dentin sialophosphoprotein (DSPP) and alkaline phosphatase (ALP) was measured by qPCR. @*Results @#Compared with the control group (1.12 ± 0.11), TGF-β1 inhibited the deposition of calcium minerals (0.67 ± 0.12) (P < 0.05) via hSCAPs and inhibited the mRNA expression of RUNX2 (0.60 ± 0.03), DSPP (0.43 ± 0.12) and ALP (0.69 ± 0.05) (P < 0.05). In contrast, overexpression of CREB attenuated the effect of TGF-β1 on hSCAPs, resulting in the development of a high number of mineralized nodules (1.27 ± 0.10) (P < 0.01) and increased RNA levels of RUNX2 (1.33 ± 0.07), DSPP (1.32 ± 0.11) and ALP (1.26 ± 0.03) (P<0.05) compared with those in the TGF-β1 group. @*Conclusion@#Overexpressed CREB promotes odontogenic differentiation of hSCAPs by interfering with TGF-β1.
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The ball has been set rolling with the isolation of dental pulp stem cells in 2000 for tissue engineering to generate dental pulp. The rationale lies in the possibility of further root development and reinforcement of dentinal walls by deposition of hard tissues which are usually a clinical problem of interest for dentists. In such scenarios, mechanical preparation of the root canal may further weaken the thin dentinal walls and increase the risk of tooth fracture. On the other hand, regenerative endodontic procedures (REPs) could be a better solution. Many exciting areas of endodontic research are emerging and contributing to an increasing momentum of activity in this discipline including regenerative procedures in permanent teeth. This review is an attempt to update on the current trends.
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Objective: To observe the differential expression of miRNAs between human dental pulp stem cells(DPSCs) and stem cells from the apical papilla(SCAPs). Methods: DPSCs and SCAPs were isolated by immune-magnetic binding specific STRO-1 antibody separation system. Osteogenic and adipogenic differentiation of DPSCs and SCAPs were tested by ALP assay, alizarin red staining(ARS) and Oil Red O staining. Differential miRNA expression of DPSCs and SCAPs was screened by the Next generation sequencing. Target genes and their possible roles of these differential miRNAs were predicted using biological information analysis. Results: The results revealed that 7 miRNAs(hsa-miR-224-5p, hsa-miR-1247-5p, hsa-miR-3065-3p, hsa-miR-452-5p, hsamiR-767-5p, hsa-miR-4284, hsa-miR-146a-5p) were downregulated while no miRNAs was upregulated in SCAPs compared with DPSCs. 27 target genes which mainly involved in the cell migration, differentiation and apoptosis were found. Conclusion: Downregulation of some specific miRNAs might be related to the stemness of SCAPs.
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Objective:To study the effect of transplantation of stem cells from the apical papilla (SCAP) to collagen-induced arthritis (CIA).Methods:Twenty DBA/1J mice with collagen Ⅱ-induced arthritis were equally divided into 2 groups and treated with human SCAP and PBS,respectively,with other 6 normal mice as negative control.Inflammation was evaluated by measuring clinical score,TNF-α and anti-CⅡ antibody levels (ELISA),histological analyses and micro-CT analyses.The levels of CD4+Th cells subsets in spleens were assessed by flow cytometric analysis.Results:System infusion of SCAP could significantly reduce the severity of CIA, and resume the balance of Th cells subsets.Conclusion:SCAP transplantation can induce immune tolerance and ameliorate the CIA in-flammation.
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Objective To compare the reprogramming effects and biological characteristics of two types of human odontogen-ic induced pluripotent stem cells(iPSCs).Methods Human dental pulp stem cells(DPSCs)and stem cells from apical papilla (SCAP)were isolated and primarily cultured.The Sendai reprogramming system was utilized to induce DPSCs and SCAP into iP-SCs.The morphology,reprogramming efficiency,reprogramming and time were compared between human DPSCs-iPSCs and SCAP-iPSCs.The SeV and exogenous transcriptional gene expression were detected by RT-PCR.Results Human DPSCs and SCAP were reprogrammed as iPSCs with classical ES-like clonal morphology.The reprogramming efficiencies were(0.68 ± 0.02)% and(0.7 ± 0.01)% respectively,the difference was not statistically significant(P>0.05).The reprogramming time was(26.0 ± 2.1)d and (27.0 ± 1.4)d respectively,the difference was not statistically significant(P>0.05).The RT-PCR results showed that no expres-sion of exogenous virus or transcriptional gene sequence in both iPSCs.Conclusion Human DPSCs and SCAP can be reprogrammed as virus-free and transgene-free iPSCs,which are the ideal sources of iPSCs.
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Objective To investigate the role of NFIC on the stimulation effects of cAMP-induced differentiation of stem cells from the apical papilla ( SCAPs) in vitro. Methods SCAPs isolated from dental papilla of human imma-ture third molars were cultured by enzyme digestion. SCAPs were transfected with lentivirus that overexpressed NF-IC gene ( ov-NFIC) or an empty vector ( LV-empty) and co-treatment with Forskolin. Mineralized nodule formation of each group was measured by alizarin red staining. Quantitative real-time reverse-transcription polymerase chain reaction was performed to test the expressions of RUNX2,ALP,OCN mRNA. Results Forskolin increased the ex-pression of Runx2, ALP, OCN mRNA as well as matrix mineralization in SCAPs, and the stimulation effects of For-skolin were enhanced by overexpressing NFIC gene. Conclusion The results indicate that NFIC can promote cAMP-induced differentiation of SCAPs.
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Objective To comparatively study the features of two reprogramming systems of induced pluripotent stem cells (iPSCs)from human dental origin.Methods Two kinds of reprogramming system,i.e.STEMCCA lentivirus /feed layer and Sen-dai virus /matrigel were used to induce human stem cells from apical papilla(SCAP)into iPSCs,respectively.The induction efficien-cies,workload of generating iPSCs,aneuploidy karyotype ratio,complexities of eliminating exogenous transcription factors and spe-cific markers expression were compared between these two systems.Results The STEMCCA reprogramming system required to prepare the feeder cell MEF.The reprogramming efficiency was 0.1%.Transcription gene-free iPSCs cells were obtained by the Cre-loxp enzyme digestion technique at the later stage.Sendai virus reprogramming system was feeder-free and the preparation of matrigel was quite simple with unified standard.The reprogramming efficiency was 0.7%,which was much higher than that of STEMCCA system(P <0.05).The exogenous virus and transgenes could be gradually eliminated after several passages of natural subclone.Conclusion The Sendai virus/matrigle reprogramming system is much more applicable for the induction of iPSCs from dental origin than the STEMCCA system.
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Autologous bone transplantation is the current gold standard for reconstruction of jawbone defects. Bone regeneration usingmesenchymal stem cells (MSC) is an interesting alternative to improve the current techniques,which necessitate a second site of surgery resulting in donor site morbidity. In this study,we compared the osteogenic ability of jawboneMSC(JB-MSC) withMSC from tissues with neural crest origin, namely, the dental pulp, apical papilla and periodontal ligament. All four types ofMSC were isolated from the same patient (n = 3 donors) to exclude inter-individual variations.TheMSCgrowth and differentiation properties were characterized. The osteogenic differentiation potential in each group of cells was assessed quantitatively to determine if there were any differences between the cell types. All cells expressed the MSC-associated surface markers CD73, CD90, CD105, and CD146 and were negative for CD11b, CD19, CD34, CD45 and HLA-DR. All cell types proliferated at similar rates, exhibited similar clonogenic activity and could differentiate into adipocytes and osteoblasts. An alkaline phosphatase assay, OsteoImage™ assay for mineralization and qRT-PCR measuring the genes runx2, ALP and OCN, indicated that there were no significant differences in the osteogenic differentiation ability between the variousMSCs. In conclusion,we show that from a small segment of jawbone it is possible to isolate sufficient quantities of MSC and that these cells can easily be expanded and differentiated into osteoblasts. JB-MSC appear to be good candidates for future bone regeneration applications in the craniofacial region.
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Humanos , Adipócitos , Fosfatase Alcalina , Regeneração Óssea , Transplante Ósseo , Polpa Dentária , Antígenos HLA-DR , Técnicas In Vitro , Células-Tronco Mesenquimais , Mineradores , Crista Neural , Osteoblastos , Ligamento Periodontal , Células-Tronco , Doadores de TecidosRESUMO
Objective SCAP are seen as seed cells of peri-apical tissue regeneration and used in periapical tissue regeneration project based on stem cells .In this study, we aim to explore the ef-fectdifferent mechanical stretch stress on the proliferation and differen-tiation potential of human stem cells from the apical papilla ( SCAP ) ,and to clarify the mechanism of how mechanical stretch stress regulate human SCAP ,which will provide theoretical guidance for ortho-dontic treatment . Methods Human SCAP was isolated , cultured and identified by combined explants method and enzymatic separa-tion method and limited dilution .MTT assay was used to detect the effect different static mechanical stretch stress stimulation have on the proliferation of SCAP .Western blot was used to detect the expression changes of SCAP osteogenesis /odontoblast differentiation-re-lated protein (ALP, OSX,DSP) under mechanical stretch stressand to detect the expression of SCAP endoplasmic reticulum stress mo -lecular chaperone GRP 78 under different static mechanical stretch stress . Results SCAP were successfully isolated and cultured , and we induced SCAP to differentiate into osteoblasts and adipocytes successfully by osteogenic medium and adipogenic medium .Flow cytometry was performed in accordance with SCAP immunophenotype .Compared with the control group , 150g mechanical stretch stress stimulation promoted SCAP proliferation first and then inhibited SCAP proliferation [(0.481±0.226),(1.375±0.104),(1.425± 0.136),(1.556±0.268),(0.589±0.29),P<0.05].It was same in the 200g group.250g mechanical stretch stress stimulation signifi-cantly inhibited SCAP proliferation [(0.373±0.146),(0.545±0.069),(0.745±0.273),(0.967±0.278),(1.060±0.362),P<0.05]. The expression levels of ALP , OSX and DSP protein in each group were higher than those in the control group (P<0.05).Compared with the control group, the expression of GRP78 protein was up-regulated (P<0.05). Conclusion Mechanical stretch stress could regulate the SCAP proliferation and osteogenesis/odontoblast differentiation .What′s more,endoplasmic reticulum stress played a role in osteogenesis/odontoblast differentiation under mechanical stretch stress and promoted SCAP osteogenesis /odontoblast differentiation .
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Objective SCAP are seen as seed cells of peri-apical tissue regeneration and used in periapical tissue regeneration project based on stem cells .In this study, we aim to explore the ef-fectdifferent mechanical stretch stress on the proliferation and differen-tiation potential of human stem cells from the apical papilla ( SCAP ) ,and to clarify the mechanism of how mechanical stretch stress regulate human SCAP ,which will provide theoretical guidance for ortho-dontic treatment . Methods Human SCAP was isolated , cultured and identified by combined explants method and enzymatic separa-tion method and limited dilution .MTT assay was used to detect the effect different static mechanical stretch stress stimulation have on the proliferation of SCAP .Western blot was used to detect the expression changes of SCAP osteogenesis /odontoblast differentiation-re-lated protein (ALP, OSX,DSP) under mechanical stretch stressand to detect the expression of SCAP endoplasmic reticulum stress mo -lecular chaperone GRP 78 under different static mechanical stretch stress . Results SCAP were successfully isolated and cultured , and we induced SCAP to differentiate into osteoblasts and adipocytes successfully by osteogenic medium and adipogenic medium .Flow cytometry was performed in accordance with SCAP immunophenotype .Compared with the control group , 150g mechanical stretch stress stimulation promoted SCAP proliferation first and then inhibited SCAP proliferation [(0.481±0.226),(1.375±0.104),(1.425± 0.136),(1.556±0.268),(0.589±0.29),P<0.05].It was same in the 200g group.250g mechanical stretch stress stimulation signifi-cantly inhibited SCAP proliferation [(0.373±0.146),(0.545±0.069),(0.745±0.273),(0.967±0.278),(1.060±0.362),P<0.05]. The expression levels of ALP , OSX and DSP protein in each group were higher than those in the control group (P<0.05).Compared with the control group, the expression of GRP78 protein was up-regulated (P<0.05). Conclusion Mechanical stretch stress could regulate the SCAP proliferation and osteogenesis/odontoblast differentiation .What′s more,endoplasmic reticulum stress played a role in osteogenesis/odontoblast differentiation under mechanical stretch stress and promoted SCAP osteogenesis /odontoblast differentiation .