Effect of Intracanal Medicaments and Irrigants on the Release of Transforming Growth Factor Beta 1 and Vascular Endothelial Growth Factor from Cervical Root Dentin

IntroductionThis study aimed to evaluate (1) the effect of irrigating solutions and intracanal medicaments on the release of transforming growth factor beta 1 (TGF-β1) and vascular endothelial growth factor (VEGF) from cervical root dentin and (2) the effect of associating triple antibiotic paste (TAP) and calcium hydroxide paste (CH) with 2% chlorhexidine (CHX) on TGF-β1 release.MethodsFirst, 119 specimens from roots (cervical thirds) were obtained and were distributed into 5 groups: 2% CHX, 2.5% sodium hypochlorite, TAP, CH, and 10% EDTA by each growth factor (TGF-β1 [n = 8] and VEGF [n = 8]). Then, specimens were distributed as follows (n = 13): TAP + 2% CHX, CH + 2% CHX, and 10% EDTA and treated with irrigating solutions and intracanal medicaments. After the treatments, the specimens were immersed in 10% EDTA (20 minutes), and the solution was analyzed using the enzyme-linked immunosorbent assay. The data were submitted to normality, homogeneity of variance, and Mann-Whitney tests (P < .05).ResultsSignificant differences were found between the irrigating solutions (P < .05) and intracanal medicaments for TGF-β1 (P < .05). No VEGF release was detected for any group. Our results showed no significant differences among the TAP + 2% CHX and EDTA groups for TGF-β1 but a significant difference between CH + 2% CHX and the other groups (P < .05).ConclusionsThe use of 2% CHX as the irrigating solution, CH as the intracanal medicament, and 10% EDTA as the final irrigation provides higher TGF-β1 release from the cervical root dentin, whereas VEGF was not detected. Moreover, TAP and 2% CHX with 10% EDTA as the final irrigation resulted in greater TGF-β1 release from cervical root dentin than CH + 2% CHX.     

Release of Transforming Growth Factor Beta 1 from Human Tooth Dentin after Application of Either ProRoot MTA or Biodentine as a Coronal Barrier

IntroductionVarious factors may influence intracanal calcification in teeth treated with regenerative endodontic procedures. Bioactive materials, including ProRoot MTA (MTA; Dentsply Tulsa Dental Specialties, Memphis, TN) and Biodentine (BD; Septodont, Saint-Maur-des-Fossés, France), have been widely used as a coronal barrier in the final step of regenerative endodontic procedures. The purposes of this study were to evaluate the effect of either MTA or BD after application as a coronal barrier on transforming growth factor beta 1 (TGF-β1) release from root canal dentin and to observe the impact of these materials on human apical papilla cell (APC) mineralization.MethodsEither MTA or BD was applied in enlarged root canals of human root segments. After storing for 14 days in phosphate-buffered saline, TGF-β1 release was evaluated using an enzyme-linked immunosorbent assay. To investigate the effect of the materials on APC mineralization, APCs were grown in the presence of either the materials alone or material-filled root segments. Cell mineralization was quantified after 14 and 21 days using alizarin red S staining. Calcium deposits were quantitatively analyzed. Statistical analysis was performed using the Kruskal-Wallis and Mann-Whitney U tests with the significance level at .05.ResultsThe greatest amount of TGF-β1 release was observed in the root segments treated with BD. BD, used either alone or as a coronal barrier, promoted greater APC mineralization than did MTA on both days 14 and 21. Interestingly, when BD was applied as a coronal barrier, the mineralization effect was significantly reduced compared with the use of the materials alone (P < .05).ConclusionsWhen used as a coronal barrier, BD promoted the release of TGF-β1 from the root canal dentin. A higher mineralization effect was observed with BD than with MTA.     

Human Dental Pulp Stem Cells Suppress Alloantigen-induced Immunity by Stimulating T Cells to Release Transforming Growth Factor Beta

Introduction

Human dental pulp stem cells (hDPSCs) are ideal candidates for regenerating damaged dental tissue. To examine the possibility that hDPSCs may be used to regenerate pulp, we tested their in vitro effects on acute allogeneic immune responses.

Melatonin Treatment Alters Biological and Immunomodulatory Properties of Human Dental Pulp Mesenchymal Stem Cells via Augmented Transforming Growth Factor Beta Secretion

IntroductionMelatonin is an endogenous neurohormone with well-reported anti-inflammatory and antioxidant properties, but the direct biological and immunomodulatory effects of melatonin on human dental pulp stem cells (hDPSCs) has not been fully elucidated. The aim of this study was to evaluate the influence of melatonin on the cytocompatibility, proliferation, cell migration, odontogenic differentiation, mineralized nodule formation, and immunomodulatory properties of hDPSCs.MethodsTo address the melatonin biological effects on hDPSCs, the cytocompatibility, proliferation, cell migration, odontogenic differentiation, mineralized nodule formation, and immunomodulatory properties of hDPSCs after melatonin treatment were evaluated. The statistical differences were evaluated using 1-way analysis of variance with the Tukey multiple comparison test.ResultsWe found that melatonin did not alter hDPSC immunophenotype or cell viability, even at the highest concentrations used. However, using intermediate melatonin concentrations (10–300 μmol/L), a significantly higher proliferation rate (P < .05 and P < .01) and migration of hDPSCs (P < .01) were observed. Importantly, melatonin treatment (100 μmol/L) significantly increased the secretion of the anti-inflammatory cytokine transforming growth factor beta (P < .05 and P < .01) and provoked a more robust antiproliferative effect on mitogen-stimulated T cells (P < .05). Finally, and unlike previous results found with mesenchymal stem cells from other sources, melatonin fails to induce or accelerate the spontaneous osteogenic differentiation of hDPSCs.ConclusionsTogether, these findings provide key data on the bioactivity of melatonin and its effects on hPDSC biological and immunomodulatory properties.     

大鼠正畸牙移动过程中转化生长因子β1在牙槽骨中的表达

目的：观察大鼠正畸牙移动过程中转化生长因子β1（TGF－β1）在牙槽骨中的表达变化,探讨TGF－β在正畸牙槽骨改建中的作用。方法：建立大鼠正畸牙移动模型,用免疫组织化学方法检测牙槽骨中TGF－β1的表达。结果：对照组正常牙槽骨组织TGF－β1呈弱阳性表达。实验组压力侧和张力侧牙槽骨组织TGF－β1阳性表达增强。牙齿移动5d组和7d组,压力侧破骨细胞和张力侧成骨细胞TGF－β1均呈阳性表达。结论：TGF－β1作为局部调控因子可能参与了正畸牙槽骨改建过程。     

Immunoexpression of Transforming Growth Factor Beta and Interferon Gamma in Radicular and Dentigerous Cysts

Introduction

The aim of this study was to evaluate and compare the immunohistochemical expression of transforming growing factor beta (TGF-β) and interferon gamma (IFN-γ) between radicular cysts (RCs) and dentigerous cysts (DCs).

Methods

Twenty RCs and DCs were selected for analysis of the immunoexpression of TGF-β and IFN-γ in the epithelium and capsule.

Results

The cell reactivity of TGF-β and IFN-γ in the lining epithelium and capsule of RCs showed no significant differences when compared with DCs (P > .05). There was a tendency of a higher expression of TGF-β in the capsule of DCs.

Conclusions

Our results showed the presence of TGF-β and IFN-γ in RCs and DCs, supporting the hypothesis that both participate in the development of these lesions, where IFN-γ usually plays a role in bone resorption, which is counterbalanced by the osteoprotective activity performed by TGF-β.     

rhBMP-2对人牙龈成纤维细胞体外矿化能力的影响

目的 探讨重组人骨形成蛋白-2(rhBMP-2)对人牙龈成纤维细胞(GF)体外矿化能力的影响。方法 以组织块培养法培养人牙龈成纤维细胞，分别以含或不含rhBMP-2的矿化培养液长期培养，以倒置显微镜观察细胞生长状况。于培养第7天检测碱性磷酸酶(ALP)活性。于培养第30天行von Kossa染色观察矿化结节。结果 rhBMP-2提高GF的ALP水平，呈剂量依赖性，并能够刺激GF在体外形成矿化结节。结论 rhBMP-2能够诱导人GF向成骨细胞表型分化。     

牙周炎大鼠正畸牙移动张力侧牙周组织转化生长因子β1表达研究冰

目的研究炎性牙周条件下牙移动及牙周改建中转化生长因子β1（transforming growth factor—β1,TGF-β1）的表达。方法90只雄性SD大鼠随机分为为正常牙移动组、牙周炎牙移动组,通过对大鼠上颌第一磨牙的近中移动,在预定的0、0．5、1、2、3、5、7、14、21d时间点处死动物,运用免疫组织化学,检测牙移动张力侧TGF—β1蛋白表达强度及时间分布特点。结果正常牙移动组中,TGF-β1在牙周膜的成纤维细胞、成骨细胞和骨髓组织呈弱阳性表达,牙移动1d后,张力侧牙周膜区免疫染色的阳性反应明显增强,2d后达到高峰;牙周炎牙移动组,TGF-β1在破骨细胞、牙周膜成纤维细胞呈阳性表达,牙移动0．5d后,张力侧牙周膜区免疫染色的阳性反应明显增强,以后逐渐下降。结论牙周炎症影响正畸力效应下的牙周组织改建。     

软骨上清液与转化生长因子诱导滑膜间充质干细胞向软骨细胞分化的对比研究

目的:对比使用软骨上清液和转化生长因子两种方法诱导滑膜间充质干细胞向软骨细胞分化。方法:分别采用消化法获取SD大鼠滑膜间充质干细胞和软骨细胞。体外扩增。实验组一:通过软骨细胞上清液诱导滑膜间充质干细胞向软骨方向分化。实验组二:通过软骨诱导液(TGF-β1,ITS+ Premix,2-磷酸抗坏血酸等)诱导滑膜间充质干细胞向软骨方向分化。培养21 d后通过形态学、免疫组织化学法检测其生物学特性,RT-PCR检测诱导后产物Ⅱ型胶原RNA含量。结果:2种诱导方法均能诱导滑膜间充质干细胞成软骨细胞方向分化。在形态学可见2组诱导后产物成软骨样结构,呈乳白色,质地韧。免疫组织化学鉴定基质能被Ⅱ型胶原染色,细胞染色呈现棕黄色。RT-PCR结果显示诱导后的微团表达软骨特异性基因Ⅱ型胶原和蛋白聚糖。结论:两组实验组均能诱导滑膜间充质干细胞向软骨方向分化。对比两种实验方法,使用上清液诱导滑膜间充质干细胞向软骨方向分化能力更强。     

胰岛素样生长因子-I对培养人牙周膜成纤维细胞生物学活性的影响

胰岛素样生长因子(IGF)可通过直接作用于成骨细胞或通过甲状旁腺素间接影响骨代谢过程而参与骨的改建。本文利用细胞培养技术观察了IGF-I对培养人牙周膜成纤维细胞(PDLFs)增殖、碱性磷酸酶(ALP)活性变化以及蛋白质含量改变的影响。结果显示,IGP-I对PDLFs有明显的促增殖作用,对PDLFs的ALP活性、蛋白质合成也具有较强的促进作用,表明IGF-I可能通过PDLFs发挥其调节牙槽骨改建的作用,从而在牙齿移动过程中起重要作用。     

Growth and Differentiation Factor-5 Regulates the Growth and Differentiation of Human Dental Pulp Cells

Introduction

Growth and differentiation factor-5 (GDF-5) is a multifunctional protein that regulates the development and repair in many tissues. The purpose of this study was to investigate whether GDF-5 may influence the proliferation, differentiation, and collagen turnover of human dental pulp cells.

Methods

Human dental pulp cells were treated with different concentrations of GDF-5 (0–500 ng/mL). Morphology of pulp cells was observed under a microscope. Cell proliferation was evaluated by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay. Immunofluorescent assay was used to observe the percentages of cell mitosis. Collagen content was measured by Sircol collagen assay. Tissue inhibitor of metalloproteinase-1 level in the culture medium was measured with enzyme-linked immunosorbent assay and Western blotting. Cell differentiation was evaluated by alkaline phosphatase (ALP) staining and ALP enzyme activity assay.

Results

After exposure of dental pulp cells to various concentrations of GDF-5, cell number was up-regulated significantly in dose-dependent manner. GDF-5 also stimulated mitosis of dental pulp cells as indicated by an increased percentage of binucleated cells from 28% to 35%–45%. GDF-5 did not affect the collagen content and tissue inhibitor of metalloproteinase-1 level of pulp cells. GDF-5 decreased the ALP activity of pulp cells as analyzed by ALP staining and enzyme activity assay, with 14%–44% of inhibition.

Conclusions

GDF-5 revealed mitogenic and proliferative activity to dental pulp cells. GDF-5 showed inhibitory effect on ALP activity but little effect on the collagen turnover. These events are crucial in specific stages of dental pulp repair and regeneration. GDF-5 may be potentially used for tissue engineering of pulp-dentin complex.     

尼古丁对大鼠牙槽骨内ALP和DMP1 C末端表达的影响

目的:观察尼古丁对大鼠牙槽骨内碱性磷酸酶(ALP)和牙本质基质蛋白1(DMP1)C末端表达的影响. 方法:将20只5周龄健康雄性Wistar大鼠随机等分为实验组和对照组,每组再随机等分为30天和60天两个亚组,实验组每天腹腔注射尼古丁0.73 mg·kg-1. 分别使用钙钴法和免疫组化法检测ALP和DMP1 C末端在大鼠牙槽骨内的表达. 结果: 对照组:ALP和DMP1 C末端都为深棕色线形表达于牙槽骨的钙化前缘, 形成明显的矿化条带;ALP和DMP1 C末端分别为棕色沉淀和棕黄色颗粒状密布于骨细胞周围.实验组:ALP和DMP1 C末端表达部位同对照组,表达强度减弱;相对于30天组,60天组变化更明显. 结论:尼古丁可能通过抑制大鼠牙槽骨内ALP和DMP1 C末端的表达,抑制大鼠牙槽骨的矿化.     

bFGF对大鼠骨髓细胞骨形成的影响

为了探讨碱性成纤维细胞生长因子对骨髓细胞系的作用采用Ｈ＾３－ＴｄＲ法,比色法和ＶｏｎＫｏｓｓａ染色法,检测ｂＦＧＦ作用下骨髓细胞增殖,分化,骨形成情况。结果持续使用ｂＦＧＦ时,其促进骨髓细胞增殖,并向成骨细胞分化。     

Collagen Membranes Adsorb the Transforming Growth Factor‐β Receptor I Kinase‐Dependent Activity of Enamel Matrix Derivative

Background: Enamel matrix derivative (EMD) and collagen membranes (CMs) are simultaneously applied in regenerative periodontal surgery. The aim of this study is to evaluate the ability of two CMs and a collagen matrix to adsorb the activity intrinsic to EMD that provokes transforming growth factor (TGF)‐β signaling in oral fibroblasts. Methods: Three commercially available collagen products were exposed to EMD or recombinant TGF‐β1, followed by vigorous washing. Oral fibroblasts were either seeded directly onto collagen products or were incubated with the respective supernatant. Expression of TGF‐β target genes interleukin (IL)‐11 and proteoglycan 4 (PRG4) was evaluated by real time polymerase chain reaction. Proteomic analysis was used to study the fraction of EMD proteins binding to collagen. Results: EMD or TGF‐β1 provoked a significant increase of IL‐11 and PRG4 expression of oral fibroblasts when seeded onto collagen products and when incubated with the respective supernatant. Gene expression was blocked by the TGF‐β receptor I kinase inhibitor SB431542. Amelogenin bound most abundantly to gelatin‐coated culture dishes. However, incubation of palatal fibroblasts with recombinant amelogenin did not alter expression of IL‐11 and PRG4. Conclusion: These in vitro findings suggest that collagen products adsorb a TGF‐β receptor I kinase‐dependent activity of EMD and make it available for potential target cells.     

矿化胶原膜浸提液对MG-63人骨肉瘤细胞分化相关基因表达的影响

目的 研究新型矿化胶原膜对MG-63人骨肉瘤细胞(简称：MG-63细胞)成骨分化相关基因表达的影响。方法 将 MG-63细胞与新型矿化胶原膜浸提液(实验组)共培养,以市售的胶原膜(Bio-gide)浸提液作为同类产品对照组,以不加材料的细胞培养液为空白对照组,采用荧光实时定量 PCR法检测碱性磷酸酶(ALP)、I型胶原(COL I)、骨保护素(OPG)和骨钙素(OC)mRNA表达水平;采用SPSS 17.0 软件对数据进行统计学分析。结果 在ALP与OPG mRNA相对表达量检测中,3组成骨细胞的表达量差异无统计学意义(P>0.05);在COL I与 OC基因相对表达量检测中,14天时Bio-gide组和矿化胶原膜组表达均明显上调,与空白对照组相比,差异均具有统计学意义(P<0.05),而Bio-gide组和矿化胶原膜组相比,差异无统计学意义(P>0.05)。结论 矿化胶原膜和Bio-gide膜的浸提液均可上调MG-63细胞COL I和OC的基因表达,在一定程度上促进了成骨细胞的分化。     

Quantitative Analysis of Interdental Gingiva in Patients With Chronic Periodontitis and Transforming Growth Factor‐β1 29C/T Gene Polymorphisms

Background: The association of transforming growth factor (TGF)‐β1 29C/T gene polymorphisms with level of tissue breakdown and periodontal disease progression is not clear. In this study, quantitative parameters of interdental papilla are investigated in patients with chronic periodontitis (CP) and TGF‐β1 29C/T gene polymorphisms. Methods: Sixty gingiva samples were included. After determination of TGF‐β1 29C/T gene polymorphisms using tetra‐primer amplification refractory mutation system/polymerase chain reaction (T‐ARMS‐PCR), 15 gingival tissue samples from patients with CP in each genotype (TT, TC, and CC) were considered as case groups. Fifteen control samples were also collected from healthy individuals. After tissue processing, interdental gingiva tissues were exhaustively sectioned into 4‐μm‐thick sections. Ten to 13 sections were sampled by systematic uniform random sampling and stained with Masson trichrome, and the volume density (V_v) of the gingival components was estimated using Cavalieri estimation. Results: Statistically significant differences were found in V_v of epithelium, connective tissue, collagenous and non‐collagenous matrix, and blood vessels between control and CP groups (P <0.0001). There was a corresponding decrease in the collagenous matrix V_v in patients with the TT genotype compared with those with CT and CC genotypes. Collagenous matrix and blood vessel V_v values were statistically correlated with the number of T alleles (r = ?0.74, r² = 54.8%, P = 0.0001 and r = 0.84, r² = 70.6%, P = 0.0001, respectively). Conclusion: This study shows that there is a strong association between TGF‐β1 29C/T gene polymorphisms and quantitative parameters of interdental papilla in patients with CP.     

TGF-β1壳聚糖微球的制备及其缓释效果研究

目的:制备转化生长因子β1(TGF-β1)壳聚糖微球,并研究其体外缓释效果。方法:采用乳化交联的方法制备壳聚糖微球,然后吸附TGF-β1获得TGF-β1壳聚糖微球。利用酶联免疫吸附的方法(ELISA)动态检测7 d内TGF-β1壳聚糖微球的缓释效果,并绘制释放曲线。结果:壳聚糖微球形态较好,粒径为(8.6±2.8)μm。TGF-β1壳聚糖微球在缓释实验的前12 h内释放速度较快,随后释放趋于平稳。7 d内微球的TGF-β1释放率为42.6%。结论:TGF-β1壳聚糖微球的制备工艺简单可行,成球性好,具有良好的缓释效果。