Recombinant human basic fibroblast growth factor (bFGF) stimulates periodontal regeneration in class II furcation defects created in beagle dogs

Several growth factors (or cytokines) have been recently investigated for their use as potential therapeutics for periodontal tissue regeneration. The objective of this study was to evaluate periodontal tissue regeneration, including new bone and cementum formation, following topical application of recombinant basic fibroblast growth factor (bFGF, FGF-2) to furcation class II defects. Twelve furcation class II bone defects were surgically created in six beagle dogs, then recombinant bFGF (30 micro g/site) + gelatinous carrier was topically applied to the bony defects. Six weeks after application, periodontal regeneration was analyzed. In all sites where bFGF was applied, periodontal ligament formation with new cementum deposits and new bone formation was observed histomorphometrically, in amounts greater than in the control sites. Basic FGF-applied sites exhibited significant regeneration as represented by the new bone formation rate (NBR) (83.6 +/- 14.3%), new trabecular bone formation rate (NTBR) (44.1 +/- 9.5%), and new cementum formation rate (NCR) (97.0 +/- 7.5%). In contrast, in the carrier-only sites, the NBR, NTBR, and NCR were 35.4 +/- 8.9%, 16.6 +/- 6.2%, and 37.2 +/- 15.1%, respectively. Moreover, no instances of epithelial down growth, ankylosis, or root resorption were observed in the bFGF-applied sites examined. The present results indicate that topical application of bFGF can enhance considerable periodontal regeneration in artificially created furcation class II bone defects of beagle dogs.     

Expression of receptors for basic fibroblast growth factor on human periodontal ligament cells

Basic fibroblast growth factor (FGF-2; bFGF) is a major mitogen for connective tissue cells, and participates in the healing process. It has already been reported that FGF-2 could be applicable to enhance periodontal regeneration. In the present study, we examined FGF receptor (FGFR) expression on human periodontal ligament (PDL) cells. The binding of [¹²⁵I]-labeled FGF-2 to human PDL cells was studied by radioreceptor assay. The binding of [¹²⁵I]-FGF-2 to PDL cells reached a plateau after 2.5 h incubation at 4°C and was inhibited by the addition of unlabeled FGF-2 and acidic FGF (FGF-1; aFGF), but not insulin-like growth factor-I, platelet-derived growth factor and transforming growth factor-β₁. Scatchard analysis revealed the presence of approximately 1.0 × 10⁵ FGF-2 binding sites per cell with an apparent Kd of 1.2 × 10-¹⁰ M. Interestingly, the binding of [¹²⁵I]-FGF-2 on PDL cells reached its maximum at d 6 of the culture and then gradually decreased. Scatchard analysis also demonstrated that the number of FGFRs on a PDL cell was altered during the course of the culture, while the affinity between FGF-2 and its receptor was not. The responsiveness of PDL cells to FGF-2, which was monitored by the inhibitory effect on alkaline phosphatase activity, was reduced in proportion to the decrease in the number of FGFRs on the PDL cells. The present study suggests that PDL cells alter the responsiveness to FGF-2 during the course of the culture by changing the density of its receptor, and that the density of FGFR expression might be a marker of the cytodiflerentiation of PDL cells into mineralized tissue forming cells.     

Expression of receptors for basic fibroblast growth factor on human periodontal ligament cells

Basic fibroblast growth factor (FGF-2; bFGF) is a major mitogen for connective tissue cells, and participates in the healing process. It has already been reported that FGF-2 could be applicable to enhance periodontal regeneration. In the present study, we examined FGF receptor (FGFR) expression on human periodontal ligament (PDL) cells. The binding of [¹²⁵I]-labeled FGF-2 to human PDL cells was studied by radioreceptor assay. The binding of [¹²⁵I]-FGF-2 to PDL cells reached a plateau after 2.5 h incubation at 4°C and was inhibited by the addition of unlabeled FGF-2 and acidic FGF (FGF-1; aFGF), but not insulin-like growth factor-I, platelet-derived growth factor and transforming growth factor-β₁. Scatchard analysis revealed the presence of approximately 1.0 × 10⁵ FGF-2 binding sites per cell with an apparent Kd of 1.2 × 10⁻¹⁰ M. Interestingly, the binding of [¹²⁵I]-FGF-2 on PDL cells reached its maximum at d 6 of the culture and then gradually decreased. Scatchard analysis also demonstrated that the number of FGFRs on a PDL cell was altered during the course of the culture, while the affinity between FGF-2 and its receptor was not. The responsiveness of PDL cells to FGF-2, which was monitored by the inhibitory effect on alkaline phosphatase activity, was reduced in proportion to the decrease in the number of FGFRs on the PDL cells. The present study suggests that PDL cells alter the responsiveness to FGF-2 during the course of the culture by changing the density of its receptor, and that the density of FGFR expression might be a marker of the cytodifferentiation of PDL cells into mineralized tissue forming cells.     

引导组织再生治疗术及碱性成纤维细胞生长因子促进牙周组织再生的实验研究

目的 了解引导组织再生治疗术(GTR)及碱性成纤维细胞生长因子(b-FGF)是否能够促进犬Ⅱ度根分叉病变牙周组织的再生。方法 将六只杂种犬的下颌双侧第二、三、四前磨牙制备慢性Ⅱ度根分叉病变模型后。随机分成两组，每组三只动物。每组中一只动物一侧进行GTR治疗，另一侧只进行翻瓣术治疗；另一只双侧都进行GTR b-FGF治疗；剩下的一只一侧进行GTR治疗，另一侧进行GTR b-FGF治疗。分别于术后第6、8周进行形态学和组织学观察。结果与翻瓣术组对比，GTR组和GTR b-FGF组都有明显的牙周组织再生；而GTR组和GTR b-FGF组之间牙周组织再生的量没有明显的差别。结论 GTR能够促进犬Ⅱ度根分叉病变牙周组织的再生，但单纯应用喷洒法使用b-FGF于牙周组织再生术中难以发挥其应有的疗效。     

碱性成纤维细胞生长因子对人牙周膜成纤维细胞内核心蛋白多糖基因表达的影响

目的研究碱性成纤维细胞生长因子（bFGF）对人牙周膜成纤维细胞内核心蛋白多糖（decorin）的影响,探讨bFGF在牙周再生中的意义。方法体外原代培养人牙周膜成纤维细胞,用外源性bFGF刺激细胞,半定量RTPCR法检测牙周膜成纤维细胞内decorin基因表达的变化。结果电泳结果表明,bFGF抑制人牙周膜成纤维细胞内decorin的mRNA合成,并且随着质量浓度的增加抑制作用减弱。结论decorin具有很多生物学功能,bFGF对decorin的抑制作用很可能是牙周炎损伤修复过程中一个重要的调节因素,为bFGF在牙周组织再生中的作用提供理论基础。     

碱性成纤维细胞生长因子对牙周膜成纤维细胞整合素β1亚单位mRNA表达的影响

目的研究碱性成纤维细胞生长因子（bFGF）对人牙周膜成纤维细胞整合素β1亚单位mRNA表达的影响,探讨bFGF在牙周组织再生中的意义。方法体外培养人牙周膜成纤维细胞,分别用质量浓度0.1、1.0、10.0 ng·mL-1的bFGF刺激细胞,培养24、48、72 h,采用实时荧光定量聚合酶链反应法检测牙周膜成纤维细胞内整合素β1亚单位mRNA表达的变化。结果bFGF可促进人牙周膜成纤维细胞内整合素β1亚单位mRNA的合成,在培养24、48、72 h时,1.0 ng·mL-1组整合素β1亚单位表达均明显高于对照组;培养72 h时各实验组整合素β1亚单位表达均明显高于培养24、48 h时。结论bFGF通过提高整合素β1亚单位mRNA的表达,促进牙周膜成纤维细胞的黏附,在牙周组织修复再生中起作用。     

体外培养碱性成纤维细胞因子对人牙周膜成纤维细胞生物学特性的影响

目的：探讨体外培养过程中,碱性成纤维细胞生长因子（bFGF）对人牙周膜成纤维细胞（hPDLFs）生物学特性的影响。方法：体外分离培养人牙周膜成纤维细胞并鉴定,加入不同浓度bFGF（1ng／ml、10ng／ml、50ng／ml、100ng／ml）培养,MTT方法检测细胞增殖情况;并对细胞进行矿化诱导,检测细胞的碱性磷酸酶活性。结果：hPDLFs呈星形或长梭形,免疫组化波丝蛋白阳性,角蛋白阴性,证实该细胞来源可靠。bFGF在一定浓度范围内,与细胞增殖成正比,而在本实验培养条件下（10％FBS）bFGF最大效应浓度为10ng／ml。矿化诱导条件下,碱性磷酸酶活性明显增加,在联合应用bFGF的情况下,100ng／ml组碱性磷酸酶活性明显高于其他组。结论：不同浓度bFGF对人牙周膜成纤维细胞生物学行为的影响不同,在一定浓度条件下,低浓度bFGF促进人牙周膜成纤维细胞的增殖,而高浓度bFGF作用于人牙周膜成纤维细胞可能更易于分化。     

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牙周疾病是以牙周支持组织破坏为特征,最终导致牙齿丧失的严重危害口腔健康的疾病,牙周疾病造成的牙周附着丧失和牙槽骨缺损是临床急需解决的一个重要问     

碱性成纤维细胞生长因子对牙周膜细胞表皮生长因子受体基因表达的影响

目的研究碱性成纤维细胞生长因子（bFGF）对人牙周膜细胞（PDLC）表达表皮生长因子受体（EGFR）的影响,探讨bFGF在牙周组织分化再生中的意义。方法体外原代培养人PDLC,有限稀释法形成单细胞克隆,用外源性bFGF刺激单细胞克隆,采用逆转录聚合酶链反应（RT-PCR）检测克隆细胞内EGFR基因表达的变化。结果bFGF促进人PDLC内EGFR mRNA的合成,并且随着质量浓度的增加促进作用增强。结论bFGF对EGFR的促进作用很可能是牙周炎损伤修复过程中一个重要的调节因素,为牙周组织分化再生提供部分理论基础。     

纳米羟基磷灰石材料复合碱性成纤维细胞因子促进牙周组织再生的实验研究

目的:探讨新型纳米羟基磷灰石材料—纳米羟晶/胶原仿生骨材料(nHAC)作为生长因子载体和牙周组织工程支架材料应用的可行性,观察评价其对牙周组织再生修复的影响和意义。方法:将人牙周膜细胞(HPDLCs)接种于nHAC三维支架上复合培养,体外扫描电镜观察HPDLCs在nHAC支架上的附着、生长情况,并将碱性成纤维细胞生长因子(bFGF)与nHAC复合植入动物人工牙周组织缺损,表面覆盖聚四氟乙烯(e-PTFE)膜,以空白对照组和单纯置膜组作为对照。术后8周进行组织学观察和测量,分析评价其牙周组织的再生情况。结果:扫描电镜可见nHAC具有良好的多孔网状结构,PDLCs在nHAC支架上贴附紧密,生长旺盛,伸展充分,动物实验结果显示nHAC/bFGF/膜复合植入组较空白对照组和单纯置膜组有更多新生的牙槽骨、牙周膜和牙骨质生成,结果具有显著性差异。结论:nHAC具有良好的三维空间结构和细胞相容性,与bFGF复合植入牙周缺损后可显著促进牙周组织再生,提示nHAC有望成为理想的生长因子载体和牙周组织工程支架材料。     

雌激素与机械张应力对碱性成纤维细胞生长因子的影响

目的研究雌激素与机械张应力共同作用对人牙周膜成纤维细胞(human periodontal ligament fibroblast,HPDLF)增殖分化相关因子碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)mRNA表达水平的影响。方法体外培养HPDLF,分为两组。实验组使用10-7mol/L雌激素干预后,使用4点弯曲加力装置对HP-DLF进行张应力加载;对照组不使用雌激素干预,只进行机械张应力加载。采用实时多聚酶链反应、琼脂糖凝胶电泳和紫外线分析检测HPDLF加力时3、6、12 h时bFGF基因的表达。结果加力0、3、6、12 h时,对照组bF-GFmRNA表达量光密度值分别为1.000,3.245±0.261,4.498±0.431,7.969±0.597,实验组bFGFmRNA表达量光密度值分别为16.736±1.003,22.542±1.768,27.923±1.914,31.556±2.142。雌激素干预与张应力加载共同作用与单纯受到机械张应力作用相比,bFGF的mRNA表达量明显上调(P<0.05)。结论雌激素对机械张应力状态下的牙周膜成纤维细胞增殖分化相关因子bFGF有较强调控作用,表现为bFGF的mRNA表达量显著上升。     

血小板衍化生长因子-BB和碱性成纤维细胞生长因子对人牙周膜细胞增殖的影响

目的探讨重组人血小板衍生生长因子（rhPDGF）-BB和重组人碱性成纤维细胞生长因子（rh—bFGF）对人牙周膜细胞（PDLC）增殖的影响。方法原代培养人PDLC,应用甲噻唑四唑氮（MTT）比色和流式细胞仪检测rh-PDGF-BB和rh-bFGF单独及联合作用下细胞的增殖能力和增殖指数。结果rhPDGF-BB和rh-bFGF联合作用后,PDLC增殖能力提高,活性增强。其中,10μg&#183;L^-1的rhPDGF—BB加10μg&#183;L^-1的bFGF为最佳显效质量浓度,第3天为最佳显效时间。结论PDGF—BB和bFGF联合能更显著促进人PDLC的增殖并且具有明显的协同效应。     

碱性成纤维细胞生长因子对人牙周膜细胞多配体蛋白聚糖-4基因表达的影响

目的 通过研究碱性成纤维细胞生长因子(bFGF)对人牙周膜细胞(PDLC)多配体蛋白聚糖-4 mRNA水平表达的影响,探讨bFGF在人PDLC迁移中的作用.方法 收集正畸拔除的12～18岁青少年健康前磨牙68颗,体外原代培养人PDLC,用外源性bFGF刺激细胞,培养24、48、72 h后,采用实时荧光定量聚合酶链反应检...     

Glucose modulates growth of gingival fibroblasts and periodontal ligament cells: correlation with expression of basic fibroblast growth factor

Diabetes mellitus is a systemic disease with profound effects on oral health and periodontal wound healing. Uncontrolled diabetes adversely affects surgical wound healing and is often associated with abnormal proliferation of fibroblasts, excessive angiogenesis and poor bone regeneration. Human gingival fibroblasts and periodontal ligament cells from both diabetics and non-diabetics were evaluated for growth responses following culture in 20 mM glucose, a concentration compatible with blood glucose levels in uncontrolled diabetics. Gingival fibroblasts derived from 9 non-diabetic patients and 3 insulin-dependent diabetics either proliferated or showed little change of growth in elevated glucose. Enhanced proliferation was observed following 1 wk of culture in glucose. Growth of periodontal ligament cells from 5 non-diabetic patients was inhibited by 20 mM glucose. Fibroblasts that were markedly growth stimulated were probed for expression of basic fibroblast growth factor (bFGF) using a reverse-transcribed polymerase chain reaction (RT-PCR). Results indicate that fibroblasts exhibiting the greatest increase in growth in response to high glucose also exhibited increased expression of bFGF. No changes were observed in mRNA expression for platelet-derived growth factor-AA, platelet-derived growth factor-BB, insulin-like growth factor and transforming growth factor-β₁. Mitogenic effects induced by the cytosol of fibroblasts exhibiting increases of growth in 20 mM glucose were abrogated by neutralizing antibodies to bFGF. In addition, some periodontal ligament cells that were growth inhibited by high glucose had reduced expression of bFGF. These data suggest that bFGF may play a role in the abnormal wound healing associated with periodontal surgery of uncontrolled diabetics.     

碱性成纤维细胞生长因子对人牙周膜细胞增殖和细胞周期的影响

目的：探讨碱性成纤维细胞生长因子（bFGF)对体外培养的人牙周膜细胞（PDLC）增殖和细胞周期的影响，且呈剂量依赖性；FCM对细胞周期时相分析结果显示，经bFGF作用后，人PDLC的DNA合成量（S%）显著升高，G1%降低，反映细胞增殖活力的增殖指数PrI值（S G2M）%增高。结论：bFGF能促进人PDLC的增殖和DNA合成，这一作用可能是通过促使处于G1期的PDLC进入S期来实现的。     

骨形成蛋白—2和成纤维细胞生长因子对人牙周膜细胞增殖的联合效应

目的：探讨重组人骨形成蛋白－2（rhBMP-2)和碱性成纤维细胞生长因子(bFGF) 单独或联合作用对人牙周膜细胞（PDLCs)增殖的影响。方法：体外培养人PDLCs,分别用不同浓度的rhBMP-2和bFGF单独或联合作用,用四唑盐比色法（MTT法）进行观察。结果：rhBMP-2和bFGF单独作用后PDLCs的增殖较对照组有明显的升高;而rhBMP-2与bFGF联合作用后PDLCs的增殖较各自单独作用有更明显的升高（P＜0．05）。结论：rhBMP-2与bFGF联合应用对于促进人PDLCs的增殖具有协同作用。