纤维蛋白粘合胶对人牙周膜细胞增殖和碱性磷酸酶活性的影响

目的：探讨纤维蛋白粘合胶（FG）对牙周膜细胞（PDLCs）增殖和碱性磷酸酶（ALP）活性的影响。方法：应用细胞培养技术,噻唑盐比色测定法（MTT法）和酶动力学方法,观察FG对PDLCs的增殖作用和时间效应以及ALP活性的作用。结果：FG组的PDLCs增值和ALP活性较对照组均有显著升高,且在作用的第1天就显著促进了细胞的增殖。结论：FG可促进PDLCs的增殖和ALP活性。     

Ultrastructural localization of alkaline and acid phosphatase activities in dental plaque

Ultrastructural cytohistochemical techniques showed presence of acid and alkaline phosphatases in dental plaque. Both phosphatases had intra- and extramicrobial localization. In the extracellular matrix, phosphatases were associated with small vesicles of bacterial origin, or were freely scattered in the matrix without apparent connection with microbial structures. Intracellularly, alkaline (AlkP) and acid (AcP) phosphatases were observed in Gram-negative and Gram-positive bacteria, showing a different localization. The AlkP was mainly located in the periplasmic space, while AcP had a double preferential localization: along the outer surface of the cell wall and in the periplasmic space. Less frequently an intracellular phosphatase reaction was seen in the cytoplasm.     

碱性成纤维细胞生长因子和胰岛素样生长因子-1对人牙髓细胞碱性磷酸酶活性及矿化能力的影响

目的探讨碱性成纤维细胞生长因子(bFGF)和胰岛素样生长因子(IGF)-1对人牙髓细胞碱性磷酸酶活性及矿化能力的影响。方法经10ng/mlbFGF(或)和100ng/mlIGF-1刺激4d后,在410nm波长,检测牙髓细胞的碱性磷酸酶活性值;经矿化液诱导后,采用VonKossa染色法检测不同组细胞的矿化能力。结果培养4d后,IGF-1组及bFGF加IGF-1组均高于对照组(P<0.05),而且bFGF加IGF-1组高于IGF-1组(P<0.05),而bFGF组吸光度值与对照组差异无显著性(P>0.05)。结论IGF-1可促进牙髓细胞向具有矿化能力的细胞分化,而bFGF可能起协同促进作用。     

人牙本质涎蛋白对体外培养的人牙乳头间充质细胞增殖和碱性磷酸酶活性的影响

目的 :观察人牙本质涎蛋白对体外培养的人牙乳头间充质细胞增殖和碱性磷酸酶活性的影响。方法 :分对照组和实验组 ,对照组是转染不含目的基因的空白质粒 pcDNA3的培养上清 ,实验组是转染pcDNA3 -hDSP重组质粒的培养上清。取第 5代人牙乳头间充质细胞 ,接种于 96孔板 ,对照组 6孔 ,实验组 10孔 ,用MTT法检测hDSP对体外培养的人牙乳头间充质细胞增殖的影响 ,用碱性磷酸酶检测试剂盒检测hDSP对体外培养的人牙乳头间充质细胞碱性磷酸酶活性的影响。结果 :hDSP能明显抑制体外培养的人牙乳头间充质细胞增殖 (P <0 .0 1) ,但能促进细胞内和培养上清中碱性磷酸酶的分泌 (P <0 .0 1)。结论 :hDSP能促进人牙乳头间充质细胞的分化和矿化     

白细胞介素-10对体外培养人牙囊细胞增殖和碱性磷酸酶活性的影响

目的:研究白细胞介素-10(Interleukin-10,IL-10)对体外培养人牙囊细胞增殖和牙囊细胞碱性磷酸酶活性的影响。方法:体外培养人牙囊细胞,取生长状态良好的第5代细胞,用四甲基偶氮唑蓝法(MTT法)检测IL-10对细胞增殖的作用;用碱性磷酸酶试剂盒检测IL-10及其抑制剂对细胞碱性磷酸酶活性的作用。结果:0、1、10、25、50、100ng/mLIL-10对人牙囊细胞的增殖影响无显著性差异(P>0.05);10ng/mLIL-10作用0、1、3、5、7d对人牙囊细胞的增殖影响无显著性差异(P>0.05)。10ng/mLIL-10作用3～7d降低人牙囊细胞的碱性磷酸酶活性(P<0.05),10ng/mLIL-10抑制剂作用5～7d增加人牙囊细胞的碱性磷酸酶活性(P<0.05)。结论:IL-10对人牙囊细胞的增殖无影响。IL-10降低人牙囊细胞的碱性磷酸酶活性,说明IL-10抑制人牙囊细胞向成骨方向分化。     

大鼠牙本质基质蛋白1对人牙髓细胞增殖和ALP活性的影响

目的 :探讨大鼠牙本质基质蛋白 1(DMP1)对体外培养的人牙髓细胞增殖和ALP活性的影响。方法 :利用MTT法研究牙髓细胞的细胞增殖情况 ,碱性磷酸酶检测试剂盒检测牙髓细胞内ALP活性。结果 :DMP1在细胞培养 3d和 5d时 ,5 μg/mL组可促进人牙髓细胞增殖 (P <0 .0 5 )。细胞培养 3d时 ,仅5 μg/mL组可促进人牙髓细胞ALP活性 (P <0 .0 5 ) ;培养 5d时 1μg/mL和 5 μg/mL均可促进人牙髓细胞ALP活性 (P <0 .0 5 ) ;培养 7d时促进作用进一步加强 (P <0 .0 5 )。结论 :DMP1在一定浓度下可促进牙髓细胞的增殖 ;DMP1对牙髓细胞ALP活性的促进作用呈浓度依赖性 ,高浓度DMP1可以促进人牙髓细胞ALP活性 ,随着时间的延长 ,促进ALP活性的作用也增强。     

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目的探讨种植体周围龈沟液(GCF)量及碱性磷酸酶(ALP)活性在种植体周围组织炎症发展中的变化。方法选择2006年3月至2009年3月于鞍山市铁东区口腔医院修复科接受牙种植术后12～36个月的患者30例(种植体40枚)。按照牙龈指数(GI)等级将受检种植体分为4组,测量并比较各组GCF量及ALP活性。结果受检种植体周围GCF量及ALP活性随着GI分级的增加而增加。GI1级的种植体周围GCF量和ALP活性较GI0级稍增加,差异无统计学意义(P>0.05);GI2级和3级的种植体周围GCF量和ALP活性与GI0级比较,差异均有统计学意义(P<0.01),且GI2级和3级的种植体周围GCF量、ALP活性差异亦有统计学意义(P<0.05)。结论种植体周围GCF量及ALP活性随其GI增加而增加。     

牙周优势菌内毒素对人牙周膜细胞增殖和碱性磷酸酶活性的影响

目的：研究牙周优势菌牙龈卟啉菌、中间普氏菌、具核梭杆菌的内毒素对人牙周膜细胞（PDL细胞）增殖和碱性磷酸酶活性（ALP）的影响。方法：采用MTT比色试验及酶动力学方法，测定PDL细胞的增殖和ALP活性。结果：内毒素在10μg/mL、100μg/mL高浓度时，可显著抑制PDL细胞增殖，而在0.01μg/mL、0.1μg/mL低浓度时，则促进PDL细胞增殖；在10μg/mL、100μg/mL可呈浓度依赖性方式抑制PDL细胞ALP活性。结论：内毒素对牙周膜细胞的抑制作用主要和其浓度有关，不同来源内毒素差异并不显著；内毒素可能通过影响PDL细胞功能而影响牙周组织的代谢和修复过程。     

DNA content and alkaline phosphatase expression in cells of different gingival overgrowths

The present study compared the alkaline phosphatase (ALPase) expression and DNA content at specific periods in cultured cells derived from non-inflamed enlarged gingivae of idiopathic gingivofibromatosis (IGF) and phenytoin-induced hyperplasia (PHG). Cultured cells from healthy gingiva or periodontal ligament (PDL) were used as controls. The DNA assay, ALPase assay and cytochemical staining for ALPase in cultured cells were performed at four, seven, and nine days. The presence of intense ALPase activity was a prominent feature in cultured IGF cells, whereas very low ALPase activity was detected in PHG cells. The cell lines tested showed no significant differences in DNA content. The expression of ALPase in these cells was population density-dependent. The observation that cells isolated from both types of gingival overgrowth exhibited a different ALPase profile at variance with normal gingival fibroblasts suggested that a distinct pathogenic mechanism may be involved in each type of gingival overgrowth.     

老年人牙周膜细胞碱性磷酸酶活性研究

目的 观察老年人牙周膜细胞碱性磷酸酶活性的变化。方法 利用原代培养的老年人牙周膜细胞，测定其碱性磷酸酶活性，并与青年人进行比较。结果 老年组牙周膜细胞碱性磷酸酶活性明显低于青年组（P＜0．001）；且对小牛血清刺激增殖的反应也明显低于青年组（P＜0．001）。结论 由于衰老的影响，老年人牙周膜细胞成骨活性降低，对外源性促细胞生长因素的敏感性降低，提示了老年人牙周膜细胞再生能力降低。     

维生素D3衍生物对人成骨细胞生长和碱性磷酸酶活性的影响

目的：使用人胎成骨细胞（ＯＢ）为体外模型，观察了１，２５－双羟维生素Ｄ３〔１，２５（ＯＨ）２Ｄ３〕、２４，２５－双羟维生素Ｄ３［２４，２５（ＯＨ）２Ｄ３）］对ＯＢ生长和代谢的影响。方法：用ＡＬＰ测定法和Ｂｒａｄｆｏｒｄ蛋白含量法。结果：１，２５（ＯＨ）２Ｄ３在浓度为１０８ｍｏｌ／Ｌ时，刺激碱性磷酸酶（ＡＬＰ）的活性。但抑制ＯＢ的生长。２４，２５（ＯＨ）２Ｄ３在１０－８ｍｏｌ／Ｌ时无上述作用。结论：１，２５（ＯＨ）２Ｄ３对ＯＢ的ＡＬＰ有直接的促活作用，其作用与时间相关。２４，２５（ＯＨ）２Ｄ３对ＯＢ的ＡＬＰ活性无关。１，２５（ＯＨ）２Ｄ３对人胎ＯＢ生长有抑制作用。     

Cell-bound and extracellular matrix-associated alkaline phosphatase activity in rat periodontal ligament

In previous studies it was noted that alkaline phosphatase (ALP) activity in periodontal ligament does not only seem to be related to cells but may also be associated with the extracellular matrix. In an attempt to clarify this we studied the distribution of the enzyme at the electron microscopic level. In addition, ALPactivity was assessed biochemically following extraction of the ligament with (i) agents dissolving the membrane or splitting the phosphatidylinositol anchor (Triton X-100 or phosphatidylinositol-phospholipase C, respectively), and (ii) a matrix-degrading enzyme cocktail (collagenase, hyaluronidase and elastase). Histochemical observations revealed (a) a heterogeneous distribution of ALP-activity, with highest activity adjacent to the alveolar bone and (b) two pools of activity; one bound to cells and one associated with the collagenous extracelluJar matrix. In line with this were the biochemical data indicating that approximately 10% of the enzyme activity was firmly bound to the extracellular matrix and 90% to plasma membranes. Isoelectric focusing did not reveal differences between the two fractions, both samples yielding a single broad band corresponding with an isoelectric point of about 4.4.     

Temporal and local appearance of alkaline phosphatase activity in early stages of guided bone regeneration. A descriptive histochemical study in humans

Alkaline phosphatase (ALP) catalyzes the hydrolysis of phosphate esters and it seems to be a prerequisite for normal skeletal mineralization. Also, ALP is the most widely recognized marker of osteoblast phenotypes. By a tissue regenerative technique called Guided Bone Regeneration (GBR), it is possible nowadays to regenerate small bony defects. The aim of the present study was to investigate early events in bone healing and neogenesis by studying histochemically the temporal and local appearance of the marker Alkaline Phosphatase (ALP) in a GBR model system. Nine healthy volunteers (5 males, 4 females, mean age 31.7 years) participated in the experiment. After raising a mucoperiosteal flap from the mandibular second molar to the retromolar area in each volunteer, a hollow titanium test cylinder was placed into a congruent bony bed and the coronal end of the cylinder was closed with an ePTFE-membrane. Then the flap was adapted and sutured to obtain primary wound closure. After 2, 7 and 12 weeks, the regenerated tissue within the cylinders was harvested. Histologically, ALP activity was observed associated with the osteoid seams in the very basal part of the regenerate where new bone trabeculae were in the process of being formed. More coronally, large round cells seemed to secrete an ALP-positive substance since in the center of such cell clusters strong ALP activity located extracellularly was detected. In the present experiment, ALP seemed to have been an early sign of osteoblast secretion of a matrix which subsequently was determined to become osteoid. ALP activity was never seen isolated within connective tissue and away from bone. This is an indication that its source is linked to existing bone. The present study has documented for the first time the appearance of ALP activity in guided bone regenerations in humans. It has revealed that: 1) Osteogenesis in guided bone regeneration is preceded by localized, marked expression of ALP in an organized connective tissue environment. 2) Bone neogenesis is an early event in this experimental setup and may be detected already 2 weeks after wounding. 3) Expression of ALP and subsequent bone neogenesis is originating from and topographically linked to pre-existing bone structures.     

Activity of alkaline and acid phosphatases in dental epithelial cells and enameloid during odontogenesis in two teleost fish,Oreochromis niloticus and Tilapia buttikoferi

The dental epithelial cells and enameloid at the stages of enameloid matrix formation, mineralization and maturation in the teleosts Oreochromis niloticus and Tilapia buttikoferi were investigated by means of enzyme histochemistry in order to identify their functions associated with the structural modification. No marked enzyme activities were found in the inner dental epithelial cells in the stage of enameloid matrix formation, although the outer dental epithelial cells often exhibited moderate alkaline phosphatase (ALPase) activity. In the stages of enameloid mineralization and maturation, the inner dental epithelial cells, which possessed a ruffled border at the distal ends, showed intense ALPase activity at their lateral and proximal cell membranes. At the same time, many acid phosphatase (ACPase)-positive vesicles and granules were localized at the distal cytoplasm of the inner dental epithelial cells. The outer dental epithelial cells, which contained well-developed labyrinthine canalicular spaces, showed neither marked ALPase nor ACPase activity. It is postulated that the dental epithelial cells in these two teleosts are mainly involved in the removal of the organic matrix from the enameloid, and in material transport to the enameloid during the later half of odontogenesis.     

胰岛素对人牙周膜细胞碱性磷酸酶活性和总蛋白含量的影响

目的：探讨胰岛素作用下，牙周膜（ＰＤＬ）细胞碱性磷酸酶（ＡＬＰ）活性和总蛋白含量的变化。方法：细胞生物学法、酶动力学法和考马斯亮蓝法。结果：胰岛素浓度超过１０－４Ｕ／ｍｌ时，明显增加ＰＤＬ细胞的ＡＬＰ活性和总蛋白含量（Ｐ＜０．０１）。其中，在１０－４～１０－１Ｕ／ｍｌ之间，效应随浓度增加而增加，超过１０－１Ｕ／ｍｌ效应趋于稳定，此时效应最大，ＡＬＰ活性和总蛋白含量分别比对照组增加１．４倍和１．２倍。结论：胰岛素明显促进ＰＤＬ细胞分化及蛋白合成功能。胰岛素作为骨代谢的重要调节因子，可能对牙周组织的再生修复有调节作用。     

碱性磷酸酶在第三期牙本质形成过程中的分布及意义

目的：观察碱性磷酸酶在第三期牙本质形成过程中的分布,探讨其在牙髓损伤修复过程中的作用及意义。方法：建立大鼠第三期牙本质形成动物模型,用免疫组织化学染色方法研究大鼠第三期牙本质形成过程中碱性磷酸酶的表达变化。结果：碱性磷酸酶于牙髓损伤早期阶段表达于穿髓点下方细胞群;随着修复性牙本质的形成其表达逐渐减弱直至消失。结论：碱性磷酸酶在牙髓损伤的早期阶段发挥着重要作用。     

Heat stress induces alkaline phosphatase activity and heat shock protein 25 expression in cultured pulp cells

AIM: To investigate the responses of cultured rat pulp cells to heat stress. METHODOLOGY: Pulp cells were obtained from rat incisors and cultured at 37 degrees C. The cells were cultured at 42 degrees C for 30 min and then cultured at 37 degrees C again. Morphology, alkaline phosphatase (ALP) activity and expression of heat shock protein 25 (HSP25) were investigated at 0, 1, 3, 5, 7, 10 and 14 days following stimulation. As a control, the cells were maintained at 37 degrees C. RESULTS: Although there were few cells of apoptosis immediately after heat stress, there were mitotic cells from day 1 after heat stress. ALP activity in the heat stress group significantly increased at days 7 and 14 compared with the control group (about 1.7-fold, P < 0.01, Friedman test). HSP25 expression increased in both groups, with HSP25 in the heat stress group being expressed earlier than in the control group, and nuclear localization of HSP25 was observed at days 0 and 1 in heat-stressed cells. CONCLUSION: These results suggest that heat stress not only induces HSP25 but also enhances ALP activity in pulp cells.     

Effective method for discriminating between oral bacterial and human alkaline phosphatase activity

Alkaline phosphatase (ALPase) activity was quantitatively compared in various kinds of oral bacteria. High ALPase activity was detected in 3 species of periodontal bacteria, Porphyromonas gingivalis. Prevotella intermedia and Capno-cvtophaga sputigena. The ALPase activity detected in these bacteria was almost completely inhibited in the presence of 1% sodium dodecyl sulfate (SDS). By contrast, the activity of mammalian ALPase isoenzymes was not inhibited at all even in the presence of 1% SDS. These results indicate that the ALPase assay in combination with 1% SDS can identify the origin of ALPase detected in gingival crevicular fluid as being from bacteria or from a host response. Clinical examination with adult periodontitis revealed that ALPase activity in gingival crevicular fluid from the patients consisted of a combination of SDS-sensitive and SDS-resistant activities. These findings indicate that ALPase activity detected in gingival crevicular fluid originates not only from bacteria but also from a host response.