Departamento de Bioquimica da Universidade Federal de São Paulo, Disciplina de Biologia Molecular, Rua 3 de Maio 100, 04044-020, São Paulo, Brazil; Universidade Guarulhos, Praça Tereza Cristina, 229, Guarulhos, SP, 07023-070, Brazil.
Departamento de Bioquimica da Universidade Federal de São Paulo, Disciplina de Biologia Molecular, Rua 3 de Maio 100, 04044-020, São Paulo, Brazil.
Arch Oral Biol. 2018 Jun;90:67-73. doi: 10.1016/j.archoralbio.2018.03.003. Epub 2018 Mar 10.
Proteoglycans are glycosylated proteins which have covalently attached highly anionic glycosaminoglycans. They can be located on the extracellular matrix, cell membrane or intracellular granules. To date, few studies have reported the presence of proteoglycans in human dental pulp.
The aim of this study was, therefore, to analyze the expression of lumican, versican and glypican proteoglycans in deciduous and permanent human dental pulp by real-time polymerase chain reaction (q-PCR) and immunofluorescence.
Healthy human dental pulps were used: 13 from permanent teeth (group 1) and eight from deciduous teeth (group 2). Versican, lumican and glypican (glypican-1 to 6) gene expressions were quantitatively evaluated by real-time PCR technique, using the expression of the endogenous gene GAPDH as control. Pulp sections were submitted to immunostaining procedure with fluorescence labelling, the tissues being fixed and incubated with well-characterized monoclonal and polyclonal antibodies against proteoglycan epitopes, including anti-versican and anti-lumican. Comparisons among the groups of the quantitative scores for each proteoglycan were analyzed using the t-test and ANOVA (P < 0.05).
The real-time PCR analysis showed expression of versican and lumican proteoglycans in the two groups, with significant predominance of lumican gene (P = 0.03). Considering the glypican genes, glypican-3 was the proteoglycan most significantly expressed in permanent pulps (P < 0.001), while glypican-2 was not expressed in this tissue. The immunofluorescence quantification exhibited no significant differences between lumican and versican among the pulps and groups.
The lumican gene was more expressed than versican and glypican-3 was the isoform more expressed in permanent pulp compared to deciduous.
蛋白聚糖是一种糖基化蛋白，其带有共价连接的高度阴离子性糖胺聚糖。它们可以位于细胞外基质、细胞膜或细胞内颗粒上。迄今为止，很少有研究报道蛋白聚糖存在于人类牙髓中。
因此，本研究旨在通过实时聚合酶链反应（q-PCR）和免疫荧光法分析乳牙和恒牙人牙髓中核心蛋白聚糖、 versican 和 glypican 蛋白聚糖的表达。
使用健康的人牙髓：来自恒牙的 13 个（第 1 组）和来自乳牙的 8 个（第 2 组）。通过实时 PCR 技术定量评估 versican、lumican 和 glypican（glypican-1 至 6）基因的表达，以 GAPDH 作为内参基因。牙髓切片进行荧光标记免疫染色，组织固定后用针对蛋白聚糖表位的特异性单克隆和多克隆抗体（包括抗 versican 和抗 lumican）孵育。使用 t 检验和 ANOVA 分析各组中每种蛋白聚糖的定量评分之间的差异（P<0.05）。
实时 PCR 分析显示两组均表达 versican 和 lumican 蛋白聚糖，lumican 基因表达显著占优势（P=0.03）。考虑到 glypican 基因，glypican-3 是恒牙牙髓中表达最显著的蛋白聚糖（P<0.001），而 glypican-2 则不在该组织中表达。免疫荧光定量分析显示，lumican 和 versican 在牙髓和组间无显著差异。
与乳牙相比，lumican 基因在恒牙中的表达高于 versican，glypican-3 是在恒牙中表达更显著的同工型。
