人糖尿病牙髓组织中的褪黑素水平及其在高血糖条件下对牙髓细胞的影响。
Melatonin levels in human diabetic dental pulp tissue and its effects on dental pulp cells under hyperglycaemic conditions.
机构信息
Department of Pharmacology in Dentistry, School of Dental Medicine, University of Belgrade, Belgrade, Serbia.
Department of Physiology and Biochemistry, School of Dental Medicine, University of Belgrade, Belgrade, Serbia.
出版信息
Int Endod J. 2018 Oct;51(10):1149-1158. doi: 10.1111/iej.12934. Epub 2018 Apr 21.
AIM
To investigate melatonin (MEL) levels in human dental pulp tissue (hDP) in type 2 diabetic (T2D) participants and the underlying molecular mechanisms of its effects in human dental pulp cells (hDPCs) under hyperglycaemia.
METHODOLOGY
The study included 16 healthy and 16 T2D participants who underwent vital pulp extirpation for hDP and four healthy participants undergoing third molar extraction for hDPCs analyses. MTT and NRU were used as tests for cytotoxicity. The pulp tissue levels of MEL, inducible NO synthase (iNOS) and superoxide dismutase (SOD) activity, as well as iNOS, histone acetyltransferase p300 (p300) and SOD activity levels in hDPCs incubated with MEL (0.1 and 1.0 mmol L ) under normoglycaemia and hyperglycaemia were measured by enzyme-linked immunosorbent assay. Comparisons between the two groups were made by unpaired t-tests or Mann-Whitney test whilst the chi-square test was used for dichotomous variables. To compare more groups, the Kruskal-Wallis test with Dunn's multiple comparison was used, whilst Spearman correlation was used to assess association between two variables.
RESULTS
Melatonin was decreased (124.30 ± 21.6 vs. 240.0 ± 19.1 pg mL , P < 0.01), whilst iNOS levels increased (0.92 ± 0.08 vs. 0.32 ± 0.09 ng mL , P < 0.01) in hDP from T2D compared to nondiabetic participants. In hDPCs, MEL (0.1 and 1.0 mmol L ) had no cytotoxicity. Incubation with 1.0 mmol L of MEL (24 h) decreased hyperglycaemia-induced increases of iNOS (0.34 ± 0.01 ng mL vs. 0.40 ± 0.01 ng mL , P < 0.01) and p300 (11.59 ± 0.58 ng mL vs. 16.12 ± 0.39 ng mL , P < 0.01), and also, increased SOD activity (87.11 ± 3.10% vs. 68.56 ± 3.77%, P < 0.01) to the levels comparable to the normoglycaemic; iNOS and p300 protein expression levels showed strong positive correlation under hyperglycaemia (Spearman r = 0.8242, P < 0.001).
CONCLUSION
Type 2 diabetic participants had decreased MEL in hDP. At pharmacological concentrations, MEL is not cytotoxic for hDPCs and normalizes iNOS and SOD activity levels in hyperglyceamic hDPCs suggesting its antioxidant and protective effects in human dental pulp tissue under hyperglycaemia.
目的
研究 2 型糖尿病（T2D）患者牙髓组织（hDP）中褪黑素（MEL）的水平，以及 MEL 在高血糖条件下对人牙髓细胞（hDPC）的作用的潜在分子机制。
方法
本研究纳入了 16 名健康和 16 名 T2D 参与者，他们因 hDP 进行活髓切除术，另有 4 名健康参与者因第三磨牙提取进行 hDPC 分析。MTT 和 NRU 被用作细胞毒性测试。通过酶联免疫吸附测定法测量 MEL（0.1 和 1.0 mmol/L）在正常血糖和高血糖条件下孵育的 hDPC 中 MEL 水平、诱导型一氧化氮合酶（iNOS）和超氧化物歧化酶（SOD）活性、iNOS、组蛋白乙酰转移酶 p300（p300）和 SOD 活性水平。两组间的比较采用配对 t 检验或曼-惠特尼检验，而二项变量采用卡方检验。为了比较更多的组，采用 Kruskal-Wallis 检验加 Dunn 多重比较，而采用斯皮尔曼相关评估两个变量之间的关联。
结果
与非糖尿病参与者相比，T2D 患者的 hDP 中 MEL 减少（124.30±21.6 与 240.0±19.1 pg/mL，P<0.01），而 iNOS 水平升高（0.92±0.08 与 0.32±0.09 ng/mL，P<0.01）。在 hDPC 中，MEL（0.1 和 1.0 mmol/L）没有细胞毒性。孵育 1.0 mmol/L 的 MEL（24 h）降低了高血糖诱导的 iNOS（0.34±0.01 ng/mL 与 0.40±0.01 ng/mL，P<0.01）和 p300（11.59±0.58 ng/mL 与 16.12±0.39 ng/mL，P<0.01）的增加，同时增加 SOD 活性（87.11±3.10% 与 68.56±3.77%，P<0.01）到与正常血糖相当的水平；iNOS 和 p300 蛋白表达水平在高血糖条件下呈强烈正相关（Spearman r=0.8242，P<0.001）。
结论
2 型糖尿病患者的 hDP 中 MEL 减少。在药理浓度下，MEL 对 hDPC 无细胞毒性，并使高血糖 hDPC 中的 iNOS 和 SOD 活性水平正常化，提示其在高血糖下人牙髓组织中的抗氧化和保护作用。
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