Exogenous sphingomyelinase increases collagen and sulphated glycosaminoglycan production by primary articular chondrocytes: an in vitro study.
Exogenous sphingomyelinase increases collagen and sulphated glycosaminoglycan production by primary articular chondrocytes: an in vitro study.
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DOI：
10.1186/ar1961
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发表时间：
2006
影响因子：
4.9
通讯作者：
Mason DJ
中科院分区：
文献类型：
--
作者：
Gilbert SJ;Blain EJ;Jones P;Duance VC;Mason DJ
我们之前确定了第二信使神经酰胺在蛋白激酶 R (PKR) 介导的关节软骨降解中的作用。已知神经酰胺在胶原蛋白基因调节中发挥双重作用，神经酰胺对胶原蛋白启动子活性的影响取决于其浓度。用低剂量的鞘磷脂酶处理细胞会导致内源性神经酰胺的少量增加。我们研究了神经酰胺是否影响关节软骨细胞基质稳态，如果影响，则研究 PKR 在此过程中的作用。用鞘磷脂酶刺激牛关节软骨细胞 7 天，以增加神经酰胺的内源水平。为了抑制 PKR，将 2-氨基嘌呤添加到重复培养物中。通过分别向培养基中添加[35S]-硫酸盐和[3H]-脯氨酸来测量从头硫酸化糖胺聚糖和胶原合成。使用 RT-PCR 和蛋白质印迹分析研究软骨细胞表型。 7 天以上，鞘磷脂酶增加了新合成的硫酸化糖胺聚糖和胶原蛋白向培养基的释放，而在鞘磷脂酶处理的细胞中抑制 PKR 降低了新合成的硫酸化糖胺聚糖和胶原蛋白的水平。鞘磷脂酶处理的软骨细胞表达 col2a1 mRNA，这表明软骨细胞表型正常；然而，检测到 II 型胶原蛋白显着减少。因此，软骨细胞中内源性神经酰胺的小幅增加似乎将稳态平衡推向细胞外基质合成，但以牺牲软骨细胞表型为代价，而软骨细胞表型部分是由 PKR 介导的。
We previously established a role for the second messenger ceramide in protein kinase R (PKR)-mediated articular cartilage degradation. Ceramide is known to play a dual role in collagen gene regulation, with the effect of ceramide on collagen promoter activity being dependent on its concentration. Treatment of cells with low doses of sphingomyelinase produces small increases in endogenous ceramide. We investigated whether ceramide influences articular chondrocyte matrix homeostasis and, if so, the role of PKR in this process. Bovine articular chondrocytes were stimulated for 7 days with sphingomyelinase to increase endogenous levels of ceramide. To inhibit PKR, 2-aminopurine was added to duplicate cultures. De novo sulphated glycosaminoglycan and collagen synthesis were measured by adding [35S]-sulphate and [3H]-proline to the media, respectively. Chondrocyte phenotype was investigated using RT-PCR and Western blot analysis. Over 7 days, sphingomyelinase increased the release of newly synthesized sulphated glycosaminoglycan and collagen into the media, whereas inhibition of PKR in sphingomyelinase-treated cells reduced the level of newly synthesized sulphated glycosaminoglycan and collagen. Sphingomyelinase treated chondrocytes expressed col2a1 mRNA, which is indicative of a normal chondrocyte phenotype; however, a significant reduction in type II collagen protein was detected. Therefore, small increments in endogenous ceramide in chondrocytes appear to push the homeostatic balance toward extracellular matrix synthesis but at the expense of the chondrocytic phenotype, which was, in part, mediated by PKR.
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影响因子：
4.1
作者：
Little, CB;Flannery, CR;Caterson, B
通讯作者：
Caterson, B
影响因子：
3.2
作者：
Farina, F;Cappello, F;Stassi, G
通讯作者：
Stassi, G
影响因子：
4.9
作者：
Gilbert SJ;Duance VC;Mason DJ
通讯作者：
Mason DJ
影响因子：
4.8
作者：
Patel, CV;Handy, I;Patel, RC
通讯作者：
Patel, RC
影响因子：
3.5
作者：
Dbaibo, GS;El-Assaad, W;Hannun, YA
通讯作者：
Hannun, YA