| Objective:Condylar cartilage cells cultured in vitro single were added into all-trans retinoic acid, and alkaline phosphatase expression and NO and two types of NOS expression level was detected at different times, to explore the role of NOS in the cell differentiation ;non-selecti-ve nitric oxide synthase inhibitor-LNMA was added into the all-trans retinoic acid induced condylar cartilage cells, changes in the level of alkaline phosphatase expression were detected, to explore the role of different NOS in it. Then, extracellular calcium channel blockers EGTA was added into the all-trans retinoic acid induced condylar cartilage cells, changes in the level of alkaline phosphatase expression were detected, to explore the role of different types of NOS in it.Materials and Methods:1．Neonatal rat condylar cartilage cells was selected as a source of the organization, to estabish limited cell lines. 10-7 mol/L all-trans retinoic acid were added into the second generation of condylar cartilage cell, activity of alkaline phosphatase, two type of nitric oxide and nitric oxide oxide synthase were measured spectively at 48,72,96,120 hours with spectrophotometer. other culture condition and detection method of control group were the same as the experimental group. 2．Cells treated by 10-7 mol/L all-trans retinoic acid for 72 hours were added into 0．25-1mmol/L non-selective nitric oxide synthase inhibitor-LNMA, and activity of alkaline phosphatase were measured again after 24 hours. the other culture condition and detection method of control group were the same as the experimental group 3．Cells treated by 10-7 mol/L all-trans retinoic acid for 72 hours were added into 2mmol/L extracellular calcium channel blockers-EGTA, and activity of alkaline phosphatase were measured again after 8,12,24 hours, the other culture condition and detection method of control group were the same as the experimental group.Results:1．Activity of ALP and two type of NOS and NO of cells increased after treated by 10-7 mol/L concentration of for 48 to 96 hours. There was Statistically significant difference between control group and experimental group. The expression level of iNOS was far higher than cNOS. After 120 hours, expression level reduced compared with 96 hours, there was still Statistically significant difference between control group and experimental group. The result Suggested that NOS may play an active role condylar cartilage in the cell differentiation 2．After adding into the non-selective NOS inhibitor L-NMA, activity of ALP gradually reduced to base level with the increasing concentration of L-NMA. This suggested that NOS may play an important role in cell differentiation. 3．After adding into extracellular calcium channel blockers EGTA for 8,12 hours,24 hour, expression levels of ALP has been reduced compared with RA group, but was still significant higher than that of control group. the result suggested both cNOS and iNOS may play a role in the regulation of cell differentiation. The role of iNOS may be even more pronounced. |
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