THE EFFECTS OF RETINOIC ACID ON ALKALINE-PHOSPHATASE ACTIVITY AND TISSUE-NON-SPECIFIC ALKALINE-PHOSPHATASE GENE-EXPRESSION IN HUMAN PERIODONTAL-LIGAMENT CELLS AND GINGIVAL FIBROBLASTS

Alkaline phosphatase (ALP) in human periodontal ligament (HPDL) cells is classified as a tissue-non-specific alkaline phosphatase (TNSALP) b y its enzymatic and immunological properties. Since retinoic acid (RA) has been shown as a potent inducer of TNSALP expression in various os teoblastic and fibroblastic cells, we investigated the effects of RA o n the level of ALP activity and expression of TNSALP mRNAs in HPDL cel ls. Cultured cells were treated with desired RA concentrations (0, 10( -7), 10(-6), 10(-5) M) in medium containing 1% bovine serum albumin wi thout serum. ALP activity was determined by the rate of hydrolysis of p-nitrophenyl phosphate and was also assayed in the presence of specif ic inhibitors. In order to identify the TNSALP mRNA type expressed by HPDL, a set of oligonucleotide primers corresponding to 2 types of hum an TNSALP mRNA (i.e. bone-type and liver-type) were designed, and mRNA isolated from HPDL was amplified by means of reverse transcription-po lymerase chain reaction (RT-PCR). After treatment with RA (10(-6) M) f or 4 d, there was a significant increase in the ALP activity of HPDL c ells. The use of inhibitors and thermal inactivation experiments showe d that the increased ALP activity had properties of the TNSALP type. R T-PCR analysis revealed that bone-type mRNA was highly stimulated in H PDL cells by RA treatment, but the expression of liver-type mRNA was n ot detected. These results indicated that the upregulation of ALP acti vity in HPDL cells by RA was due to the increased transcription of bon e-type mRNA of the TNSALP gene.