INCREASED LYSYL OXIDASE ACTIVITY IN FIBROBLASTS CULTURED FROM ORAL SUBMUCOUS FIBROSIS ASSOCIATED WITH BETEL NUT CHEWING IN TAIWAN

Growth characteristics and lysyl oxidase activity of fibroblasts deriv ed from human normal mucosa (NM) and oral submucous fibrosis (OSF) ass ociated with betel nut chewing were compared in cell cultures. The gro wth rates of cultured cells were identified by plating 5X10(5) cells/3 5 mm culture dish (Day 0) and every 24 hours cell proliferation was de termined by quantifying the cell number (using a hemocytometer). The t hird to seventh passages were used. A medium without serum but supplem ented with 5 mg/ml bovine serum albumin was substituted for the origin al medium at the subconfluent period and cultured for an additional 24 h. The medium was collected and used for assays of protein content an d lysyl oxidase activity. Lysyl oxidase activity was assayed with [4,5 -H-3] - lysine labelled purified chick - embryo aorta elastin substrat e. After incubation for 10 h at 37 degrees C, the enzyme activity was measured from 3HHO (tritiated water) separated by ultrafiltration usin g Amicon C-10 micro-concentrators. The results showed the mean doublin g time of OSF fibroblasts was 3.2 days and of NM fibroblasts was 3.6 d ays. NM fibroblasts became confluent at day 6 as determined by cell nu mber, while OSF fibroblasts were confluent by Day 5. Furthermore, the immunoenzymatic assay for BrdUrd incorporation revealed that OSF fibro blasts proliferate significantly faster than NM fibroblasts under stan dard culture conditions. Both total protein content (10.84+/-1.15 mg/m l) and lysyl oxidase activity (3558.6+/-345.5 cpm/10(6) cell) in OSF f ibroblasts were greater than in NM fibroblasts (6.35+/-0.96 mg/ml and 2436.0+/-352.6 cpm/10(6) cell). The results of this study provide evid ence that fibroblasts derived from oral submucous fibrosis (OSF) tissu e and normal mucosa (NM), although similar in many respects, exhibit s pecific differences in proliferation rates and lysyl oxidase activity. Moreover, collagen deposition in OSF tissue may, at least in part, be ascribed to increased lysyl oxidase activity.