The effect of mitomycin-c in keloid fibroblast cultures

Abstract

Keloid occurs due to hyperactivity of keloid fibroblast (KF) in proliferation, migration, collagen deposition, together with low rates of collagen degradation. These are under the responsibility of TGF-b. Mitomycin C (MC) is used for treating keloid by a topical application during surgery at the level of 0.02% to 0.08%. Unfortunately, the lowest effective level of MC for keloid has not been determined yet. We aimed to determine the lowest effective level of MC in the suppression of KF activities. Various levels of MC diluted in growth medium were administered on KF that were isolated from six patients. After 24 hours and 72 hours of incubation, cellular proliferation, collagen deposition, cellular migration and level of TGF-b, were analyzed. Application of 120 uM MC on KF culture for 24 hours could significantly reduce TGF-b production from 1265.74 ± 274.81 pg/mL to 265.17 ± 12.20 pg/mL; proliferation index from 100% to 84.01 ± 12.91%; inhibit cellular migration to 64.38 ± 3.66%; but reduce collagen depositions from 100% to only 91.13 ± 10.19%. The lowest MC level is on 30 uM or equal with 0.001%. In conclusion, the lowest level of MC can suppress the activities of KF is 0.001%. Moreover, due to low activity in inhibiting collagen deposition, MC would be better as an adjuvant drug for keloid surgery. ABSTRAKKeloid timbul karena proliferasi, migrasi, dan sintesis kolagen oleh fibroblas keloid (FK) secara berlebihan diikuti dengan rendahnya degradasi kolagen. Semua itu terjadi karena pacuan TGF-b. Olesan mitomycin C (MC) 0,02% sampai 0,08% digunakan untuk mengobati dan mencegah kekambuhan keloid yang dioperasi. Sayangnya, dosis terendah yang masih efektif belum pernah diteliti. Penelitian ini bertujuan untuk menentukan kadar terendah MC yang masih efektif untuk mengobati keloid. MC dilarutkan dalam medium pertumbuhan dalam berbagai kadar dan diberikan pada biakan FK yang diisolasi dari material keloid enam orang pasien. Setelah inkubasi 24 jam dan 72 jam, proliferasi, migrasi, timbunan kolagen dan kadar TGF-b, dianalisis dan dibandingkan. Pemberian 120 uM MC pada biakan FK ternyata dapat menurunkan produksi TGF-b dari 1265,74 ± J Med Sci, Volume 48, No. 3, 2016 168 274,81 pg/mL menjadi 265.17 ± 12.20 pg/mL; indek proliferasi dari 100% menjadi 84,01 ± 12.91%; menghambat migrasi sampai 64,38 ± 3.66%, tetapi daya hambat timbunan kolagen hanya dari 100% menjadi 91.13 ± 10.19%. Kadar MC terendah yang masih efektif adalah 30 uM atau setara dengan 0.001%. Kadar MC di bawah itu tidak lagi efektif. Kesimpulan, kadar MC terendah yang masih efektif sebesar 0,001% dan karena daya hambat timbunan kolagen yang rendah, MC sebaiknya digabung dengan operasi keloid.

The effect of 5α-oleandrin on keloid fibroblast activities

“…For example, Richard et al [20] indicated that mitomycin-C could inhibit KF proliferation by suppressing DNA synthesis. Dachlan et al [17] showed that 30 uM mitomycin-C could suppress keloid fibroblast proliferation and collagen synthesis. Both studies are based on Stewart et al [8] who used topical application of mitomycin-C for clinical wound repair trials to prevent keloid.…”

“…Both materials were diluted in DMEM containing 10% fetal bovine serum plus 1% penicillin/streptomycin in various concentrations. We used the lowest effective dose of mitomycin-C (30 μM) as previously reported by Dachlan et al [17] as the positive control. To determine the effective dose of 5α-oleandrin, we diluted 5α-oleandrin into various concentrations, and the highest dose was considered to be half of the dose of mitomycin-C.…”