Pengaruh Tetrahydrozoline HCl terhadap viabilitas sel fibroblast Gingiva secara in vitro :: Uji Sitotoksisitas

DEWI R., Tri Purnami, Dr.drg. Widowati Siswomihardjo, MS

2004 | Tesis | S2 Ilmu Kedokteran Gigi

Bahan kimia yang digunakan untuk retraksi gingiva pada prosedur pembuatan gigi tiruan cekat, harus efektif menghasilkan retraksi sementara jaringan gingiva bebas ke arah lateral dan apikal, tanpa menyebabkan kerusakan jaringan yang ireversibel. Bahan retraksi juga harus bersifat aman, dan seminimal mungkin menimbulkan efek samping yang merugikan, baik lokal maupun sistemik. Penelitian ini bertujuan untuk mengetahui pengaruh konsentrasi dan waktu paparan tetrahydrozoline HCl terhadap viabilitas sel fibroblast gingiva secara in vitro (efek sitotoksik) dalam aplikasinya sebagai bahan retraksi gingiva. Penelitian ini mempergunakan 192 sumur kultur sel fibroblast gingiva manusia. Setiap sumur berisi 100μL suspensi sel fibroblast gingiva dengan kepadatan 2x104 sel/100 μL media kultur sel. Seratus sembilan puluh dua sampel tersebut dibagi menjadi 3 kelompok, terdiri dari kelompok kontrol positif diberi epinefrin 0,1%, kelompok kontrol negatif tanpa perlakuan dan kelompok perlakuan diberi tetrahydrozoline HCl dengan konsentrasi dan waktu paparan yang berbeda. Kelompok kontrol positif dan kelompok kontrol negatif, masing-masing dibagi lagi menjadi 8 sub kelompok sesuai dengan waktu paparan, yaitu 1, 3, 5, 7, dan 10 menit serta 24, 48, dan 72 jam. Setiap sub kelompok terdiri dari 3 sumur sampel. Kelompok perlakuan dibagi menjadi 8 sub kelompok sesuai waktu paparan, kemudian tiap sub kelompok dibagi lagi menjadi 6 sub sub-kelompok sesuai dengan konsentrasi paparan yang digunakan dalam penelitian ini, yaitu 0,05%, 0,1%; 1%;10%; 20%; dan 30%. Setiap sub sub-kelompok terdiri dari 3 sumur sampel. Perhitungan persentase kematian kelompok perlakuan dilakukan berdasarkan hasil OD dan penghitungan jumlah sel yang viabel. Pada kelompok perlakuan, rerata persentase kematian sel terendah, yaitu 0% dihasilkan pada konsentrasi tetrahydrozoline HCl 0,05%; 0,1% dan 1% dengan waktu paparan 1 dan 3 menit, dan tertinggi yaitu 88,5% dihasilkan pada konsentrasi tetrahydrozoline HCl 30% dengan waktu paparan 72 jam. Probabilitas koefisien regresi 0,001 menunjukkan bahwa konsentrasi dan waktu paparan tetrahydrozoline HCl berpengaruh secara signifikan terhadap viabilitas sel fibroblast gingiva. Hasil uji Univariate ANOVA menunjukkan adanya perbedaan viabilitas sel yang bermakna antar kelompok (p<0,05). Hasil uji LSD menunjukkan perbedaan viabilitas sel yang bermakna antara beberapa sub-kelompok. Analisis probit menghasilkan estimasi nilai efektif median, seperti konsentrasi IC50 tetrahydrozoline HCl untuk waktu paparan 5 menit (1630,5x103μM), 7 menit (1464,7x103μM), 10 menit (1384,9x103μM), 24 jam (507,8 x103μM), 48 jam (502,7x103μM) dan 72 jam (40,25x103μM). Penggunaan tetrahydrozoline HCl dengan waktu paparan dan konsentrasi terapeutik tidak menimbulkan efek sitotoksik terhadap viabilitas sel fibroblast gingiva, sebaliknya epinefrin memiliki potensi sitotoksik.

The use of chemical agent as gingival retraction in the procedure of making fixed prosthodontic impression should be effective to produce temporary lateral and apical retraction of the free gingival tissue, without creating irreversible tissue damage. The retraction agent must be save and as minimum as possible creating either local or systemic harmful side effects. The aim of this study was to investigate the effect of tetrahydrozoline HCl towards the viability of human gingival fibroblast cells at different concentrations and exposure times. This study utilized one hundred and ninety two wells cultured human gingival fibroblast cell. Each well contained 100μL gingival fibroblast cell suspension with the density of 2x104 cells/100 μL cultured cell medium. All wells were divided into 3 sample groups, the first group as positive control, was treated with 0.1% epinephrine, the second group without treatment as negative control, and the last one was treated with tetrahydrozoline HCl. Both positive and negative control groups, were divided into 8 sub groups, in accordance to the exposure time: 1, 3, 5, 7, and 10 minutes, then 24, 48 and 72 hours. Each sub group consisted of 3 wells. The treated group were divided into 8 sub groups in accordance to the exposure time, and each sub group were divided into 6 sub sub-groups, in accordance with the concentrations of tetrahydrozoline HCl that was used in this study, such as 0.05%, 0.1%, 1%, 10%, 20% and 30%. Each sub sub-group consisted of 3 wells. The percentage calculation of the viability was determined by the result of OD and the calculation of viable cells number. In the treated groups, the lowest mean of cell death percentage was 0%, were resulted at 0.05%, 0.1%, 1% concentration of tetrahydrozoline HCl with exposure time: 1 and 3 min, and the highest mean was 88.5%, were resulted at 30% concentration of tetrahydrozoline HCl with exposure time: 72 h. The probability of regression coefficient 0.001 showed that the influence of concentration and exposure time of tetrahydrozoline HCl to the viability of human gingival fibroblast cell was statistically significant. The result of Univariate ANOVA indicated that there was a significant difference in cell viability between groups. The result of LSD test indicated that there was a significant difference in cell viability within group (between sub-groups). Probit analysis reported estimation of median effective value, such as IC50 concentration, at different exposure times of tetrahydrozoline HCl: 5 min (1630.5x103μM), 7 min (1464.7x103μM), 10 min (1384.9x103μM), 24h (507.8x103 μM), 48h (502.7x103μM) and 72h (40.25x103μM). The use of tetrahydrozoline HCl had no cytotoxic effect to the viability of human gingival fibroblast cell at the therapeutic concentration and exposure time; on the contrary epinephrine had cytotoxic potency.

Kata Kunci : Tetrahydrozoline HCl, retraksi gingiva, viabilitas, fibroblast gingiva, gingival retraction, viability, gingival fibroblast