The role of ranibizumab as an adjunctive agent in mitomycin-C-treated glaucoma tube implantation on conjunctival fibroblast tissue culture

Yusof, Siti Fairuz Mohd (2018) The role of ranibizumab as an adjunctive agent in mitomycin-C-treated glaucoma tube implantation on conjunctival fibroblast tissue culture. Masters thesis, Universiti Sains Malaysia.

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Elevated expression of vascular endothelial growth factor (VEGF) usually occurs after the glaucoma surgery. The elevated expression of VEGF is unnecessary as it contributes to excessive scarring which will fail the surgery. Therefore, this study investigated the potential role of ranibizumab, an anti – vascular endothelial growth factor (anti – VEGF), as anti – adjunctive agent in preventing the excessive fibrosis. The aim of this study is to determine the effect of adjunctive use of ranibizumab with mitomycin – C on proliferation, cytotoxicity and migration of human conjunctival fibroblasts (HConFs) cell line. In proliferation and cytotoxicity study, HConFs were cultured in fibroblast medium and treated with 0.40 mg/ml mitomycin – C (MMC). The cultures were then administered with ranibizumab at different concentration (0 mg/ml, 0.30 mg/ml, 0.45 mg/ml and 0.60mg/ml). Viability and proliferation of HConFs were assessed at 24, 48 and 72 hours by alamarBlue assay. For cytotoxicity, the apoptosis rate of treated HConFs was evaluated via flow cytometry using Annexin-V FITC and propidium iodide staining after 72 hours treatment. Meanwhile, for morphology and migration study, in vitro model of glaucoma tube implantation was used. HConFs were treated according to 4 groups; control, MMC 0.40 mg/ml, ranibizumab 0.45 mg/ml and MMC 0.40 mg/ml +ranibizumab 0.45 mg/ml. Migration of HConFs onto the tube was observed at day 7,14 and 21 through alamarBlue assay and scanning electron microscope (SEM). Application of ranibizumab following MMC caused a dose – dependent inhibition of HConF’s viability and proliferation which significant after 48 and 72 hours. It was shown that HConF’s proliferation was significantly reduced by ranibizumab at concentration 0.45 mg/ml, (p<0.05). In fact, surprising higher number of HConF was noted in 0.60 mg/ml ranibizumab group in contrast to other treatment groups. Concurrently, treatment of ranibizumab also found to induce apoptosis and reduced necrosis among HConFs. The highest level of apoptotic HConFs and the least necrotic HConFs were observed at concentration 0.45 mg/ml. While in tube migration study, 7 days treatment resulted in lower percentages of HConFs migrated on to the tube observed in MMC and MMC+ranibizumab treated group compared to control, (p<0.05). Treatment of MMC showed lower percentage of migrated fibroblasts than MMC+ranibizumab group, (p=0.792). In contrast, higher percentage of migrated fibroblasts was observed in ranibizumab group with respect to control, (p=0.985). Whereas, at day 14 and 21, percentage of migrated fibroblasts is almost the same in all groups, (p<0.05) and (p=0.248), respectively. In conclusion, application of ranibizumab could further inhibit the proliferation of MMC treated HConFs by increasing the apoptosis and reducing the necrosis. It was also discovered that ranibizumab alone was not effective in suppressing the migration of HConFs onto the glaucoma tube. Instead, the adjunctive treatment of MMC and ranibizumab could significantly affect the migration of fibroblasts on to the glaucoma tube.

Item Type: Thesis (Masters)
Uncontrolled Keywords: Glaucoma, Therapy
Subjects: R Medicine
Divisions: Kampus Kesihatan (Health Campus) > Pusat Pengajian Sains Perubatan (School of Medical Sciences) > Thesis
Depositing User: Mr Abdul Hadi Mohammad
Date Deposited: 16 Feb 2021 08:43
Last Modified: 16 Feb 2021 08:43

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