Oküler Yüzey Uygulamaları İçin Foto-Çapraz Bağlanmış Jelatin Metakriloil–Amniyotik Membran Hidrojeli Geliştirilmesi

Öz

Amniyotik membran (AM), plasentanın en iç tabakası olup kalın bir bazal membran ve avasküler bir stromal matriksten oluşmaktadır. İlk olarak 20. yüzyılın başlarında deri grefti olarak kullanılan AM, 1940 yılında De Rotth tarafından oküler yüzey rekonstrüksiyonu amacıyla oftalmoloji alanına kazandırılmıştır. Bu çalışmada, klinik kullanımda olan amniyotik membran toz formuna dönüştürülmüş, jelatin metakriloil (GelMA) ile birleştirilerek oküler yüzey uygulamasına uygun, sürekli salınımlı bir göz damlası formülasyonu geliştirilmiştir. AM–GelMA formülasyonu %5 ve %10 son konsantrasyonlarda hazırlanmış ve yapısal bütünlüğün sağlanması amacıyla foto-çapraz bağlama uygulanmıştır. Reolojik analizlerde, tüm çapraz bağlama süreleri boyunca elastik davranışın (G′ > G″) baskın olduğu gözlenmiş, viskozitenin ölçüm aralığında belirgin değişiklik göstermediği ve kayma incelmesi (shear-thinning) özelliği gösterdiği belirlenmiştir. İnsan limbal stromal mezenkimal kök hücreleri kullanılarak yapılan in vitro sitotoksisite değerlendirmesinde, 24. saatte mitokondriyal aktivitede artış gözlenmiş; hücre canlılığı %5 AM grubunda %180’e, %10 AM grubunda ise %160’a ulaşmıştır. Buna karşılık, 48 ve 72. saatlerde her iki grupta da hücre canlılığı %100’ün altında ölçülmüştür. Büyüme faktörü analizlerinde, %5 AM grubunda 48. saatte epidermal büyüme faktörü (EGF) düzeyi 313 pg/mL, dönüştürücü büyüme faktörü-beta (TGF-β) düzeyi ise 350 pg/mL olarak ölçülmüştür. Bu bulgular, geliştirilen AM–GelMA formülasyonunun in vitro koşullarda biyolojik olarak aktif faktör salımı sağlayabildiğini ve oküler yüzey uygulamalarında kullanılabilecek bir sürekli salım sistemi olarak değerlendirilebileceğini düşündürmektedir.

Anahtar Kelimeler

• Amniyotik membran
• Transforme edici büyüme faktör-beta
• Epidermal büyüme faktörü

Development of Photo-Crosslinked Gelatin methacryloyl –Amniotic Membrane Hydrogel for Ocular Surface Applications

Öz

The amniotic membrane is the innermost layer of the placenta, composed of a thick basal membrane and an avascular stromal matrix. Initially utilized as a skin graft in the early 20th century, it was later introduced to ophthalmology in 1940 when De Rotth proposed its use for ocular surface reconstruction. In this study, clinically used amniotic membrane was converted into a powder form, combined with gelatin methacryloyl (GelMA), and developed as a sustained-release eye drop formulation for ocular surface application. The amniotic membrane–GelMA formulation was prepared at final concentrations of 5% and 10% and photo-crosslinked to support structural integrity. Rheological analysis showed that the elastic component (G′) exceeded the viscous component (G″) across all crosslinking durations, with no marked variation in viscoelastic parameters within the tested range. The formulation exhibited shear-dependent viscosity reduction under increasing shear rates. In vitro cytotoxicity assessment using human limbal stromal mesenchymal stem cells revealed increased mitochondrial activity at 24 hours, with cell viability values of 180% in the 5% amniotic membrane group and 160% in the 10% group relative to control. At 48 and 72 hours, viability values decreased to below 100% while remaining within a non-cytotoxic range. Growth factor analysis demonstrated measurable release of epidermal growth factor (EGF) and transforming growth factor-beta (TGF-β), reaching 313 pg/mL and 350 pg/mL, respectively, at 48 hours in the 5% group. These findings suggest that the amniotic membrane–GelMA formulation exhibits sustained factor release and biocompatible characteristics under in vitro conditions. The system may represent a non-surgical, drop-based approach for delivering amniotic membrane–derived bioactive components to the ocular surface.

Anahtar Kelimeler

• Amniotic membrane
• Transforming growth factor-bet
• Epidermal growth factor

Kaynakça

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