Research On Ectatic Corneal Disorders In Mechanobiology

Ectatic corneal disorders(including keratoconus and ectasia after LASIK)are the second most common indications for keratoplasty.After corneal refractive surgery,corneal tensile strain will increase if the cornea becomes thin or if intraocular pressure increases that caused by forceful eye-rubbing.Meanwhile,the inflammatory factor IL-1?,which is released by the epithelium and caused by dry eye disease will also increase after keratorefractive surgery.Therefore,stromal cells are in a complex mechanical and biological environment after surgery.After stromal injury,corneal stromal cells can differentiate into fibroblasts for corneal repairing and wound healing.With the increase of TNF-? in tears and in epithelium of keratoconus,as well as abnormal eye rubbing,keratoconus fibroblasts are also in a complex mechanical and biological environment.To investigate pathogenetic mechanism of estatic corneal disorders,different amplitudes of mechanical stretch were applied to corneal fibroblasts in the presence of IL-1?,the cell morphology of corneal fibroblasts and the ECM changes were observed.On the other hand,keratoconus fibroblasts were extracted to investigate the effects of TNF-? on the expression of the matrix metalloproteinases and their inhibitors.The main work and findings are as follows:(1)In vitro,different amplitude of mechanical stretch(5%,10% or 15%)was applied to corneal fibroblasts to analyse the effects of the mechanical stimulation on the cell morphology of corneal fibroblasts respectively.It was shown that mechanical stretch changed the cell morphology by increasing the contractility of F-actin fibres.Mechanical stretch increased the contractility of F-actin and restrained the migration of corneal fibroblasts.(2)To investigate effects of mechanical stretch on the expression of Collagen? and Lumican,corneal fibroblasts were subjected to 5% or 10% cyclic equibiaxial stretch for 12 h,24 h or 36 h in the presence of IL-1?.At the same time,the expression of MMP2 and MMP9 in cell supernatant was detected by gelatin zymography and western blot under different amplitude stretch for 36 h.The results showed that mechanical stretch and IL-1? reduced the synthesis of Collagen?at 12 h alone.Low stretch(5%)increased the expression of Collagen?at 24 h or 36 h but large stretch decreased it.Mechanical stretch decreased the expression of Collagen ? in the presence of IL-1?.Low mechanical stretch alone promoted the expression of Lumican and large mechanical stretch restrained it at 12 h.The expression of Lumican was increased by mechanical stretch in the presence of IL-1?.IL-1? induced the expression of MMP9 and increased the expression of MMP2 which can be increased by mechanical stretch in the presence of IL-1?.Therefore,it indicates that inflammatory factor and mechanical stretch can induce the synthesis of Collagen?and increase the degragation of extracellar matrix,which maybe play an important role in the development of iatrogenic corneal ectasia.(3)The elevated expression of TNF-? in tears and in keratoconus epithelium puts keratoconus fibroblasts in the condition of inflammatory environment.Keratoconus fibroblasts were extracted and cultured in the medium containing TNF-?.The results showed that TNF-? promoted the expressions of MMPs but restrained the expressions of TIMPs.The increased MMPs and the decreased TIMPs may increase the degradation of the extracellular matrix and destroy the integrity of cornea structure.(4)In the presence of TNF-?,mechanical stretch was applied to keratoconus fibroblasts.It was shown that mechanical stretch and TNF-? restrained the proliferation ofkeratoconus fibroblasts,and did not affect the apoptosis of keratoconus fibroblasts.The decrease in the number of corneal fibroblasts may reduce the synthesis of extracelluar matrix,changed the biomechanical property of cornea.In conclusion,inflammatory factor and mechanical stretch may play an important role in the occurrence and development of ectatic corneal disorders by regulating the metabolism of extracelluar matrix and the proliferation of corneal fibroblasts.