Effect Of Ad-CNTF On The Retinal Ganglion Cells In Rat With Experimental Glaucoma

Backgrounds and purpose: Glaucoma is the second leading cause of blindness worldwide. The path physiology of glaucoma and its optimal treatment are still under investigation, although it is widely accepted that the level of intraocular pressure (IOP) is a consistent risk factor in its incidence, severity, and progression. Moreover, the main approach to therapy for glaucoma is IOP reduction. recently, it was reported in many studies that apoptosis is the mainly death way of retinal ganglion cells (RGCs) of glaucoma. Following the deep researches of glaucoma pathological mechanism, the protective effects of neurotrophic were more and more important for therapy of glaucoma.ciliary neurotrophic factor (CNTF) can induce differentiation of nerve cells, decrease apoptosis of nerve cells and accelerator regeneration of cells. Some experiments showed that CNTF enhance the survival of RGCs, but not report in the animals. The purpose of this study was to observe the effects of Ad-CNTF on the retinal ganglion cells in experimental rat glaucoma. At first, we studied the variation of CNTF expression in the rat retina with chronic ocular hypertension. then observed the effects of Ad-CNTF on the retinal ganglion cells in rat with experimental glaucoma. we use the following. Method: 1. The CNTF recombinant Adenovirus was constructed using the Adenovirus system. 2. Using a diode laser with wavelength of 532 nm aimed at the trabecular meshwork and made 60-80 burns and 5 burns at 3-5 episcleral veins. The intraocular pressure (IOP) was measured by a Tonopen. The retina was dissected after 3days 14days and 35days. The expression of CNTF mRNA was compared using a semi-quantitative RT-PCR. 3. The Ad-CNTF or saline for control were injected in vitreous after the laser treated. The intraocular pressure (IOP) was measured by a Tonopen. The retina was dissected after 3days 14days and 35days. Pattern electroretinogram (PERG) were measured before the animal were excuted.Vertical semithin sections of the retinal were stained by methylene blue and the CNTF immunohistochemistry was measured in the retinal sections. Expression of CNTF mRNA was compared using a semi-quantitative RT-PCR,the thickness of the inner nuclear layer (INL) and inner plexiform layer to internal limiting membrane(IPL-ILM)were measured, the number of RGCs were counted. Results: 1.The CNTF recombinant Adenovirus were constructed successfully, The titer of CNTF Adenovirus was 1.6×1012 PFU. The Adenovirus can mediate the expression of CNTF in NIH3T3 cells by infection study. 2.IOP was 18.1±2.24mmHg in control eyes, At the 3d,14d and 35d, the IOP of laser treated eyes were 21.1±2.08 mmHg,34.8±3.41 mmHg and 32.4±3.01 mmHg respectively. Minimum expression of CNTF mRNA was found in the control retina,and there was an increase of 45.54% and 160.31% in 3d and 14d after laser treated. However, at the 35d there was a CNTF expression decreased and similar as that of control. But after were injected Ad-CNTF, and there was an increase in 14d and 35d after laser treated. mean and HIOP group. after 35d the amplitudes of b wave from Ad-CNTF treated and control were 90.28% and 66.12%,(P<0.05),control retinal thinned in the IPL-ILM and reduced cell numbers in the GCL,some dying RGCs in the experimental retinal showed morphologic features of necrosis, Ad-CNTF ameliorated the degenerative changes of retinal, the IPL-ILM layer was far damaged and more ganglion cells were present than in saline injected eyes. The numbers of dead cells were much reduced than those in control eyes. Conclusions: 1.The CNTF recombinant Adenovirus were constructed successfully and it can expression in the rat's retinal during 35 days. 2. The glaucoma model in rat was made successful,The CNTF expression increased at the start of IOP elevated in rat retina. It is a self-defend mechanism by up-regulation of CNTF mRNA expression when retinal neurons respond to chronic ocular hypertension. But the CNTF expression was exhausted during IOP keeping high for month. It implied CNTF as a foreign agent can be used for neuroprotection against ocular hypertension. 3. Our study support to the concept that the choric high intraocular pressure in the rat eyes induced the retinal ganglion cells apoptosis and the Ad-CNTF can prevent it.