[Expression of substance P in experimental fungal keratitis].

ABSTRACT

OBJECTIVE: To detect the change process of substance P (SP)expression and the difference between fungus and bacterial keratitis and explore effects of SP on the damage and repair process of keratitis model of Wistar rats. METHODS: Wistar rats were divided randomly as blank controlled group, fungal keratitis group and bacterial keratitis group. Inoculate fusarium and staphylococcus into cornea of Wistar rats to make animal keratitis model. Choose the right eye as experimental eye and the left eye as control. After the model was created successfully, 25 rats were executed randomly at 1, 5, 8, 10 and 14 days after the surgery. Detailed record and hematoxylin-eosin dye were done to each group to examine the process of ulcer. Expression of SP were detected through immunohistochemistry and reverse transcription polymerase chain reaction. Use variance analysis, T test and Q test to evaluate the results. RESULTS: One day after infection, the cornea of experimental group showed ulcer with disordered shallow matrix layer collagenous fibre arrangement and neutral granular cell infiltration; after 5 days, ulcer worsened and granular cells infiltrated through the whole layer; after 8 days, ulcer shrinked and fibroblast began to increase with new vessels formed; after 10 days, new vessels began to decrease; after 14 days, the matrix level textile fiber assumes scar type restructuring. The expression of SP increased in endothelium cells, inflammatory cells, fibroblast cells and endothelium of new vessels 1 d after surgery (absorbance for fusarium and staphylococcus group were 0.3313 ± 0.0133 and 0.3995 ± 0.0191 respectively; mRNA were 0.4525 ± 0.0170 and 0.5532 ± 0.0258), peaked at 8 d (for fusarium and staphylococcus group were 0.5525 ± 0.0171 and 0.7050 ± 0.0119 respectively; mRNA were 0.5975 ± 0.0221 and 0.7150 ± 0.0238), began to decrease at 10 d (for fusarium and staphylococcus group were 0.1533 ± 0.0176 and 0.3125 ± 0.0170 respectively; mRNA were 0.2416 ± 0.0082 and 0.4835 ± 0.0082) and dropped to lower than normal at 14 d (for fusarium and staphylococcus group were 0.1150 ± 0.0128 and 0.1675 ± 0.0126 respectively; mRNA were 0.1275 ± 0.0126 and 0.2325 ± 0.0171), which had significant difference (the absorbance of SP for fusarium group, F = 832.24, q = 35.3675, 12.9044, 27.3621, 34.6506, 22.4632, 62.7296, 70.0182, 40.2664, 47.5550, 7.2886, P < 0.01;for staphylococcus group F = 636.17, q = 38.7494, 11.4245, 10.8298, 28.6082, 27.3249, 49.5791, 67.3575, 22.2543, 40.0327, 17.7784, P < 0.01. mRNA expression for fusarium group F = 658.60, q = 18.9941, 9.5132, 27.4719, 42.0007, 9.4809, 46.4661, 60.9948, 36.9852, 51.5139, 14.5287, P < 0.01; for staphylococcus group F = 335.13, q = 16.9266, 4.1677, 7.1081, 32.4724, 12.7589, 24.0347, 49.3990, 11.2759, 36.6402, 25.3643, P < 0.01). Two experimental groups showed similar changes as time changes, the bacterial group showed more SP expression than fungal group, which had significant difference (absorbance of SP t = 6.5493, 7.3867, 16.0505, 14.5479, 6.5360, P < 0.05; mRNA expression t = 7.2878, 9.5232, 8.8149, 43.6256, 11.1269, P < 0.05). Controlled eyes showed increased SP expression at 1 d (absorbance was 0.2840 ± 0.0212; mRNA was 0.3950 ± 0.0129) and dropped to normal at 5 d (absorbance was 0.2125 ± 0.0174; mRNA was 0.3321 ± 0.0041). There was significant difference between bacterial group, fungal group and controlled eyes (absorbance of SP compared with fusarium group t = 4.2261, 18.3314, 35.5163, 5.3609, 13.4826; compared with staphylococcus group t = 9.0508, 25.6639, 63.4924, 7.4828, 7.1301, P < 0.05. mRNA expression compared with fusarium group t = 6.0249, 31.9158, 26.0413, 9.1550, 19.1741; compared with staphylococcus group t = 12.2636, 53.4404, 36.8727, 15.8687, 8.2939, P < 0.05). CONCLUSION: SP possibly participated in the early time damage and the later period repair process in fungus keratitis and its lower expression level possibly participated in the mechanism of lighter ache in fungus keratitis.