| ObjectiveThere are 60 kinds of drugs inducing ototoxicity now days, and most of these drugs are aminoglycosidic antibiotics : such as streptomycin, gentamicin and so on. The main expression of drug deaf is the chronic toxicity of hearing system, and it can induce the damage of vestibular and cochlea hair cell, and can induce dizziness and non - reversible hearing damage. There are many deaf patients in clinic, and about 20,000 30,000 newborn deafness were born, 50% of these was induced by heredity. drug deafness is the main reason of newborn congenital deafness and adults postnatal deafness. there are many hypothesis about aminoglycosidic antibiotics ototoxicity in recently years, for example, drugs storing up in inner hear, and drugs interfering protein compose, and so on. While these hypothesis can not explain completely the mechanism of ototoxicity, which made it difficult to prevent ototoxicity. The US scholar Shacht put forward a new hypothesis - free radical damage hypothesis, that is aminoglycosidic antibiotics and iron ion can compound, which has the activity of oxidizing and reducing reaction, and can induce the form of free radical, and cause hair cells lack, hearing hinder. Among the free radical, NO is paid close attention to. The free radical damage induced by AmAn has strong toxicity to renal besides ototoxicity. The expression of nephrotoxicity is the injury of uriniferous tubules and acute renal failure (ARF). Using free radical elimination VitE can protect the damage of lysosome induced by GM nephrotoxicity, which imply themore free radical is the reason of GM nephrotoxicity. While the more and more NO can induce amount of free radical having toxicity, so infer there are relations between nephrotoxicity and NO.Salvia Miltiorrhiza Injection has the function of invigorating the blood and removing extravasated blood, and is used widely in clinic. It can relieve the vas-ospasm, improve microcirculation, eliminate free radical and improve the energy metabolism of hair cells, in the same time it can eliminate free radical during renal damage, and improve the anti - oxidize function. The recently study of our lab implied Salvia Miltiorrhiza Injection can antagonist the GM ototoxicity , and it can decrease the activity of NOS, decrease the product of NO.The study takes the guinea pigs as experimental object, and takes the cochlea and renal as experimental specimen, investigate the iNOS expression after u-sing SM, and the related protein expression, and investigate the apoptosis, in order to study the mechanism of SM toxicity damage. Meanwhile use Salvia Miltiorrhiza Injection antagonist, in order to impose a useful protect drug for SM toxicity damage.Methods1. Animal grouping, model metabolism and study(1) In this experiment we used 20 healthy , white red - eyes, otomicro-scopically normal, ABR thresthold of double ears ≦ 5dB ,adult guinea pigs either female or male with body weights in the range 200 250g (the second affiliated hospital animal lab , China Medical University). The animals were divided in random four groups, each had 10 animals. ①the control group ;②SM group; ③DS + SM group;④DS group. The control group: injected normal saline 2. 5ml/kg/d by abdominal cavity; SM group: injected SM 150mg/kg/d; DS + SM group: injected DS 3g/kg/d and SM 150mg/kg/d in the two side of abdominal separately; DS group: injected DS 3g/kg/d. each group was administrated continuously 10d, and adjusted the drug dose by weighting. Every animal was exam ABR before administration and the 1st stopping drugs, in order to supervise the toxicity damage.(2) after recorded ABR, beheaded the guinea pig rapidly, toke out two sides of otocyst, renal and urinary specimen. opened the otocyst, exposed the cochlea, put the cochlea into 4% paraformaldehyde and 2.5% glutaric dialde-hydes fixators to fix. Investigated the expression of iNOS, AChE, bFGF using the method of immunohistochemical staining, and the normal HE staining. Investigated the apoptosis with terminal doxynucleotidyl transferase - mediated dUTP nick end labeling (TUNEL) method. observed the change of marphy under light microscope, transmission electron microscope ( TEM ). put the renal specimen into 4% paraformaldehyde and 2. 5% glutaric dialdehydes fixators to fix. Investigated the expression of iNOS, bFGF using the method of immunohistochemical staining, and the normal HE staining. Investigated the apoptosis with TUNEL method. observed the change of morphology under light microscope, transmission electron microscope (TEM).2. image analysescochlea: Used the MetaMorph - Coolsnapfx - AX70 micro image analysis system to determine the average gray value of iNOS positive reaction of each group. When exam, took 10 piece of the slide of each group , each slide took 4 slices randomly , each slice took each chapter of cortis, stria vascularis, spiral ganglion , nerve fibers as the field of view. Put the slice under the microscope (10 × 40) ,after presented the image, photographed with the digital camera, image input computer. The gray value smaller, the expression of positive reaction was stronger. Apoptosis: apoptosis positive cell nucleus is expressed by yellowi-shbrown pellet, and the normal cell nucleus is blue. One positive cells expressed in the outer hair cells of Corti's is positive expression, more than two positive cells is thought strong positive expression, and the non - typical case is little positive expression.Renal : Used the MetaMorph - Coolsnapfx - AX70 micro image analysis system to determine the average gray value of iNOS and bFGF positive reaction of each group. When exam, placed the slide under the microscope(10 40) , after presented the image, photographed with the digital camera, image input computer. measuring average gray of iNOS and bFGF in cortial nephron and jux-ta medullary nephron in the same condition, the gray value smaller, the expres-sion of positive reaction was stronger. counting the number of apoptosis and the sum in the continuity un - repeatedly in visual fields for calculating the percentage of the number of apoptosis occupies total cells.3. statistics processingThe result indicated with the mean value standard difference ((x|-) ± s) , and average comparison of each group was examined with t values and the correlation values , carried on the statistical analysis using the Excel statistics software.Results1. the cochlea: Each group of ABR threshold value did not have remarkable difference before administration ( P >0. 05 ) , but the ABR threshold value of SM group after administration elevated remarkably, and compared with the control group, the difference is remarkable (P <0.01) , though the ABR threshold value of DS + SM group is higher than the control, it is lower than that of SM group, and the difference is remarkable ( P < 0. 01 ) ; the DS group has no difference before and after administration ( P >0. 05 ). Observed under the light microscope and transmission electron microscope (TEM) , the change of DS + SM group is more slight than that of SM group, the change of apoptosis cells is less than that of SM group. Meanwhile the iNOS expression of SM group is more strong , compared with the control and DS + SM group , there is remarkable difference ( P < 0. 01) , the AchE expression is also stronger than that of the control and DS + SM group, and the difference is remarkable( P <0.01) , while the bFGF expression is lower than DS + SM group ( P <0. 05). the number of apoptosis cells of DS + SM group is little than that of SM group.2. renal: observed under the light microscope and transmission electron microscope (TEM) , the change of SM group is more serious, and the apoptosis is also more serious than DS + SM group. The iNOS expression of SM group is stronger than that of the control and DS + SM group, and the difference is remarkable ( P < 0. 01) , bFGF expression is lower than that of DS + SM group, and the difference is remarkable ( P < 0. 01) , the percentage of the number of apoptosis occupies total cells of DS + SM group is lower than SM group. The...
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