【精品论文】Effects of dibutyl phthalate on ovarian active substance of mice.doc
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1、精品论文Effects of dibutyl phthalate on ovarian active substance of miceWANG Yu, WANG Shengqing, YE Wengbin, HE Jiujun5(Department of Biology and Chemistry, Longnan Teachers College, GanSu ChengXian 742500) Abstract: With superoxide dismutase(SOD), catalase(CAT), alkaline phosphatase(AKP), acid phosphat
2、ase(ACP), malondialdehyde(MDA) and Bax protein expression as characteristic indexes, the effects of dibutyl phthalate(DBP) on ovarian active substance of mice were studied, and the toxic mechanisms were discussed. One hundred sixty mice were given intragastric administration of DBP (0,10100, 250, or
3、 500 mg/(kgd), respectively, for 30 days. At 5, 10, 20, 30 days after DBP exposure, the ovarian examinations were taken. The results revealed that administration of DBP decreased theactivities of SOD, CAT, AKP and ACP, while MDA content and Bax protein expression increased. Two-way analysis of varia
4、nce indicated that dose contributed more to DBP-induced ovarian damage than treatment time. The present study demonstrated that DBP promoted apoptosis and inhibited15antioxidant capacity of the ovarian.Keywords: Dibutyl phthalate(DBP); Ovarian; Antioxidant capacity; Bax protein; Mice0IntroductionIt
5、has caused serious concern about dibutyl phthalate, being added to certain foods and20benerages manufactured in Taiwan. DBP is used extensively in the manufacture of plastics (such as PVC piping and carpet backing), various paints, varnishes and lacquers, medical supplies (such as transfusion and de
6、nta l materials), safety glass for automobiles, food packaging, cosmetics (such as nail polishes and perfume oils), textiles (as a lubricant and as an insect repellant forimpregnation in clothing), and paper coatings1. Although used as an insect repellant, dibutyl25phthalate is not considered to be
7、as effective as dimethyl phthalate. Because of this worldwide use, there is a high potential for human exposure through direct sources in the workplace and the home environment. In addition, numerous indirect sources exist in water, air, and foodstuffs due to the pervasiveness of DBP in the environm
8、ent and by contamination through leachable products. The potential for long-term human exposure to environmental chemicals such as pesticides and30plasticizers, particularly in utero and in neonates, has raised additional concerns2.The antioxidant defense system is an important active oxygen scaveng
9、ing system in organisms, and plays a crucial role in the protective defense response3-4. When an antioxidant defense reaction occurs in an organism in response to toxicants, superoxide dismutase (SOD) is the first antioxidant to react with the reactive oxygen species, causing the superoxide anion35r
10、adical (O-2) to disproportionately form H2O2 and O2. Subsequently, H2O2 is decomposed bycatalase (CAT) into H2O and O2 to decrease the accumulation of H2O2 in organisms 5. Malondialdehyde (MDA) can be formed in the lipid peroxidation reaction. Thus MDA content can be used as an index of the extent o
11、f lipid peroxidation in the tissue, which indirectly represents the level of cell injury. It has been demonstrated that DBP can intensify the tissues40(liver, kidney, testis, lung) lipid peroxidation6, inducing damage to the antioxidant defensesystem and the histological structure of the mouse thymu
12、s, liver, kidney, testis and lung 7-13. However, the mechanisms of DBP toxicity remain unclear. Most previous studies of DBP-related reproductive system damage have focused on the effects of DBP on the histological structure of the testis. The present study hypothesized that DBP inducesFoundations:
13、the science fund of GanSu province (No. 1107RJZK243) and Gansu Provincial EducationCommittee(No.1128B-01).Brief author introduction:Wang Yu, (1973-), master, Associate professor, Research Direction: cell and developmental biology. E-mail: gswangyu- 8 -45pathological damage in the ovarian of mice. We
14、 sought to investigate the effects of DBP on the antioxidant capacity of the ovarian with SOD, CAT, ACP, AKP and MDA as indices of antioxidant function, observe cell apoptosis by Bax protein levels.1Materials and methods1.1 Materials501.1.1AnimalsA total of 160 healthy, adult, Kunming female mice, r
15、egardless of gender, weighing 20-22g, were provide by the Experimental Animal Center of Lanzhou University, China (Certification No.14-006). The experimental procedures were conducted in accordance with the GuidanceSuggestions for the Care and Use of Laboratory Animals, formulated by the Ministry of
16、 Science55and Technology of the Peoples Republic of China.1.1.2ReagentsThe dibutyl phthalate(DBP) samples(a purity of 99%) were provided by Hengxing Chemical Co., Ltd., Tianjin, China. Rabbit anti-Bax protein polyclonal antibody and normal goat serum were purchased form Wuhan Boster Bioengineering I
17、nstitute, China. SOD, CAT, ACP, AKP and60MDA determination kits were provided by Nanjing Jiancheng Bioengineering Institute, Nanjing, China. All the other reagents were of analytical grade or guaranteed grade.1.2Methods1.2.1Establishing the animal modelA total of 160 mice were equally and randomly a
18、ssigned to four groups (40 mice per group):65control, low-dose DBP(100 mg/kg), middle-dose DBP (250 mg/kg), and high-dose DBP (500 mg/kg). The control group was treated with deionized water. The other three groups were treated respectively with different concentrations (100, 250, or 500 mg/(kgd) of
19、DBP6-7. Each group received intragastric administration for 50 consecutive days, once per day, 0.5 mL once. All 160 mice were included in the final analysis.701.2.2ImmunocytochemistryAt 5, 10, 20, 30 days after DBP exposure, five mice were randomly taken from each group. Tissue samples from the ovar
20、ian were obtained after opening the peritoneal. The tissues were fixed with 20% formalin, followed by dehydration, waxing, embedding, and sectioning (6 m thick). The slices were dewaxed with xylene, hydrated in gradient alcohol, boiled in citrate buffer75solution (pH 6.00.1) in a heated pressure coo
21、ker for 1.5 minutes to retrieve antigen, andincubated for 10 minutes at room temperature with 3% hydrogen peroxide to block endogenous peroxidase activity. The slices were incubated with rabbit anti-Bax protein polyclonal antibody (1:200) at 4 overnight, followed by incubation in quick MaxvisionTM t
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