动物造模,尿道疾病动物模型 z-bio 2021-10-13 658

　　A mouse hypospadias model induced by flutamide (1) Reproduction method 10 week-old pure Kunmei mice were kept in a cage overnight, and the female mice after vaginal embolism were raised the next morning In a cage. On the first day of pregnancy, flutamide was orally administered to mice on the 15th day of pregnancy, and the flutamide tablets were crushed into fine powder before administration. The drug was dissolved in 0.4ml of distilled water and administered continuously for 5 days. The doses were 133.3 mg/kg body weight, 200.0 mg/kg body weight, 266.7 mg/kg body weight and 333.3 mg/kg body weight. Male mice were collected on the 21st day after the offspring were born, and the morphological changes of the male mice's external genitalia in each dose group were observed with naked eyes and a dissecting microscope, and judged according to the following four items. Grade: Grade 0: No deformation, the size and shape of the external genitalia are basically normal, and there are no obvious cracks. Grade 1: Slight deformation, obvious cracks in the upper part of the genital protrusions, and the appearance is "M". Grade 2: Moderate strain, with significantly smaller genital protrusion. Cracks, hair on the scrotum, non-protruding testicles, cryptorchidism, female appearance; Grade 3: Severe abnormality, genital protrusions are completely torn, urethra open to the anus, cryptorchidism has a pseudo-vaginal depression structure.

　　(2) Model characteristics When observing the external genitalia of model animals, the external genitalia of male mice weighing 133.3 mg/kg has no obvious abnormalities, while the external genitalia of 200.0 mg/kg has no obvious abnormalities. There are no obvious abnormalities in kilogram male mice. The kilogram recombination is prominent, the upper part is obviously cracked, and the deformation rate can reach 86.6%, which is mainly mild abnormality. On the other hand, the reproductive organs of male mice weighing 266.7 mg/kg are very similar to female mice. Hair on the scrotum, without protruding testicles. Faintly deformed, with a deformation rate of 83.3%; 333.3 mg/kg recombined male mice have completely divided external genital protrusions, forming a vagina-like structure on the anal side. , All of them show obvious moderate to severe deformation. According to observation and statistical analysis, oral flutamide was administered to model mice at a dose of 200.0 to 333.3 mg/kg body weight from the 15th day after conception. Continuous administration for 5 days can cause more than 80% of males. . There was congenital hypopnea in mice. The model creation method is simple and convenient, the model creation time is short, the cost is low, and the model formation rate is high. (3) Comparative medicine Hypospadias is one of the common clinical congenital hip dysplasias, and its incidence has been increasing in recent years. The establishment of an ideal congenital hypospadias animal model can provide valuable experimental materials for the etiology and drug treatment of the disease. The factors leading to the development of the disease include environmental factors, endocrine factors and genetic factors. The method of creating this model is based on the principle of androgen receptor interference. Flutamide can be used to interfere with the normal action of androgens in the body and successfully replicate the congenital androgen rupture mouse model. During this period, the androgens and urethra of male mice begin to develop on the 17th day of the embryonic period and complete on the 19th day. Therefore, testosterone and its biotransformer dihydrotestosterone (DHT) are present in the mice. Androgens. Very important role in male reproductive differentiation. However, this effect must be mediated by the androgen receptor (AR) in the body, and the AR function of the target cell is normal to ensure the normal expression of androgen efficacy. Therefore, the differentiation and development of male animal (human) reproductive organs are closely related to AR. The flutamide used in the modeling process is an androgen receptor antagonist. Excessive flutamide may interfere with the action of androgens in the development of the external genitalia and urethra of male fetal mice, which may interfere with the action of androgens and lead to male puppies. The incidence of hypospadias is low. This model can be used for clinical research on the etiology and drug treatment of upper hypospadias. 2 Establish a rat model of hypospadias with diethylstilbestrol (1) Copy method Place adult rats weighing 200 to 250 g in a cage overnight and mate. By the next morning, female rats were observed in a cage after vaginal reproduction. This is considered the first day of pregnancy. Grind diethylstilbestrol (DES) tablets into powder to make 1.5 ml of salt-containing DES suspension, and shake it well during tube feeding. From the 12th day to the 17th day of pregnancy, 1.5 ml of DES was taken orally at a dose of 100 μg/kg body weight every day for a total of 6 times. The pregnant mice are pregnant until the 21st. After spontaneous birth, count the newborn rats and observe the appearance of the foreskin, the position of the urethra, urination, body weight, and anogenital distance under a magnifying glass. Then use an dissecting microscope (AGD) to determine whether there is an abnormality in the hypospadias and take pictures. (2) Model features DES injection can cause hypospadias in male offspring, and the model incidence is about 30%. The urethra of the offspring of normal newborn males is opened in the upper part of the penis, and the foreskin is intact. When stimulated, urination becomes linear. The urethra of the newborn male offspring with hypospadias is opened at the junction of the penis and the perineum or on the ventral side of the penis, and the foreskin is torn on the ventral side of the penis, exposing the glans. In addition, the urethra of some hypospadias offspring is open on the ventral side of the cavernous body or at the base of the penis. It is similar to the human penile hypospadias or perineal hypospadias. The AGD of model male puppies is shorter than that of normal puppies, which indicates that DES has a feminizing effect on males. At the same time, under the action of DES, the weight of male puppies was significantly lower than that of normal puppies. The second and second trimesters are periods of rapid weight gain in the uterus.

　　(3) Comparative medicine Hypospadias is one of the most common congenital malformations in clinical pediatric surgery, and its incidence is increasing, but its cause and mechanism are still unclear. Therefore, the establishment of an animal model similar to clinical pediatric congenital hypospadias has very important clinical research value. This model uses rats with a low incidence of spontaneous malformations. By administering DES to pregnant rats during pregnancy, newborn male hypospadias rats can be successfully prepared. The model method is simple, short in time, low in cost, and easy to observe. Compared with mice, the incidence of spontaneous teratogenesis in rats is very low, the rate of spontaneous conception is higher than that of mice, the drug resistance is very strong, and the size of the offspring of rats is also larger. The mouse deformation is easy to observe. This model is easier to observe and operate than the mouse hypospadias model, and is an excellent animal model for studying the etiology, prevention and treatment of hypospadias. 3 Rat model of hypopnea caused by dibutyl phthalate (1) Replication method Adult female rats weighing about 220g were mated with male rats overnight, and their presence was observed in the early morning of the next day. The day I saw the vaginal plug was the first day of pregnancy, and the female mice were placed in a cage regularly, drinking and eating at will. Dibutyl phthalate (DBP) was added to refined soybean oil at a weight of 500 mg/kg, and it was taken orally once a day from the 12th to the 19th day of pregnancy. The dose of tube feeding is only 1 ml/day. Get pregnant naturally at the end of the gestation period. Seventy days after birth (the rats were sexually mature), the penis length, curvature and urethral orifice of male mice were examined and measured one by one. Use a microscope with a micrometer to determine the distance between the anus and the genitals (AGD) and whether there is a spine. On day 19, some pregnant mice underwent a cesarean section, and testicular tissue was collected from male mice. Reverse transcription polymerase chain reaction (RT-PCR) is used to detect the rate-determining enzyme cytochrome P450 cholesterol in the process of testosterone synthesis. mRNA horizontal chain lyase (P450SCC), 3β-hydroxysteroid dehydrogenase (3β-HSD) and cytochrome P45017 (α-hydroxylase (P450c17). Standard for normal male offspring: normal male rat urethra is located above the testis When opened, the anterior skin stimulates intact and urination can be linear; in males, the urinary tract of hypospadias rats is the junction of the penis and testis or the ventral side of the penis body, and the tears and particles on the anterior skin are bare This indicates that a certain dose of DBP can cause hypopnea in rats. At this time, the genitourinary system of the offspring of male rats in the uterus of pregnant rats has been formed and started to develop on the 12-19th day of pregnancy. At this time, the DBP (500 mg /kg body weight) The gene expression levels of P450SCC, 3β-HSD and P450c17 in the testis tissues of fetal rats in the DBP-induced hypopnea group were lower than those of normal fetuses. Conclusion DBP interferes with the organism of testosterone in the testis tissues of male offspring and sexually developed mice The synthetic pathway lowered testosterone levels, resulting in hypopnea after birth in rats.

　　(3) Comparative medicine lyase is a clinical cause of anterior urinary tract dysplasia: the deformation that makes the penile urinary tract open to its normal position is usually related to penile flexion, and is mainly found in men. This is a childhood urinary tract disease The common congenital malformations are increasing year by year. Abnormal appearance and urination patterns seriously affect the physical and mental development of children. This disease is usually combined with other variants such as invisible testicular disease and indirect hernia, which may cause difficulty in sexual life. Adults will cause problems for children and families, and life will bring serious consequences. The cause of hypopnea is so complicated that its cause has not yet been clarified. Environmental pollutants are one of the factors that increase the incidence of hypopnea. A cheap and practical plasticizer, widely used in people's daily life and production. It is also an environmental pollutant. It belongs to anti-androgen environmental endocrine disrupting substances. Its mechanism of action is similar to that of flutamide, but it is not perfect. Well. it's the same. DBP has obvious testicular toxicity, embryo toxicity and reproductive abnormalities to animals. This can significantly reduce testosterone levels in the body, leading to testicular atrophy, testicular atrophy and epididymal dysplasia. Secondary gonads and external genitalia. Pathological damage to the male reproductive system, such as absenteeism and lack of breathing. In addition to flutamide, DES and DBP can also cause hypopnea in animals, and benzoic acid and aspirin can also cause hypopnea in mice. The hypothalamic model is mainly related to the effect of drugs on animal hormone levels, because patients with hypospadias caused by clinical urethra are often related to sex hormones, especially the abnormal metabolism of androgens in the body. The hypothalamus is not only a local malformation of the body, but also a local symptom of endocrine disorders. Hypospadias caused by drugs may be mainly due to the changes in the apoptosis and proliferation activity of the ventral urethral epithelial cells of the intrauterine device. Therefore, the location of inducing hypospadias is closely related to the gestational age at the time of drug exposure. The rat model of DBP-induced hypospadias provides scientific animal experimental evidence for the relationship between the etiology and environmental pollutants of congenital hypospadias and the occurrence of hypospadias in children for clinical research.