Mesangial proliferative glomerulonephritis (MsPCN) is one of the most common pathological types of primary glomerulonephritis in China, accounting for about 30% of the primary glomerular diseases in China e. The development of an animal model of mesangial proliferative glomerulonephritis of clinical utility is of great significance for the study of pathogenesis, pathology and preventive measures of the disease; it also provides a research tool for the study of cytokines of various glomerular diseases similar to that of the human body. The production of animal models with practical clinical value is important for the in-depth study of the pathogenesis of nephritis, lesion patterns and preventive measures; it also provides a research tool similar to the human body for the study of the cytokines of various glomerular diseases. The commonly used animal model of thylakoid proliferative glomerulonephritis is the rat Thy-l thylakoid proliferative glomerulonephritis model. In this section, a modified chronic serum sickness nephritis model is presented, and an animal model of thylakoid proliferative glomerulonephritis with homogeneous pathologic types was obtained.
Principle
Depending on the experimental method, the corresponding principles differ:
The basic principle of the animal model of the mesangial proliferative nephritis:
When the foreign protein enters the body, it stimulates the immune system to produce antibodies, and the combination of antigen and antibody produces immune complexes and deposits them in the kidneys, thus producing immune damage. The titer of antibody production was increased by multiple subcutaneous injections of Fuchs' adjuvant containing BSA and intraperitoneal injections of high concentration of BSA.
Tail vein injections and intraperitoneal injections were alternated on alternate days to maintain a high antigen concentration in the animals.
Appliance
The common applications of the animal model of thylakoid nephritis are as follows: This modified thylakoid glomerulonephritis model is very similar to the human thylakoid glomerulonephritis. Compared with the traditional model. The model has a high morbidity rate, short cycle time, uniform lesion degree and pathology, and can be used for basic and clinical nephritis research in human thylakoid proliferative glomerulonephritis.
Operation method
Animal model of mesangial proliferative nephritis
Principle
When the foreign protein enters the body, it stimulates the immune system to produce antibodies, and the combination of antigen and antibody produces immune complexes that are deposited in the kidneys, resulting in immune damage. The titer of antibody production was increased by multiple subcutaneous injections of Fuchs' adjuvant containing BSA and intraperitoneal injections of high concentration of BSA. Tail vein injections and intraperitoneal injections were alternated on alternate days to maintain a high antigen concentration in the animals.
Materials and Instruments
Experimental subjects: rats. Move The basic process of the animal model of thylakoid proliferative nephritis can be divided into the following steps: (I) Process I A. Preparation of the model (1) Experimental animals: 30 male SD rats, weighing 160--200 g. After ether anesthesia, the left kidney was removed via abdominal cavity and rested for 1 week. (2) Pre-immunization: 26 experimental rats were injected subcutaneously with Fuchs' complete adjuvant 0.1 mg plus t 3 mg of bovine serum albumin (BSA) at the end of the week of 1.2. At the end of the week of 3, 4 consecutive injections of BSA were given intraperitoneally at 1-hour intervals at the doses of 0.5, 1.0, 1.5, and 3.0 mg per rat, respectively; and then the dose was reinforced by 1 injection the next morning at the rate of 2.0 mgo per rat. (3) Immunization:The experimental group was randomly divided into the entry group, B, group and B, group, each group was 7 animals. Experimental group A:BSA tail vein injection was alternated with intraperitoneal injection on alternate days. The dose of tail vein injection started from 0.5 mg per animal, and increased by 0.5 mg each time to 2.5 mg, and continued to increase by 0.5 mg to 5 mg per week. The amount of intraperitoneal injection was twice the amount of tail vein injection, and 100 mg of Escherichia coli endotoxin was injected into the tail vein after 2 weeks of immunization, and the animals were killed after 10 weeks of observation. Experiment B, Group B: The immunization method was the same as that of Group A, and the animals were killed after 7 weeks of observation. Experiment B, Group B: BSA was injected into the tail vein alone at the same dosage as in Group A. The animals were not injected with Escherichia coli endotoxin and were observed for 7 weeks and then killed. Control group(5pcs):Pre-immunization and immunization methods were the same as in group A. The same volume of isotonic saline was used to replace the BSA, and the animals survived for 7 weeks and were killed. B. Observation indexes (l) Blood . Urine examination: blood creatinine test with picric acid method; blood urea nitrogen with IFCC rate method. The total amount of urinary protein, urinary creatinine, urea nitrogen and urine sediment smear were measured by Hitachi 7150 automatic hematology analyzer in 24 hours. Urine samples were collected before immunization, once a week after immunization and before execution. (2) Pathological examination: Kidney specimens were examined by light microscopy, projection electron microscopy and direct immunofluorescence. The PAS-stained thin sections of kidneys were analyzed by AXIO HOME image analyzer. Three animals were randomly selected from each group, and one section of the right side of the kidney was taken from each animal. 6 glomeruli (3 cortical and 3 paramedullary) were randomly selected from each section. The total area of the glomeruli and the area of each mesangium were measured directly, and the number of glomerular mesangial cells was counted. Caveat 1、Blood and urine examination results of each group of rats plasma urea nitrogen, blood creatinine ratio difference was not significant.①Hematuria: 2 weeks after immunization, 8 animals showed microscopic hematuria; 3 weeks after immunization, 3 animals showed microscopic hematuria.Urine protein: all animals were negative for urine protein before immunization. After 2 weeks of immunization, micro-proteinuria began to appear, dynamic observation of the total amount of urinary protein in Group A rats weekly 24 changes in immunization 2 weeks, 3 weeks, 7 weeks and 10 weeks, there were two times rise in the total amount of urinary protein compared with the beginning of immunization, the difference was significant.Urine biochemical examination: the rise and fall of urinary creatinine and urinary protein in experimental group A were basically parallel, and the difference between the experimental groups and the control group in terms of urea nitrogen and urinary creatinine was not significant. 2. Pathologic examination(1) Light microscopy: 3 weeks after immunization, glomerular capillaries were obviously dilated, endothelial cells were swollen, neutrophils were infiltrated, plasma proteins and erythrocytes leaked out of the renal capsule; epithelial cells of the renal tubules were granular and vacuolated, and erythrocyte tubular and hyaline tubular forms were observed. At 4 weeks of immunization, there was segmental hyperplasia of the mesangial cells, and some of the renal vesicles were adherent. At 7 weeks of immunization, there was diffuse mild to moderate proliferation of glomerular mesangial cells.At 10 weeks of immunization, the proliferation of mesangial cell nuclei and mesangial stroma was more obvious, and the renal interstitium was congested, edematous and infiltrated by focal mononuclear cells and lymphocytes; the basement membranes of glomerular capillaries had no obvious lesions; the lesions of group B were lighter than those of groups A and B, and the structure of the glomeruli in the control group was normal.(2) Immunofluorescence examination: at 3 weeks of immunization, there were fine granular IgG (++~+++) and C3 (+~++) deposits along the mesangial area in the glomeruli of some animals.At 4-7 weeks of immunization, all cases showed positive reactions (IgG ++~10++, C3++++, FRA++~4++), which were manifested as coarse granular or clumped deposits in the thylakoid zone.At 10 weeks of immunization, lgG and C3 were strongly positively deposited in large clumps in the thylakoid membranes, with or without deposition in the glomerular capillary wall. Immunofluorescence was negative in the control group.(3) Electron microscopy: 4 weeks after immunization, the glomerular cells in the glomerular membrane showed mild stage hyperplasia, and the epithelial cells were fused with pedunculated cells in stages. After 7-10 weeks of immunization, the proliferation of mesangial cells and mesangial stroma was more obvious, and cloudy electron-dense material was deposited in the mesangial area, occasionally seen in the subendothelium. For more product details, please visit Aladdin Scientific website.
Experimental reagents: ether, bovine serum albumin.