动物造模,心肌病模型 z-bio 2021-04-07 522

　　Cytoskeleton protein genes: Studies have shown that the genes encoding myocardial skeletal proteins are closely related to the pathogenesis of dilated cardiomyopathy. The cytoskeleton system plays a role in transmitting contractile force and mechanical support. Studies have found that many dilated cardiomyopathy are related to the defect of force conduction protein. The defect of force conduction process and the skeletal protein cannot effectively maintain the cell membrane structure during myocardial contraction, which will lead to chronic mechanical damage to myocardial cells. , And then interstitial fibrosis, heart enlargement. Related cytoskeletal proteins include laminin, desmosomal protein, delta-sarcoglycan, anti-dystrophin and vinculin.

　　Dystrophin gene dystrophin gene, located at Xp21.2, its mutation causes X-linked dilated cardiomyopathy. Dystrophin is mainly expressed in skeletal muscle, myocardium, and smooth muscle, including 4 independent domains (N-terminal domain, long repeat domain, cysteine-rich domain and C-terminal domain), which are involved in power Conduction and mechanical support function to disperse the force to the surrounding adjacent tissues and make the muscle fibers resist the stress during contraction. Mutations in the dystrophin gene are mostly located at the 5'end of the dystrophin gene, which affects the amino end of dystrophin and actin. Dystrophin is a larger cytoskeletal protein, its amino terminal is connected to actin, and the carboxy terminal is connected to a larger transmembrane glycoprotein complex (DAG). Therefore, dystrophin plays an important role in establishing the connection between the intracellular skeleton and sarcomere and the extracellular matrix, and provides structural support for the cell membrane. In addition, dystrophin also participates in cell signal transduction, especially through its interaction with NOS. It is known that in all striated muscles, the contractile force is transmitted from the musculature to the extracellular matrix by the DAG complex. Mutations in the dystrophin gene will inevitably lead to the destruction of DAG, loss of the tightness of the plasma membrane and fiber necrosis. Some mouse animal models are used to simulate human Duchenne muscular dystrophy and Baker's muscular dystrophy. Mainly mdx mice with dystrophin deletion, dystrophin/utrophin double gene knockout (u-dko) mice and m-dko mice. ①mdx mouse: The naturally occurring mdx mouse is caused by a meaningless mutation in exon 23 of the dystrophin gene. The heart of mdx mice younger than 3 months has few or only mild lesions: small focal degeneration and local inflammation. By 6 to 8 months, the heart of mdx mice begins to show moderate myocardial necrosis and fibrosis. The typical physical changes in heart disease do not appear until December, and they become more and more serious with age. There is no obvious pathological change in the heart of mdx mice less than 3 months old, but in vitro experiments have confirmed that the myocardial contractile function of mdx mice aged 8 to 14 weeks is reduced but it has not affected hemodynamics until the stress state occurs. Electrocardiographic studies. Old mdx mice showed deep Q waves as reported in DMD/BMD patients, increased R/S, frequent ventricular premature contractions, and decreased heart rate variability. Gene defects similar to mdx mice also exist in humans, so mdx mice are a good animal model for studying cardiomyopathy, especially for gene therapy of cardiomyopathy. ②U-dko mouse: The utrophin gene is located at 6q24 and is 1Mb long. The cDNA is about 13kb in length and contains 73 exons. It is the second largest gene discovered by humans after dystrophin. The utrophin protein has extensive homology with dystrophin, with a relative molecular mass of 395,000 Da. It is located under the muscle cell membrane in some myopathy and embryonic development, and is confined to the neuromuscular junction and tendon junction after birth. The utrophin protein located on the muscle fiber membrane of the neuromuscular junction (NMJ) connects the extracellular matrix with the skeletal protein under the cell membrane, and plays an important role in cell transmembrane signal transduction. utrophin is up-regulated in mdx mice. In order to study whether the up-regulation of utrophin gene can compensate for the loss of dystrophin, u-dko (utrophin/dystrophin double gene knockout) mice were produced. Compared with light mdx mice, u-dko mice are weaker and smaller in size. They exhibit progressive muscle atrophy, growth retardation, weight loss, curvature and deformation of the spine. Died 8-10 weeks after birth. The laboratories of Grady and Deconinck established models of U-dko rats. However, due to different genetic backgrounds or different gene knockout methods, the performance of U-dko mice established by these two laboratories is not completely the same. The U-dko strain of Deconinck laboratory is comparable to that of wild animals in terms of heart weight and body weight. Similar to the litter mice, the 10-week-old Deconinck u-dko strain with a cardiology score is similar to mdx mice. All utrophin subtypes in the Grady strain are alive, while only most utrophin in the Deconinck strain is inactivated. ③M-dko mouse: MyoD is one of the four members of the myogenic regulatory factor family. It has a basic helix-loop-helix (bHLH) domain, which can transform many types of cells into myoblasts and promote the fusion of myoblasts into Myotube. In the transcriptional regulation of muscle-specific genes, MyoD acts as a master switch, acting on the promoter or enhancer regions of many genes, and with the assistance of other factors, promoting their transcriptional activity, insulin-like growth factor-Ⅱ, transcription Co-activator p300/CBP, nuclear factor 90, nuclear hormone receptor complex activator, interferon-related developmental regulator 1, etc. can promote MyoD's transcriptional regulation of genes; inhibin 2, myogenic somatostatin, histone A Base transferase Suv39h1, DNA binding inhibitor Id, tumor necrosis factor-α, etc. can inhibit MyoD's transcriptional regulation and activity of genes, so that MyoD can adapt to the needs of internal environment and physiological functions. MyoD is mainly degraded through the ubiquitin proteasome pathway to meet the needs of molecular pathway regulation. In order to study whether muscle regeneration has an effect on relatively healthy mdx mice, Megeney, L.A's laboratory established m-dko mice. As expected, m-dko mice developed severe muscular dystrophy symptoms similar to those of patients. Surprisingly, m-dko mice developed dilated cardiomyopathy.

　　Lamin A/C gene (UV-NA) lamina protein (Lamin A/C) coding gene LMNA is located at 1p1-q21. Lamin A/C is a dimer located in the inner layer of the nuclear membrane. As a nuclear membrane intermediate filament protein, Lamin A/C maintains the structural integrity of the nuclear membrane and provides mechanical support. Arbustini et al. found that 5 mutations (K97E, E111X, R190W, E317K, 1712 4 base pair insertion) are related to dilated cardiomyopathy, and the results suggest that about 33% of dilated cardiomyopathy with atrioventricular block There are LMNA mutations, but the specific mechanism is unclear. Karkkainen et al. found the mutation Ser143Pro in 24 people and 1 sporadic patient in 5 families. Most patients have sinus node or atrioventricular node dysfunction and require permanent pacemaker implantation. It is speculated that it is of great significance to investigate the lamin A/C gene, especially the Ser143Pro mutation in patients with dilated cardiomyopathy with conduction system disorder. LMNA knockout mice (sullivan, 1999) showed rapidly progressive dilated cardiomyopathy, similar to the human autosomal dominant disease Emery-Dreifuss muscular dystrophy (Emery-Dreifuss muscular dystrophy), according to Emery-Dreifuss muscle In a mouse model of malnourished LMNA gene mutation site with L530P gene knock-in, heterozygous LmnaL530P/wt showed no significant difference from wild-type mice, and homozygous showed a phenotype similar to Hutchinson-Gilford Progeria, after birth 4 Severe developmental delay began to occur within ~5 days, and death occurred within 4 to 5 weeks.

　　"DES", the gene encoding desmin, is located on chromosome 2q35. Desmin is a specific intermediate filament protein in muscle. Thornell L et al. introduced the mutant desmin into the germline of mice, and the homozygous mutant mice developed pathological manifestations of cardiomyopathy after 10 days of birth. Carlsson L knockout (K/O) desmin gene does not affect myogenesis and muscle fiber formation, but develops cardiomyopathy and skeletal muscular dystrophy after birth. Therefore, the consequence of the lack of desmin gene is that muscle fibers are more sensitive to damage. Gene targeting rupture mouse muscle LIM protein (MLP) is a new model of heart failure. MLP is a regulator of muscle tissue formation or differentiation. MLP knockout homozygous mice developed dilated cardiomyopathy with cardiac hypertrophy, interstitial cell proliferation, and fibrosis. Adult mice have hemodynamics and clinical signs similar to human heart failure. Due to the similarity between the two, it is suggested to scholars that the molecular mechanism that causes MLP dysfunction may be involved in the occurrence of human dilated cardiomyopathy.

　　Syrian hamster BIO14.6, BIO14.6, BIO50.54, BIO82.62, BIO53.58, UM-X7.1 and other strains (cardiomyopathic hamster, CM) are hamsters that can develop spontaneous cardiomyopathy, and the general lesions appear in 30 ～40-day-old hamsters, female rats were found as early as 10 days, ventricular hypertrophy and myocardial degenerative changes were visible at about 60 days of age. The pathological changes included cardiomyocyte lysis. The course of the disease continued to be seen in the myocardium with mononuclear inflammatory cell infiltration and connective tissue replacing degenerative muscles. cell. At the age of 100 days, extreme dilation of the heart cavity, pulmonary edema, and liver congestion can be seen. The pathological changes of this kind of mouse myocardium are more serious than that of skeletal muscle, and it is generally considered to be auto-recessive inheritance due to mutations in the sarcoglycan gene (SGCD gene). δ-sarcoglycan (SG) is a component of the sarcoglycan subcomplex in the dystrophin-related glycoprotein complex (DGC). δ-sarcoglycan is involved in the stability and signal transduction of the muscle cell membrane and is located on chromosome 5q33-34. It connects cytoplasmic actin, cell membrane and cardiomyocyte matrix angiotensin-converting enzyme through alpha2-laminin. The introduction of δ-sarcoglycan gene through adenovirus can improve the heart function of hamsters with cardiomyopathy.

　　Previous studies have shown that mutations in the sarcomere protein gene mainly cause hypertrophic cardiomyopathy. In recent years, studies have found that patients with dilated cardiomyopathy also have mutations in sarcomere components, including actin, myosin, binding protein, and β-myosin. Protein heavy chain, troponin, tropomyosin and myoglobin, etc. Cardiomyopathy was also observed in tropomodulin overexpression models and transgenic mice expressing Epstein-Barr virus antigen-leader protein or polyoma virus large T-antigen. Tropomodulin is a component of thin filament protein that determines the length of sarcomere-actin filaments. Recently, it was reported that transgenic mice overexpressing tropomodulin developed dilated cardiomyopathy, weakened myocardial contraction and heart failure 2 to 4 weeks after birth.