Mutations in MYH9 encoding nonmuscle myosin heavy chain II-A (NMHC II-A) have been identified a group of disorders distinguished by different combinations of pathologies, including deafness, nephritis, cataracts, large but fewer platelets and Dohle-like leukocyte inclusions. To date more then 20 single amino mutations in MYH9 have been reported in humans. The 98% identity between human and mouse NMHC II-A suggests that mouse can be a useful experimental model to verify the genotype-phenotype connection and to understand the molecular pathogenesis of this syndrome. Such a model may lead to new diagnostic tools and treatments. We have successfully generated mice with a point mutation, D1424N, in exon 31 of the Myh9 gene using homologous recombination with a neomycin cassette inserted in the intron between exons 31 and 32. We are breeding the first generation of heterozygous mice and testing them for the phenotype. Blood test using Giemsa staining will be used for primary evaluation of the presence of macrothrombocytopenia in both homozygous and heterozygous mice compared to wild-type mice. Hearing and vision tests will be performed on the same groups of mice.? ? Dilated cardiomyopathy (DCM) leads to an increase in cardiac size and accumulation of scar tissue (collagen). DCM threatens affected patients with sudden death or progression to overt heart failure. Early treatment may prevent illness and death, but pre-symptomatic detection is limited by the sensitivity and specificity of diagnostic tests. Histological examination of cardiac-myocyte specific NMHC II-B ablated mice (BMHC/BMHC mice) showed cardiac myocyte hypertrophy starting from the first postnatal day, however echocardiography and ECG did not detect compromised cardiac function or abnormal contractions until 6 months of age. Starting at 8 months of age the mice begin dying. This phenotype mimics the asymptomatic human DCM. ? To examine whether the functional cardiac defects can be detected at an earlier age and perhaps predict the appearance and severity of DCM, dobutamine stress tests were performed on BMHC/BMHC mice at age 3.5-4.5 months followed by repeat testing at age 8-9 months. Dobutamine is a beta-adrenergic agent that induces positive inotropy, raising heart rate similar to the effect of exercise on the heart. MRI didn't detect any difference between mutant and wild type mice without dobutamine at age 3.5-4.5 months. However, increased end-diastolic and end-systolic volumes and a reduced ejection fraction in 75% (3 of 4) of the conditional BMHC/BMHC mice compared to 12.5% wild type and heterozygote mice was detected after the administration of dobutamine. ECGs on stress-induced animals showed bradyarythmia and ventricular fibrillations in 50% of the BMHC/BMHC mice. By age 8-9 months all BMHC/BMHC mice and 75% of BFLOX/BMHC mice developed defects in cardiac function shown by MRI and tolerated dobutamine stress significantly less well than BFLOX/BFLOX mice. Higher end diastolic volume was found to be associated with the worst prognosis. Affected mice started to hunch and had breathing problems at an earlier age. Conditional BMHC/BMHC mice had no significant systolic, diastolic and main arterial pressure differences from BFLOX/BFLOX and BFLOX/BMHC mice. Histological analysis at 8-9 months of age of all BMHC/BMHC mice and some BFLOX/BMHC mice showed cardiomyocyte hypertrophy, collagen accumulation, vacuolization and dilated cardiomyopathy. Additionally pulmonary blood vessel hypertrophy was found in cardiac hypertrophic mice. Atrial thrombosis found in some of the mice and may be a result of disrupted, turbulent flow caused by arrhythmic contractions. Electron microscopy findings supported the histology and showed collagen accumulation surrounding nerve cells, which may prevent appropriate signal transduction necessary for contractions. Our results suggest that the dobutamine stress test can be a successful diagnostic tool for early prognosis of non-hypertensive cardiac hypertrophy.
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