Marcia C. Willing

Mutations in Transforming Growth Factor-β Receptor Type II Cause Familial Thoracic Aortic Aneurysms and Dissections

Background—A genetic predisposition for progressive enlargement of thoracic aortic aneurysms leading to type A dissection (TAAD) is inherited in an autosomal-dominant manner in up to 19% of patients, and a number of chromosomal loci have been identified for the condition. Having mapped a TAAD locus to 3p24–25, we sequenced the gene for transforming growth factor-&bgr; receptor type II (TGFBR2) to determine whether mutations in this gene resulted in familial TAAD. Methods and Results—We sequenced all 8 coding exons of TGFBR2 by using genomic DNA from 80 unrelated familial TAAD cases. We found TGFBR2 mutations in 4 unrelated families with familial TAAD who did not have Marfan syndrome. Affected family members also had descending aortic disease and aneurysms of other arteries. Strikingly, all 4 mutations affected an arginine residue at position 460 in the intracellular domain, suggesting a mutation “hot spot” for familial TAAD. Despite identical mutations in the families, assessment of linked polymorphisms suggested that these families were not distantly related. Structural analysis of the TGFBR2 serine/threonine kinase domain revealed that R460 is strategically located within a highly conserved region of this domain and that the amino acid substitutions resulting from these mutations will interfere with the receptor’s ability to transduce signals. Conclusion—Germline TGFBR2 mutations are responsible for the inherited predisposition to familial TAAD in 5% of these cases. Our results have broad implications for understanding the role of TGF-&bgr; signaling in the pathophysiology of TAAD.

Mutations in Smooth Muscle Alpha-Actin (ACTA2) Cause Coronary Artery Disease, Stroke, and Moyamoya Disease, Along with Thoracic Aortic Disease

The vascular smooth muscle cell (SMC)-specific isoform of alpha-actin (ACTA2) is a major component of the contractile apparatus in SMCs located throughout the arterial system. Heterozygous ACTA2 mutations cause familial thoracic aortic aneurysms and dissections (TAAD), but only half of mutation carriers have aortic disease. Linkage analysis and association studies of individuals in 20 families with ACTA2 mutations indicate that mutation carriers can have a diversity of vascular diseases, including premature onset of coronary artery disease (CAD) and premature ischemic strokes (including Moyamoya disease [MMD]), as well as previously defined TAAD. Sequencing of DNA from patients with nonfamilial TAAD and from premature-onset CAD patients independently identified ACTA2 mutations in these patients and premature onset strokes in family members with ACTA2 mutations. Vascular pathology and analysis of explanted SMCs and myofibroblasts from patients harboring ACTA2 suggested that increased proliferation of SMCs contributed to occlusive diseases. These results indicate that heterozygous ACTA2 mutations predispose patients to a variety of diffuse and diverse vascular diseases, including TAAD, premature CAD, ischemic strokes, and MMD. These data demonstrate that diffuse vascular diseases resulting from either occluded or enlarged arteries can be caused by mutations in a single gene and have direct implications for clinical management and research on familial vascular diseases.

Bone Mineral Density and Its Change in White Women: Estrogen and Vitamin D Receptor Genotypes and Their Interaction

Low bone mineral density (BMD) is a major risk factor for development of osteoporosis; increasing evidence suggests that attainment and maintenance of peak bone mass as well as bone turnover and bone loss have strong genetic determinants. We examined the association of BMD levels and their change over a 3‐year period, and polymorphisms of the estrogen receptor (ER), vitamin D receptor (VDR), type I collagen, osteonectin, osteopontin, and osteocalcin genes in pre‐ and perimenopausal women who were part of the Michigan Bone Health Study, a population‐based longitudinal study of BMD. Body composition measurements, reproductive hormone profiles, bone‐related serum protein measurements, and life‐style characteristics were also available on each woman. Based on evaluation of women, ER genotypes (identified by PvuII [n = 253] and XbaI [n = 248]) were significantly predictive of both lumbar spine (p < 0.05) and total body BMD level, but not their change over the 3‐year period examined. The VDR BsmI restriction fragment length polymorphism was not associated with baseline BMD, change in BMD over time, or any of the bone‐related serum and body composition measurements in the 372 women in whom it was evaluated. Likewise, none of the other polymorphic markers was associated with BMD measurements. However, we identified a significant gene × gene interaction effect (p < 0.05) for the VDR locus and PvuII (p < 0.005) and XbaI (p < 0.05) polymorphisms, which impacted BMD levels. Women who had the (−/−) PvuII ER and bb VDR genotype combination had a very high average BMD, while individuals with the (−/−) PvuII ER and BB VDR genotype had significantly lower BMD levels. This contrast was not explained by differences in serum levels of osteocalcin, parathyroid hormone, 1,25‐dihydroxyvitamin D, or 25‐dihydroxyvitamin D. These data suggest that genetic variation at the ER locus, singly and in relation to the vitamin D receptor gene, influences attainment and maintenance of peak bone mass in younger women, which in turn may render some individuals more susceptible to osteoporosis than others.

Osteogenesis imperfecta: translation of mutation to phenotype.

By the end of the recently completed Fourth International Conference on Osteogenesis Imperfecta (Pavia, Italy, 9-12 September 1990) more than 70 mutations in the two genes that encode the chains of type I collagen, the major protein of bone, had been identified as the molecular cause of different forms of osteogenesis imperfecta (OI). Although by no means complete, the set of mutations in hand provides a rough guide to how to predict the phenotypic effects of mutations in type I collagen genes, predicts that certain classes of mutations will give rise to very mild phenotypes that will blend with common disorders, such as osteoporosis, and clarifies the genetic aspects of the widely used clinical classification of O.1

Mapping a Locus for Familial Thoracic Aortic Aneurysms and Dissections (TAAD2) to 3p24–25

Background—Familial thoracic aortic aneurysms and dissections (TAAD) occur as part of known syndromes such as Marfan syndrome but can also be inherited in families in an autosomal dominant manner as an isolated condition. Previous studies have mapped genes causing nonsyndromic familial TAAD to 5q13–15 (TAAD1) and 11q23.2-q24 (FAA1). Further genetic heterogeneity for the condition was evident by the presence of TAAD in some families not linked to these known loci. Methods and Results—A 4-generation family with dominant mode of inheritance of TAAD was studied. Affected status was determined by dilation of the ascending aorta, surgical repair of an aneurysm or dissection, or death as the result of aortic dissection. None of the family members evaluated met the diagnostic criteria for Marfan syndrome. After exclusion of known loci for familial TAAD, a genome-wide scan was carried out to map the defective gene causing the disease in the family. A locus was mapped to a 25-cM region on 3p24–25 with a maximum multipoint logarithm of the odds score of 4.28. Conclusions—A third locus for nonsyndromic TAAD was mapped to 3p24–25 and termed the TAAD2 locus. This locus overlaps a previously mapped second locus for Marfan syndrome, termed the MFS2 locus. Future characterization of the TAAD2 gene will determine if TAAD2 is allelic to MFS2. In addition, identification of the TAAD2 gene will improve the presymptomatic diagnosis of individuals with this life-threatening genetic syndrome and provide information concerning the pathogenesis of the disease.

Reduced penetrance and variable expressivity of familial thoracic aortic aneurysms/dissections

Autosomal dominant inheritance of thoracic aortic aneurysms and dissections occurs in subjects with Marfan syndrome, which results from mutations in the FBN1 gene on chromosome 15. A second chromosomal locus on 3p24-25 has been identified for a Marfan-like condition with thoracic aortic aneurysms. We describe here 6 families with multiple members with thoracic aortic aneurysms and dissections in the absence of the ocular and skeletal complications of Marfan syndrome. Medical records and autopsy reports on affected subjects in families with multiple members with thoracic aortic aneurysms and dissections were reviewed. Subjects in these families at risk for developing aortic disease underwent echocardiography to evaluate the aorta. The pattern of inheritance of thoracic aortic aneurysms and dissections was autosomal dominant in these families. Most affected subjects presented with aortic root dilatation or acute type I dissection, but the age of onset of disease was variable and there was decreased penetrance of the disorder. In 2 of the families, the syndrome was not linked to FBN1 or 3p24-25. Familial thoracic aortic aneurysm and dissection is an autosomal dominant condition with marked variability in the age of onset of aortic disease and decreased penetrance, making identification of affected subjects difficult. This condition is not due to mutations in the FBN1 gene or the unidentified gene on 3p24-25.

Analysis of multigenerational families with thoracic aortic aneurysms and dissections due to TGFBR1 or TGFBR2 mutations

Background: Mutations in the transforming growth factor β receptor type I and II genes (TGFBR1 and TGFBR2) cause Loeys–Dietz syndrome (LDS), characterised by thoracic aortic aneurysms and dissections (TAAD), aneurysms and dissections of other arteries, craniosynostosis, cleft palate/bifid uvula, hypertelorism, congenital heart defects, arterial tortuosity, and mental retardation. TGFBR2 mutations can also cause TAAD in the absence of features of LDS in large multigenerational families, yet only sporadic LDS cases or parent–child pairs with TGFBR1 mutations have been reported to date. Methods: The authors identified TGFBR1 missense mutations in multigenerational families with TAAD by DNA sequencing. Clinical features of affected individuals were assessed and compared with clinical features of previously described TGFBR2 families. Results: Statistical analyses of the clinical features of the TGFBR1 cohort (n = 30) were compared with clinical features of TGFBR2 cohort (n = 77). Significant differences were identified in clinical presentation and survival based on gender in TGFBR1 families but not in TGFBR2 families. In families with TGFBR1 mutations, men died younger than women based on Kaplan–Meier survival curves. In addition, men presented with TAAD and women often presented with dissections and aneurysms of arteries other than the ascending thoracic aorta. The data also suggest that individuals with TGFBR2 mutations are more likely to dissect at aortic diameters <5.0 cm than individuals with TGFBR1 mutations. Conclusion: This study is the first to demonstrate clinical differences between patients with TGFBR1 and TGFBR2 mutations. These differences are important for the clinical management and outcome of vascular diseases in these patients.

Sustained Effect of Early Physical Activity on Body Fat Mass in Older Children

BACKGROUND Physical activity is assumed to reduce excessive fatness in children. This study examined whether the benefits of early childhood moderate-to-vigorous physical activity (MVPA) on fatness are sustained throughout childhood. METHODS MVPA minutes per day (min/d) and fat mass (kilograms; kg) were measured using accelerometry and dual-energy x-ray absorptiometry in 333 children aged 5, 8, and 11 years who were participating in the Iowa Bone Development Study. Mixed regression models were used to test whether MVPA at age 5 years had an effect on fat mass at age 8 years and age 11 years, after adjustment for concurrent height, weight, age, maturity, and MVPA. The analysis was repeated to control for fat mass at age 5 years. Using mixed-model least-squares means, adjusted means of fat mass at age 8 years and age 11 years were compared between the highest and lowest quartiles of MVPA at age 5 years. Data were collected between 1998 and 2006 and analyzed in 2008. RESULTS For boys and girls, MVPA at age 5 years was a predictor of adjusted fat mass at age 8 years and age 11 years (p<0.05). In girls, the effect of MVPA at age 5 years was not significant when fat mass at age 5 years was included. Boys and girls in the highest quartile of MVPA at age 5 years had a lower fat mass at age 8 years and age 11 years than children in the lowest MVPA quartile at age 5 years (p<0.05; mean difference 0.85 kg at age 8 years and 1.55 kg at age 11 years). CONCLUSIONS Some effects of early-childhood MVPA on fatness appear to persist throughout childhood. Results indicate the potential importance of increasing MVPA in young children as a strategy to reduce later fat gains.

Genetic Markers, Bone Mineral Density, and Serum Osteocalcin Levels

We evaluated five genetic markers for products that contribute to skeletal mineralization including the Sp1 polymorphism for type I collagen Ai (COLIA1), the vitamin D receptor (VDR) translation initiation site polymorphism, the promoter of the osteocalcin gene containing a C/T polymorphism, the estrogen receptor (ER) gene containing a TA repeat, and the polymorphic (AGC)n site in the androgen receptor. These markers were evaluated for their potential relationship with bone mineral density (BMD), measured by dual‐energy X‐ray densitometry, or its 3‐year change. Additionally, potential associations of these genotypes and with baseline osteocalcin concentration or its 3‐year change (assessed using radioimmunoassay) were evaluated. The study was conducted in 261 pre‐ and perimenopausal women of the Michigan Bone Health Study, a population‐based longitudinal study of musculoskeletal characteristics and diseases. The polymorphic (AGC)n site in the androgen receptor showed a strong association with BMD of the femoral neck (FN) and lumbar spine and remained highly significant after adjusting for body mass index (BMI), oophorectomy/hysterectomy, oral contraceptive (OC) use and hormone replacement use (p < 0.001). The TA repeat at the 5′ end of the ER gene was associated with total body calcium (p < 0.05) after adjusting for BMI, oophorectomy and hysterectomy, and OC use. The frequency of oophorectomy and hysterectomy within selected genotypes explained much of the statistically significant association of the ER genotypes with BMD of the FN and spine. There was no association of measures of BMD or bone turnover with the Sp1 polymorphism for COLIA1, the VDR translation initiation site polymorphism, or the C/T promoter polymorphism of the osteocalcin gene. These findings suggest that sex hormone genes may be important contributors to the variation in BMD among pre‐ and perimenopausal women.

Early Physical Activity Provides Sustained Bone Health Benefits Later in Childhood

PURPOSE This study examined the potential effect of early childhood moderate and vigorous physical activity (MVPA) on later bone health. METHODS Three hundred and thirty-three children, participating in the Iowa Bone Development Study, were studied at ages 5, 8, and 11 yr. MVPA (min x d(-1)) was measured using an accelerometry-based physical activity monitor. Bone mineral content (BMC; g) of the whole body, lumbar spine, and hip was measured using dual-energy x-ray absorptiometry. Mixed regression models were used to test whether MVPA at age 5 yr had an effect on BMC at ages 8 and 11 yr after adjustment for concurrent height, weight, age, maturity, and MVPA. The analysis was repeated to control for bone outcomes at age 5 yr. Mixed-model least-squares mean values at the person level of covariates for age group were used to compare the BMC at ages 8 and 11 yr of children in the highest and lowest quartiles of MVPA at age 5 yr. RESULTS For boys and girls, MVPA at age 5 yr predicted BMC adjusted for concurrent height, weight, age, maturity, and MVPA at ages 8 and 11 yr (P < 0.05). When the analysis was repeated to also control for BMC at age 5 yr, the effect of MVPA at age 5 yr was significant for boys but not for girls. Boys and girls in the highest quartile of MVPA at age 5 yr had 4%-14% more BMC at ages 8 and 11 yr than those in the lowest quartile of MVPA at age 5 yr (P < 0.05). CONCLUSIONS These results provide support for the benefits of early MVPA on sustained bone health during childhood especially for boys. Results indicate the importance of increasing MVPA as a strategy to improve BMC later in childhood.