Maria Carmen Fernández

The anatomical elements that in humans prevent blood backflow from the aorta and pulmonary artery to the left and right ventriclesare the aortic and pulmonary valves, respectively. Each valve regularly consists of three leaflets (cusps), each supported by its valvular sinus. From the medical viewpoint, each set of three leaflets and sinuses is regarded as a morpho-functional unit. This notion also applies to birds and non-human mammals. However, the structures that prevent the return of blood to the heart in other vertebrates are notably different. This has led to discrepancies between physicians and zoologists in defining what a cardiac outflow tract valve is. The aim here is to compare the gross anatomy of the outflow tract valvular system among several groups of vertebrates in order to understand the conceptual and nomenclature controversies in the field.

The abnormal origin of a coronary artery from the ascending aorta, above the sinotubular junction or high take‐off (HTO) is a rare congenital anomaly, which is associated with sudden cardiac death in approximately 1.5% of cases. HTO has been reported in 62.8% of mice from the C57Bl/6 (B6) strain, while it is absent in the Balb/c (BC) mouse strain. We have observed that the incidences of HTO in hybrid mice are intermediate between those of the parental animals (F1: 26.32%; F1xF1: 20%; F1xB6: 46.34%; F1xBC: 9.80%). In addition, we found that these incidences fit with a mode of inheritance consisting in the influence of a major causal dominant allele, present in the B6 strain, and a modifier dominant allele in the BC strain that reduces by half the incidence of HTO. In order to identify chromosomal regions including genes responsible for HTO development, we have carried out a genetic linkage study through the generation of a congenic strain, using microsatellite genetic markers. A region of 2,699,738 bp in chromosome 18 between the markers MIT‐49 and MIT‐106 was found statistically associated with HTO occurrence. This chromosomal region included a total of 9 genes ( Smad2, Skor2, Ier3pi, Hdhd2, Katnal2, Pias2, St8sia5, Loxhd1, Rnf165 ). Among these, Smad2 was selected as a candidate gene, due to its multiple biological functions related to cardiovascular system development, specifically to aortic root formation and coronariogenesis. The comparative analysis of the Smad2 sequence between B6 and BC mouse strains revealed the presence of an intronic SNP ( Smad2 ^{rs29725537:C>A} or Smad2 ^C>A ) located in the 5’ splicing zone of intron 10‐11. Sequence analysis predicted a possible error in Smad2 10‐11 intron splicing caused by the B6 missense variant ( Smad2 ^C>A ). After genotyping a total of 104 B6xBC F2 to F5 hybrid individuals of the developing congenic strain, by means of PCR‐RFLP using Smad2 specific primers, we found a statistically significant (p Smad2 ^C>A and HTO occurrence. We conclude that HTO is an inherited trait in mice, probably caused by a dominant allele and a dominant modifier allele. Our genotype‐phenotype association study points to the Smad2 ^{rs29725537:C>A} allele as the main causal factor for HTO in mice. Further studies are required to 1) confirm the role of the Smad2 ^C>A SNP in murine HTO development; 2) check whether human HTO development is also influenced by the Smad2 ^C>A allele; and 3) investigate the possible involvement of the Smad2 ^C>A allele in the development of additional traits, particularly those differentially expressed between mice of the B6 and BC strains.

Summary: The concept that anatomical variations in the coronary artery tree might be influenced by genes is relatively old. However, empirical evidence on the effect of genotype on the coronary morphology is still scarce. In the Syrian hamster, there is a septal coronary artery which arises from the left or from the right coronary artery and supplies most of the interventricular septum. The aim was to decide whether the anatomical origin of the septal artery is subject to inheritance factors. Overall, 483 internal casts of the heart and coronary arteries were examined. All the hamsters included in this study had normal coronary arteries. The results of 74 crosses were compared statistically to seek for any significant difference between the phenotypes of the offspring and the phenotypes of the parents. The left septal artery was over‐represented in the offspring of crosses between parents having both a left septal artery (p < .01), while the right septal artery was over‐represented in the offspring of crosses between parents, one with a right and the other with a left septal artery (p < .001), and, more markedly, in the offspring of crosses between parents both with a right septal artery (p < .001). These results are the first to reveal that the coronary artery pattern is influenced by genetic factors, at least in its proximal portion with regard to the aorta.

Bicuspid aortic valve (BAV) is the most prevalent human congenital cardiac malformation. It may appear isolated, associated with other cardiovascular malformations, or forming part of syndromes. Cranial neural crest (NC) defects are supposed to be the cause of the spectrum of disorders associated with syndromic BAV. Experimental studies with an inbred hamster model of isolated BAV showed that alterations in the migration or differentiation of the cardiac NC cells in the embryonic cardiac outflow tract are most probably responsible for the development of this congenital valvular defect. We hypothesize that isolated BAV is not the result of local, but of early alterations in the behavior of the NC cells, thus also affecting other cranial NC-derived structures. Therefore, we tested whether morphological variation of the aortic valve is linked to phenotypic variation of the mandible and the thymus in the hamster model of isolated BAV, compared to a control strain. Our results show significant differences in the size and shape of the mandible as well as in the cellular composition of the thymus between the two strains, and in mandible shape regarding the morphology of the aortic valve. Given that both the mandible and the thymus are cranial NC derivatives, and that the cardiac NC belongs to the cephalic domain, we propose that the causal defect leading to isolated BAV during embryonic development is not restricted to local alterations of the cardiac NC cells in the cardiac outflow tract, but it is of pleiotropic or polytopic nature. Our results suggest that isolated BAV may be the forme fruste of a polytopic syndrome involving the cranial NC in the hamster model and in a proportion of affected patients.

Bicuspid aortic valve (BAV) is the most frequent congenital cardiac malformation in humans, and appears frequently associated with dilatation of the ascending aorta. This association is likely the result of a common aetiology. Currently, a Syrian hamster strain with a relatively high (∼40%) incidence of BAV constitutes the only spontaneous animal model of BAV disease. The characterization of molecular alterations in the aorta of hamsters with BAV may serve to identify pathophysiological mechanisms and molecular markers of disease in humans. In this report, we evaluate the expression of ten candidate reference genes in aortic tissue of hamsters in order to identify housekeeping genes for normalization using quantitative real time PCR (RT-qPCR) assays. A total of 51 adult (180–240 days old) and 56 old (300–440 days old) animals were used. They belonged to a control strain of hamsters with normal, tricuspid aortic valve (TAV; n = 30), or to the affected strain of hamsters with TAV (n = 45) or BAV (n = 32). The expression stability of the candidate reference genes was determined by RT-qPCR using three statistical algorithms, GeNorm, NormFinder and Bestkeeper. The expression analyses showed that the most stable reference genes for the three algorithms employed were Cdkn1β, G3pdh and Polr2a. We propose the use of Cdkn1β, or both Cdkn1β and G3pdh as reference genes for mRNA expression analyses in Syrian hamster aorta.

A previous manuscript [Fernández B, et al. (2008) J Anat 212, 12] reported on the unusual coronary artery patterns in mice belonging to the C57BL/6 strain. The aim here was to elucidate whether this pattern is unique to C57BL/6 mice or appears in other laboratory mouse strains and in wild‐living mice. Stereomicroscopy, scanning electron microscopy, light microscopy and a corrosion cast technique were used to examine 597 adult mice belonging to three inbred strains (C57BL/6, Balb/c, DBA/2), three outbred stocks (CD1, OF1, NMRI) two hybrid lines (129sv × BL/6, CD2F1) and wild mice. It was shown that lock‐like ostium is an exclusive trait of C57BL/6 mice, whereas left septal artery, accessory ostium, high take‐off, intramural course and solitary ostium in aorta are all present in the different laboratory strains and wild mice included in the present study. However, each mouse population shows a specific incidence of these coronary conditions. Several clinically relevant human coronary artery anomalies are present in healthy mice from different strains that may serve as animal models for humans. These results should be taken into consideration in research concerning the murine coronary system, especially in coronary artery occlusion experiments and in studies on cardiovascular developmental biology using murine mutant lines.

During a 4-year period (January 2009–December 2012), the ⁷Be, ²¹⁰Pb, and ⁴⁰K activity concentrations in airborne particulate matter were weekly determined at the Málaga (Spain) located in the southern Iberian Peninsula. Totally 209 polypropylene filters were analyzed in the mentioned period. In 100 % of the filters, ⁷Be and ⁴⁰K activity concentrations were detected while ²¹⁰Pb activity concentration was detected in 96 % of the filters. The results from individual measurements of ⁷Be, ²¹⁰Pb, and ⁴⁰K concentrations were analyzed to derive the statistical estimates characterizing the distributions. Principal components analysis (PCA) was applied to the datasets and the results of the study reveal that aerosol behavior is represented by two principal components which explain 73.2 % of total variance. Components PC1 and PC2 respectively explain 46.0 and 27.2 % of total variance. PC1 was related positively to dust content, ⁷Be and ⁴⁰K concentrations and negatively to sunspot numbers. In contrast, PC2 was related positively to temperature and ²¹⁰Pb activity and negatively to precipitation and relative humidity. The ⁷Be levels showed a significant correlation with sunspot numbers due to the cosmogenic origin. ⁴⁰K activities showed a good correlation with dust deposition in filters mainly because it was transported to the air as resuspended particle from the soil. An inverse relationship was observed between the ²¹⁰Pb concentrations and monthly rainfall, indicating washout of atmospheric aerosols carrying these radionuclides and a pronounced positive correlation with the average monthly temperature of air.