M. Carmen Fernández

Bicuspid aortic valves with different spatial orientations of the leaflets are distinct etiological entities.

OBJECTIVES The aim of this study was to decide whether bicuspid aortic valves (BAVs) with fused right and noncoronary leaflets (R-N) and BAVs with fused right and left leaflets (R-L) have different etiologies or are the product of a single diathesis. BACKGROUND The BAV is the most common congenital cardiac malformation. The R-N and R-L BAVs are the most frequent BAV subtypes. METHODS The study was carried out in adult and embryonic hearts of endothelium nitric oxide synthase knock-out mice and inbred Syrian hamsters with a high incidence of R-N and R-L BAVs, respectively. The techniques used were histochemistry, immunohistochemistry, and scanning electron microscopy. RESULTS The R-N BAVs result from a defective development of the cardiac outflow tract (OT) endocardial cushions that generates a morphologically anomalous right leaflet. The left leaflet develops normally. The R-L BAVs are the outcome of an extrafusion of the septal and parietal OT ridges that thereby engenders a sole anterior leaflet. The noncoronary leaflet forms normally. CONCLUSIONS The R-N and R-L BAVs are different etiological entities. The R-N BAVs are the product of a morphogenetic defect that happens before the OT septation and that probably relies on an exacerbated nitric oxide-dependent epithelial-to-mesenchymal transformation. The R-L BAVs result from the anomalous septation of the proximal portion of the OT, likely caused by a distorted behavior of neural crest cells. Care should be taken in further work on BAV genetics because R-N and R-L BAVs might rely on different genotypes. Detailed screening for R-N and R-L BAVs should be performed for a better understanding of the relationships between these BAV morphologic phenotypes and other heart disease.

Anatomy and Formation of Congenital Bicuspid and Quadricuspid Pulmonary Valves in Syrian Hamsters

Congenital bicuspid and quadricuspid pulmonary valves have received little attention because of their limited clinical relevance. However, knowledge of the mechanisms by which these anomalous valves develop is essential to obtain a more accurate survey of the etiological factors implicated in the malformations of the cardiac outflow tract in mammals. The present study was designed to assess the anatomical features of bicuspid and quadricuspid pulmonary valves in Syrian hamsters as well as to elucidate the mechanisms involved in the formation of these defective valves.

Genetically alike Syrian hamsters display both bifoliate and trifoliate aortic valves

The bifoliate, or bicuspid, aortic valve (BAV) is the most frequent congenital cardiac anomaly in man. It is a heritable defect, but its mode of inheritance remains unclear. Previous studies in Syrian hamsters showed that BAVs with fusion of the right and left coronary leaflets are expressions of a trait, the variation of which takes the form of a phenotypic continuum. It ranges from a trifoliate valve with no fusion of the coronary leaflets to a bifoliate root devoid of any raphe. The intermediate stages are represented by trifoliate valves with fusion of the coronary aortic leaflets, and bifoliate valves with raphes. The aim of this study was to elucidate whether the distinct morphological variants rely on a common genotype, or on different genotypes. We examined the aortic valves from 1 849 Syrian hamsters belonging to a family subjected to systematic inbreeding by full‐sib mating. The incidence of the different trifoliate aortic valve (TAV) and bifoliate aortic valve (BAV) morphological variants widely varied in the successive inbred generations. TAVs with extensive fusion of the leaflets, and BAVs, accounted for five‐sixths of the patterns found in Syrian hamsters considered to be genetically alike or virtually isogenic, with the probability of homozygosity being 0.999 or higher. The remaining one‐sixth hamsters had aortic valves with a tricuspid design, but in most cases the right and left coronary leaflets were slightly fused. Results of crosses between genetically alike hamsters, with the probability of homozygosity being 0.989 or higher, revealed no significant association between the valvar phenotypes in the parents and their offspring. Our findings are consistent with the notion that the BAVs of the Syrian hamster are expressions of a quantitative trait subject to polygenic inheritance. They suggest that the genotype of the virtually isogenic animals produced by systematic inbreeding greatly predisposes to the development of anomalous valves, be they bifoliate, or trifoliate with extensive fusion of the leaflets. We infer that the same underlying genotype may account for the whole range of valvar morphological variants, suggesting that factors other than genetic ones are acting during embryonic life, creating the so‐called intangible variation or developmental noise, and playing an important role in the definitive anatomic configuration of the valve. The clinical implication from our study is that congenital aortic valves with a trifoliate design, but with fusion of coronary aortic leaflets, may harbour the same inherent risks as those already recognised for BAVs with fusion of right and left coronary leaflets.

Osteoglycin deficiency does not affect atherosclerosis in mice

OBJECTIVE The small leucine-rich proteoglycan Osteoglycin/Mimecan (OGN) is a component of the extracellular matrix, where it regulates collagen fibrillogenesis and cytokine availability. OGN is abundant in normal vessels and in atherosclerotic and restenotic lesions of rat, rabbit and human arteries. Osteoglycin-null mice show alterations in the thickness of collagen fibers of the cornea and the skin. In this work, we inspect the possible involvement of OGN in the atherosclerosis progression using a double knockout mouse model. METHODS In order to examine the progression of atherosclerosis in the absence of OGN, we developed double Apoe and Ogn knockout mice and performed a comparative histomorphological and immunofluorescence study of the atherosclerotic lesions of Apoe(-/-)Ogn(-/-) and Apoe(-/-)Ogn(+/+) mice. RESULTS We demonstrate the presence of Ogn transcript in the aorta of wildtype mice, its absence in Ogn(-/-) mice, and the normal histomorphology of arteries of Ogn(-/-) mice. The composition of the extracellular matrix and also the cellular content and distribution were similar in atherosclerotic lesions of Apoe(-/-)Ogn(-/-) and Apoe(-/-)Ogn(+/+) mice. Quantification of the lesion size revealed no significant differences between double and single knockout mice. The incidence, size and distribution of calcium deposits were similar in both groups of mice. CONCLUSIONS The lack of the proteoglycan OGN does not affect the progression of atherosclerosis in mice. Possible causes for the absence of phenotype in the Apoe/Ogn double mutants are discussed.

Genetic contribution of bicuspid aortic valve morphology.

Genetic Contribution of Bicuspid Aortic Valve Morphology Borja Fern andez,* Ana C. Dur an, M. Carmen Fern andez, Josep M. Arqu e, Miguel Such, and Valent ın Sans-Coma Faculty of Science, Department of Animal Biology, University of M alaga, M alaga, Spain Department of Cardiac Surgery, Hospital Quir on, M alaga, Spain Department of Cardiovascular Surgery, University Hospital Virgen de la Victoria, M alaga, Spain

Anatomical, histochemical and immunohistochemical characterisation of the cardiac outflow tract of the silver arowana, Osteoglossum bicirrhosum (Teleostei: Osteoglossiformes)

The cardiac outflow tract of chondrichthyans and actinopterygians is composed of a myocardial conus arteriosus and a non-myocardial bulbus arteriosus. In teleosts, the conus has been subjected to a reduction in size over the evolution in conjunction with the further development of the bulbus. Most studies on the outflow tract of the teleost heart refer to species of modern groups and are mainly devoted to the bulbus. Knowledge on the outflow tract of species belonging to early teleost groups is scarce. The aim here was to characterise the structure of the cardiac outflow tract of the silver arowana, a representative of the ancient teleost clade of the Osteoglossomorpha. The material consisted of hearts from six juvenile animals. The cardiac outflow tract of the silver arowana is composed of a conus, which supports two conal valves, and a bulbus. Both components are lined externally by the epicardium and internally by the endocardium. The conus is immunoreactive to antibodies against myosin heavy chains and is composed of compact myocardium, thus contrasting with the ventricle, which has exclusively trabeculated myocardium. The bulbus is immunoreactive to antibodies against smooth muscle α-actin and mainly consists of elastic fibres and smooth muscle cells, both arranged in three layers, outer, middle and inner. The most remarkable feature of the bulbus is the presence of two prominent longitudinal ridges, dorsal and ventral, at the luminal side, which serve to anchor the commissures of the conal valves. This arrangement has not been described so far in any fish species. Pigment cells, presumably of neural crest origin, are present in the subepicardium of the bulbus and anterior part of the ventricle.

Unusual anatomical origins of the coronary arteries in C57BL/6 mice. Are they strain‐specific?

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.

Long-term data from a small mammal community reveal loss of diversity and potential effects of local climate change

Abstract Climate change affects distribution and persistence of species. However, forecasting species’ responses to these changes requires long-term data series that are often lacking in ecological studies. We used 15 years of small mammal trapping data collected between 1978 and 2015 in 3 areas at Doñana National Park (southwest Spain) to (i) describe changes in species composition and (ii) test the association between local climate conditions and size of small mammal populations. Overall, 5 species were captured: wood mouse Apodemus sylvaticus, algerian mouse Mus spretus, greater white-toothed shrew Crocidura russula, garden dormouse Eliomys quercinus, and black rat Rattus rattus. The temporal pattern in the proportion of captures of each species suggests that the small mammal diversity declined with time. Although the larger species (e.g., E. quercinus), better adapted to colder climate, have disappeared from our trapping records, M. spretus, a small species inhabiting southwest Europe and the Mediterranean coast of Africa, currently is almost the only trapped species. We used 2-level hierarchical models to separate changes in abundance from changes in probability of capture using records of A. sylvaticus in all 3 areas and of M. spretus in 1. We found that heavy rainfall and low temperatures were positively related to abundance of A. sylvaticus, and that the number of extremely hot days was negatively related to abundance of M. spretus. Despite other mechanisms are likely to be involved, our findings support the importance of climate for the distribution and persistence of these species and raise conservation concerns about potential cascading effects in the Doñana ecosystem.

Factors other than genotype account largely for the phenotypic variation of the pulmonary valve in Syrian hamsters

Understanding of the aetiology of congenitally anomalous pulmonary valves remains incomplete. The aim of our study, therefore, was to elucidate the degree to which the phenotypic variation known to exist for the pulmonary valve relies on genotypic variation. Initially, we tested the hypothesis that genetically alike individuals would display similar valvar phenotypes if the phenotypic arrangement depended entirely, or almost entirely, on the genotype. Thus, we examined pulmonary valves from 982 Syrian hamsters belonging to two families subject to systematic inbreeding by crossing siblings. Their coefficient of inbreeding was 0.999 or higher, so they could be considered genetically alike. External environmental factors were standardized as much as possible. A further 97 Syrian hamsters from an outbred colony were used for comparative purposes. In both the inbred and outbred hamsters, we found valves with a purely trifoliate, or tricuspid, design, trifoliate valves with a more or less extensive fusion of the right and left leaflets, bifoliate, or bicuspid, valves with fused right and left leaflets, with or without a raphe located in the conjoined arterial sinus, and quadrifoliate, or quadricuspid, valves. The incidence of the different valvar morphological variants was similar in the outbred and inbred colonies, except for the bifoliate pulmonary valves, which were significantly more frequent in the hamsters from one of the two inbred families. Results of crosses between genetically alike hamsters revealed no significant association between the pulmonary valvar phenotypes as seen in the parents and their offspring. The incidence of bifoliate pulmonary valves, nonetheless, was higher than statistically expected in the offspring of crosses where at least one of the parents possessed a pulmonary valve with two leaflets. Our observations are consistent with the notion that the basic design of the pulmonary valve, in terms of whether it possesses three or two leaflets, relies on genotypic determinants. They also denote that the bifoliate condition of the valve is the consequence of complex inheritance, with reduced penetrance and variable expressivity. Moreover, in showing that the incidence of the bifoliate pulmonary valve significantly differs in two different isogenetic backgrounds, our data suggest that genetic modifiers might be implicated in directing the manifestation of such specific pulmonary valvar malformations. Finally, our findings indicate that factors other than the genotype, operating during embryonic life and creating developmental noise, or random variation, play a crucial role in the overall phenotypic variation involving the pulmonary valve.