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PubMed1997

Distribution of fibronectin, type I collagen, type IV collagen, and laminin in the cardiac jelly of the mouse embryonic heart with retinoic acid-induced complete transposition of the great arteries.

BACKGROUND: In the mouse model of complete transposition of the great arteries (TGA) produced by all-trans retinoic acid (RA), parietal and septal ridges in the outflow tract (OT) are hypoplastic. At first, these ridges are generated by an expanded cardiac jelly (mainly myocardial basement membrane). Thereafter, endothelial cells delaminate and invade into the adjacent cardiac jelly to form endocardial cushion tissue (formation of cushion ridge). During cushion tissue formation, basement membrane antigens play an important role in the regulation of this endothelial-mesenchymal transformation. METHODS: To examine whether the myocardial basement membrane components are altered in the RA-treated heart OT, immunohistochemistry for fibronectin, type I collagen, type IV collagen, and laminin was carried out in mouse embryonic hearts at 9.5 and 10.5 ED (embryonic day; vaginal plug = day 0) with or without prior exposure to RA. RESULTS: Particulate/fibrillar fibronectin and fibrillar type I collagen were observed in the thick cardiac jelly of the control heart at the onset of mesenchymal formation. In the RA-treated heart, an intermittent patchy staining for fibronectin and a sparse distribution of type I collagen were observed in the thin cardiac jelly. Laminin and type IV collagen were distributed continuously on the basal surface (layer adjacent to the basal plasma membrane) of endocardium and myocardium in both control and RA-treated hearts. CONCLUSIONS: The alterations in the antigens of the myocardial basement membrane (cardiac jelly) may be responsible for the hypoplasticity of parietal and septal ridges that characterizes RA-induced TGA morphology. This may be one of the reasons why mesenchymal cell formation is inhibited in the RA-induced TGA.

Animals↗
PubMed1996

Multiple fetal intracardiac echogenic foci: not always a benign sonographic finding.

Isolated left-sided echogenic foci in the fetal heart are considered as a benign condition, probably representing a normal variant of the development of papillary muscles. The objective of the present study was to evaluate the incidence and significance of multiple or diffuse echogenic foci in the fetal heart. We analysed retrospectively 25 725 ultrasound examinations conducted for fetal malformations between 12 and 24 weeks' gestation. In the study group, echogenic intracardiac foci were observed in 44 cases (0.17 per cent). In 35 fetuses, these foci were confined to the region of the papillary muscles/chordae tendinae as an isolated finding in the left side of the heart. All these fetuses had an uneventful neonatal follow-up. However, in nine of these cases, diffuse echogenic foci were demonstrated in various regions of the fetal heart, five of them with involvement of the right ventricle. In five of the nine cases, other major pathologies were found. In one case, a missed abortion occurred and in one case of early termination of pregnancy, calcifications were demonstrated in the endocardium on histological examination. Only in two of the nine fetuses was the outcome uneventful. Our findings suggest that diffuse echogenicity in the fetal heart, especially when the right ventricle is also involved, may signal a poor prognosis and deserves a further search for associated pathologies. This is in contrast to the benign character of an isolated left-sided echogenic focus.

Adult↗
PubMed1999

Bone morphogenetic protein-2 acts synergistically with transforming growth factor-beta3 during endothelial-mesenchymal transformation in the developing chick heart.

In the early embryonic heart, endothelial cells in atrioventricular (AV) and outflow tract (OT) regions are transformed into the invasive mesenchymal cells that form endocardial cushion tissue (endothelial-mesenchymal transformation). It has been reported that bone morphogenetic proteins (BMPs) are transcribed in the AV and OT regions of the embryonic mouse heart. We previously reported that transforming growth factor beta 3 (TGFbeta3) triggers the initial phenotypic changes seen in endothelial-mesenchymal transformation. We cloned BMP2 from embryonic chick hearts and examined its functional role during endocardial cushion tissue formation. In situ hybridization showed BMP2 transcripts in the myocardium of the AV and OT regions, but not in endothelial/mesenchymal cells. Antisense oligodeoxynucleotides to BMP2 inhibited mesenchyme formation in AV endocardium cocultured with associated myocardium. This inhibitory effect was reversed by the addition of recombinant BMP2. In cultured AV endothelial monolayers, recombinant BMP2 did not induce any cellular phenotypic changes characteristic of endothelial-mesenchymal transformation. However, BMP2 enhanced the TGFbeta-induced initial phenotypic changes associated with endothelial-mesenchymal transformation. These results suggest that BMP2 1) plays an important role in the formation of endocardial cushion tissue and 2) acts synergistically with TGFbeta3 in the regulation of this developmental event.

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PubMed1998

Common role for each of the cGATA-4/5/6 genes in the regulation of cardiac morphogenesis.

The GATA-4/5/6 genes encode transcription factors implicated previously in the regulation of cardiac-specific differentiation programs. However, recent analyses of mouse GATA-4 null mutations found evidence for function in endoderm development (in vitro) and embryonic morphogenesis (in vivo). Whether each of the three cardiac-associated GATA factors function within distinct or common developmental programs was previously untested; past studies defined specific and distinct roles for each of the GATA-1/2/3 genes in embryonic hematopoiesis. In this study, we compare the transcript patterns of cGATA-4/5/6 during chick embryogenesis. Each of the three GATA factors is expressed in a similar pattern within gastrulating cells of the primitive streak, prior to determination of the cardiomyocyte progenitors, and later within the lateral plate mesoderm and associated endoderm layer. The patterns overlap but extend beyond the presumptive cardiomyocyte population expressing cNkx-2.5. Later in development, cGATA-4/5/6 are all transcribed throughout the differentiating heart, in similar but not identical patterns, within the endocardium, myocardium, and great vessels. In order to test the function of GATA factors during chick cardiogenesis, embryos were cultured in vitro in the presence of antisense oligomers designed to deplete specifically transcripts encoding cGATA-4/5/6, beginning around stage 7. When oligomers are used to target transcripts for all three genes, a high percentage of the embryos develop abnormal hearts related to the failure to form a normal primitive heart tube. In the most severe phenotype, cardiac bifida results in two bilateral beating hearts. In some embryos, the paired heart primordia undergo partial fusion but fail to form a single looping heart tube. In all cases, cellular differentiation is not obviously affected, as the abnormal hearts form beating tissue. Depletion of transcripts encoding any single GATA factor, or any combination of two GATA factors, does not affect development. The partial depletion of all three genes in chick results in a remarkably similar phenotype compared to the null GATA-4 mutation in mouse. Therefore, in the chick, each of the GATA-4/5/6 genes functions in a common pathway, at the time of cardiac crescent formation, for regulating early embryonic cardiac morphogenesis, apparently associated with embryonic folding or the migration of primordia to form a primitive tube.

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PubMed1999

An efficient MR phosphorous spectroscopic localization technique for studying ischemic heart.

To obtain the spatially resolved (31)P spectroscopic image from myocardium during an acute myocardium ischemia at a high signal-to-noise ratio (SNR) in a very limited time window, we have exploited the spatial variation of the radiofrequency (RF) field produced by a single loop transmit/receive (TR) RF coil along its axis for spatial discrimination. By incrementally lengthening the duration of a square RF excitation pulse, the positional information can be systematically encoded as harmonics of various orders in MR signal. In the in vivo open-chest animal experiment, this RF coil was surgically sutured onto the epicardial surface of the left ventricular (LV) wall over the region perfused by the left anterior descending coronary artery. Using only 17 encoding steps, we have obtained one-dimensional (31)P spectroscopic images from both a multiple-layer phosphor phantom and an in vivo LV myocardium. In the animal study, the cardiac gating is used with respiratory synchronization. The MR data were only collected during the end diastole phase of the cardiac cycle (cardiac and respiratory synchronized) with an effective sequence repetition time (TR) of 6 seconds (to ensure the complete relaxation of the phosphorous magnetization). The total acquisition time for a complete experiment is about 10 minutes. Prior to the CSI reconstruction process, the raw data matrix was zero-filled in the spatial dimension. The spatially resolved metabolite map exhibited all the metabolite peaks including creatine phosphate and adenosine triphosphate. At the layer of endocardium, two peaks corresponding to 2, 3-diphosphoglycerate, which is contained in the erythrocytes, were clearly seen in the LV wall. Also, the method allows compensation in both volume and coil sensitivity variations for the resulting spectra. All results have demonstrated that it is an efficient nuclear magnetic resonance method capable of obtaining high-quality (31)P spectroscopic images with both excellent spatial localization and SNR in the research of cardiac ischemia. J. Magn. Reson. Imaging 1999;10:892-898.

Animals↗
PubMed2000

A multicenter validation of an active contour-based left ventricular analysis technique.

Quantitative analysis of functional cardiac magnetic resonance (MR) images has been limited by the lack of well-validated, semiautomatic, methods for rapid analysis. We describe the evaluation of a DICOM-compatible PC-based parallel-processing tool, for cardiac magnetic resonance analysis (CAMRA), which supports semiautomatic image mensuration using an active contour model-based algorithm. The CAMRA software was used to analyze data from 12 patients in a multicenter acquisition and analysis trial to compare semiautomatic contour detection with manual planimetry of the left ventricular endocardium from short-axis, breath-held, cine gradient-echo images. There was excellent agreement between the manual and semiautomatic measurements of global left ventricular function, with no significant (P = 0.32) difference in the determination of ejection fraction (-0.9 +/- 3.1% [mean difference +/- 1 standard deviation]). There was no significant interobserver difference in the semiautomatically measured ejection fraction. Additionally, a single observer completed the analysis on data from 30 patients and found no significant (P = 0.05) difference in the determination of ejection fraction (-1.3 +/- 3.5% [mean difference +/- 1 standard deviation]). The CAMRA software demonstrates the capability for the reproducible evaluation of global left ventricular function in cardiac patients, with adequate interobserver reproducibility for use in multicenter trials.

Adult↗
PubMed1975

Scanning electron microscopy of cardiac endothelium of the dog.

Tissue obtained from young dogs was fixed in buffered aldehydes by vascular perfusion or direct immersion. Selected hearts were maintained in modified mammalian Ringer's solution for three to five minutes prior to fixation. The chambers of the heart and related valves were exposed by dissection and prepared by routine techniques for observation by scanning electron microscopy (SEM). Subsequent to SEM studies, selected specimens were embedded in Epon 812 and sectioned for transmission electron microscopy (TEM). The cardiac endothelium, when fixed immediately in buffered aldehydes, presents an essentially invariable surface throughout the interior of the heart. The predominant nuclear bulges and attenuated peripheral plasmatemma are consistently smooth, with occasional marginal ruffles, scattered microvilli and small blebs. Apart from the higher population of nuclear bulges on valvular surfaces, local variations in SEM of endocardium occur in response to the various stages of systole and diastole encountered. These physiological changes do not produce microappendages. Immersion for three to five minutes in (Chenoweth's) Ringer's solution, prior to fixation, produces a substantial population of microappendages. The cellular surface acquires a swirled appearance erupting in microvilli, blebs and ruffles. These exhibit considerable pleomorphism. There is great lability of the endocardial surface in response to a classic "holding solution" widely used in preparatory techniques. In preparing soft tissues for SEM caution must be used if physiological "holding solutions" are used prior to fixation.

Animals↗
PubMed1979

Morphogenesis of the truncus arteriosus of the chick embryo heart: the formation and migration of mesenchymal tissue.

The appearance and migration of mesenchymal cushion tissue within the truncus arteriosus of the normal 2.5 to 6-day chick embryo heart was surveyed systemically with the light microscope. Series of cross-sections taken from replicate hearts at successive developmental stages allowed comparison of the following qualitative and quantitative aspects of early truncal morphogenesis. Mesenchyme within the truncus was derived from two distinct sources. The first mesenchyme appeared to migrate caudally into the cardiac jelly of the distal truncus from the nearby aortic arch region, coincident with slowing of the anterior elongation of the heart tube (Hamburger-Hamilton Stage 17-18). A second, separate mesenchymal population, derived from endocardium, began to fill the conus and proximal truncus in a radial direction, coicident with expansion of the bulbs cordis (Stage 12-19). The measured kinetics of relative cell numbers, distributions, and mitotic indices suggest substantial contributions from both sources. By Stage 26, the conotruncal region was filled with mesenchyme, which then condensed to form the anlagen of three future structures: the semilunar valves, the aorticopulmonary septum, and the tunica media of the great arteries.

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PubMed1979

Morphogenesis of the truncus arteriosus of the chick embryo heart: tissue reorganization during septation.

Septation of the truncus arteriosus of the normal chick embryo heart was surveyed systematically with the light microscope. Tissue from replicate samples at successive periods of development was sectioned within an arbitrary coordinate system based on positional reference points along the external surface of the heart. Correlation of several aspects of tissue morphology within this spatial and temporal reference-frame yielded a new description of tissue associations and kinetics during septation. A stable complex of tissue structures appeared in the downstream, distal truncus at Stage 25 and persisted throughout the septation process. This complex consisted of (1) the cephalic margin of the myocardial sheath, and (2) the adjacent bifurcation of the vascular lumen, linked together by (3) the newly condensed Y-shaped strap of cells forming the aorticopulmonary septum. The apparent motion of this septation-complex toward the ventricle(s), the appearance within the thoracic cavity of the adjacent segments of the aortic arches, and measures of tissue length and width suggested that septation was accompanied, and perhaps initiated, by increased tension along the truncus. The truncal ridges remained upstream from the complex, with mesenchymal condensations beneath the endocardium differentiating into the definitive semilunar valves. Downstream from the bifurcation, mesenchyme in the aortic arch region condensed around the separate lumens to form the smooth muscular tunica media of the great arteries. The epicardium developed in a caudocephalic direction along the heart tube. Vagal innervation approached the heart cephalocaudally. Capillaries formed along the dividing truncus in both directions. Autoradiography following 3H-thymidine labelling demonstrated reduced DNA synthetic activity in the cephalic margin of the myocardium and aorticopulmonary septum, compared with the associated loose mesenchyme.

Animals↗
PubMed1984

Anatomy and vasculature of a minke whale heart.

The heart from a 4-m-long minke whale (Balaenoptera acutorostrata) was studied to determine the details of its anatomy and to consider cardiac adaptations to diving. The volume fraction (Vvc) of capillaries in the wall of the left ventricle was determined at different levels from base to apex and at different depths from epi- to endocardium using a light microscopic stereologic technique. Typical of cetaceans, this minke whale heart was distinctly flattened dorsoventrally. A moderator band, characteristic of ungulate hearts, spanned the right ventricle. The right and left atrioventricular valves were tricuspid and bicuspid, respectively. The right coronary artery supplied the dorsal and right lateral myocardium. The left coronary artery supplied the ventral and left lateral myocardium. An anastomosis between the dorsal and ventral interventricular arteries occurred in the dorsal interventricular groove. Stereologically, a decreasing transmural gradient in Vvc was identified between the epicardium and the subepicardium at 15 cm from the apex. Our results, however, did not reveal any significant deviations in the pattern of capillary distribution in the wall of the left ventricle between this baleen whale and terrestrial mammals. Measurements of the heart, great vessels, coronary vasculature, and ventricular walls are also given, and they suggest a physiologic and adaptive right ventricular hypertrophy. Based on these and other observations, we propose that the relatively great thickness of the right ventricle and the distinctive shape of the cetacean heart are adaptations to the hemodynamic changes and collapse of the thorax associated with apneic diving.

Animals↗
PubMed1989

Lectin histochemistry of the embryonic heart: expression of terminal and penultimate galactose residues in developing rats and chicks.

Rat embryos at days 10-18 of gestation and chicken embryos at days 3-6 of incubation were fixed and processed for lectin histochemistry. The distribution of binding sites for a lectin from the peanut Arachis hypogaea (PNA) conjugated to horseradish peroxidase (HRP) was determined on tissue sections both before and after enzymatic cleavage of sialic acid with neuraminidase (sialidase). Endocardial cushion tissue in the rat, but not in the chick, reacted with PNA-HRP prior to digestion with sialidase. Endocardium of both species (12 and 13 days in rat, 5 and 6 days in chick), particularly at the level of endocardial cushions, reacted strongly with the sialidase-PNA sequence; this staining decreased markedly after day 14 of gestation in the rat. PNA binding sites capped by sialic acid were most abundant in the developing rat heart during the critical period of endocardial cushion formation and decreased as development proceeded. The marked changes in the appearance and distribution of cardiac cell and tissue glycoconjugates during cardiogenesis support the concept that rapid changes occur in the structure of complex carbohydrates during embryonic and fetal development. The findings also suggest that such glycosylation-related events may be species specific.

Animals↗
PubMed1992

Urokinase activity in the developing avian heart: a spatial and temporal analysis.

Early events in cardiac morphogenesis are characterized by cell migrations and extensive tissue remodeling. This study was undertaken to determine the levels of urokinase in specific regions of the avian heart during early stages of development. Urokinase has previously been shown to be involved in both cell migration and matrix turnover. Elevated urokinase activity and mRNA levels were associated with the onset of ventricular trabeculation and mesenchymal cell migration in the endocardial cushion tissues. Urokinase was localized by immunostaining to the endocardial and mesenchymal cells of the developing atrioventricular canal (AVC) and outflow tract (OFT) as well as with evaginating ventricular endocardium. No immunoreactivity was seen associated directly with the matrix, suggesting that the enzyme remained mostly cell associated, a finding which was confirmed in isolated endocardial cells. Results from this study suggest a role for urokinase in the tissue remodeling and cell migration that occurs during the early stages of cardiac morphogenesis.

Animals↗
PubMed1994

Elastic extracellular matrix of the embryonic chick heart: an immunohistological study using laser confocal microscopy.

The "elastic matrix" constitutes a specialized component of the extracellular matrix which confers resiliency to tissues and organs subjected to repeated deformations. The role of the elastic matrix in living organisms appears to be of key importance since diseases characterized by expression of defective inherited genes which encode components of the elastic matrix lead to premature death. While the elastic matrix of adult organs has received a great deal of attention, little is known about when it first appears in embryonic tissues or its possible role in developing organs. In the present study we have performed an immunohistochemical study of the distribution of elastin and three additional components often associated with elastic matrices in adult tissues (i.e., fibrillin, emilin, and type VI collagen) during the development of the chicken embryonic heart. The three-dimensional arrangement of these components was established through the observation of whole-amount specimens with scanning laser confocal microscopy. Our results revealed three different periods of heart development regarding the composition of the elastic matrix. Prior to stage 21 the embryonic heart lacks elastin but exhibits a matrix scaffold of fibrillin and emilin associated with the endocardium and the developing cardiac jelly. Between stages 22 and 29 the heart shows a transient elastic scaffold in the outflow tract which contains elastin, fibrillin, and emilin. Elastin-positive fibrillar material is also observed during these stages in the base of the atrioventricular cushion adjacent to the myocardial wall. In addition, emilin-positive material appears to be associated with the zones of formation of ventricular trabeculae. Collagen type VI was not detected during these early stages. From stage 30 to stage 40 a progressive modification of the pattern of distribution of elastin, fibrillin, emilin, and collagen type VI is observed in association with the formation of the definitive four-chambered heart. The distribution of the elastic scaffold in the outflow tract appears to be rearranged and becomes restricted to the roots of the main arteries. Each of the components studied here is also deposited at increasing levels in the developing valvular apparatus including the valve leaflets and the chordae tendinea. The components are also present in the subendocardial space where they form aligned fibrillar tracts, an arrangement suggestive of a role in ventricular contractile function. The epicardium constitutes an additional region of elastic matrix deposition during these later stages and contains elastic, fibrillin, and collagen type VI.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1984

Morphogenesis of chordae tendineae. I: Scanning electron microscopy.

The formation of the chordae tendineae of the left atrioventricular valve in the chick embryo is described using scanning electron microscopy. These supportive structures for the valve cusps develop between days 6 and 13 of incubation. Elevations which represent the primitive papillary muscles form on the ventricular wall. These elevations bifurcate into thin, web-like folds which are attached to the primitive valve cusps. The folds are the primordia of the chordae tendineae. Linear ridges develop on the web between the cusp and papillary muscle. These ridges alternate with depressions. The depressions become perforate to create the individual chorda from the linear ridges. Multiple perforations form initially but they typically consolidate to create one large aperture between two chordae. Some interchordal connections of tissue do persist throughout the period studied. During the period of perforation, prominent rounded cells are typical of the endocardium between the chordae. These cells are similar at the scanning electron microscope level to those present in the formation of the foramina secunda of the atrial septum. Primary, secondary, and tertiary chordae tendineae appear to develop in the same manner. First order chordae (those attached at the free margin of a cusp) are not found in the chick embryo. The majority of the chordae are second order, which insert into the ventricular surface of the cusp a short distance from the free edge. These chordae typically have a horizontal banding or grooving along their length. Third order chordae which extend from the papillary muscle to the ventricular wall are also present. It is suggested that chordal development is a programmed cellular and hemodynamic event.

Animals↗
PubMed1993

Early formation of the vascular system in quail embryos.

The relation between vascular development and translocation of the splanchnic mesodermal layers was studied in presomite to 20-somite quail embryos by scanning electron microscopy. In addition, serially sectioned embryos were stained immunohistochemically with monoclonal antibodies (alpha QH1 or alpha MB1) specific for endothelial and hemopoietic cells. By the formation of the foregut the anterior borders of the two splanchnic mesodermal layers of a presomite embryo are translocated to the lateral and ventral sides of the foregut and fuse in the ventral midline of a 4-somite embryo. Meanwhile the splanchnic mesoderm differentiates into a splanchnic mesothelial layer and a plexus of endothelial cells, facing the endoderm. From 4 somites onward the foregut is covered by a single endothelial plexus. At first the endothelial precursors bordering the anterior intestinal portal and those in the area of the ventral mesocardium lumenize, subsequently giving rise to the endocardium of the heart tube. Hereafter, the pharyngeal arch arteries and the dorsal aortae develop from the remaining precursors. During formation of the pharyngeal arches, the pharyngeal arch arteries maintain their connections with the splanchnic plexus through the developing ventral pharyngeal veins. After disappearance of the dorsal mesocardium, the midpharyngeal endothelial strand, which is a longitudinal strand of proendocardial cells, remains connected to the foregut. This strand will contribute to the formation of the pulmonary venous drainage into the left atrium. A bilateral accumulation of cardiac jelly developing between the promyocardium and proendocardial plexus only suggests that the heart develops from two tubes. The proendocardial layer, however, is not divided by the ventral mesocardium but initially forms just one endocardial heart tube.

Animals↗
PubMed1994

Early development of macrophages in intact and organ cultured hearts of rat embryos.

BACKGROUND: Macrophage precursors are present early in embryonic life, being demonstrable in placental and embryonic connective tissues of rats at the neurula stage and as potential macrophages in the brain, liver, and lungs near the onset of organogenesis. We examined the development of macrophages in the heart and the possibility that they initially appear at sites of programmed cell death (apoptosis). METHODS: Precursors were recognized by the binding of peroxidase-coupled Griffonia simplicifolia isolectin B4 (GSA) on the cell membrane. Their capacity for conversion into macrophages was assayed in organ cultures; confirmation of the progeny as bona fide macrophages was obtained from their responses to particle exposure and macrophage colony-stimulating factor (M-CSF). RESULTS: GSA+cells were first seen on gestational day 12 (4 mm embryos) as 2-3 cycling, nonvacuolated cells located in cardiac tissue outside the blood vessels. This population increased to approximately 12 cells by day 14 (9 mm embryos). Two-thirds were distributed along the bulbus cordis in the jellylike endocardium and a more densely cellular connective tissue closer to the aortic arches where apoptotic sites are expected to develop. Such sites were not found in serial glycol methacrylate sections through our 14-day specimens, although in whole heart explants of this age an area of necrosis developed along the prospective line of bulbar endocardial fusion on the second day of organ culturing, and by then macrophages were fairly abundant. Organ culturing of 13-day embryonic hearts also yielded large, highly vacuolated, GSA+mononuclear phagocytes. After a few days in culture most of the macrophages migrated onto the medium where they formed a tight corona of cells about the explants. They readily ingested iron oxide particles and concentrated supravitally administered neutral red in their vacuoles. Macrophages from 14-day cultures exposed to M-CSF developed significantly larger coronas than macrophages from explants grown in serum-rich control medium (p < 0.001). In the presence of cytokines, moreover, these cardiac macrophages survived as many as 100 (92 "postnatal") days. CONCLUSIONS: Macrophage precursors first appear in embryonic rat hearts well before they are needed to clear debris generated by programmed cell death and are capable of rapid conversion into outright phagocytic cells as early as the 13th prenatal day.

Animals↗
PubMed1995

Developmental expression of connexins in the chick embryo myocardium and other tissues.

BACKGROUND: Connexins are cell surface proteins that form specialized regions of cell-cell communication called gap junctions. These allow impulse conduction in in voluntary muscle tissue such as the heart, but also allow the formation of communities of like cells during development of organs. METHODS: We used an antipeptide antibody to connexin 43 in immunolocalization studies and an anti-peptide antibody to an external loop domain common to most connexins in Western blotting of total heart protein to measure the accumulation of connexins in the heart as it develops from 33 hours to 21 days (hatching), and in the adult. RESULTS: Immunolocalization revealed that connexin 43 is widely distributed in the earliest organ rudiments. It is especially prominent in the neural tube and its derivatives, in the lens and nasal placodes, in the foregut and its derivatives, in the somites, in the mesonephric tubules, and in the heart and major arteries. Heart tissue staining grew more intense with development through day 8. However, at day 11 and day 15, and in the adult, heart staining diminished. Endocardium and valve tissue did not stain. Western blotting of heart homogenates with the antibody directed against the external loop domain peptide showed 26, 32, 43, 45, and 56 kilodalton connexins, which changed in relative abundance, displaying unique patterns during development. CONCLUSIONS: Our results show patterns of connexin immunolocalization in early germ layers and organ rudiments that are similar to those known in the mouse, but with certain differences. Our results show a distinctive pattern of multiple connexin gene expression in the developing heart from days 2-21.

Animals↗
PubMed2003

Neurocan in the embryonic avian heart and vasculature.

The chondroitin sulfate proteoglycan (CSPG) neurocan was previously considered to be nervous-system specific. However, we have found neurocan in the embryonic heart and vasculature. In stage 11 quail embryos, neurocan was prominently expressed in the myocardium, dorsal mesocardium, heart-forming fields, splanchnic mesoderm, and vicinity of the extraembryonic vaculature, and at lower levels in the endocardium. A comparison of neurocan staining with QH1 staining of vascular endothelial cells demonstrates that neurocan is frequently expressed by cells adjacent to endothelial cells, rather than by endothelial cells themselves. In some cases, a dispersed subset of cells are neurocan-positive in a field of cells that otherwise appear uniform in morphology. Later in development, neurocan expression becomes relatively limited to the nervous system. However, even in 10-day embryos, neurocan is expressed in the chorio-allantoic membrane in the tissue that separates closely packed, small-diameter blood vessels. In summary, our results suggest that neurocan may function as a barrier that regulates vascular patterning during development.

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