Glucose-regulated protein 78 (GRP78) is elevated in embryonic mouse heart and induced following hypoglycemic stress

This study investigates the distribution and heart levels of glucose regulated protein (GRP) 78 during normal development and in response to hypoglycemia in the mouse. Results demonstrate that GRP78 is strongly expressed with in the heart, neural tube, gut endoderm, somites, and surface ectoderm of mouse embryos during early organogenesis, and GRP78 staining remains prominent in the heart from gestational days 9.5 through 13.5. Cardiac myocytes are the primary site of GRP78 expression within the heart. GRP78 levels are highest in the heart during early organogenesis and levels decrease significantly by the fetal period. GRP78 expression is increased after 24 h of hypoglycemia in the early organogenesis-stage heart. Considering the tissue specific pattern of GRP expression and changes during development of the heart, GRPs may play significant roles in the normal differentiation and development of cardiac tissue. GRP induction may also be involved in hypoglycemia-induced cardiac dysmorphogenesis. Accepted: 25 January 2000     

Wnt6 expression in epidermis and epithelial tissues during Xenopus organogenesis.

Here, we report the localization within embryonic tissues of xWnt6 protein; together with the temporal and spatial expression of Xenopus laevis Wnt6 mRNA. Wnt6 expression in Xenopus embryos is low until later stages of neurulation, when it is predominantly found in the surface ectoderm. Wnt6 expression increases during early organogenesis in the epidermis overlaying several developing organs, including the eye, heart, and pronephros. At later stages of development, Wnt6 mRNA and protein generally localize in epithelial tissues and specifically within the epithelial tissues of these developing organs. Wnt6 localization correlates closely with sites of both epithelial to mesenchymal transformations and mesenchymal to epithelial transformations. Xenopus Wnt6 sequence and its expression pattern are highly conserved with other vertebrates. Xenopus embryos, therefore, provide an excellent model system for investigating the function of vertebrate Wnt6 in organ development and regulation of tissue architecture.     

Expression of bone marrow stromal cell specific antigen during murine development:Its expression in embryonic hematopoietic tissues as well as in other developing tissues

Monoclonal antibody R4-A9 demonstrated specificity for a cell surface antigen of stromal cells in murine bone marrow and spleen. In order to Identify patterns of expression that may elucidate the potential role of R4-A9 antigen, the developmental expression of this antigen in mouse embryos from 8 days post-coitum to 5 days post-partum was investigated by immunohistochemistry. At an early developmental stage, weak staining for R4-A9 antigen could be detected in the yolk sac. At later stages, strong staining of this antigen was detected predominantly in the embryonic liver, the main site of embryonic hematopoiesis. However, concomitant with the decreased staining in the liver, increased expression of this antigen was observed in bone marrow and spleen. Therefore, the changes in expression in those hematopofetic tissues suggest that its expression is coordinately regulated during the developmental stage of the sites of embryonic hematopoiesis. Compared with the distribution of R4-A9 antigen in adult tissues as previously reported, the expression of this antigen in fetal tissues was more widespread during the period of organogenesis, and was most abundant in other developing tissues, including the heart, skin, and lung. In contrast, fetal expression detected in hematopoietic and other developing tissues was lost after birth. These results taken together show a marked gradient of R4-A9 antigen expression, with the highest level at the peak of organ development, raising the possibility that this molecule may act as a growth/differentiation factor both in hematopoietic and other developing tissues in a fetus.     

葡萄糖调节蛋白94在人胚胎发育中的表达

目的 为探讨葡萄糖调节蛋白９４（ＧＲＰ９４）在胚胎发育中的作用,研究了１０－３１周人胎组织ＧＲＰ９４的定位及表达。方法 用ＳＰ免疫组织化学方法。结果 结肠粘膜上皮,肾小管上皮及肝细胞的胞质内,在１０周已有ＧＲＰ９４的表达,１８－２０周时表达增多,一直维持较高水平。胃粘膜上皮及心肌细胞也有较强的表达。     

Determination of hypoxic region by hypoxia marker in developing mouse embryos in vivo: a possible signal for vessel development.

Hypoxia is a well-known signal for angiogenesis, but the recent proposal that hypoxia exists in developing embryonic tissues and that it induces vascular development remains to be proven. In the present study, we demonstrate the presence of hypoxia in normal developing embryos by means of a hypoxia marker, pimonidazole, and its associated antibody. Our data clearly show that hypoxia marker immunoreactivity was highly detected in developing neural tubes, heart, and intersomitic mesenchyme at an early stage of organogenesis, suggesting that hypoxia may exist in the early stages of embryo development. We also found that hypoxia inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) were spatiotemporally co-localized with possible hypoxic regions in embryos. Investigation of platelet endothelial cell adhesion molecule (PECAM) expression provides evidence that endothelial cells proliferate and form the vessels in the hypoxic region in developing organs. Furthermore, we found that hypoxia induced both HIF-1alpha and VEGF in F9 embryonic stem and differentiated cells. Thus, we suggest that hypoxia may exist widely in developing embryonic tissues and that it may act as a signal for embryonic blood vessel formation in vivo.     

Xenopus Hand2 expression marks anterior vascular progenitors but not the developing heart.

We have isolated cDNAs encoding the bHLH protein Hand2 in the amphibian Xenopus laevis and analysed Hand2 expression in early development from the onset of gastrulation to feeding tadpole stages. XHand2 is expressed in the branchial arch mesenchyme and also in small bilateral populations of cells in the anterior, ventrolateral region of early tailbud embryos. At later stages, these punctate Hand2-expressing cells are located at the sites of the forming common cardinal veins, suggesting that they may constitute progenitors of vascular smooth muscle cells. Other Hand2-expressing cells are also associated with further components of the forming anterior vasculature but are not detected in mature blood vessels. Interestingly, no myocardial expression of XHand2 can be detected in the developing tadpole heart, in marked contrast to results obtained with chick and mouse embryos.     

Overexpression of GRP78 and GRP94 are markers for aggressive behavior and poor prognosis in gastric carcinomas

Glucose-related proteins (GRPs) are ubiquitously expressed in endoplasmic reticulum and able to assist in protein folding and assembly; consequently, they are considered as molecular chaperones. GRP78 and GRP94 expression was induced by glucose starvation and up-regulated in the malignancies. To clarify the roles of both molecules in tumorigenesis and progression of gastric carcinomas, immunohistochemistry was used on tissue microarray containing gastric carcinomas, adenomas, and nonneoplastic mucosa using the antibodies against GRP78 and GRP94, with a comparison of their expression with clinicopathological parameters of carcinomas. Gastric carcinoma cell lines (MKN28, AGS, MKN45, KATO-III, and HGC-27) were studied for both proteins by immunohistochemistry and Western blot. There was more expression of both proteins in gastric carcinoma and adenoma than in nonneoplastic mucosas (P < .05). All gastric carcinoma cell lines showed their expression at different levels. They were positively correlated with tumor size, depth of invasion, lymphatic and venous invasion, lymph node metastasis, and Union Internationale Contre le Cancer staging (P < .05), with positive relationship between both proteins (P < .05). Univariate analysis indicated the postsurgical cumulative survival rate of patients with positive GRP78 or GRP94 expression to be lower than that in those without GRP78 or GRP94 expression (P < .05), but the close link disappeared if stratified according to depth of invasion (P > .05). Multivariate analysis showed that age, depth of invasion, lymphatic invasion, lymph node metastasis, Union Internationale Contre le Cancer staging, and Lauren classification (P < .05), but not GRP78 and GRP94 expression, were independent prognostic factors for carcinomas (P > .05). Up-regulated expression of GRP78 and GRP94 was possibly involved in pathogenesis, growth, invasion, and metastasis of gastric carcinomas. They were considered objective and effective markers for the aggressive behavior and poor prognosis in gastric carcinomas.     

Pulmonary Neuroendocrine Cells and Lung Development

Pulmonary neuroendocrine cells produce bioactive peptides such as gastrin-releasing peptide (GRP) at high levels in developing fetal lung. The role of GRP and other peptides in promoting branching morphogenesis, cell proliferation, and cell differentiation during lung organogenesis is reviewed. Possible roles for bioactive peptides derived from these cells in the pathophysiology of perinatal lung disorders are discussed.     

Bves expression during avian embryogenesis.

Bves (blood vessel/epicardial substance) is a transmembrane protein postulated to play a role in cell adhesion. While it is clear that Bves and gene products of the same family are expressed in adult striated muscle cells, the distribution of these proteins during development has not been critically examined. An understanding of the expression pattern of Bves is essential for a determination of protein function and its role in embryogenesis. In this study, we present an expression analysis of Bves during chick gastrulation and germ layer formation. Our data show that Bves is expressed in epithelia of all three germ layers early in development. Furthermore, Bves protein is observed in epithelial tissues during organogenesis, specifically the developing epidermis, the gut endoderm, and the epicardium of the heart. These data support the hypothesis that Bves may play a role in cell adhesion and movement of epithelia during early embryogenesis.     

Spred2 expression during mouse development

SPREDs (Sprouty‐related proteins with Ena/Vasodilator‐stimulated phosphoprotein homology‐1 domain) are known membrane‐associated modulators of receptor tyrosine kinases by inhibiting the mitogen‐activated protein kinase (MAPK) signaling pathway. Although Spred2^?/? mice exhibit dwarfism and increase of early haematopoiesis, the precise expression and role of SPRED2 in mouse development remains unknown. Here, we demonstrate a detailed Spred2 expression pattern during mouse development using X‐Gal stainings from samples of a gene‐trapped Spred2 mouse line. In early stages, Spred2 was highly expressed in ectodermal and mesodermal tissues, and later on in developing neural tissue, heart, lung, intestine, urogenital tract, and limbs. Strikingly, we observed that Spred2 was mainly expressed at leading edges of further outgrowing structures and in folds of newly forming grooves. Therefore, SPRED2 is likely involved in the regulation of dynamic developmental processes. These new data provide valuable information for further studies regarding the still enigmatic physiological SPRED functions during mouse development. Developmental Dynamics 239:3072–3085, 2010. © 2010 Wiley‐Liss, Inc.     

The oncoproteins c-erb-B2, c-fos and the tumour suppressor protein p53 in human embryos and fetuses

 Oncoproteins and tumour-suppressor proteins are thought to possess an antagonistic function in the regulation of growth and differentiation processes during embryonic and fetal development. In contrast, in the adult, tumour growth is associated with the overexpression of oncoproteins or the malfunction of tumour-suppressor proteins. We examined the occurrence of the tumour proteins c-erb-B2 and c-fos and the tumour-suppressor protein p53 in 17 human embryos and fetuses with the help of immunohistochemistry. C-erb-B2 was detected mainly in embryonic tissue that are not known for c-erb-B2-overexpression in tumours in the adult. In contrast, c-fos was almost always located in fetal tissues corresponding to its location in adult tumours. Staining for p53 was found in a wide variety of embryonic and fetal tissues. C-erb-B2 and p53 were localized in the same tissue structures of the developing skin, heart and muscle. In other tissues, e.g. muscle and bone, c-fos was found together with p53, suggesting an antagonistic action of these proliferative and antiproliferative factors. Furthermore, c-erb-B2, c-fos and p53 appear to be important for growth and differentiation processes in human development as the occurrence of these proteins was not only restricted to specific tissues but also to specific stages of development of these tissues. Accepted: 30 September 1996     

Versican expression during skeletal/joint morphogenesis and patterning of muscle and nerve in the embryonic mouse limb

Versican, an extracellular matrix proteoglycan, has been implicated in limb development and is expressed in precartilage mesenchymal condensations. However, studies have lacked precise spatial and temporal investigation of versican localization during skeletogenesis and its relationship to patterning of muscle and nerve during mammalian limb development. The transgenic mouse line hdf (heart defect), which bears a lacZ reporter construct disrupting Cspg2 encoding versican, allowed ready detection of hdf transgene expression through histochemical analysis. Hdf transgene expression in whole mount heterozygous embryos and localization of versican relative to cartilage, muscle, and nerve tissues in paraffin-embedded limb sections of wild-type embryos from 10.5-14 days postcoitum were evaluated by lacZ histochemistry, immunohistochemistry, and in situ hybridization. Versican was localized within precartilage condensations and nascent cartilages with expression diminishing during maturation of the cartilage model at later time points. Interestingly, versican remained highly expressed in developing synovial joint interzones, suggesting potential function for versican in joint morphogenesis. Isolated myoblasts, incipient skeletal muscle masses, and neurites were not present in areas of strong versican expression within the developing limb. Versican-expressing tissues may reserve space for the future limb skeleton and developing joints and may aid in patterning of muscle and nerve by deterring muscle migration and innervation into these regions.     

MAP kinase activation in avian cardiovascular development.

Signaling pathways mediated by receptor tyrosine kinases (RTK) and mitogen-activated protein kinase (MAPK) activation have multiple functions in the developing cardiovascular system. The localization of diphosphorylated extracellular signal regulated kinase (dp-ERK) was monitored as an indicator of MAPK activation in the forming heart and vasculature of avian embryos. Sustained dp-ERK expression was observed in vascular endothelial cells of embryonic and extraembryonic origins. Although dp-ERK was not detected during early cardiac lineage induction, MAPK activation was observed in the epicardial, endocardial, and myocardial compartments during heart chamber formation. Endocardial expression of dp-ERK in the valve primordia and heart chambers may reflect differential cell growth associated with RTK signaling in the heart. dp-ERK localization in the epicardium, subepicardial fibroblasts, myocardial fibroblasts, and coronary vessels is consistent with MAPK activation in epicardial-derived cell lineages. The complex temporal-spatial regulation of dp-ERK in the heart supports diverse regulatory functions for RTK signaling in different cell populations, including the endocardium, myocardium, and epicardial-derived cells during cardiac organogenesis.     

Embryopathy in experimental diabetic gestation: assessment of oxidative stress and antioxidant defence

Maternal diabetes is associated with an increased rate of congenital fetal anomaly. In the present study, diabetes was induced by streptozotocin in female rats one week prior to conception and the embryos were examined during organogenesis. Experimental diabetes is associated with over-production of free radicals and disturbed antioxidant defence, particularly in malformed embryos. Oxidative stress is demonstrated by increased MDA accumulation and reduced glutathione levels. Despite large differences in the reduced/oxidised glutathione ratios during organogenesis in the control, diabetic non-malformed and malformed embryo groups, the half-cell redox potential was constant for each group during the experimental period. Calculated redox potentials indicated that although embryo cells from the control and diabetic mother groups were of the same chronological age, the stages of development were different. Increased oxidative stress in rat embryos was associated with increased glutathione peroxidases and glutathione-S-transferase activity. This may, in part, provide an explanation for the observed accumulation of oxidised glutathione in malformed embryos. Moreover, decreased levels of vitamin C and selenium were observed. Increased oxidative stress and perturbations in antioxidant defence contribute to the high incidence of congenital anomalies in experimental diabetic gestation.