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1.
During the process of endochondral ossification chondrocytes progress through stages of terminal differentiation culminating in apoptotic death. We have developed a serum-free suspension culture that allows terminal differentiation and facilitates the investigation of factors affecting chondrocyte apoptosis. We have found that chondrocytes not committed to terminal differentiation, i.e., those from the caudal region of chick embryo sterna, a region that remains cartilaginous for some months after the chick hatches, maintained high viability in serum-free suspension culture. A strong dependence of viability on culture density and sensitivity to induction of apoptosis with the protein kinase inhibitor, staurosporine, was consistent with the proposal that these chondrocytes, like nearly all cells, require intercellular communication for survival. Chondrocytes that were committed to terminal differentiation, i.e., those from the cephalic region of chick embryo sterna, a region that is replaced by bone before the chick hatches, expressed the hypertrophic phenotype but maintained their viability in culture for only approximately 6 days. Subsequent cell death was very consistent between cultures and shown to occur by an apoptotic process by analysis of DNA fragmentation and cell morphology. Short-term viability of hypertrophic chondrocytes was independent of culture density and relatively resistant to treatment with staurosporine. Induction of the hypertrophic phenotype in immature chondrocytes committed them to cell death and prevention of expression of the hypertrophic phenotype prevented cell death. We conclude that commitment of chondrocytes to terminal differentiation is associated with a commitment to apoptosis and apoptosis of hypertrophic chondrocytes in growth cartilage does not require initiation by external signals.  相似文献   

2.
A serum-free organ culture model for chondrocyte maturation, using the Avian sternum, was developed. Day-14 chick embryo sterna were placed in organ culture in the presence of defined medium. The optimal medium for chondrocyte terminal differentiation contained specific concentrations of dexamethasone, insulin, thyroid hormone and ascorbic acid. Three parameters, including sternal growth, cell diameter and type X collagen production, were analyzed as indicators of chondrocyte terminal differentiation. These parameters were analyzed in cephalic, middle and caudal regions of the organ-cultured chick sterna and compared to sterna grown in ovo. This study demonstrates that the organ-cultured tissue maintains normal morphological characteristics and terminal differentiation in the cephalic region only, similar to in ovo development, while maintaining normal cell-matrix relationships.  相似文献   

3.
Myocardial hypertrophy in different cardiac diseases is considered to be an adaptive mechanism to the increase of hemodynamic load which might restore to normal radius/wall thickness ratio and consequently to normalize wall stress. However, it has been widely demonstrated that beside the hemodynamic load, other factors contribute to the development of myocardial hypertrophy. It has been shown that in hypertensive patients, functional abnormalities (increased contribution of atrial systole to total diastolic filling, increased isovolumic relaxation period, prolonged diastolic duration, slowed ventricular filling and altered diastolic distensibility) precede the development of myocardial hypertrophy. Thus, in hypertensive patients, sign and symptoms of heart failure could be manifested in absence of myocardial hypertrophy, and might be exclusively due to diastolic dysfunction (with normal systolic function). Systolic function might be involved and compromised late when focal myocardial cell death and fibrosis occur and consequently ?adequate? hypertrophy is shifted to ?inadequate?. This evolution is accompanied by morphological and functional changes of the myocardium similar to those encountered in dilated cardiomyopathy. Impairment of systolic function in ?inadequate? hypertrophy is also due to structural changes; altered ratio between sarcomers and mitochondria, increased intercapillary distance, sarcoplasmatic reticulum dysfunction, increase of collagene component with a consequent increment of wall rigidity, hypertrophy of arterial tunica media, which alters coronary flow and coronary reserve. The progression of these morpho-functional abnormalities is a very slow process, in which adaptive mechanism mediated by several enzymes and contractile protein, contribute to maintain myocardial viability. However, over the long course, disseminated focal myocardial cell necrosis and fibrosis, which is an evolving process, is considered to be the main responsible factor for the irreversible myocardial damage and systolic dysfunction in advanced myocardial inadequate hypertrophy.  相似文献   

4.
Endochondral ossification in growth plates proceeds through several consecutive steps of late cartilage differentiation leading to chondrocyte hypertrophy, vascular invasion, and, eventually, to replacement of the tissue by bone. It is well established that the subchondral vascular system is pivotal in the regulation of this process. Cells of subchondral blood vessels act as a source of vascular invasion and, in addition, release factors influencing growth and differentiation of chondrocytes in the avascular growth plate. To elucidate the paracrine contribution of endothelial cells we studied the hypertrophic development of resting chondrocytes from the caudal third of chick embryo sterna in co-culture with endothelial cells. The design of the experiments prevented cell-to-cell contact but allowed paracrine communication between endothelial cells and chondrocytes. Under these conditions, chondrocytes rapidly became hypertrophied in vitro and expressed the stage-specific markers collagen X and alkaline phosphatase. This development also required signaling by thyroid hormone in synergy. Conditioned media could replace the endothelial cells, indicating that diffusible factors mediated this process. By contrast, smooth muscle cells, fibroblasts, or hypertrophic chondrocytes did not secrete this activity, suggesting that the factors were specific for endothelial cells. We conclude that endochondral ossification is under the control of a mutual communication between chondrocytes and endothelial cells. A finely tuned balance between chondrocyte-derived signals repressing cartilage maturation and endothelial signals promoting late differentiation of chondrocytes is essential for normal endochondral ossification during development, growth, and repair of bone. A dysregulation of this balance in permanent joint cartilage also may be responsible for the initiation of pathological cartilage degeneration in joint diseases.  相似文献   

5.
The earliest stages of development in most animals are under the control of maternally inherited information. The initiation of embryonic gene expression has been reported at the mid-blastula in amphibians and the mid-2-cell stage to the early morula in mammals. In chick embryos, embryonic gene expression was detectable at stage X (morula) and showed marked activation at stage XIII (blastula) with a gradual increase thereafter. Synthesis of rRNA and tRNA was low at stage X and was already the major class of RNA at stage XIII in chick embryos. The observed upregulation of RNA synthesis seems to coincide with a period of extensive fine structural differentiation when the first major cellular migrations start and signal the formation of the primitive streak in the chick embryo.  相似文献   

6.
To investigate the roles of the enzymes butyryl- and acetylcholinesterase (BChE and AChE) in retinal proliferation and differentiation, we use reaggregated spheres from retinal cells of the 6-day-old chick embryo, forming cellular and fibrous areas homologous to all layers of a normal retina. Recently, we could suppress BChE expression by transfecting these so-called retinospheroids during their proliferation period with a pSVK3 expression vector containing a 5' fragment of the rabbit BChE gene in antisense orientation. Along with morphological changes, proliferation was significantly decreased. Here, we have studied the effect of antisense BChE suppression during the differentiation period of retinospheroids. As BChE is suppressed, the differentiation of AChE-positive cells is increased, whereas the immunoreactivities for red and green cone-specific opsins are strongly reduced. Concomitantly, the rate of apoptosis as determined by propidium iodide uptake, by increased CPP 32-like caspase expression, and by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling and DNA fragmentation assays is roughly doubled, predominantly at the expense of degenerating photoreceptor precursors. This is further strong evidence that the proliferation marker BChE regulates an intricate balance between cell proliferation, cell differentiation, and programmed cell death in this in vitro retinal system.  相似文献   

7.
8.
PURPOSE: To investigate the associations between specific preoperative 12-lead electrocardiogram (ECG) abnormalities, perioperative ischemia, and postoperative myocardial infarction or cardiac death in major vascular surgery. METHODS: Two prospective studies on perioperative myocardial ischemia performed in two tertiary university hospitals were combined to include 405 patients. All preoperative ECGs were analyzed according to the Sokolow-Lyon criteria for left ventricular hypertrophy by investigators who were blinded to the patients' perioperative clinical course. Perioperative myocardial ischemia was detected by continuous ECG recording, and postoperative cardiac complications included myocardial infarction and cardiac death. RESULTS: A total of 19 postoperative cardiac complications occurred (two cardiac deaths and 17 myocardial infarctions). Voltage criteria for left ventricular hypertrophy (78 patients, 19%) and ST segment depression greater than 0.5 mm (98 patients, 24.2%) on preoperative ECGs were both significantly associated with postoperative myocardial infarction or cardiac death (odds ratio, 4.2 and 4.7; p = 0.001 and 0.0005, respectively) and with longer intraoperative and postoperative myocardial ischemia. In each of the two study groups, a preoperative ECG abnormality that involved voltage criteria, ST segment depression, or both (134 patients, 33.1%) was more predictive of postoperative cardiac complications than any other preoperative clinical variable, including a history of myocardial infarction or angina pectoris, diabetes mellitus, pathologic Q-wave by ECG, or preoperative myocardial ischemia. The combined duration of intraoperative and postoperative ischemia and the preoperative ECG with either voltage criteria or ST segment depression were the only independent factors associated with adverse cardiac events by multivariate analysis (p < or = 0.0001 and p = 0.02, respectively). CONCLUSION: Left ventricular hypertrophy and ST segment depression on preoperative 12-lead ECGs are important markers of increased risk for myocardial infarction or cardiac death after major vascular surgery.  相似文献   

9.
Platelet-Activating Factor (PAF) is a potent lipid mediator involved in physiological and pathological events in the nervous tissue where it can be synthesized by two distinct pathways. The last reaction of the de novo pathway utilizes CDPcholine and alkylacetylglycerol and is catalyzed by a specific phosphocholinetransferase (PAF-PCT) whereas the remodelling pathway ends with the reaction catalyzed by lyso-PAF acetyltransferase (lyso-PAF AcT) utilizing lyso-PAF, a product of phospholipase A2 activity, and acetyl-CoA. The levels of PAF in the nervous tissue are also regulated by PAF acetylhydrolase that inactivates this mediator. We have studied the activities of these enzymes during cell proliferation and differentiation in two experimental models: 1) neuronal and glial primary cell cultures from chick embryo and 2) LA-N-1 neuroblastoma cells induced to differentiate by retinoic acid (RA). In undifferentiated neuronal cells from 8-days chick embryos the activity of PAF-PCT was much higher than that of lyso-PAF AcT but it decreased during the period of cellular proliferation up to the arrest of mitosis (day 1-3). During this period no significant changes of lyso-PAF AcT activity was observed. Both enzyme activities increased during the period of neuronal maturation and the formation of cellular contacts and synaptic-like junctions. The activity of PAF acetylhydrolase was unchanged during the development of the neuronal cultures. PAF-PCT activity did not change during the development of chick embryo glial cultures but lyso-PAF AcT activity increased up to the 12th day. RA treatment of LA-N-1 cell culture in proliferation decreased PAF-PCT activity and had no significant effect on lyso-PAF AcT and PAF acetylhydrolase indicating that the synthesis of PAF by the enzyme catalyzing the last step of the de novo pathway is inhibited when the LA-N-1 cells are induced to differentiate. These data suggest that: 1) in chick embryo primary cultures, both pathways are potentially able to contribute to PAF synthesis during development of neuronal cells particularly when they form synaptic-like junctions whereas, during development of glial cells, only the remodelling pathway might be particularly active on synthesizing PAF; 2) in LA-N-1 neuroblastoma cells PAF-synthesizing enzymes coexist and, when cells start to differentiate the contribution of the de novo pathway to PAF biosynthesis might be reduced.  相似文献   

10.
Behaviour of 6-day chick embryo thigh skin in vitro in two different nutrients was studied electron microscopically. In explants supported with chicken serum containing medium epithelium keratinized faster than and in a similar way to that in ovo, except for the absence of corpuscola cribriformia in pericytes. It did not in explants supported with chick embryo extract containing medium, but underwent a noticeable ultrastructural evolution, mainly of granular reticulum and Golgi complexes. In the two series of cultures mesenchyme presented the same ultrastructural characteristics, especially as far as collagen fibers were concerned. The above data rule out the suggested regulatory role of collagen fibers in epidermal differentiation and support a possible epidermal two-step differentiative process. They are discussed in relation to the general mechanisms implicated in skin evolution.  相似文献   

11.
Differentiation of biliary epithelial cells from hepatic endodermal cells of the mouse embryo was examined with a special attention to the role of the connective tissue. When the whole liver primordium of the 9.5-day mouse embryo was cultured in vitro for 5 days, the endodermal cells differentiated into mature hepatocytes expressing carbamoylphosphate synthetase I (CPSI) and accumulating glycogen. Intrahepatic bile duct cells and connective tissue were poorly developed in this culture. However, when the hepatic endoderm was recombined with the 4-day embryonic chick lung mesenchyme and cultured in vitro, the endodermal cells differentiated into many ductal epithelial cells as well as mature hepatocytes with abundant connective tissue development. These results suggest that the ducts might be bile ducts, and that connective tissue is very important for bile duct development. In addition, this in vitro culture system might be useful for the study of mechanisms of bile duct differentiation and congenital biliary atresia.  相似文献   

12.
13.
When cultures of skeletal muscle cells of the chick embryo are subjected to repetitive, electrical stimulation, the contractions increase the amount of protein produced by these cells. The increase is greater for contractile proteins such as myosin heavy chain than for total cellular protein. This demonstrates that in a culture system of skeletal muscle cells that have differentiated in the absence of innervation, one can elicit the protein synthetic response associated with skeletal muscle hypertrophy in vivo.  相似文献   

14.
The functional status of brachially innervated hindlimbs, produced by transplanting hindlimb buds of chick embryos in place of forelimb buds, was quantified by analyzing the number and temporal distribution of spontaneous limb movements. Brachially innervated hindlimbs exhibited normal motility until E10 but thereafter became significantly less active than normal limbs and the limb movements were more randomly distributed. Contrary to the findings with axolotls and frogs, functional interaction between brachial motoneurons and hindlimb muscles cannot be sustained in the chick embryo. Dysfunction is first detectable at E10 and progresses to near total immobility by E20 and is associated with joint ankylosis and muscular atrophy. Although brachially innervated hindlimbs were virtually immobile by the time of hatching (E21), they produced strong movements in response to electrical stimulation of their spinal nerves, suggesting a central rather than peripheral defect in the motor system. The extent of motoneuron death in the brachial spinal cord was not significantly altered by the substitution of the forelimb bud with the hindlimb bud, but the timing of motoneuron loss was appropriate for the lumbar rather than brachial spinal cord, indicating that the rate of motoneuron death was dictated by the limb. Measurements of nuclear area indicated that motoneuron size was normal during the motoneuron death period (E6-E10) but the nuclei of motoneurons innervating grafted hindlimbs subsequently became significantly larger than those of normal brachial motoneurons. Although the muscle mass of the grafted hindlimb at E18 was significantly less than that of the normal hindlimb (and similar to that of a normal forelimb), electronmicroscopic examination of the grafted hindlimbs and brachial spinal cords of E20 embryos revealed normal myofiber and neuromuscular junction ultrastructure and a small increase in the number of axosomatic synapses on cross-sections of motoneurons innervating grafted hindlimbs compared to motoneurons innervating normal forelimbs. The anatomical data indicate that, rather than being associated with degenerative changes, the motor system of the brachial hindlimb of late-stage embryos is intact, but inactive.  相似文献   

15.
Male and female gonads from 7- to 9-day-old chick embryos were cultured for 6 days in Sertoli cell-conditioned medium or in serum-free medium to investigate the possible effect of substances secreted by rat Sertoli cells on chick gonad development. Histological analysis showed that whereas all female gonads proceed through normal ovarian development in both culture media, most of male gonads showed clear feminization only when cultured in Sertoli cell-conditioned medium; male gonads cultured in serum-free medium developed as normal testes. Because the only substance detected in our conditioned medium with the potential to cause these effects was sex-specific antigen (Sxs), our results provide further evidence that Sxs antigen may play a role in sexual differentiation in birds, and probably in mammals.  相似文献   

16.
Aborted sudden death as the presenting manifestation of hypertrophic cardiomyopathy in a 14-year-old child is reported. Documented ventricular fibrillation was the cause of cardiac arrest. No ventricular arrhythmia was induced during programmed electrical stimulation. An implantable cardioverter-defibrillator was indicated. As the patient had a family history of myocardial disease, he had undergone a cardiovascular evaluation 4 years before the major event, and was found normal. It is suggested that normal physical examination, ECG, echocardiogram should not rule out the diagnosis of hypertrophic cardiomyopathy when a family history is present. Left ventricular hypertrophy may develop during childhood in patients with hypertrophic cardiomyopathy.  相似文献   

17.
The interaction between chick embryo fibroblasts and various lectins has been studied at different stages of embryo development. There is evidence that Robinia lectin, Dolichos lectin, and Concanavalin A decrease cell number and proportion of cells incorporating [3H] thymidine in case of 8- and 10-day-old chick embryo fibroblasts, whereas they stimulated the proliferation of 16-day-old embryo cells. No effect was noticed in 12-day cells. These results suggest that some cell surface changes occur during embryo development. The site number of Dolichos lectin remains the same during embryo development, and the affinity constant decreases. The site number of Robinia lectin and Concanavalin A decreases from the 8th to the 12th day of development, and slowly increases on the 16-day cells, the affinity constant remaining rather constant. The results indicate that the age-dependent effect of lectin on embryo cells could not be directly related to the number of lectin-binding sites. Competitive binding experiments revealed that Dolichos receptor sites were distincts from binding sites of Robinia lectin and Concanavalin A, and Robina receptor sites distinct from those of concanavalin A. Lectin effects on embryo fibroblasts were very specific as determined by inhibitory assays.  相似文献   

18.
We have previously shown that feather formation in chick embryonic skin depends on accumulation of sulphated glycosaminoglycans in the underlying mesenchyme, and that addition of spermidine to chick embryo fibroblasts increases the extracellular sulphated glycosaminoglycans. In the present work, using histological, histochemical and biochemical procedures, we have investigated the effects on glycosaminoglycan accumulation and on epithelial differentiation of adding spermidine and bis-cyclohexylammonium sulphate, a spermidine inhibitor, to embryonic chick skin cultures. Our results demonstrate that spermidine induces an accumulation of sulphated glycosaminoglycan and an increase in feather formation, suggesting that the morphogenetic effect of spermidine may be dependent on specific glycosaminoglycan accumulation.  相似文献   

19.
During early heart development the expression pattern of N-cadherin, a calcium-dependent cell adhesion molecule, suggests its involvement in morphoregulation and the stabilization of cardiomyocyte differentiation. N-cadherin's adhesive activity is dependent upon its interaction with the intracellular catenins. An association with alpha-catenin and beta-catenin also is believed to be involved in cell signaling. This study details the expression patterns of alpha-catenin, beta-catenin, and gamma-catenin, during definition of the cardiac cell population as distinct compartments in the anterior regions of the chick embryo between stages 5 and 9. The restriction of N-cadherin/catenin localization at stage 5+ from a uniform pattern in vivo, to specific cell clusters that demarcate areas where mesoderm separation is initiated, suggests that the N-cadherin/catenin complex is involved in boundary formation and in the subsequent cell sorting. The latter two processes lead to the specification and formation of the somatic and cardiac splanchnic mesoderm. N-cadherin colocalized with alpha- and beta-catenin at the cell membrane before and during the time that its expression becomes restricted to the lateral mesoderm and continues cephalocaudad into stage 8. These proteins continue to colocalize in the myocardium of the tubular heart. Plakoglobin is not expressed in this region during stages 6-8, but is detected in the myocardium later at stage 13. The observed in vivo expression patterns of alpha-catenin, beta-catenin, and plakoglobin suggest that these proteins are directly linked with the developmental regulation of cell junctions, as cardiac cells become stably committed and phenotypically differentiated to eventually form a mature myocardium. The localization of N-CAM also was analyzed during these stages to determine whether the N-cadherin-catenin localization was unique or whether other cell adhesion molecules were expressed similarly. The results indicate that the unique pattern of N-cadherin expression is not shared with N-CAM. We also show that perturbation of N-cadherin using a function perturbing N-cadherin antibody (NCD-2) inhibits normal early heart development and myogenesis in a cephalocaudad, stage-dependent manner. We propose a model whereby myocardial cell compartmentalization also defines the endocardial population. The presence of beta-catenin suggests that a similar signaling pathway involving Wnt (wingless)-mediated events may function in myocardial cell compartmentalization during early vertebrate heart development, as in Drosophila contractile vessel development.  相似文献   

20.
A gene encoding a zinc finger protein of the Snail family, cSnR, is expressed in the right-hand lateral mesoderm during normal chick development. Antisense disruption of cSnR function during the hours immediately preceding heart formation randomized the normally reliable direction of heart looping and subsequent embryo torsion. Implanted ectopic sources of intercellular signal proteins that are involved in establishing normal left-right information randomized the handedness of heart development and also altered the asymmetry of cSnR expression. cSnR thus appears to act downstream of these signals, or perhaps in parallel with the latest expressed of them, the Nodal protein, in controlling the anatomical asymmetry.  相似文献   

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