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Capitation Fee↗
Biomedical subjects
G Gibson
Publications and source records attributed to G Gibson.
Apoptosis of terminally differentiated chondrocytes in culture.
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.
Animals↗
Angioplasty and stenting of an unprotected left main bifurcation lesion.
A case of unprotected bifurcation left main disease treated on 2 occasions by angioplasty and stenting is presented. This case demonstrates the 2 main features of salvage angioplasty, namely medically refractory rest angina and refused bypass surgery. In addition, it presents short inflation time angioplasty for sole or main conduits and treating the left main as a bifurcation lesion.
Aged↗
In vitro formation of mineralized cartilagenous tissue by articular chondrocytes.
Study of the deep articular cartilage and adjacent calcified cartilage has been limited by the lack of an in vitro culture system which mimics this region of the cartilage. In this paper we describe a method to generate mineralized cartilagenous tissue in culture using chondrocytes obtained from the deep zone of bovine articular cartilage. The cells were plated on Millipore CMR filters. The chondrocytes in culture accumulated extracellular matrix and formed cartilagenous tissue which calcified when beta-glycerophosphate was added to the culture medium. The cartilagenous tissue generated in vitro contains both type II and type X collagens, large sulfated proteoglycans, and alkaline phosphatase activity. Ultrastructurally, matrix vesicles were seen in the extracellular matrix. Selected area electron diffraction confirmed that the calcification was composed of hydroxyapatite crystals. The chondrocytes, as characterized thus far, appear to maintain their phenotype under these culture conditions which suggests that these cultures could be used as a model to examine the metabolism of cells from the deep zone of cartilage and mineralization of cartilagenous tissue in culture.
Alkaline Phosphatase↗
Selective changes in cell bodies and growth cones of nerve growth factor-differentiated PC12 cells induced by chemical hypoxia.
Cytosolic free Ca2+ concentration ([Ca2+]i) was measured in differentiated PC12 cells to test whether chemical hypoxia selectively alters intracellular Ca2+ in growth cones and cell bodies. Hypoxia increased [Ca2+]i and exaggerated its response to K+ depolarization in both parts of the cells. [Ca2+]i in the cell bodies was greater than that in the growth cones under resting conditions and in response to K+ or hypoxia. Ca2+-channel blockers selectively altered these responses. The L-channel blocker nifedipine reduced [Ca2+]i following K+ depolarization by 67% in the cell bodies but only 25% in the growth cones. In contrast, the N-channel blocker omega-conotoxin GVIA (omega-CgTX) diminished K+-induced changes in [Ca2+]i only in the growth cones. During hypoxia, nifedipine was more effective in the cell bodies than in the growth cones. During hypoxia, omega-CgTX diminished K+-induced changes by 50-75% in both parts of the cell, but only immediately after depolarization. The combination of nifedipine and omega-CgTX diminished the [Ca2+]i response to K+ with or without hypoxia by >90% in the cell body and 70% in the growth cones. Thus, the increased Ca2+ entry with K+ during hypoxia is primarily through L channels in the cell bodies, whereas in growth cones influx through L and N channels is about equal. The results show that chemical hypoxia selectively alters Ca2+ regulation in the growth cone and cell body of the same cell.
Animals↗
Hepatic enzyme induction and mutagenicity of airborne particulate matter from Santiago, Chile in the nourished and malnourished rat.
1. Respirable, airborne particles in the ambient air in downtown Santiago, Chile, have been characterized for the seasonal variation in total polycyclic aromatic hydrocarbon content, 13 of which have been identified including the mutagens (benzo(a)pyrene, dibenzo(a,h)anthracene, benzo(a)anthracene, benzo(b)fluoranthene and indeno(1,2,3, c,d)pyrene amongst others. 2. Organic extracts derived from these particles were administered to both the nourished and malnourished rat and resulted in modulation of the hepatic mixed function oxidase system including induction of NADPH-cytochrome P450 reductase, cytochrome P4501A as determined by Western blot analysis and the associated ethoxyresorufin O-deethylase and aryl hydrocarbon hydroxylase activities. 3. The cytochrome P4504A1-dependent 12-hydroxylation of lauric acid was induced in the malnourished state, but this activity was significantly inhibited by treatment of the animals with particle extracts in both nutritional states. 4. The particle extracts contained both direct and indirect-acting mutagens in the Ames test, and depending on the relative complement of both, resulted in either increased or decreased mutagenicity in the presence of S9 activation systems derived from both nourished and malnourished animals. 5. These results are discussed in the context of the interindividual risk assessment of airborne, particulate matter to man.
Air Pollutants↗
Is function of the Drosophila homeotic gene Ultrabithorax canalized?
Genetic variation affecting the expressivity of an amorphic allele of the homeotic gene Ultrabithorax, (Ubx1) was characterized after 11 generations of introgression into 29 different isofemale lines. Heterozygotes display a range of haploinsufficient phenotypes, from overlap with wild-type halteres to dramatic transformations such as a 50% increase in area and the presence of over 20 bristles on the anterior margin of each haltere. In both the wild-type and mutant genetic backgrounds, there is moderate genetic variance and low environmental variance/developmental asymmetry, as expected of a trait under stabilizing selection pressure. Surprisingly, there is little evidence that mutant halteres are more variable than wild-type ones, so it is unclear that haltere development is also canalized. The correlation between wild-type and Ubx haltere size is very low, indicating that interactions among modifiers of Ubx are complex, and in some cases sex-specific. The potential quantitative genetic contributions of homeotic genes to appendage morphology are discussed, noting that population-level effects of variation in key regulatory genes may be prevalent and complex but cannot be readily extrapolated to macroevolutionary diversification.
Animals↗
Pulmonary artery smooth muscle cell [Ca2+]i and contraction: responses to diphenyleneiodonium and hypoxia.
To investigate mechanisms of inhibition of hypoxic pulmonary vasoconstriction (HPV), we studied pulmonary artery smooth muscle cell (PASMC) responses to hypoxia, utilizing diphenyleneiodonium (DPI), which blocks HPV. We measured cell contraction in primary cultures of rat PASMC grown on collagen gels and cytosolic free Ca2+ concentration ([Ca2+]i) in PASMC grown on glass. DPI (5 and 20 microM) caused contraction of PASMC and increased [Ca2+]i. Omission of extracellular Ca2+ diminished the DPI-induced PASMC contraction and greatly reduced the increase in [Ca2+]i. DPI substantially inhibited KCl-induced PASMC contraction (1 microM DPI) and the increase in [Ca2+]i (5 microM DPI). Severe hypoxia contracted PASMC and quadrupled [Ca2+]i. DPI, 1 microM, substantially inhibited hypoxic contraction, but neither 1 nor 5 microM DPI diminished the hypoxia-induced increase in [Ca2+]i, which was greatly attenuated by 20 microM DPI. These data show 1) that DPI increases [Ca2+]i, accounting for DPI-induced PASMC contraction and 2) that 1 and 5 microM DPI inhibit the hypoxia-induced contraction but not the hypoxia-induced increase in [Ca2+]i, suggesting that DPI inhibits hypoxic PASMC contraction downstream of the Ca2+ signal by desensitizing the contractile apparatus and indicating a potential control point for modulation of HPV.
Animals↗
Parathyroid hormone and parathyroid hormone-related peptide in venous umbilical cord blood of healthy neonates.
Objective of this study was to assess the role umbilical cord serum parathyroid hormone (PTH), and parathyroid hormone-related peptide (PTH-RP) may play in maintaining the maternal-fetal calcium (Ca) gradient. PTH and PTH-RP and total and ionized Ca levels were measured in blood samples of 20 neonates immediately after birth. Maternal peripheral blood total Ca was measured simultaneously. Mothers were free of hypertension, diabetes or Ca disorders. Neonates were healthy term babies with Apgar scores of eight or greater at one and at five minutes post delivery. PTH was measured using an immunoradiometric double antibody assay that recognizes intact PTH (1-84). PTH-RP was measured by an immunoradiometric double antibody assay that recognizes only PTH-RP (1-74). There was no overlap between the two assays. Ca levels in the neonates were higher than those in their mothers (p < 0.01), confirming a maternal-fetal Ca gradient. However in 18 out of 20 neonates PTH levels in cord blood were below the detection limit (3 pg/ml) and PTH-RP levels also were below detection limit (0.2 pmol/L). PTH-RP and PTH levels in the other two neonates were 0.5 and 0.6 pmol/L (PTH-RP) and 3 pg/ml (PTH) which are in the low normal range for normal adults. We conclude that these data do not support a role for either PTH or PTH-RP in venous cord blood in maintaining the maternal-fetal Ca gradient. They are, however, compatible with a paracrine role for these hormones.
Calcium↗
Identification and immunolocalization of type X collagen at the ligament-bone interface.
In some ligaments, ligamentous collagen fibrils attach to bone by first passing through non-mineralized and mineralized fibrocartilage present at the ligament-bone interface. To understand better the function of these fibrocartilages, collagens present at the femoral insertion of the bovine medial collateral ligament were isolated and characterized. Types II and IX collagens were identified in pepsin digests of the tissue in addition to type X collagen originally thought to be associated with the cartilages undergoing endochondral bone formation. Presence of type X collagen was confirmed by immunoblotting and by immunofluorescence localization using laser confocal microscopy. Type X collagen was localized predominantly in the mineralized zone of the ligament insertion. These data indicated that type X collagen may play a role in ligament attachment to bone.
Animals↗
Effect of polymorphism in the Drosophila regulatory gene Ultrabithorax on homeotic stability.
Development is buffered against unpredictable environmental and genetic effects. Here, a molecular genetic analysis of one type of developmental homeostasis, the establishment of thoracic segmental identity under the control of the Ultrabithorax (Ubx) gene in Drosophila melanogaster, is presented. Flies were artificially selected for differential sensitivity to the induction of bithorax phenocopies by ether vapor. The experiments demonstrated that increased sensitivity to ether correlated with a loss of expression of UBX in the third thoracic imaginal discs and that a significant proportion of the genetic variation for transcriptional stability can be attributed to polymorphism in the Ubx gene.
Alleles↗
Epistasis and pleiotropy as natural properties of transcriptional regulation.
A statistical thermodynamic model of transcriptional regulation is employed to investigate the likely effects of genetic variation on the stabilization of gene expression. The model is tailored to empirical data on the control of transcription of the hunchback gene by the morphogen Bicoid during Drosophila embryogenesis. Variable parameters include the number of binding sites for activator protein and the DNA-protein and protein-protein cooperative binding energies. Recursions are performed to derive transcriptional response curves over a range of concentrations of activator. Sigmoidal responses are indicative of threshold-dependent activation of gene expression, and the effects of variation of the parameters on the width and location of the threshold are considered. It is shown that there is a minimum threshold width (maximum switch sensitivity) that is a function of the number of binding sites and the level of response desired, but independent of the binding energies. This places a constraint on the evolution of sensitive genetic switches that generate discrete cell types. Inevitable trade-offs between threshold widths and locations for multiple target genes of most transcriptional activators are found to occur. These naturally lead to epistatic and pleiotropic effects, and may favor the generation of networks of compensatory mutations that together produce homeostatic developmental pathways.
Animals↗
Altered oxidation and signal transduction systems in fibroblasts from Alzheimer patients.
Abnormalities in calcium regulation, amyloid-beta-protein (A beta) production and oxidative metabolism have been implicated in Alzheimer's disease (AD). The use of cultured fibroblasts complement post-mortem and genetic approaches in clarifying the interaction of these processes and the underlying mechanism for the changes in AD. Definition of gene defects in particular Alzheimer families (FAD) permits elucidation of the role of those genetic abnormalities in altered signal transduction in cell lines from those families. Abnormalities in calcium regulation, ion channels, cyclic AMP, the phosphatidylinositide cascade and oxidative metabolism are well documented in fibroblasts from patients with primary genetic defects in the presenilins. Recent studies in AD fibroblasts that demonstrate abnormal secretion of A beta, a protein known to form the characteristic extracellular amyloid deposits in AD brain, further supports the use of these cells in AD research. Comparison of changes in calcium signaling, mitochondrial oxidation and A beta production in these cells suggests that changes in signal transduction including calcium may be a more consistent observation than altered A beta production in fibroblasts from some FAD families. An understanding of these abnormalities in fibroblasts may provide further insights into the pathophysiology of AD, new diagnostic measures and perhaps innovative therapeutic approaches.
Aging↗
Regulation of bradykinin-induced Ins(1,4,5)P3 formation by protein kinase C in human fibroblasts.
To better understand the molecular mechanisms that underlie the exaggerated bradykinin (BK)-stimulated release of Ins(1,4,5)P3 in fibroblasts from Alzheimer patients, the role of G-proteins, protein kinase C (PKC) and cyclic AMP in BK-induced Ins(1,4,5)P3 formation was determined. A role for G-proteins in the coupling of the BK receptor to intracellular signals was indicated by guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) enhanced BK-stimulated Ins(1,4,5)P3 release. The coupling of G-proteins to Ins(1,4,5)P3 formation was sensitive to cholera toxin (CTX), but not pertussis toxin (PTX), and was not altered by PKC activation. The inhibition by CTX appeared to be secondary to its ability to increase cyclic AMP, because forskolin also inhibited the BK-mediated Ins (1,4,5)P3 release. Activation of PKC with TPA diminished the number of BK receptors by 33% and proportionally decreased BK-mediated Ins(1,4,5)P3 formation by 28%. The latter response was abolished by PKC inhibitors. Depletion of PKC by prolonged TPA treatment did not further alter the number of BK receptors but further decreased the Ins(1,4,5)P3 response by 65%. Thus, changes in PKC probably do not underlie the enhanced BK-induced Ins(1,4,5)P3 formation in AD fibroblasts, because both activation and depletion of the PKC diminished the Ins(1,4,5)P3 response.
Bradykinin↗
Case report: the CT demonstration of soft tissue involvement in multicentric reticulohistiocytosis.
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Adipose Tissue↗
Type X collagen is colocalized with a proteoglycan epitope to form distinct morphological structures in bovine growth cartilage.
Using antibodies raised against type X collagen, isolated from deer antler, we have immunolocalized type X collagen in growth plate and epiphyseal cartilage from fetal and ambulatory calves. In ambulatory calf growth plate, type X collagen was demonstrated to be present in longitudinal septa that extend from the resting zone into the underlying trabecular bone. The much more restricted distribution of type X collagen seen in fetal growth plate, both here and previously, suggested that prominent localization of type X collagen to the longitudinal septa was stimulated by weight bearing. The location of type X collagen has been compared with that of an epitope (7D4), composed of specific sulfation patterns in chondroitin sulfate of some aggrecan molecules. Colocalization of type X collagen with the 7D4 epitope in longitudinal septa of growth plates from ambulatory calves suggested these molecules contribute to the formation and function of longitudinal septa. Immunolocalization of type X collagen and 7D4 in calf epiphyseal cartilage demonstrated colocalization at an interface that appears to represent the junction of articular cartilage and cartilage associated with formation of the secondary ossification center. Immunolocalization of type X collagen and the 7D4 proteoglycan epitope in these structures suggested that type X collagen and the 7D4 epitope form structures with important mechanical roles and capacity to influence the morphology of associated bony structures.
Animals↗
Molecular cloning and expression of cDNAs for rat and human kidney cysteine conjugate beta-lyase.
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Animals↗