Biomedical subjects
Publications and source records attributed to R H Chen.
We report a case of complete resection of a very large intracavitary metastatic melanoma of the right atrium. We describe a technique for resecting without damaging the atrium and for preventing the narrowing of the right atrium. In such cases, complete resection may be indicated despite widespread disease.
When consulted emergently by another surgeon in the operating room, we accomplished repair of a major laceration of the posterior wall of the distal trachea with associated avulsion of the left upper-lobe bronchus via the existing left thoracotomy exposure in a 7-year-old girl. Mobilization of the descending aorta anteriorly provided adequate exposure of the tracheal injury.
The scientific rationale for avoiding the use of calcium-enriched cardioplegic solutions and calcium supplementation during cardioplegic induction and the early phase of reperfusion in open heart surgical procedures is reviewed. The role of the extracellular and intracellular free ionized calcium concentrations during ischemia and reperfusion is explored and the biochemical effects of ischemia on calcium fluxes, adenosine triphosphate levels, and mitochondrial function are discussed. The role of calcium in causing myocardial stunning and the biochemical basis of reperfusion injury are also addressed. Both prolonged ischemia and an increased concentration of Ca2+ during reperfusion have proved to be deleterious. I conclude on the basis of my review that there is no justification for the use of calcium chloride before and during the early phase of reperfusion and that hypocalcemic perfusion is an effective and easily controllable means of myocardial protection.
The novel calcineurin inhibitor, dibefurin, has been isolated from the fungal culture AB 1650I-759. The isolation was bioactivity-directed fractionation using an assay which measures the phosphatase activity of calcineurin. The compound was purified by countercurrent, reverse phase and gel filtration chromatographies. Several studies, including crystallographic, NMR and MS, revealed that dibefurin is a novel dimeric compound of a unique structural type.
Two novel triterpene sulfates have been isolated from Fusarium compactum by bioactivity-directed fractionation using an assay which measures the inhibition of proteolytic activity of rhinovirus 3C protease on a fluorogenic peptide substrate. The compounds were purified by countercurrent and reverse phase chromatographies. NMR, MS, UV and IR studies revealed two triterpene sulfates, uncommon metabolites of terrestrial fungi.
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Insulin-dependent diabetes mellitus (IDDM) is a slowly progressive autoimmune disease. In the prediabetic phase of IDDM, the intravenous glucose tolerance test (IVGTT) demonstrates a progressive decline in the first-phase insulin response. The first-phase insulin response to intravenous glucose may enable individuals at risk of IDDM to be identified and preventive therapy to be instituted. However, the reproducibility of the first-phase insulin response is controversial. The aim of this study was to determine the variability of the first-phase insulin response to intravenous glucose. Ten healthy normal volunteers underwent two IVGTT within a 1 week interval, using the Islet Cell Antibody Register User's Study (ICARUS)-recommended standard protocol. The first-phase insulin response to intravenous glucose was expressed as the sum of the 1- and 3-minute insulin concentrations (muU/mL) and the total 0 to 10 minute insulin area under the curve (muU.min.mL-1). Variability was represented by the coefficient of variation (CV). The within-subject median CV was 21.2% for the sum of the 1- and 3-minute insulin concentrations and 23.9% for the total 0 to 10 minute insulin area under the curve. In conclusion, the reproducibility of the first-phase insulin response to intravenous glucose is poor. Hence, caution should be exercised when the IVGTT is applied to prediabetic IDDM patients for longitudinal study.
The nuclear oncoprotein v-Myb is a transcriptional activator in both animal cells and the budding yeast Saccharomyces cerevisiae. Previous studies have suggested that an acidic domain of approximately 50 amino acids (amino acids 204-254 of v-Myb) is necessary and sufficient for transcriptional activation by v-Myb, c-Myb and GAL4-Myb fusion proteins. However, we find that first, none of the acidic residues within this region is essential for transcriptional activation in either animal cells or yeast. Second, transcriptional activation requires cooperation among multiple domains of v-Myb. In animal cells, transcriptional activation by v-Myb requires a central domain (amino acids 234-295), a C-terminal domain (amino acids 295-356), plus either of two more N-terminal domains (amino acids 163-197 or 198-232); in yeast, it requires the central domain plus either the C-terminal domain or a more N-terminal domain (amino acids 163-233). Third, although various subsets of these domains are sufficient for transcriptional activation by v-Myb, all of the domains must be present for transformation of primary hematopoietic cells. These results demonstrate that transcriptional activation by v-Myb is not sufficient for oncogenic transformation.
Transforming growth factor-beta (TGF-beta) is the prototype for a family of extracellular polypeptides that affect cell proliferation and differentiation, and tissue morphogenesis. TGF-beta signalling is mediated by two types of serine/threonine kinase receptors, the type I and II receptors, which are able to form a heteromeric complex. No cytoplasmic proteins that associate with these receptors in vivo, or are their kinase targets, have yet been described. We have now identified a WD-domain-containing protein, TRIP-1, which specifically associates with the type II TGF-beta receptor in a kinase-dependent way. TRIP-1 does not interact with the type II activin or type I receptors, but associates with the heteromeric TGF-beta receptor complex. TRIP-1 is phosphorylated on serine and threonine by the receptor kinase, strongly suggesting that it has a role in TGF-beta signalling. This is supported by coexpression of TRIP-1 and type II receptor during development. The existence of TRIP-1 homologues in plant and yeast suggests a conserved function in all eukaryotes.
Transforming growth factor-beta (TGF-beta) transduces signals through its type I and type II receptors. Both receptor types have previously been shown to interact in a heteromeric complex in the presence of TGF-beta. We have now characterized these interactions between both receptor types using a combination of yeast two-hybrid interaction assays and coimmunoprecipitation analyses. Our results indicate a direct association between the cytoplasmic domains of the two receptor types. Mutation analysis of these cytoplasmic domains reveals that this direct interaction requires kinase activity and, thus, depends on phosphorylation, probably via a transphosphorylation mechanism. Furthermore, the two receptor types already have an inherent affinity for each other in the absence of TGF-beta, and the heteromeric complex can be detected in coimmunoprecipitations under these conditions. Taken together, our results reveal a novel mechanism of receptor complex formation, whereby two different cytoplasmic domains directly associate with each other. This interaction may play a major role in activation of serine/threonine kinase receptors.
Recently, we were unable to use the left internal mammary artery for coronary artery bypass grafting in 2 patients who had undergone successful late correction of coarctation of the aorta. In both patients, the mammary arteries were severely atherosclerotic and calcified; this may have resulted from prolonged and severe obstructive hypertension, which both patients had sustained before repair of the coarctation. Thus, in patients who have undergone late repair of coarctation, a bypass conduit other than the mammary artery may be needed.
Bleeding remains an important complication after repeat and complicated cardiac surgery. Although aprotinin has recently been approved by the Food and Drug Administration for use as an antifibrinolytic agent, many surgeons continue to have concerns about its added cost and potential side effects. We review here the current state of antifibrinolytic therapy for excessive bleeding in cardiothoracic surgery and suggest the use of a single intravenous dose of 10 g of epsilon-aminocaproic acid immediately before cardiopulmonary bypass as a safe, inexpensive, and effective alternative to aprotinin. Further clinical and laboratory studies are needed to confirm or modify this protocol.
Transforming growth factor-beta (TGF-beta) binds with high affinity to three cell-surface receptors. Both type I and II receptors are transmembrane serine/threonine kinases and thought to mediate TGF-beta responses by forming a heteromeric complex in the presence of TGF-beta. We investigated whether the type II receptors form a homomeric complex in the presence or absence of ligand. Double immunoprecipitation analyses were performed using lysates from metabolically labeled cells cotransfected with differentially epitope-tagged type II receptors. We demonstrate that the type II receptors can form a homomeric complex even in the absence of their ligand, TGF-beta. This pre-existing type II receptor complex has the ability to bind TGF-beta. Moreover, in addition to the extracellular and transmembrane domains, the cytoplasmic portions of the receptors are also able to interact with each other, indicating that multiple contact points are involved in the formation of the homomeric type II receptor complex. Our results suggest a novel mechanism of complex formation and receptor activation of the serine/threonine kinase receptor family.
Human platelets provide an excellent model system for the study of phosphorylation events during signal transduction and cell adhesion. Platelets are terminally differentiated cells that exhibit rapid phosphorylation of many proteins upon agonist-induced activation and aggregation. We have sought to identify the kinases as well as the phosphorylated substrates that participate in thrombin-induced signal transduction and platelet aggregation. In this study, we have identified two forms of mitogen-activated protein kinase (MAPK), p42mapk and p44mapk, in platelets. The data demonstrate that p42mapk but not p44mapk becomes phosphorylated on serine, threonine, and tyrosine during platelet activation. Immune complex kinase assays, gel renaturation assays, and a direct assay for MAPK activity in platelet extracts all support the conclusion that p42mapk but not p44mapk shows increased kinase activity during platelet activation. The activation of p42mapk, independently of p44mapk, in platelets is unique since in other systems, both kinases are coactivated by a variety of stimuli. We also show that platelets express p90rsk, a ribosomal S6 kinase that has previously been characterized as a substrate for MAPK. p90rsk is phosphorylated on serine in resting platelets, and this phosphorylation is enhanced upon thrombin-induced platelet activation. Immune complex kinase assays demonstrate that the activity of p90rsk is markedly increased during platelet activation. Another ribosomal S6 protein kinase, p70S6K, is expressed by platelets but shows no change in kinase activity upon platelet activation with thrombin. Finally, we show that the increased phosphorylation and activity of both p42mapk and p90rsk does not require integrin-mediated platelet aggregation. Since platelets are nonproliferative cells, the signal transduction pathways that include p42mapk and p90rsk cannot lead to a mitogenic signal and instead may regulate cytoskeletal or secretory changes during platelet activation.
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Transforming growth factor-beta (TGF-beta) is a multifunctional protein that modulates cell proliferation and interaction with the extracellular matrix. Three common TGF-beta receptors are found on the cell surface. The type III receptor is a transmembrane proteoglycan with a short cytoplasmic domain and is thought not to be involved in TGF-beta induced signalling. In contrast, the type II and type I receptors are transmembrane serine/threonine kinases. The type II receptor determines the ligand specificity, whereas the type I receptor interacts with the type II receptor and may not have a ligand binding specificity by itself. Both type II and type I receptors are involved in TGF-beta induced signalling. The type II receptor, likely in conjunction with the type I receptor, is required for the antiproliferative effect of TGF-beta, whereas the type I receptor is the likely mediator of the effects of TGF-beta on the expression of several genes including some extracellular matrix proteins. To address the role of TGF-beta signalling in myoblast differentiation, we transfected a dominant negative mutant of the type II receptor in myoblasts, thus inhibiting type II receptor mediated signalling. These cells not longer had the ability to differentiate in vitro or in vivo, suggesting that TGF-beta signalling through the type II receptor provides competence for myoblastic differentiation. These studies also indicate that there are several signalling pathways involved in myoblastic differentiation, one of which is modulated by the TGF-beta signalling.
Phosphorylation of the C terminus of c-Fos has been implicated in serum response element-mediated repression of c-fos transcription after its induction by serum growth factors. The growth-regulated enzymes responsible for this phosphorylation in early G1 phase of the cell cycle and the sites of phosphorylation have not been identified. We now provide evidence that two growth-regulated, nucleus- and cytoplasm-localized protein kinases, 90-kDa ribosomal S6 kinase (RSK) and mitogen-activated protein kinase (MAP kinase), contribute to the serum-induced phosphorylation of c-Fos. The major phosphopeptides derived from biosynthetically labeled c-Fos correspond to phosphopeptides generated after phosphorylation of c-Fos in vitro with both RSK and MAP kinase. The phosphorylation sites identified for RSK (Ser-362) and MAP kinase (Ser-374) are in the transrepression domain. Cooperative phosphorylation at these sites by both enzymes was observed in vitro and reflected in vivo by the predominance of the peptide phosphorylated on both sites, as opposed to singly phosphorylated peptides. This study suggests a role for nuclear RSK and MAP kinase in modulating newly synthesized c-Fos phosphorylation and downstream signaling.
Transforming growth factor-beta (TGF-beta) and activin signal primarily through interaction with type I and type II receptors, which are transmembrane serine-threonine kinases. Tsk 7L is a type I receptor for TGF-beta and requires coexpression of the type II TGF-beta receptor for ligand binding. Tsk 7L also specifically bound activin, when coexpressed with the type IIA activin receptor. Tsk 7L could associate with either type II receptor and the ligand binding specificity of Tsk 7L was conferred by the type II receptor. Tsk 7L can therefore act as type I receptor for both activin and TGF-beta, and possibly other ligands.