Biomedical subjects

Q C Yu

Publications and source records attributed to Q C Yu.

47 records · Page 3Linked to original sources

PubMed1992

Cachexia and graft-vs.-host-disease-type skin changes in keratin promoter-driven TNF alpha transgenic mice.

Tumor necrosis factor alpha (TNF alpha) orchestrates a wide range of effects that combat severe infections in animals. At lower levels, TNF alpha plays an important protective role in stimulating chemotaxis and antimicrobial activity of neutrophils, macrophages, and eosinophils. During chronic illness, TNF alpha secretion can be elevated markedly, giving rise to cachexia, hemorrhage, necrosis and, ultimately, death. Although TNF alpha may mediate many of its effects through macrophages, 30% of TNF alpha injected into animals concentrates in the skin. In recent years, it has been shown that keratinocytes can be induced to synthesize TNF alpha. To explore the role of TNF alpha synthesis in keratinocytes, we used a keratin-14 (K14) promoter to target human TNF alpha expression in the epidermis and other stratified squamous epithelia of transgenic mice. Most mice expressing the K14-TNF alpha transgene stopped gaining weight within 1 week postbirth, and exhibited retarded hair growth. In the skin, adipose production was profoundly inhibited, whereas signs of fibrosis and immune infiltration were evident in the dermis. Over time, the epidermis exhibited an increased stratum corneum, as signs of necrosis began to appear in the skin. Within 3-5 weeks, the mice displayed features characteristic of cachexia and necrosis. Our results suggest that TNF alpha expression by keratinocytes not only plays a role in inflammatory and graft-versus-host-disease-like responses in the skin, but also in other tissues, apparently by virtue of stratified squamous epithelial-derived TNF alpha entering the bloodstream. Our results have enabled the first evaluation of many of the effects of TNF alpha in transgenic animals.

Animals↗

PubMed1992

The C terminus of human immunodeficiency virus type 1 matrix protein is involved in early steps of the virus life cycle.

Deletion mutations at the C terminus of the matrix (MA) protein of human immunodeficiency virus type 1 (HIV-1) were generated by site-directed mutagenesis. The resultant mutant viruses had a severe defect in virus infectivity. This defect did not involve late steps of the virus life cycle, as the synthesis and processing of the Gag polyprotein and the assembly and release of mutant virions were not greatly affected. The incorporation of viral proteins and the viral RNA genome was similar for mutant and wild-type virions. In contrast, the early steps of the virus life cycle were severely affected, as the synthesis of viral DNA postinfection was dramatically reduced in mutant-virus-infected cells. One stretch of amino acids that was deleted in one of the mutants has significant homology with a region in VP1 of the picornavirus family. This region of VP1 is presumably involved in poliovirus penetration into cells. These results suggest that in addition to its functional role in virus assembly, the MA protein of HIV-1, and possibly of other retroviruses, plays an important role in virus entry.

Amino Acid Sequence↗

PubMed1992

Transient disruptions of aortic endothelial cell plasma membranes.

Cells of gut, skin, and muscle frequently suffer transient survivable plasma membrane disruptions ("wounds") under physiological conditions, but it is not known whether endothelial cells of the aorta, which are constantly exposed to hemodynamically generated mechanical forces, similarly are injured in vivo. We have used serum albumin as a molecular probe for identifying endothelial cells of the rat aorta that incurred and survived transient plasma membrane wounds in vivo. Such wounded endothelial cells were in fact observed in the aortas of all rats examined. However, the percentage of wounded cells in the total aortic endothelial population varied remarkably between individuals ranging from 1.4% to 17.9% with a mean of 6.5% (+/- 4.6% SD). Wounded endothelial cells were heterogeneously distributed, being found in distinct clusters often in the shape of streaks aligned with the long axis of the vessel, or in the shape of partial or complete rims surrounding bifurcation openings, such as the ostia of the intercostal arteries. Physical exercise (running) did not increase the frequency of aortic endothelial cell membrane wounding, nor did spontaneous hypertension. Surprisingly, 80% of mitotic endothelial cell figures were identified as wounded. This article identified a previously unrecognized form of endothelial cell injury, survivable disruptions of the plasma membrane, and shows that injury to the endothelial cells of the normal aorta is far more commonplace than previously suspected. Plasma membrane wounding of endothelial cells could be linked to the initiation of atherosclerosis.

Animals↗

PubMed1991

Role of conserved gp41 cysteine residues in the processing of human immunodeficiency virus envelope precursor and viral infectivity.

All animal retroviruses whose nucleotide sequences have been determined contain two or three closely spaced cysteine residues in the extracellular domain of the env-encoded transmembrane protein. Using human immunodeficiency virus type 1 gp41 as a working model, the functional significance of these highly conserved cysteines was investigated. We report here that substituting the two conserved cysteine residues in this domain of gp41 with glycine residues resulted in the loss of viral infectivity, which could be attributed to severe impairment in the processing of gp160 precursor to gp120.

Amino Acid Sequence↗

PubMed1991

The vpx gene of simian immunodeficiency virus facilitates efficient viral replication in fresh lymphocytes and macrophage.

vpx is a unique open reading frame found in simian immunodeficiency virus (SIV) and human immunodeficiency virus type 2 (HIV-2) but not in HIV-1. It encodes a 12- to 16-kDa virion-associated protein. Although vpx is dispensable for viral replication in several established human lymphocyte cell lines, there is no consensus regarding whether this gene is required for efficient viral replication in freshly isolated lymphocytes. We report here that the vpx mutant of SIVmac exhibits different degrees of impairment from wild-type SIVmac in freshly isolated lymphocytes. This defect is more pronounced in macrophages from the same donors. Our findings suggest that vpx is required for efficient viral replication in fresh lymphocytes and macrophages.

Cells, Cultured↗

PubMed1991

Link between cell apical morphology and H+ secretion in salamander small intestine.

To determine the morphological basis for the cellular secretion of H+ by in vitro salamander jejunum the transepithelial proton flux (JH) was measured under different conditions and compared with the ultrastructure of the villus absorptive cells using transmission electron microscopy in conjunction with morphometric analysis. Norepinephrine (NE) stimulated JH and significantly increased the density of small cytoplasmic vesicular and tubular profiles in the region immediately below the apical plasma membrane of the absorptive cells. Cytoplasmic tubular elements were often observed in direct contact with the plasma membrane at the base of the microvilli, suggesting that expansion of the plasma membrane had occurred. The alpha 2-antagonist, yohimbine, blocked NE-stimulated H+ secretion and reduced the number of tubulovesicles. Theophylline reduced the rate of spontaneous acid secretion and the density of apical tubules and vesicles. Exposure to horseradish peroxidase revealed a physical connection between the extracellular space and the apical tubulovesicular compartment. The direct relationship between the number of apical vesicular and tubular profiles and JH suggests that the luminal membrane plays a fundamental role in the cellular events in H+ secretion. The membranes of the tubules and vesicles may be elements of a dynamic pool of transporting membranes under the control of exogenous effectors such as catecholamines.

Animals↗

PubMed1990

Multitubular bodies in intestinal cells of Amphiuma means/tridactylum (Urodela): ultrastructural characterization.

The jejunal absorptive cells of the salamander Amphiuma, when examined using transmission electron microscopy, were found to possess a unique type of intracellular vacuole containing membranous tubules. These vacuoles, tentatively named multitubular bodies, were located in the cytoplasm between the nucleus and the brush-border membrane, and were seen with greatest frequency in the summer and fall. The vacuoles containing multitubular bodies had an average diameter of 0.6 microns, and the membranous tubules within had an average diameter of 30 nm. The tubules differed morphologically from the vesicles in the multivesicular bodies, and from the primary lysosomes in the polylysosomal vacuoles. The tubules did not exhibit acid phosphatase activity, and were of similar diameter and membrane thickness as the Golgi saccules. In contrast to the multivesicular bodies, the multitubular bodies did not take up exogenous horseradish peroxidase. Early forms of autophagosomes resembling these vacuoles were often seen in the para-Golgi region of the cell. The multitubular bodies may represent a distinct type of autophagosome. Although the exact origin of the tubules as well as their role in cellular activity is unclear, their seasonal appearance within the multitubular bodies of the absorptive cells suggests a unique means of selective down-regulation of Golgi-like organelles.

Acid Phosphatase↗

PubMed1990

Postmortem viability and early changes in organ culture of human and rabbit aortic endothelial cells.

In order to understand the role of the postmortem interval (PMI) on endothelial cell changes both in human and rabbit aortas, we have examined the ultrastructural cytomorphologic alterations of these cells. Human aorta (HA) and rabbit aorta (RA) were maintained in calcium-free, glucose-supplemented Hank's balanced salt solution (HBSS). Rabbit endothelial cells (REC) on the aorta (organ culture) assayed morphologically survive for at least 12 h in culture solution. The predominant morphological change in the RA was the formation of multiple subendothelial vacuoles (SEV). These vacuoles may form as the results of increased permeability of endothelial cells to ions and fluid or cell contraction. Cell to cell connection remained intact. Individual and dispersed endothelial cells were observed 8 h after removal from the animal when incubated in calcium-free HBSS. These necrotic endothelial cells were scattered among viable endothelial cells. Human aortic endothelial cells were also well preserved in the same media for periods of 6-8 h postmortem. Increased extracellular calcium (1.3 mM) in the incubation media caused accelerated cell death. These findings suggest that aortic endothelial cells can be preserved for longer periods of postmortem time than would be expected and that the use of calcium-free HBSS media supplemented with glucose improves endothelial cell viability in vitro.

Adult↗

PubMed1988

Response of human hepatocyte lysosomes to postmortem anoxia.

We studied the ultrastructure and degradative activity of lysosomes in human livers after somatic death due to cerebral necrosis secondary to shock and/or head trauma. The livers were obtained at autopsy after varying postmortem intervals. Liver ultrastructure was studied in intact liver, in hepatocyte suspension obtained by collagenase perfusion of the liver, and in cultured hepatocytes. Lysosomal protein degradation was measured in a case 24 hours after hepatocyte isolation. By standard ultrastructural criteria, all the livers demonstrated typical reversible manifestations of cell injury. Most hepatocytes demonstrated varying degrees of irreversible cell injury. The ultrastructural alterations were less severe in the isolated hepatocytes in suspensions, and further improvement in morphologic appearance occurred in the cultured monolayers. The most striking alteration in the liver lysosomes was the increase in the numbers of lipofuscin granules (a type of residual body) compared with control liver. The hepatocyte lysosomes constituted an average 3.1% of cytoplasmic volume. There was no correlation between the volume density of lysosomes and either the duration of postmortem anoxia, clinical course, or patient's age. There was no increase in the number of autophagic vacuoles or of secondary lysosomes in postmortem liver. Autophagic vacuoles were more frequently seen in isolated and cultured hepatocytes. Cultured hepatocytes isolated within one hour of clinical death and tested 24 hours later degraded cell proteins at a rate of 1.7% per hour. Protein degradation was stimulated by a physiologic signal (Dibutyryl cAMP, 1 mmol/L) and was inhibited by microtubule poison (vinblastine). We conclude that (1) viable hepatocytes can be isolated even after prolonged postmortem intervals (range, 30 minutes to seven hours), (2) trauma and shock cause an expansion of hepatocyte lysosomes due to accumulation of lipofuscin, and (3) autophagy is blocked by postmortem anoxia and resumes in the recovery phase from anoxic injury in hepatocyte suspensions and in culture.

Cell Survival↗

PubMed1988

Response of autophagic protein degradation to physiologic and pathologic stimuli in rat hepatocyte monolayer cultures.

The lysosomes of hepatocytes increase in numbers and size during acute cell injury in vivo. To elucidate the mechanism of this change, we have studied in vitro the response of the autophagic lysosomal system to several physiologic mediators of autophagy, and to agents known to induce injury and/or the accumulation of lysosomes in vivo. To this end, monolayer cultures of rat hepatocytes were labeled with [14C]leucine to measure hepatocyte protein degradation; ultrastructural analyses were carried out to measure the volume fraction of lysosomes in the hepatocytes. Dibutyryl cyclic AMP increased protein degradation in the hepatocytes either in the presence or absence of serum and insulin. Deprivation of serum and insulin also increased hepatocyte protein degradation. Morphometric analysis indicated parallel increases in the volume fraction of lysosomes in the hepatocytes. The calcium ionophore ionomycin (5 microM), in the presence of 1.3 mM extracellular calcium, increased protein degradation (but not the volume fraction of lysosomes), and this increase was abolished by the addition of ethyleneglycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid. On the other hand, vasopressin (5 nM) caused an increase in protein degradation coupled with an increase in volume fraction of lysosomes. The microtubule depolymerizer vinblastine decreased protein degradation. The microtubule stabilizer taxol did not prevent the inhibitory effects caused by vinblastine. Parallel decreases in the lysosomal compartment were found in the hepatocytes exposed to vinblastine or taxol. Dimethylnitrosamine inhibited protein degradation as well as decreased the volume fraction of lysosomes. Finally, carbon tetrachloride also decreased protein degradation. These data indicate that in physiologic conditions, increases in numbers of hepatocyte lysosomes reflect increased sequestration and degradation of cytoplasmic proteins in response to changes in the levels of hormones, serum factors and nutrients as well as cyclic AMP. The induction of acute cell injury in vitro by calcium ionophore, microtubule active agents, and hepatotoxins inhibits lysosomal proteolysis and causes a decrease in the volume fraction of lysosomes. We conclude that the increase in lysosomal size and numbers occurring in acutely injured hepatocytes in vivo is induced primarily by altered levels of nutritional and hormonal regulators of lysosomal protein degradation.

Animals↗

PubMed1986

Modification of lysosomal proteolysis in mouse liver with taxol.

Perturbation of lysosomal degradation pathways by drugs such as chloroquine and vinblastine has given us important insights into the means for segregation and degradation of cytoplasm by the lysosomes. Vinblastine (VBL) is a microtubule depolymerizer which increases the number of autophagosomes and also impairs the maturation of the autophagosomes into secondary lysosomes. The authors have tested the effects of the microtubule stabilizer taxol on the proteolytic activity of the lysosomal system in VBL-treated mice. Liver proteins of CD-1 mice were radioisotopically labeled. Taxol and/or vinblastine were subsequently administered. Proteolysis in liver homogenates was determined during in vitro incubation by measuring the release of trichloroacetic acid-soluble radioactivity over time. Taxol enhanced proteolysis in homogenates from VBL-treated mice. The site of the enhanced proteolysis was lysosomal, because it was demonstrated in a crude lysosomal fraction, it manifested an acid pH optimum, and it could be suppressed with cycloheximide. Morphometric analysis indicated that taxol caused a shift in hepatocyte lysosomes of VBL-treated mice toward the later stages of the degradation cycle, coupled with a decrease in the number of small dense body profiles. The results indicate that taxol enhances the maturation of the autophagosomes into proteolytically active secondary lysosomes. It is concluded that the microtubule stabilizer taxol enhances fusion between VBL-induced autophagosomes and lysosomes (small dense bodies) and does not affect the segregation of cytoplasm into autophagosomes.

Alkaloids↗