Biomedical subjects
Publications and source records attributed to R T Huang.
Influenza viruses bind to annexin V, a widely spread non-glycosylated phospholipid-binding protein. Externally added phospholipids as well as antiserum against this protein specifically inhibit infection of these viruses in cell cultures. We conclude that annexin V plays an important role in the entry of these viruses.
OBJECTIVE: To investigate the correlation of intratumoral blood flow in uterine myoma with cell proliferation, angiogenesis, tumor size, and tumor volume. METHODS: Thirty-nine patients who had been scheduled for surgery because of symptomatic uterine myomas were evaluated by transvaginal sonography and color Doppler ultrasound before surgery. The largest dimension of each tumor and the volumes of myomas were determined ultrasonographically. Pulsatility index (PI) was determined by color Doppler ultrasound according to the maximum systolic, end-diastolic, and the mean flow velocities measured within the uterine nodules. After surgery, the paraffin-embedded slides containing representative leiomyoma tissues were stained with hematoxylin and eosin, proliferating cell nuclear antigen for measurement of cell proliferation, and factor VIII for quantitation of microvessel density. The ultrasonographic findings were correlated postoperatively with pathologic findings, and the data were analyzed by simple linear regression and Fisher r to z transformation. RESULTS: Simple regression analysis of the intratumoral PI values on the sizes of myomas showed a negative correlation (r = -0.47, P = .003; n = 39), whereas a less significant correlation between PI values and tumor volumes was observed (r = -0.42, P = .008). In contrast, no statistically significant correlation was observed between the intratumoral PI values and the values of the proliferating cell nuclear antigen index (r = 0.10, P = .547) or microvessel density counts (r = 0.18, P = .282). CONCLUSION: The intratumoral blood flow by transvaginal color Doppler ultrasound correlated with a reduced tumor size and tumor volume, but did not correlate with cell proliferation or angiogenesis.
The effects of two newly synthesized sphingomyelin analogues on phosphatidylcholine biosynthesis were investigated in the immortalized human keratinocyte cell line HaCaT. N-Acetyl-erythro-sphingosine-1-phosphocholine (AcSM) and N-octanoyl-erythro-sphingosine-1-phosphocholine (OcSM) inhibited the incorporation of choline into phosphatidylcholine with half-inhibitory concentrations (IC50) of 6 micrograms/ml and 10 micrograms/ml respectively. Further experiments revealed that AcSM and OcSM interfered with the translocation of the rate-limiting enzyme of phosphatidylcholine biosynthesis, CTP:phosphocholine cytidylyltransferase (EC 2.7.7.15), in HaCaT cells and inhibited cytidylyltransferase activity in vitro. Despite the fact that OcSM was a potent inhibitor of cytidylyltransferase in vitro, its effects on phosphatidylcholine biosynthesis and translocation of cytidylyltransferase in HaCaT cells were less pronounced as compared with AcSM. Finally, we showed that the comparatively strong effects of AcSM in cell culture experiments were due to the uptake of large amounts of this sphingomyelin analogue into the cells. The results presented demonstrate that the activity of cytidylyltransferase may be negatively regulated by a high ratio of choline head group-containing sphingolipids.
The clinical utility of transvaginal ultrasonography in the differentiation of adenomyoma from leiomyoma was evaluated in 147 patients who had been scheduled for surgery due to symptomatic uterine masses. In all subjects, ultrasonographic images obtained preoperatively were correlated postoperatively with surgicopathological findings. Pathological findings showed that 110 patients proved to have fibroids, while 30 had adenomyomata. For the diagnosis of adenomyoma, transvaginal ultrasonography attained a sensitivity of 80%, a specificity of 94.3%, a positive predictive value of 85.7% and a negative predictive value of 90.9%, compared with a sensitivity of 94.3%, a specificity of 80%, a positive predictive value of 90.9% and a negative predictive value of 85.7% for leiomyoma diagnosis. Further to assess which characteristic used in ultrasonography was useful in the differential diagnosis, five characteristics were analyzed and compared by chi 2 test. These were position, number, margin and echogenicity of the uterine masses and the presence or absence of hypoechoic spaces (lacunae). Margin, echogenicity, mass number and lacunae were significantly different between both conditions. A stepwise logistic regression procedure revealed that margin, lacunae and echogenicity were good parameters for differentiating adenomyoma from leiomyoma. If we selected the features of distinct margin and absence of hypoechoic lacunae within the masses for analysis, leiomyoma could be correctly predicted in 97% of patients.
The binding of three influenza A and one influenza B virus strains to proteins of three continuously cultured cell lines was studied using protein overlay and immunostaining methods. The results obtained indicated the presence of both sialic acid-dependent and -independent binding of the virus strains; virus binding to proteins in the molecular mass range from about 40 to 103 kDa was dependent on sialic acid, whereas binding to the 33 kDa protein was independent of sialic acid. The 33 kDa binding protein was identified as annexin, a widely distributed non-glycosylated calcium-dependent phospholipid-binding protein.
Maternal phenylketonuria (PKU) is associated with significant complications such as mental retardation, microcephaly and congenital heart defects in nonphenylketonuric offspring. Dietary control with a low phenylalanine diet during the gestation period is effective in improving perinatal outcome in these cases. We present the case of a 27-year-old woman with classical features of PKU who had previously given birth to three babies, all of whom died of congenital heart disease. A low phenylalanine diet was started one month prior to the pregnancy and satisfactory fetal outcome was achieved.
This paper describes isolation and identification of a virus (termed strain 87A) which has the cytopathic effect and haemagglutination properties of hepatitis E virus (HEV). This virus was isolated by tissue culture from the faeces of a patient with acute non-A, non-B enteric hepatitis in Xinjiang, China. The isolated virus was neutralized by acute phase sera obtained from other patients with acute non-A, non-B enteric hepatitis. The virus particles also could be specifically aggregated with acute phase sera from patients with known HEV hepatitis in China, Burma, India and the U.S.S.R., and with acute and convalescent sera from an HEV-infected chimpanzee. Crystalline arrangements of virus particles in the cytoplasm were observed by electron microscopy in ultrathin sections of infected cells. The sedimentation coefficient of the strain 87A virus particles in sucrose gradients was 176S. Purified virus particles revealed a protein band of about 76K on SDS-PAGE and Western blotting. The evidence indicates that the strain 87A virus is an HEV. Our ability to propagate HEV in cell culture should facilitate research on this hepatotropic virus.
Fluorescent dansyl derivatives of 3 natural sphingolipids (gangliosides, cerebroside and sphingomyelin) were shown to be readily taken up by culture cells (HeLa-, MDCK- and primary rat brain cells). A part of the incorporated fluorescent sphingolipids remained associated with the cells after incubation in a culture medium containing serum, showing a cellular integration of these lipids. Microscopical studies indicated a localization of incorporated lipids in distinct subcellular regions; whereas dansyl cerebroside densely stained structures suggestive of the cytoskeleton and the actin filament, dansyl sphingomyelin and dansyl gangliosides were primarily associated with the plasma membrane. The findings are consistent with the current views on the arrangement of sphingolipids in animal cells.
The envelopes of influenza viruses contain in addition to lipids also two glycoproteins, the hemagglutinin and the neuraminidase, that are responsible for the adsorption, receptor splitting, penetration and budding processes of these viruses. In this article, hypotheses presented in the past with regard to the virus penetration are reconsidered. Based on results obtained with the fowl plague virus (influenza A/FPV/Rostock/34, H7N1) and MDCK-cells, we conclude that a fusion between the viral envelope and the plasma membrane is the initial step of virus entry.
We have devised a simple method for achieving 890-fold purification of ceramide glycanase with 17% recovery from a North American leech, Macrobdella decora. The method includes water extraction, ammonium sulfate fractionation, and chromatography on octyl-Sepharose, Matrex gel blue A, and Bio-Gel A-0.5m columns. The final preparation showed one major protein band at 54 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By using Bio-Gel A-0.5m filtration, the native enzyme was found to have a molecular mass of 330 kDa. With GM1 as substrate, the optimum pH of this enzyme was determined to be 5.0; the enzyme was stable between pH 4.5 and 8.5. Zn2+ at 5 mM and Cu2+, Ag+, and Hg2+ at 1 mM strongly inhibited the hydrolysis of GM1 by ceramide glycanase. The ceramide glycanase released the intact glycan chain from various glycosphingolipids in which the glycan chain is linked to the ceramide through a beta-glucosyl linkage. This enzyme also cleaved lyso-glycosphingolipids such as lyso-GM1 and lyso-LacCer and synthetic alkyl beta-lactosides. Among seven alkyl beta-lactosides tested, the enzyme only hydrolyzed the ones with an alkyl chain length of four or more carbons. The enzyme also hydrolyzed 2-(octadecylthio)ethyl O-beta-lactoside and 2-(2-carbomethoxyethylthio)ethyl O-beta-lactoside. p-Nitrophenyl, benzyl, and phytyl beta-lactosides, on the other hand, were not hydrolyzed. These results suggest that the enzyme can recognize the hydrophobic portion of glycolipid substrates. The fact that 2-(2-carbomethoxyethylthio)ethyl O-beta-N-acetyllactosaminide and DiGalCer were refractory to the enzyme indicated that in the substrate the first sugar attached to the hydrophobic chain cannot be N-acetylglucosamine and galactose. Furthermore, dodecyl maltoside, Gal alpha 1----6Glc beta Cer, and the LacCer in which the --CH2OH of the galactose was converted into --CHO were also resistant to the enzyme, and Man beta 1----4 Glc beta Cer was hydrolyzed at a much slower rate than LacCer. These results indicate that the nature and the linkage of the sugar attached to the glucose have a profound effect on the action of this enzyme. The hydrolysis of glycosphingolipids by ceramide glycanase is stimulated by bile salts. Among various bile salts tested, sodium cholate at a concentration of 1 microgram/microliter was found to be most effective in stimulating the hydrolysis of various glycosphingolipids with the exception of LacCer. For LacCer, sodium taurodeoxycholate at a concentration of 2-3 micrograms/microliters was most effective. Tween 20, Nonidet P-40, and Triton X-100 did not stimulate the hydrolysis of GM1.(ABSTRACT TRUNCATED AT 400 WORDS)
The roentgenologic findings of 76 workers exposed to vanadium were presented. The chest X-ray showed increase in lung markings in 58 cases, diffuse streaks and reticular shadows in 15 cases, interstitial pulmonary fibrosis. The clinical features of the 71 workers were as follows: 63 cases had coughs, 53 cases had coughs with expectorations, 27 cases showed difficulty in breathing, and 31 cases had wheezing or whistling sounds in the lungs. The workers were exposed to vanadium pentoxide fume at concentration ranging from 1.2 to 18 mg/m3 and dust from 1.27 to 27.7 mg/m3.
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Since mixtures of lipids alone are known to elicit membrane fusion without participation of fusion proteins, the role of viral lipids in the so-called virus-induced hemolysis and cell fusion has been investigated, using as a model the fowl plague virus (influenza A/FPV/Rostock/H7N1). The experiments were planned in a way that allowed quantitative modification of viral lipids without changing envelope glycoproteins. Under the conditions employed, cholesterol oxidase of Nocardia erythropolis and phospholipase C of Bacillus cereus were shown to completely modify their substrates in the virus without altering virus-associated hemagglutinating and neuraminidase activities. It was found with such enzyme treatment that virus-induced hemolysis and cell fusion are greatly influenced by cholesterol and phospholipids of the envelope. It became clear, that hemolysis and fusion are differently dependent on the nature of lipid components even though mediated by the same viral glycoproteins.
The role of neuraminidase and the mechanism of low pH dependence in influenza virus-induced membrane fusion have been studied further using fowl plague virus (FPV, H7N1). Two specific anti-FPV neuraminidase antisera obtained from chickens immunized with recombinant virus strains inhibited viral neuraminidase activity without influencing its haemagglutinating activity. These sera totally inhibited the FPV-induced fusion of erythrocytes and partially reduced haemolysis. But both fusion and haemolysis activities could be restored by external addition of Vibrio cholerae neuraminidase, indicating participation of neuraminidase in FPV-induced membrane fusion. With regard to low pH-dependent fusion by influenza virus, it was found that erythrocytes of various species showed different pH optima for haemolysis by FPV and that erythrocytes could be sensitized for fusion and haemolysis by FPV at neutral pH if they had been pretreated with a low pH buffer. These results demonstrated that surface properties of erythrocytes rather than that of the virus are critical in the low pH-dependent fusion and haemolysis by influenza viruses.
The lytic activity of secondary cytotoxic lymphocytes against influenza A virus was tested on cells which had been fused with liposomes containing the haemagglutinin and the neuraminidase of an avian influenza A virus (fowl plague virus, FPV). Fusion was obtained solely by the activity of the haemagglutinin and neuraminidase incorporated into the liposomes, without the need for any additional fusion factor. Highly reproducible lysis of these FPV-liposome target cells by influenza A-specific cytotoxic cells was found. In contrast, target cells containing the glycoproteins HN and F of Newcastle disease virus (NDV) were not lysed. In almost all experiments effector cell populations capable of lysing target cells also lysed the natural killer cell (NK)-sensitive cell line YAC-1. However, high NK activity alone was not sufficient to lyse target cells fused with liposomes containing the viral surface glycoproteins. To our knowledge this is the first report where after artificial introduction of viral surface components into cell membranes (either by fusion or by transfection) lysis of target cells was monitored also for non-specific lysis mediated by NK-like cells. Both the H-2 restriction and the virus specificity of lysis of FPV-liposome target cells indicate that influenza virus haemagglutinin and possibly neuraminidase do function as target antigens for influenza-specific T cells.
The small chain of influenza virus haemagglutinin, HA2 was isolated by a selective enzymic removal of HA1 or by preparative SDS-polyacrylamide gel electrophoresis. Anti-HA2 specific antisera and monoclonal antibodies were subtype-specific in immunodiffusion tests and radioimmunoassays. These antibodies did not inhibit haemagglutination or haemolysis, did not prevent virus release, did not neutralize infectivity, and HA2 did not induce a protective immunity. HA2-specific antigenic determinants could not be demonstrated on the surface of infected cells. Lymphocytes from pre-immunized mice could not be stimulated by HA2 to exert a cytotoxic effect.
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Myxoviruses (influenza virus and paramyxovirus) enter host cells by two successive steps consisting of attachment and fusion between viral and cellular membranes. The initial attachment is known to occur through specific binding of the viruses with the neuraminic acid-containing receptors of cellular membranes. Evidence is presented here that, in the following step of membrane fusion, neutral glycolipids terminating in galactose and certain phospholipids (primarily lecithin and sphingomyelin) interact with the viral envelopes and that this interaction may be fundamental to the fusion process.