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D X Liu

Publications and source records attributed to D X Liu.

At least 73 records · Page 4Linked to original sources
PubMed1995

Identification, expression, and processing of an 87-kDa polypeptide encoded by ORF 1a of the coronavirus infectious bronchitis virus.

Nucleotide sequence analysis has shown previously that the genomic-length mRNA (mRNA1) of the coronavirus infectious bronchitis virus (IBV) contains two large open reading frames (ORFs), 1a and 1b, with the potential to encode polyproteins of approximately 441 and 300 kDa, respectively. We have characterized the specificity of a set of region-specific antisera raised against the 5'-portion of ORF 1a by immunoprecipitation of in vitro-synthesized, C-terminally truncated 1a polypeptides and used these antisera to detect virus-specific proteins in IBV-infected Vero cells. Two antisera, which had specificity for IBV sequences from nucleotides 710 to 2079 and 1355 to 2433, respectively, immunoprecipitated a polypeptide of approximately 87 kDa from IBV-infected Vero cells. In vitro translation of ORF 1a sequence terminating at nucleotide 5763 did not produce this protein unless the in vitro translation products were incubated with Vero cell S10 extracts prepared from either IBV-infected or mock-infected Vero cells. However, processing of the 87-kDa protein was also observed when the same region was expressed in Vero cells using the vaccinia virus/T7 expression system. This observation indicates that the 87-kDa polypeptide is encoded within the 5'-most 3000 nucleotides of mRNA 1 and that it might be cleaved from the 1a polyprotein by viral and cellular proteinases.

Animals↗
PubMed1995

Chronic pulmonary infection caused by Mycoplasma pneumoniae leading to pulmonary arteriole remodeling and pulmonary hypertension in rats.

The pulmonary arteriole remodeling in Wistar rats with respiratory infection induced by mycoplasma pneumoniae was observed using light microscopy and morphometry. The pulmonary artery pressure (PAP) and index of right ventricular hypertrophy (RVHI) were measured. The intimal and medial hypertrophy can be seen in the pulmonary arterioles, leading to vessel wall thickening and narrowing of the lumina. The total number of the pulmonary arterioles decreased (P < 0.01), and both pulmonary hypertension (Ppa 4.11 +/- 0.19 kPa) and right ventricular hypertrophy (RVHI = 34.96 +/- 3.91%) occurred. In addition, an interstitial pulmonary fibrosis (IPF) was found, in which the content of collagen in the lung tissue changed, i. e., type I collagen increased whereas type III one decreased, and the ratio of type I collagen to type III one increased. It suggested that respiratory infection induced by repeated MP may result in remodeling of pulmonary arterioles and are closely related to pulmonary hypertension.

Animals↗
PubMed1994

A 100-kilodalton polypeptide encoded by open reading frame (ORF) 1b of the coronavirus infectious bronchitis virus is processed by ORF 1a products.

The genome-length mRNA (mRNA 1) of the coronavirus infectious bronchitis virus (IBV) contains two large open reading frames (ORFs), 1a and 1b, with the potential to encode polypeptides of 441 and 300 kDa, respectively. The downstream ORF, ORF 1b, is expressed by a ribosomal frameshifting mechanism. In an effort to detect viral polypeptides encoded by ORF 1b in virus-infected cells, immunoprecipitations were carried out with a panel of region-specific antisera. A polypeptide of approximately 100 kDa was precipitated from IBV-infected, but not mock-infected, Vero cells by one of these antisera (V58). Antiserum V58 was raised against a bacterially expressed fusion protein containing polypeptide sequences encoded by ORF 1b nucleotides 14492 to 15520; it recognizes specifically the corresponding in vitro-synthesized target protein. A polypeptide comigrating with the 100,000-molecular-weight protein (100K protein) identified in infected cells was also detected when the IBV sequence from nucleotides 8693 to 16980 was expressed in Vero cells by using a vaccinia virus-T7 expression system. Deletion analysis revealed that the sequence encoding the C terminus of the 100K polypeptide lies close to nucleotide 15120; it may therefore be generated by proteolysis at a potential QS cleavage site encoded by nucleotides 15129 to 15135. In contrast, expression of IBV sequences from nucleotides 10752 to 16980 generated two polypeptides of approximately 62 and 235 kDa, which represent the ORF 1a stop product and the 1a-1b fused product generated by a frameshifting mechanism, respectively, but no processed products were observed. Since the putative picornavirus 3C-like proteinase domain is located in ORF 1a between nucleotides 8937 and 9357, this observation suggests that deletion of the picornavirus 3C-like proteinase domain and surrounding regions abolishes processing of the 1b polyprotein. In addition, the in vitro translation and in vivo transfection studies also indicate that the ORF 1a region between nucleotides 8763 and 10720 contains elements that down-regulate the expression of ORF 1b.

Base Sequence↗
PubMed1994

[Myelographic diagnosis and treatment of lumbosacral nerve root anomalies].

Altogether 480 patients were examined with myelography in the study from 1987 to 1992 and 28 cases having anomalous lumbosacral nerve roots with other concomitant diseases were found. The anomalies were classified three types based on myelographic findings. Type 1, root sleeve displacement; Type 2, a, roots through one foramen; b, roots through two foramina; c, roots through three foramina; Type 3, double roots through one foramen. All patients were proved and treated during operation. The results of treatment were good (92.8%). We considered that most lumbosacral nerve roots anomalies were asymptomatic, however, when they were compressed by degenerative canal tissue, classic nerve root pain appeared eventually. Myelography is helpful to its diagnosis and treatment.

Adult↗
PubMed1994

[Specific binding sites of [3H] cortisol and [3H] dexamethasone in rat liver cell membrane].

By using [3H] cortisol and [3H] dexamethasone (DEX), a class of specific binding sites of glucocorticoids (GC) in rat liver cell membrane was observed. These binding sites had saturability, high affinity and low capacity. The kinetic parameters of [3H] cortisol and [3H] DEX were: Kd = 12.84 +/- 6.58 nmol/L (n = 4) and 40.27 +/- 23.44 nmol/L (n = 3) respectively; Bmax = 2.57 +/- 1.84 pmol/mg protein and 0.64 +/- 0.18 pmol/mg protein respectively (mean +/- SE). The Kd obtained from the kinetic studies was corresponding to the results of Scatchard analysis. Scatchard plots from the equilibrium experiments showed the straight line. The Hill coefficients were 0.9880 and 0.9990 respectively. The competitive displacement experiments indicated that cortisol possessed the higher specificity for the [3H] cortisol binding sites than other tested steroids (prednisolone, progesterone, RU486, DEX). The competitive ability of cortisol was at least 40 folds higher than mentioned steroids. By means of autoradiography technique, the existence of [3H] cortisol labelled particles on the liver cell membrane was found.

Animals↗
PubMed1994

Correlative study on findings of dynamic myelography and surgical operation in non-bony lumbar spinal canal stenosis.

The authors performed dynamic lumbar myelography with Omnipaque on 110 patients from 1990 to 1992, of them, 33 cases were diagnosed as non-bony lumbar spinal canal stenosis according to contrast medium defect seen on the lateral view of the myelogram. All such cases were operated on and their dynamic pathological findings during the operation were recorded to compare with the abnormal findings observed on the myelograms, there was a high coincidence rate of 88.7%. The authors believe that the fibrous degenerative changes make up the basis of dural sac compression and the dynamic compression in the spinal canal plays a very important role in causing the severity of the stenosis. As the dynamic pathological findings on the myelograms can not be well demonstrated on CT scanning and MR imaging, the dynamic lumbar myelography should be the method of choice for use in some cases.

Adult↗
PubMed1993

Human herpesvirus-6 glycoprotein H and L homologs are components of the gp100 complex and the gH external domain is the target for neutralizing monoclonal antibodies.

Previous studies have shown that monoclonal antibody (MAb) 2E4 neutralizes infectivity of human herpesvirus-6 (HHV-6) and also inhibits virus-induced T-lymphocyte syncytia formation. Here we characterize two additional MAbs, 1D3 and 5E7, which have similar properties, and identify the glycoprotein targets. The MAbs could immunoprecipitate and immunofluorescence glycoprotein from both A and B variant strain groups of HHV-6. In reactions with infected cells the MAbs immunoprecipitated a complex of glycoproteins, the "gp100" complex, composed of a major glycoprotein species of 100,000 M(r) and minor components of 80,000 M(r) and 32,000 M(r). We show that the 100,000 M(r) product and most likely the 80,000 M(r) correspond to the HHV-6 homologue of herpes simplex virus-1 (HSV-1) glycoprotein H (gH) while the 32,000 M(r) species corresponds to the glycoprotein L (gL) equivalent. All three MAbs could specifically immunoprecipitate either gH expressed on its own in fibroblasts or a complex of gH and gL co-expressed, but could not immunoprecipitate gL expressed on its own. Consistent with these results, the MAbs could recognize gH in an immunofluorescence assay but not gL. Therefore although the MAbs recognized the complex of glycoproteins, they appeared specific for gH. The HHV-6 glycoproteins were produced in a transient expression system induced by T7-vaccinia virus. Immunoprecipitations were carried out in comparisons with an "epitope-tagged" gH, a recombinant glycoprotein designed to contain at the N-terminus the linear epitope for MAb LP14, raised originally against HSV-1 glycoprotein gD. The epitope-tagged gH was also used as a positive control in determining the domain of HHV-6 gH to which MAbs 2E4, 1D3 and 5E7 were directed. Two gH deletions were constructed, one deleting sequences which may serve as a transmembrane and cytoplasmic anchor domains, the second deleting also part of the external domain. MAb LP14 could immunoprecipitate both HHV-6 gH deletions but the gp100 MAbs recognized only the full-length product or the intact external domain minus the transmembrane and cytoplasmic domains. This indicated the epitopes for these MAbs are contained in the external domain of gH, consistent with the MAbs action in neutralization of virion infectivity and inhibition of virus to cell spread by T-lymphocyte fusion.

Amino Acid Sequence↗
PubMed1993

Identification and expression of the human herpesvirus 6 glycoprotein H and interaction with an accessory 40K glycoprotein.

In herpes simplex virus (HSV) the small secreted glycoprotein gL forms a heterodimer with the transmembrane envelope glycoprotein gH. Here we identify the human herpesvirus 6 (HHV-6) gL gene, express HHV-6 gL and gH homologues, and examine interactions between HHV-6 gH and gL. The HHV-6 gL gene encoded a glycoprotein with an amino acid sequence which showed closest similarity to the human cytomegalovirus (HCMV) gL homologue (18% identity). Products of HHV-6 gH and gL genes were characterized in an in vitro transcription-translation system and in a transient in vivo expression system. Both gH and gL were transcribed and translated in vitro to give products of apparent M(r) of 65K and 28K in SDS-PAGE, and these could be processed by addition of microsomes to 110K and 40K, respectively. To study gH/gL interactions, gH was tagged with the nine amino acid epitope for monoclonal antibody LP14 (anti-HSV-1 gD). LP14 and a human serum sample specifically immunoprecipitated gH and a stable complex of gH and gL co-expressed in an in vivo vaccinia virus-T7 system. The gH and gL produced in this in vivo expression system corresponded to the M(r)s of the fully processed glycoproteins identified in the in vitro system. The gH expressed together with gL was recognized by human sera more easily than when examined on its own in immunofluorescence assays. Dual expression of gH and gL in transfected T lymphocytes (JJhan) caused reactions with 75% of human sera tested (12 HHV-6-positive, HCMV-negative serum samples), but gL expressed alone was not recognized by these sera. The immunofluorescence studies also showed that the glycoproteins were localized in Golgi-like bodies in fibroblasts, but occurred throughout the endoplasmic reticulum in T lymphocytes, the normal cellular target for HHV-6. These results show the identification of the HHV-6 homologue to the HCMV and HSV gL genes, identification and production of HHV-6 gH and gL expressed both in vitro and in vivo, complex formation between these glycoproteins, and evidence that this complex may be localized differently in fibroblasts as compared to T lymphocytes and that it is immunogenic.

Adult↗
PubMed1992

Identification of two new polypeptides encoded by mRNA5 of the coronavirus infectious bronchitis virus.

The second smallest subgenomic messenger RNA, mRNA5, of the coronavirus infectious bronchitis virus includes in its "5' unique region" two separate open reading frames (5a and 5b), whose coding function has not so far been established, and thus it may represent a dicistronic messenger RNA. We report here that two polypeptides with the sizes expected for the 5a and 5b products can be synthesised by in vitro translation of a single artificial mRNA containing both the 5a and 5b ORFs. To establish whether these polypeptides represent genuine virus gene products, both the 5a and 5b coding sequences were expressed as bacterial fusion proteins, and these were used to raise monospecific antisera. Antisera raised against both the 5a and 5b-specific sequences recognized specifically proteins of the expected size in infectious bronchitis virus-infected chicken kidney and Vero cells, indicating that 5a and 5b do represent genuine virus genes, and suggesting that mRNA5 is indeed functionally dicistronic.

Base Sequence↗
PubMed1992

Potentiation of the humoral response of intravenous antigen by splenotropic liposomes.

We have recently described that large liposomes composed of egg phosphatidylcholine (PC), cholesterol (chol) and monosialoganglioside GM1 show elevated accumulation in the red pulp of the spleen when they are i.v. administered into mice. Up to 50% of the injected dose was found in spleen at 4 h post injection. In this report, we have investigated the potential application of such liposomes in the stimulation of anti-lysozyme response in mice. Lysozyme entrapped in the splenotropic liposomes composed of PC/chol/GM1 showed higher efficiency in potentiating the humoral response than that of either free lysozyme or lysozyme entrapped in hepatotropic liposomes composed of PC/chol. The results demonstrate that high levels of i.v. antigen delivery by liposomes to the splenic macrophage instead of the liver Kupffer cells is important in the liposomal adjuvanticity. The antibody elicited by the liposome entrapped antigen was mainly IgG1 subtype.

Animals↗
PubMed1992

Internal entry of ribosomes on a tricistronic mRNA encoded by infectious bronchitis virus.

mRNA3 specified by the coronavirus infectious bronchitis virus appears to be functionally tricistronic, having the capacity to encode three small proteins (3a, 3b, and 3c) from separate open reading frames (ORFs). The mechanism by which this can occur was investigated through in vitro translation studies using synthetic mRNAs containing the 3a, 3b, and 3c ORFs, and the results suggest that translation of the most distal of the three ORFs, that for 3c, is mediated by an unconventional, cap-independent mechanism involving internal initiation. This conclusion is based on several observations. A synthetic mRNA whose peculiar 5' end structure prevents translation of the 5'-proximal ORFs (3a and 3b) directs the synthesis of 3c normally. Translation of 3c, unlike that of 3a and 3b, was insensitive to the presence of the 5' cap analog 7-methyl-GTP, and it was unaffected by alteration of the sequence contexts for initiation on the 3a and 3b ORFs. Finally, an mRNA in which the 3a/b/c infectious bronchitis virus coding region was placed downstream of the influenza A virus nucleocapsid protein gene directed the efficient synthesis of 3c as well as nucleocapsid protein, whereas initiation at 3a and 3b could not be detected. Expression of the 3c ORF from this mRNA, however, was abolished when the 3a and 3b coding region was deleted, indicating that 3c initiation is dependent on upstream sequence elements which together may serve as a ribosomal internal entry site similar to those described for picornaviruses.

Base Sequence↗
PubMed1991

A polycistronic mRNA specified by the coronavirus infectious bronchitis virus.

The third largest of the nested set of subgenomic mRNAs (mRNA3) from the coronavirus infectious bronchitis virus (IBV) contains three separate open reading frames (3a, 3b, and 3c) which are not present on the next smallest of the mRNAs, suggesting that this mRNA may be functionally polycistronic. However, although a protein product has been identified from the 3c open reading frame, to date the coding function of 3a and 3b has not been established. We present nucleotide sequence data suggesting that each of the three open reading frames is conserved in a variety of different IBV strains and further show, through the preparation of monospecific antisera against bacterial fusion proteins, that IBV-infected cells contain small amounts of the products of these ORFs. In vitro translation studies using synthetic mRNAs containing the 3a, 3b, and 3c open reading frames suggest strongly that all three proteins can be translated from a single molecular species, and expression studies carried out in intact cells support this conclusion. Thus mRNA3 of IBV appears to be functionally tricistronic.

Amino Acid Sequence↗
PubMed1991

Association of the infectious bronchitis virus 3c protein with the virion envelope.

A highly purified radiolabeled preparation of the coronavirus infectious bronchitis virus (IBV) was analyzed, by immunoprecipitation with monospecific antisera, for the presence of a series of small virus proteins recently identified as the products of IBV mRNAs 3 and 5. One of these, 3c, a 12.4K protein encoded by the third open reading frame of the tricistronic mRNA3 was clearly detectable and was found to cofractionate with virion envelope proteins on detergent disruption of virus particles. These results, together with the hydrophobic nature of 3c and its previously demonstrated association with the membranes of infected cells, suggest strongly that 3c represents a new virion envelope protein, which may have counterparts in other coronaviruses.

Amino Acid Sequence↗
PubMed1991

A study on the mutagenicity of 102 raw pharmaceuticals used in Chinese traditional medicine.

The mutagenic potential of traditional Chinese drugs was studied. After extraction with boiling water and frozen vacuum drying, preparations from 102 raw drugs were tested with the Ames test, and the micronucleus and chromosomal aberration assays in mice in vivo. Astragalus mongholicus Bunge (+S9), Sophora japonica L. (+/- S9) and Eucommia ulmoides Oliv. (+/- S9) were found to significantly induce His+ revertants in Salmonella typhimurium TA98 and/or TA100. Besides these 3, 10 other extracts were positive in the chromosomal aberration and micronucleus assays in mice. They are Datura metal L., Artemisia capillaris Thunb., Carthamus tinctorius L., Forsythia suspensa Thumb., Rehmannia glutinosa f. Hueichingensis (fermented), Paeonia suffruticosa Andr., Platycodon grandiflorum Jacq. DC., Cinnamomum mairei Levl., Notopterygium incisium Ting., and Sophora flavescens Ait. The relevance of these findings is discussed.

Animals↗
PubMed1990

Proenkephalin A and proopiomelanocortin peptides in human cerebrospinal fluid.

Precursors to beta-endorphin (BE) and methionine enkephalin (ME), and proteolytic enzymes that cleave those BE and ME precursors to BE and ME, were determined in several milliliters of human cerebrospinal fluid. Endogenous peptides were purified by reversed-phase high-performance liquid chromatography (HPLC), and were detected with radioreceptor assay (RRA), radioimmunoassay, and mass spectrometry (MS). Total opioid receptor activity measurements and the profile of HPLC-receptor activity of human CSF samples were both used to monitor neuropeptide metabolism. MS data linked the molecular ion of ME to a unique fragment ion. A later-eluting fraction (84 min) in a 90-min HPLC gradient appeared in all HPLC-RRA profiles, contained opioid receptor activity that displaced [3H]etorphine, and the quantitative and qualitative patterns of opioid receptor activity in those profiles both changed within the few minutes that elapsed between acquiring the first and second cerebrospinal fluid samples. That 84-min fraction contained precursors to opioid peptides and was fractionated further with a more shallow 120-min HPLC gradient into three sections that displayed delta-opioid receptor-preferring activity, using [3H]ME as ligand. These three sections were hydrolyzed separately with human cerebrospinal fluid as the source for endogenous neuropeptides to yield products that correlated to immunoreactive BE in section I and immunoreactive ME in section III.

Chromatography, High Pressure Liquid↗