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L F Wang

Publications and source records attributed to L F Wang.

At least 109 records · Page 6Linked to original sources
PubMed1996

[Cholecystokinin-octapeptide (CCK-8) reverses experimental endotoxin shock].

The effects of CCK-8 on the mean arterial pressure (MAP), blood components and path-morphological changes during endotoxin shock (endotoxin, 8 mg/kg b.w.iv.) in rats, as well as on the 24 h mortality rate (MR) of the lead-sensitized rats in endotoxin shock (endotoxin 1 microgram/100 g b.w.iv.) were observed. The results showed that injection of CCK in ES rats led to an increase in MAP, and a reduction of hematocrit and platelet as well as white cell count; the mortality rate decreased and the lesion in main organs lessened. It is suggested that release of endogenous CCK has important protective action during endotoxin shock.

Animals↗
PubMed1995

A single amino-acid change between the antigenically different extracellular serine proteases V2 and B2 from Dichelobacter nodosus.

Dichelobacter nodosus (Dn), the causative organism of ovine footrot, secrets three distinct types of extracellular serine proteases which have been implicated in virulence. Southern analyses have shown that the proteases are encoded by three separate genes, and the genes encoding an acidic protease V5 and a basic protease have already been characterised from virulent Dn strain 198. The gene encoding the third protease type, as represented by acidic protease V2, was isolated from an EcoRI-BamHI library of strain 198 genomic DNA by probing with a polymerase chain reaction (PCR) fragment generated with oligodeoxyribonucleotides based on protease V2 amino acid (aa) sequences. A further clone from an RsaI library was isolated to complete the 5' region of the gene to yield an ORF of 1803 bp encoding a protein precursor of 601 aa. The acidic protease V2 gene, aprV2, shows the same precursor structure as the bprV and aprV5 genes with 72% and 69% similarity at the nucleotide (nt) level and with 73% and 69% similarity at the aa level, respectively. As monoclonal antibodies consistently distinguish the virulent (V) and benign (B) forms of this protease, the gene encoding the acidic protease B2 from benign Dn strain 305 was isolated using the PCR and characterized to investigate the molecular basis for this difference in antigenicity. A 2-bp substitution in a single codon was identified which appeared to be responsible for a change of epitope.

Amino Acid Sequence↗
PubMed1995

Use of a gene-targeted phage display random epitope library to map an antigenic determinant on the bluetongue virus outer capsid protein VP5.

We describe the use of a gene-targeted random epitope library for the mapping of antigenic determinants. A DNA clone encoding the target antigen was digested randomly with DNase I to generate a population of DNA fragments of different sizes and sequences. After size fractionation, small DNA fragments (100-200 bp) were isolated and cloned into the phage expression vector fUSE2 to form an expression library displaying random polypeptide sequences as fusion proteins at the N terminus of the phage gene III protein. This library, termed a gene-targeted random epitope library to distinguish it from totally random synthetic epitope libraries, was then screened by affinity selection for recombinant phages which were specifically bound by the antibody of interest. Using this approach, we have mapped a monoclonal antibody (mAb)-defined epitope on the bluetongue virus outer capsid protein VP5. This epitope is not accessible on the intact virus surface, but is recognised by the immune system of sheep and cattle during virus infection. Although the example given here utilised a DNA fragment of known sequence and the library was screened for a mAb-defined epitope, the strategy described should be equally applicable to genes of unknown sequence and for screening of epitopes using polyclonal antibodies. The approach can also be extended to identify immunodominant epitope from much more complex genome-targeted random epitope library for virus, bacteria and eukaryotic organisms. Other applications of recombinant phages expressing defined immunodominant epitopes include serodiagnosis and vaccine development.

Amino Acid Sequence↗
PubMed1995

Self-assembly, antigenicity, and immunogenicity of the rabbit haemorrhagic disease virus (Czechoslovakian strain V-351) capsid protein expressed in baculovirus.

Rabbit haemorrhagic disease virus (RHDV) capsid protein was expressed in a baculovirus system. Analysis of the expressed product showed that the recombinant protein, which is 60 kDa in size, was antigenic as revealed by its reactions in ELISA and Western blot with the antibodies raised against RHDV. Direct electron microscopy of the cell culture supernatant and the purified protein demonstrated that the capsid protein expressed in insect cells self-assembled to form empty virus-like particles (VLP) which are similar in size and morphology to that of native virus. These particles were immunoreactive with polyclonal anti-RHDV antibodies and with four monoclonal antibodies which recognise conformational epitopes of the virus. The results indicated that the VLPs were morphologically and antigenically indistinguishable from native virus. The recombinant VLPs induced high levels of RHDV-specific antibodies in rabbits and mice following immunisation. The immune response to the VLPs protected the rabbits following challenge with the virulent RHDV. In haemagglutination assays, the VLPs bound to human red blood cells similar to the native virus particles. The recombinant protein and or VLPs is suitable for the development of a rapid, sensitive and reliable test for detection of antibodies to RHDV and for use as a vaccine for domestic rabbits.

Animals↗
PubMed1995

Differentiation of cognate dsRNA genome segments of bluetongue virus reassortants by temperature gradient gel electrophoresis.

The analysis of reassortant viruses has been a valuable tool in the investigation of protein interaction and function in double-stranded (ds) RNA virus research. The differentiation of cognate dsRNA genome segments of reassortants is conventionally achieved by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). However, due to a high degree of sequence homology among different bluetongue virus (BTV) serotypes, it is not uncommon to find that certain cognate dsRNA segments cannot be differentiated by SDS-PAGE. Temperature gradient gel electrophoresis (TGGE) has been shown to be a much more sensitive method of differentiating RNA or DNA fragments of high sequence homology. Here we report the preliminary application of TGGE in analysis of genomic reassortants of two BTV serotypes, 1 and 23. While six out of ten genome segments between BTV-1 and BTV-23 could not be resolved by SDS-PAGE, all of them were differentiated by TGGE. The ability of TGGE to distinguish between dsRNA segments of high sequence homology may also make it useful in the search for BTV genes responsible for defined characteristics, such as virulence, by differentiating wild-type and mutated gene segments of viruses displaying altered phenotypes.

Bluetongue virus↗
PubMed1995

Outline of control practice of endemic fluorosis in China.

Endemic fluorosis is prevalent in China covering 29 provinces, municipalities and autonomous regions. The endemically affected areas can be divided, according to the sources of fluoride, into three types: high fluoride water, pollution from coal burning, and drinking brick tea in excess. Since the 1960's, several pilot surveys of the disease have been made and control programmes carried out in some of the areas. An Expert Consultation Committee on Endemic Fluorosis Control of the Ministry of Public Health was formally established in 1979. A national survey programme and series of working criteria for the disease were drawn up at the First National Congress of Endemic Fluorosis Control in 1981. Under the Central Government of China, administrative organizations and institutions concerned at all levels have been set up, forming a nation-wide network of control. Cooperation and coordination among such departments as health, water conservancy, geology and finance have been achieved in the planning and implementation of control programmes. Since 1980, many projects for improving drinking water quality through de-fluoridation have been completed. At the same time, new methods and technologies for improving stove and grain baking have become widely used in some of the areas where environmental fluoride pollution exists from burning coal. After all the control programmes had been introduced, the incidence of the illness was reduced with some patients making a complete recovery.

China↗
PubMed1995

[Effects of long-term atenolol treatment on beta-adrenoceptor subtypes in rat heart].

The effects of long-term beta 1-AR selective antagonist atenolol treatment on beta-adrenoceptor subtypes were studied by radioligand binding assay, function determination and cAMP accumulation measerment in rat heart. The reasults showed that during long-term administration of atenolol: (1) The density of total beta-AR was increased by approximately 57%; the positive inotropic response and cAMP formation induced by activation of beta-AR were also enhanced. (2) The 125I-pindol competitive inhibition curve for CGP20712A showed that there were no significant difference in the percentage of beta 1- and beta 2-AR sites between the atenolol treated rats and the control rats; pA2 values for selective beta 1-AR antagonist CGP20712A and pKB values for selective beta 1-AR antagonist ICI 118, 551 were not significantly different in the two groups. The results suggested that beta 1- and beta 2-adrenoceptors were upregulated not only in density but also in positive inotropic effect to the same extent.

Animals↗
PubMed1994

Fine mapping of a surface-accessible, immunodominant site on the bluetongue virus major core protein VP7.

The 349-amino-acid major core protein VP7 of bluetongue virus (BTV) is both the most abundant viral structural protein and the major immunogenic serogroup-reactive viral antigen. Previous studies indicated that a conformation-dependent antigenic site, defined by the VP7-specific monoclonal antibody 20E9/B7/G2(20E9), was accessible from the virus surface and that the binding of the monoclonal antibody to this epitope could be blocked specifically by antisera raised against different serotypes of bluetongue virus, suggesting it is a serogroup-specific immunodominant epitope. Using a combination of three different mapping strategies, we have located the 20E9 binding site at the N-terminus of the molecule, between amino acids 30 and 48. The fine mapping of the 20E9 immunodominant epitope will facilitate structure-function analyses of the major core protein and provide new opportunities to improve existing BTV serodiagnosis methods based on this immunogenic site.

Amino Acid Sequence↗
PubMed1994

Fine mapping of a continuous epitope on VP7 of bluetongue virus using overlapping synthetic peptides and a random epitope library.

Two complementary techniques have been used to delineate an epitope on VP7 of bluetongue virus. Two MAbs (F10 and D11), both of which bound within a region spanning amino acids 255 to 274 in the 349 amino acid protein, were used to probe overlapping synthetic peptides covering this region. A pentapeptide, QYPAL, and a hexapeptide, QY-PALT (amino acids 259-264), preferentially bound both MAbs. MAb F10 also reacted with a heptapeptide (TAEIFNV) immediately adjacent to QYPALT. The MAbs were also used to affinity-purify fusion phages from a random hexapeptide library. All phage peptides selected were similar to QYPALT. Comparison of the peptides suggested that residues Q and P at positions 1 and 3 were critical for recognition. Some affinity-purified phages displayed the hexapeptide QYPSLL, which is similar to a sequence in VP7 of another orbivirus, epizootic hemorrhagic disease virus. This finding allowed a potentially cross-reactive site to be identified.

Amino Acid Sequence↗
PubMed1994

Major core protein VP7 of Australian bluetongue virus serotype 15: sequence and antigenicity divergence from other BTV serotypes.

Full-length cDNA of the RNA genome segment coding for the major core protein VP7 of Australian bluetongue virus serotype 15 (BTV-15) has been isolated by reverse transcription-PCR cloning. Comparative analysis indicated that the BTV-15 VP7 sequence had diverged significantly from that of other members of the BTV serogroup. At the amino acid level, BTV-15 VP7 exhibited sequence identities of 80 to 84% with VP7 molecules of other serotypes, significantly lower than the sequence identities of between 93 and 100% observed among other serotypes characterized to date. This was consistent with previous observations that there were significant immunological differences between BTV-15 and other BTV serotypes and that monoclonal antibodies raised against BTV-1 VP7 failed to react with BTV-15 VP7. Recombinant BTV-15 VP7 protein produced from Escherichia coli was largely insoluble, but maintained its immunogenicity. Polyclonal mouse sera raised against the recombinant VP7 protein reacted strongly with VP7 of BTV-15, but weakly with that of BTV-1.

Amino Acid Sequence↗
PubMed1994

Expression in Escherichia coli of the extracellular basic protease from Dichelobacter nodosus.

Dichelobacter nodosus, a Gram-negative obligate anaerobe and the causative agent of ovine footrot, secretes a number of extracellular proteases, one of which is highly basic in nature. The gene (bprV) encoding this basic protease, from virulent strain 198, has been cloned and sequenced. Clone pBR3KB contained the complete bprV gene which constitutively expressed an active protease using its own promoter, when cloned in Escherichia coli. However, levels of protease expression were low and unstable when the clone was expressed in liquid culture. A range of E. coli strains were examined for stable expression; strains NH274 and SURE were found to be better hosts for stable expression than other commonly used E. coli host strains. Stabilization and enhancement of expression was achieved by deletion of the native promoter region and expression from plasmid promoter or promoters, and by modification of culture conditions. The recombinant protease obtained from E. coli was indistinguishable from the native enzyme in size, activity, isoelectric point and immunological properties.

Amino Acid Sequence↗
PubMed1994

The treatment of autoimmune disease in (NZB/NZW)F1 mice with syngeneic photomodulated splenocytes.

(NZB x NZW)F1 (B/W) mice spontaneously develop a disease which is remarkably similar to systemic lupus erythematosus (SLE) in humans. This disease is characterized by the appearance of autoantibodies to double-stranded (ds)DNA and the subsequent development of fatal glomerulonephritis. The prophylactic treatment of B/W mice with syngeneic photomodulated autoimmune spleen cells was found to significantly improve survival, and to inhibit the outgrowth of autoreactive B cells and the production of high-titre IgG anti-dsDNA antibodies. The function of the autoreactive T cells in vivo, however, did not change significantly. Our findings suggested a novel treatment for spontaneously occurring autoantibody-related autoimmune diseases.

Animals↗
PubMed1994

Gene encoding a mammalian epididymal protein.

Polyclonal antibodies raised against a 20 kD epididymal protein (EP20) were used to isolate the cDNA from a human testis lambda gt11 expression library. The nucleotide sequence of the cDNA consisted of 1908 base pairs (bp) containing an open reading frame composed of 1479 bp encoding a polypeptide of 493 amino acid residues. The nucleotide sequence of EP-20 cDNA had 97% identities (282/288) with ESTO 0991 Homo Sapiens cDNA clone HHC M14 in the reverse orientation. The HHC M14 sequence corresponded to a segment in the non-translatable 3'end of EP-20 cDNA. The amino acid sequence of the deduced polypeptide showed no homology with reported polypeptides. The epididymal protein may be involved in sperm maturation and/or capacitation.

Amino Acid Sequence↗
PubMed1994

[Reversal of malignant phenotype of human hepatoma cells by antisense c-ets-2, c-myc and N-ras].

A recombinant retroviral vector was constructed which expressed antisense RNA of c-ets-2, c-myc and N-ras. The pseudotype virus was packaged and rescued by transfection in PA317 cells and used to infect human hepatoma cell line SMMC-7721. After selection with G418, resistant colonies were obtained. Stable integration of retrovirus in infectants was shown by Southern hybridization of genomic DNA and the presence of antisense RNA was detected by RNA dot blot hybridization. It was demonstrated that the antisense RNAs did inhibit the growth of human SMMC-7721 hepatoma cells. The ability to form colony in soft agar and tumorigenicity in nude mice of SMMC-7721 were significantly suppressed by the antisense RNAs. The result implicates the potential value in future cancer gene therapy.

Animals↗
PubMed1994

Calpastatin gene in human testis.

Antibodies present in a serum obtained from an infertile woman interacted with a 17.5 kD glycoprotein (BS-17 component) extracted from human sperm by Western blot. Polyclonal antibodies were raised against the BS-17 component and used to identify positive staining clones from a human testis lambda gt11 expression library. The cDNA encoding the BS-17 component was isolated and its nucleotide sequence determined. The BS-17 cDNA contained 758 nucleotides with an open reading frame of 558 nucleotides encoding a polypeptide consisting of 186 amino acids. The BS-17 cDNA showed 99.7% homology in 758 nucleotides overlap with the 3' terminus of the gene coding calpastatin and 99.5% identity in 186 amino acid overlap with the carboxyl terminus of calpastatin. The BS-17 component of human sperm corresponds to the carboxyl terminus of calpastatin. This conclusion is supported by the finding that the polyclonal antibodies also interacted with a 84 kD protein corresponding to the M(r) of calpastatin.

Amino Acid Sequence↗
PubMed1994

Immunization with synthetic peptide segments of a sperm protein impair fertility in rats.

The nucleotide sequence of the cDNA encoding a sperm protein (rSMP-B) was determined in a previous study. Two peptide segments corresponding to the extracellular domain of the deduced sperm polypeptide were synthesized as multiple antigen peptide (MAP) and designated as rSMP-229 and rSMP-230. Polyclonal antibodies were raised against the two MAPs. Sera obtained from rabbits immunized with rSMP-230 interacted with human and rabbit sperm membrane proteins with estimated molecular sizes of 72 and 20.1 kD, respectively. Adult female and male rats were immunized with the MAPs and their fertilities determined. Immunization of female rats with rSMP-229 and rSMP-230 induced infertility in 25% and 83% of the treated animals, respectively. All male rats immunized with rSMP-229 remained fertile; whereas animals immunized with rSMP-230 did not mate with normal cycling female rats. Three impotent male rats were found to regain their mating potency 45 days after the last booster injection. These findings demonstrated that immunization with rSMP-230 induced a reversible impotency in male rats. Serum testosterone and LH levels were reduced in rSMP-230-immunized male rats and were elevated in rSMP-229-immunized animals. Histopathological examination of sections of testes from male rats immunized with rSMP-230 showed impairment of spermatogenesis and sloughing of germ cells into the lumen of the seminiferous tubules. The testes of male rats immunized with rSMP-229 showed normal morphology and active spermatogenesis with scattered foci of nodular hyperplasia of Leydig cells in the interstitial areas. In conclusion, immunization with synthetic peptide segments corresponding to different domains of a deduced sperm protein induced infertility in a significant number of female rats and transient impotency in male rats.

Amino Acid Sequence↗