Biomedical subjects

W Jia

Publications and source records attributed to W Jia.

At least 55 records · Page 3Linked to original sources

PubMed1999

[Preparation of oral microspheres carrying V. cholera vaccine and its target's distribution].

To prepare oral biodegradable microspheres carrying V. cholera vaccine, the major outer membrane protein (OMP, MW = 41 kd) as a common antigen of cholera Vibriae was obtained from the classical strain Inaba 569 B, and the OMP was encapsulated in the biodegradable delivery system comprising Poly (DL-Lactide)-Co-Poly(ethylene glycol)microspheres. The average size of the microspheres was less than 5 microns, the amount of OMP encapsulated in microspheres was 15.3%. It was found that microspheres were taken up in Peyer's patches and then distributed in spleen, liver and mesenteric lymph nodes after oral administration.

Animals↗

PubMed1999

[Construction of Streptococcus mutans surface protein antigen eukaryotic expression vector pcDNA3-PAc. I. Extraction and purification of plasmid DNA pPC41 and pcDNA3].

OBJECTIVE: To Extract and purify plasmid DNA pPC41 and pcDNA3. METHODS: Bacteria were collected and lyzed by the alkaline lysis method. The plasmid pPC41 carrying structural gene pac and clone expression vector pcDNA3 were extracted and purified from E. coli clone by the polyethylene glycol precipitating method, the electroelution into dialysis bags method, the low melting-temperature agarose gel and glass fiber column chromatography. The concentration, purity and molecular weight were determined by spectrophotometry, restriction enzyme and agarose gel electrophoresis. RESULTS: The concentration of plasmid DNA of this experiment in general was 0.12-0.24 g/L, and A260/A280 of the polyethylene glycol precipitating method, the electroelution into dialysis bags method, the low metling-temperature agrose gel and glass fiber column chromatography were respectively 1.9, 2.2, 2.2 and 2.6. Molecular weight of pPC41 and pcDNA3 was respectively 10.6 kb and 5.4 kb. CONCLUSION: The four methods can obtain pure plasmid DNA. Purity of plasmid DNA obtained by glass fiber column chromatography method is the highest in the four methods. Glass fiber column chromatography is one of the effective methods getting highly purified plasmid DNA.

Antigens, Surface↗

PubMed1999

[Study of bacterial adhesion to prosthetic valve materials in vitro].

This article reports a study of the bacterial adhesion to prosthetic valve materials in vitro. The method for assessing the degree of bacterial adhesion to prosthetic valve materials was established primarily. The capacities of staphylococcus aureus(SA), staphylococcus epidermidis(SE), Escherichia Coli(EC) and Pseudomonas aeruginosa(PA) for adhesion to Dacron, Pyrolytic carbon and polytetrofluoroethylene (PTFE) were quantitatively determined by the plate counting and Gamma-ray counting of 125I radiolabeled bacteria in vitro. The results showed that the capacities of four types of bacteria for adhesion to Dacron, Pyrolytic carbon and PTFE coincided with the bacterial-growth curve. The capacities of four bacteria for adhering to Dacron were stronger. The adhesion of SE to Pyrolytic carbon was the strongest. The adhesion of PA Kept up a high level. The capacities of EC and PA for adhering to PTFE were the strongest. The results indicate that the capacities of different types of bacteria for adhesion to the same prosthetic valve material are different, and the capacities of one type of bacteria for adhesion to different materials are also different.

Bacterial Adhesion↗

PubMed1998

[Effects of Trp64Arg mutation in the beta 3-adrenergic receptor gene on body fat, plasma glucose level, lipid profile, insulin secretion and action in Chinese].

OBJECTIVE: To ascertain the association of the Trp64Arg mutation in the beta 3-adrenergic receptor (ADRB3) gene with the total body fat and fat distribution, plasma glucose level, lipid profile, insulin secretion and action in Chinese. METHODS: Genotyping of the Trp64Arg mutation in the ADRB3 gene as well as measurment of clinical variables body mass index(BMI), percent of desirable weight(PDW), area of regional adipose tissue depots, fasting lipid profile, and also, plasma glucose, insulin, C-peptide, free fatty acids levels before and after glucose challenge and blood pressure were performed in 308 Chinese, of whom, 161 were nondiabetics and 147, type 2 diabetic subjects. RESULTS: (1) The frequency of Arg64 allele of ADRB3 gene in Chinese was similar to that in Japanese but slightly higher than that in Caucasians. (2) No association of any of the clinical varaibles with Trp64Arg mutation was observed in nondiabetics. (3) The Trp64Arg mutation in the ADRB3 gene was associated with BMI(P=0.019), PDW(P=0.001) and waist circumference (P=0.045) in type 2 diabetic subjects. In female diabetic subjects, more significant association with BMI(P=0.001), PDW(P=0.001), waist circumference(P=0.023), and also, association with abdominal and femoral subcutaneous fat area(P=0.032 and P=0.023) were observed. The frequencies of Arg64 allele were 21%, 14% and 4% in female diabetics with obesity, over-weight and normal-weight respectively (P=0.026). Logistic regression analysis showed that BMI was the only independent clinical variable related to the Trp64Arg mutation in the ADRB3 gene (P=0.0027). CONCLUSION: The Trp64Arg mutation in ADRB3 gene contributes to the variation of total body fat in Chinese type 2 diabetic subjects.

Adipose Tissue↗

PubMed1998

Mini-proinsulin and mini-IGF-I: homologous protein sequences encoding non-homologous structures.

Protein minimization highlights essential determinants of structure and function. Minimal models of proinsulin and insulin-like growth factor I contain homologous A and B domains as single-chain analogues. Such models (designated mini-proinsulin and mini-IGF-I) have attracted wide interest due to their native foldability but complete absence of biological activity. The crystal structure of mini-proinsulin, determined as a T3R3 hexamer, is similar to that of the native insulin hexamer. Here, we describe the solution structure of a monomeric mini-proinsulin under physiologic conditions and compare this structure to that of the corresponding two-chain analogue. The two proteins each contain substitutions in the B-chain (HisB10-->Asp and ProB28-->Asp) designed to destabilize self-association by electrostatic repulsion; the proteins differ by the presence or absence of a peptide bond between LysB29 and GlyA1. The structures are essentially identical, resembling in each case the T-state crystallographic protomer. Differences are observed near the site of cross-linking: the adjoining A1-A8 alpha-helix (variable among crystal structures) is less well-ordered in mini-proinsulin than in the two-chain variant. The single-chain analogue is not completely inactive: its affinity for the insulin receptor is 1500-fold lower than that of the two-chain analogue. Moreover, at saturating concentrations mini-proinsulin retains the ability to stimulate lipogenesis in adipocytes (native biological potency). These results suggest that a change in the conformation of insulin, as tethered by the B29-A1 peptide bond, optimizes affinity but is not integral to the mechanism of transmembrane signaling. Surprisingly, the tertiary structure of mini-proinsulin differs from that of mini-IGF-I (main-chain rms deviation 4.5 A) despite strict conservation of non-polar residues in their respective hydrophobic cores (side-chain rms deviation 4.9 A). Three-dimensional profile scores suggest that the two structures each provide acceptable templates for threading of insulin-like sequences. Mini-proinsulin and mini-IGF-I thus provide examples of homologous protein sequences encoding non-homologous structures.

Amino Acid Sequence↗

PubMed1997

Comparison of the neuroprotective effects of APV and bcl-2 in glutamate-induced cell death.

The neuroprotective effects of the NMDA receptor blocker APV were compared with those of bcl-2 in a glutamate-induced excitotoxic cell death model in cultured rat cortical neurons. Exposure to 100 microM glutamate for 5 h caused approximately 95% of the cultured neurons to die in 24 h. The NMDA-selective antagonist D-(-)-2-amino-5-phosphonopentanoate (APV) protected the neurons effectively when applied prior to or soon after glutamate treatment. However, infection with a viral vector expressing the proto-oncogene bcl-2 strongly protected neurons even if applied as late as 8 h following the glutamate insult. These data provides evidence that APV blocks an early stage of the death cascade in response to elevations of glutamate. By contrast bcl-2 appears to act at a fairly late stage in the cell death process and these results suggest a possible clinical role in treatment of ischemic brain disorders.

2-Amino-5-phosphonovalerate↗

PubMed1997

Sequence analysis of gene 3, gene 4 and gene 5 of avian infectious bronchitis virus strain CU-T2.

We have previously reported the nucleotide sequences of gene 2 (spike (S) protein gene), gene 6 (nucleocapsid (N) protein gene), and the 3' end untranslated region of a novel avian infectious bronchitis virus (IBV) strain, CU-T2 [Jia et al. (1995) Arch. Virol. 140, 259 271]. In the present report we describe the sequences of the remaining genes of this strain (gene 3, 4 and 5) with the exception of gene 1 (RNA polymerase gene). Gene 3 contained three open reading frames (ORFs), 3a, 3b and 3c of 174, 195 and 282 nucleotides (nt), respectively. Gene 4 (membrane (M) protein gene) consisted of 749 nt with a single ORF of 687 nt. Gene 5 contained two ORFs, 5a and 5b, with 198 and 249 nt, respectively. Thus, in total, there were 7349 nt from the 5' end of S protein gene to the 3' end of the CU-T2 genome. The overall nt sequence homologies between gene 3, 4, and 5 of CU-T2 and those of other strains were between 84.1-90.8%, 85.8-88.8% and 90.4 96.4%, respectively. The predicted amino acid (aa) sequence homologies revealed that gene 3b and 5b were more conserved than 3a, 3c and 5a. Each individual gene of CU-T2 strain (with the exception of the RNA polymerase gene) had a different level of homology with the homologous gene of other strains, suggesting that the evolution of IBV strains in general has been a complex, and as yet, poorly understood process.

Amino Acid Sequence↗

PubMed1997

Intracerebral recombinant HSV-1 vector does not reactivate latent HSV-1.

Herpes simplex virus type-1 (HSV-1) has been used for gene delivery in the nervous system for the treatment of brain tumors and other neurological diseases. In most protocols, recombinant viruses containing the gene of interest are directly injected into the brain. Since many people harbor latent wild-type HSV-1 virus in sensory ganglia and other regions of the nervous system, there is a potential risk that the injected recombinant virus may reactivate the latent wild-type virus to cause severe encephalitis. The present study used two rat latent infection models to evaluate this risk. Adult rats were infected with wild-type kos by cornea scarification or by intracerebral injection, and after the establishment of latency, the ICP6(-) strain hrR3 was injected intracerebrally. In the control group, the latent virus was reactivated by treatment of cadmium sulfate. Viral shedding from tears was detected by incubation with Vero cells, and the trigeminal ganglia, cortical tissue and the eyes were collected to detect reactivated wild-type virus by RT-PCR. Our results showed that while the reactivated wild-type virus was readily detectable in the cadmium-sulfate-treated animals, intracerebrally infected hrR3 did not reactivate the latent virus in either the corneal model or the cerebral model. These results indicate that intracranial injection of partially defective recombinant virus may bear little risk of reactivating latent wild-type virus harbored in the sensory ganglia or the brain in our animal model.

Animals↗

PubMed1997

Elevated nonesterified fatty acid concentrations in severe preeclampsia shift the isoelectric characteristics of plasma albumin.

We previously hypothesized that the endothelial cell dysfunction observed in women with preeclampsia might be caused by an imbalance between circulating very low density lipoproteins and a cytoprotective pI 5.6 isoform of albumin, referred to as toxicity preventing albumin (TxPA). An accurate simplified method was developed to quantify TxPA in small volumes of pregnancy plasma by gel electrofocusing. This assay revealed that circulating TxPA concentrations in women with severe preeclampsia were significantly reduced compared to those in normal pregnant women and women with benign transient hypertension of pregnancy. Nonesterified fatty acids (NEFA) and triglycerides were elevated in plasma from women with severe preeclampsia compared to those in plasma from the two control groups. The inverse correlation between TxPA and NEFA values led us to analyze the NEFA bound to plasma albumin. Gas chromatography and mass spectrometry demonstrated no qualitative differences in the specific fatty acids bound to plasma albumin in severe preeclamptic and normal pregnant women. However, the quantity of NEFA bound to albumin was greater in preeclampsia plasma (2.5 mol NEFA/mol albumin) compared to that in normal pregnancy plasma (0.8 mol NEFA/mol albumin), accounting for the acidic pI shift observed in albumin from the former patients. Functional assays demonstrated that human very low density lipoprotein particles were toxic to human umbilical vein endothelial cells in vitro, but this toxicity was prevented by the addition of TxPA albumin to the culture medium.

Adult↗

PubMed1997

Mitochondrial tRNA(Leu(UUR)) gene mutation diabetes mellitus in Chinese.

OBJECTIVE: To ascertain the prevalence and clinical features of mitochondrial tRNALeu(UUR) gene nucleotide (nt) 3243 A-->G mutation diabetes mellitus in Chinese and to establish the approaches for genetic diagnosis of this subtype of diabetes during routine daily clinical practice. METHODS: Mitochondrial nt 3243 A-->G mutation was screened among 207 unrelated non-insulin-dependent diabetes mellitus (NIDDM) patients by using polymerase chain reaction (PCR)/Apa I restriction endonuclease digestion. Samples with positive result were confirmed by DNA sequence analysis. Genetic and clinical analysis were carried out in family members of the proband with positive genetic diagnosis. RESULTS: Positive results were found in two subjects during screening of unrelated NIDDM patients, which account for 2.4%-11.1% of NIDDM subjects with uncommon clinical features (with early age-of-onset, with low body-mass-index and on insulin therapy) or with positive family history of diabetes consistent with or unable to exclude from maternal inheritance. Genetic diagnosis was positive in 10 out of 25 family members in the pedigrees of these two probands. CONCLUSIONS: Mitochondrial tRNALeu(UUR) gene nt 3242 A-->G mutation should be considered in diabetic patients with the above-mentioned clinical features. The genetic diagnosis in daily clinical practice can be performed with allele specific priming amplification, or PCR/Hae III or Apa I digestion which are technically simple to perform and diagnostically easy to define.

Adolescent↗

PubMed1996

Mapping the functional surface of insulin by design: structure and function of a novel A-chain analogue.

Functional surfaces of a protein are often mapped by combination of X-ray crystallography and mutagenesis. Such studies of insulin have yielded paradoxical results, suggesting that the native state is inactive and reorganizes on receptor binding. Of particular interest is the N-terminal alpha-helix of the A-chain. Does this segment function as an alpha-helix or reorganize as recently proposed in a prohormone-convertase complex? To correlate structure and function, we describe a mapping strategy based on protein design. The solution structure of an engineered monomer ([AspB10, LysB28, ProB29]-human insulin) is determined at neutral pH as a template for synthesis of a novel A-chain analogue. Designed by analogy to a protein-folding intermediate, the analogue lacks the A6-A11 disulphide bridge; the cysteine residues are replaced by serine. Its solution structure is remarkable for segmental unfolding of the N-terminal A-chain alpha-helix (A1 to A8) in an otherwise native subdomain. The structure demonstrates that the overall orientation of the A and B chains is consistent with reorganization of the A-chain's N-terminal segment. Nevertheless, the analogue's low biological activity suggests that this segment, a site of clinical mutation causing diabetes mellitus, functions as a preformed recognition alpha-helix.

Amino Acid Sequence↗

PubMed1996

Native and non-native structure in a protein-folding intermediate: spectroscopic studies of partially reduced IGF-I and an engineered alanine model.

The structure of a metastable folding intermediate of human insulin-like growth factor I (IGF-I) and an engineered model are investigated by circular dichroism and two-dimensional 1H NMR spectroscopy. The intermediate, which contains two of three native disulfide bonds, was trapped by acid quenching and isolated by reverse-phase HPLC. The reduced cysteine residues were mapped to residues 47 and 52 (corresponding to A6-A11 in insulin). In the native state this disulfide bridge anchors an adjoining amphipathic alpha-helix (helix 2; residues 42 to 49) against the hydrophobic core. Comparison of CD and 1H-NMR spectra demonstrates that the acid-quenched intermediate is partially folded and contains elements of native secondary and tertiary structure. Spectra are similar to those of an equilibrium model in which the reduced cysteine residues are replaced by alanine. Complete 1H-NMR sequential assignment of the alanine model has been obtained and demonstrates that removal of the disulfide bond is associated with local unfolding of the adjoining alpha-helix. Native secondary structure (including helices 1 and 3) is otherwise retained and defines a folded subdomain. Long-range nuclear Overhauser effects (NOE) within this subdomain are similar to those of native IGF-I; no non-native NOE is observed. Our results support the hypothesis that folding of the insulin motif is directed by a subset of native structural elements and that these elements form at an early step in the pathway. Formation of helix 2, despite its prominence in the native state, is likely to represent a late step. Hydrophobic collapse of this segment appears to precede helix formation.

Alanine↗

PubMed1996

Analysis of the serotype-specific epitopes of avian infectious bronchitis virus strains Ark99 and Mass41.

The Ark and Mass serotype-specific epitopes of infectious bronchitis virus were studied by immunofluorescence and immunoprecipitation of mutant and recombinant spike glycoproteins (S protein) expressed in mouse L cells. Serotype-specific monoclonal antibodies could bind to the recombinant proteins of Ark99 and Mass41 expressed from the chimeras in which the N-terminal thirds of the S1 sequences were reciprocally exchanged. Therefore, it appears that the respective serotype-specific epitopes of both strains were localized within the C-terminal two-thirds of the S1 proteins. Deletion and insertion of a five-amino-acid fragment on the S1 proteins of Ark99 and Mass41, altered the serotype-specific epitopes. This result implies that the five-amino-acid insertion on the S1 protein of the Ark serotype was involved in determining the conformation of the protein, probably acting as a spacer. In addition, it appears that an interaction between sequences of the N-terminal third and the remaining portion of the S1 protein determines the tertiary structure of the protein as well as the conformation of the serotype-specific epitope.

Animals↗

PubMed1995

A novel variant of avian infectious bronchitis virus resulting from recombination among three different strains.

An antigenic variant of avian infectious bronchitis virus (IBV), a coronavirus, was isolated and characterized. This strain, CU-T2, possesses a number of unusual features, which have not been previously observed in IBV. The S1 glycoprotein of CU-T2 carries virus-neutralizing and serotype-specific epitopes of two IBV serotypes, Arkansas (Ark) and Massachusetts (Mass). Sequence analysis revealed that the virus, originally an Ark serotype, has acquired the Mass-specific epitope by mutation(s). This provides evidence that point mutations may lead to generation of IBV antigenic variants in the field. It was further observed that two independent recombination events involving three different IBV strains had occurred in the S2 glycoprotein gene and N protein gene of CU-T2, indicating that genomic RNA recombination in IBV may occur in multiple genes in nature. It was especially significant that a sequence of Holland 52 (a vaccine strain) had replaced half of the N gene of CU-T2. This proves that recombination among vaccine strains is contributing to the generation of IBV variants in the field. Based on these observations it is predicted that every IBV field isolate could have unique genetic nature. Therefore, several recently reported diagnostic and serotyping methods of IBV which are based on dot-blot hybridization, restriction fragment length polymorphism (RFLP), and polymerase chain reaction (PCR), may not reveal the true antigenic and/or genetic nature of IBV isolates, and may in fact yield misleading information.

Animals↗

PubMed1995

Color changes in dry-pigmented maxillofacial elastomer resulting from ultraviolet light exposure.

Five dry pigments and one maxillofacial elastomer were evaluated for color changes (delta E) resulting from prolonged exposure to two types of ultraviolet light. The elastomer, pigments, and pigmented elastomers were subjected to each ultraviolet light source for 400, 600, and 1800 hours, and delta E color shifts were determined. The unpigmented elastomer underwent minimally perceptible color change after 600 hours of exposure to both types of ultraviolet light. Two pigments underwent substantial color change after 400 hours, whereas the remaining three pigments remained color stable after 1800 hours. It was concluded that for the materials tested, early color changes in a prosthesis may be the result of degradation of certain ultraviolet light-susceptible pigments, whereas longer term color shifts may be caused by color changes within the elastomer.

Analysis of Variance↗

PubMed1995

[Culture of Streptomyces and purification of streptavidin].

For preparation and purification of Streptavidin, two synthetic mediums and a procedure for purifying Streptavidin were studied. We found that Streptomyces avidinii grew vigorously and showed typical colony characterization. At the sametime, these synthetic mediums were in favour of producing Streptavidin with a maximal production of 15.2 micrograms/ml by using DEAE 52 column for purification of Streptavidin. This purification procedure is very simple and easy, and could be widely used.

Bacterial Proteins↗

PubMed1995

[Isolation and identification of extrem thermophilic bacteria from hot springs of Sichuan and Tibet].

Eleven strains of extrem thermophilic bacteria belonging to the Bacillus Stearothermophiles were isolated from the hot springs of Sichuan and Tibet. The cells were gram-positive, sporulating and motile. The optimum temperature for growth was between 65 degrees C to 70 degrees C; the maximum 94 degrees C, and minimum 40 degrees C. The colour of colony was yellow to bright orange. The G+C content of the DNA in strains has been found to be in the range 48.4-53.15 mol%.

China↗

PubMed1995

[Identification of streptavidin].

In this report, streptavidin (SA) was further identified on the basis of our previous researches in SA isolation and purification. 23.2 mg of SA productivity could be obtained after purification by DEAE-52 column from 1 L of culture supernatant. The spectrum scanning of ultraviolet absorption was congruent with that of the Sigma product. 2 bands appeared on the SDS-PAGE electrophoresis of SA. The molecular weight of SA was 72 881 Dalton with PI 6.0; the averaged biotin-binding activity was 18 micrograms/mg. The results confirmed that the methods of isolation and purification of SA developed by authors were reliable for immunological application.

Bacterial Proteins↗