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Biomedical subjects

Q Gao

Publications and source records attributed to Q Gao.

At least 181 records · Page 10Linked to original sources
PubMed1994

Identification of novel mutations that confer drug resistance in the human immunodeficiency virus polymerase gene.

We generated variants of the human immunodeficiency virus type 1 (HIV-1) that are resistant to 2',3'-dideoxycytidine (ddC) and 2',3'-didehydro-3'-deoxythymidine (d4T) by in vitro selection in MT-4 cells. Portions of flanking protease and integrase sequences as well as the complete reverse transcriptase (RT) open-reading frame of these viruses were cloned and sequenced, using polymerase chain reaction (PCR)-based methods. Mutations were observed at amino acid position 65 (Lys-->Arg; AAA-->AGA) when ddC was employed in the selection procedure and at site 50 (Ile-->Thr; ATT-->ACT) when d4T was used. We confirmed the ability of these mutations to confer diminished sensitivity for these compounds by site-directed mutagenesis, in which these mutations were inserted into the pol gene of infectious recombinant HXB2-D DNA. Viruses that contained the site 65 mutation possessed approximately 5-10 fold resistance against ddC when compared with wild-type HXB2-D. The site 50 mutation conferred approximately 30-fold resistance to d4T in these same assays. Similar results were obtained using primary cord blood lymphocytes in drug resistance assays, indicating that these mutations could confer drug resistance in more than one cell type and that the respective mutations could be expressed in cells of primary origin. No cross-resistance against 3'-azido-3'-deoxythymidine (AZT) was noted for either the site 65 or 50 mutations.

Drug Resistance↗
PubMed1994

[Influence of anterior occlusal contacts of complete dentures on mastication].

The photocclusion and mastication analysis of the complete dentures with or without the anterior occlusal contacts were made. The results indicated that there was no obvious differences among the maximum occlusal contact intensities of each tooth position in central occlusion. Under the functional condition the activity of the anterior temporal muscles was influenced and the mastication efficiency was decreased by the relatively stronger anterior occlusal contacts.

Aged↗
PubMed1994

A clinical study on treatment of vascular complications of diabetes with the sugar--reducing and pulse--invigorating capsule.

The capsule is effective in replenishing qi, nourishing yin, activating blood, and resolving stasis. It can correct abnormalities in blood rheology, improve fat metabolism, enhance functioning of the islets of Langerhans, lower blood sugar, and alleviate clinical symptoms. Efficacious also against the chief vascular complications of diabetes, it helps in abating myocardial anoxia, improving left heart function, stimulating blood circulation to the brain, resisting coagulation and resolving thrombosis, also dilating the arteries of the legs. It is of some benefit in early diabetic retinopathy and renal diseases.

Adult↗
PubMed1993

Immortalization with herpesvirus saimiri modulates the cytokine secretion profile of established Th1 and Th2 human T cell clones.

T blasts of six established human CD4+ T cell clones with defined Ag specificity and cytokine secretion profile (3 Th1 and 3 Th2) were immortalized with Herpesvirus saimiri (HVS) and compared with their uninfected counterparts for their ability to proliferate, produce cytokines, and express cytolytic activity. HVS-transformed Th1 and Th2 clones neither substantially changed their original surface markers nor lose their ability to proliferate in response to their specific Ag but did acquire the ability to proliferate in response to contact signals delivered by SRBC or autologous APC alone. In addition, transformation by HVS substantially enhanced the lectin-dependent cytolytic activity of Th1 clones and enabled noncytolytic Th2 clones to exert cytolytic activity. HVS-transformed Th1 clones but not their uninfected counterparts spontaneously transcribed and secreted Th1-type cytokines (IL-2, IFN-gamma, and TNF-beta) and such a production was further enhanced by stimulation with either SRBC or PMA plus anti-CD3 mAb. HVS transformed but not uninfected Th2 clones constitutively expressed both IL-4 and IL-2 mRNA and secreted IFN-gamma. Stimulation with PMA plus anti-CD3 mAb induced uninfected Th2 clones to secrete high amounts of IL-4 and IL-5 but not Th1-type cytokines, whereas the same HVS-transformed Th2 showed minimal IL-4 and IL-5 secretion with concomitant high production of IL-2, IFN-gamma, and TNF-beta. Transformation by HVS also resulted in up-regulation of TNF-alpha and IL-3 production by both Th1 and Th2 clones. The ongoing proliferation of HVS-transformed clones was partially inhibited by either anti-IL-2 or anti-IL-3 antibodies and virtually abolished by the combined addition of the two anticytokine antibodies, suggesting that both IL-2 and IL-3 can function as autocrine growth factors for HVS-transformed Th1 and Th2 clones.

Cell Transformation, Viral↗
PubMed1993

Increased percentage of CD3+, CD57+ lymphocytes in patients with rheumatoid arthritis. Correlation with duration of disease.

OBJECTIVE: To determine whether a small CD3+ lymphocyte population expressing 110-kd CD57 antigens (HNK1) is expanded in patients with rheumatoid arthritis (RA), as it is in patients who have undergone bone marrow transplantation and patients with the acquired immunodeficiency syndrome, and to investigate whether it is involved in the pathogenesis of RA. METHODS: The phenotype of CD3+, CD57+ lymphocytes was analyzed by flow cytometry, and correlations between the percentage of these cells in the blood and various clinical and biologic parameters were investigated. RESULTS: The percentage of CD3+, CD57+ lymphocytes was increased in RA patients compared with controls. These lymphocytes expressed T cell receptor alpha/beta. Eighty percent expressed the CD8 accessory molecule, and 20% expressed the CD4 accessory molecule. The leukocyte common antigen CD45RA isoform was expressed by these CD3+, CD57+ lymphocytes in blood. The HLA-DR antigen was expressed in synovial fluid but not in blood. Finally, the percentage of these lymphocytes in the blood correlated with the duration of RA. CONCLUSION: The expansion of the CD3+, CD57+ lymphocyte population and their activation in the synovial fluid of RA patients suggest that these cells are involved in the inflammatory process.

Adult↗
PubMed1993

Contrasting effect of transforming growth factor type beta 1 (TGF-beta 1) on proliferation and interleukin-2 receptor expression in activated and rapidly cycling immature (CD3-CD4-CD8-) thymocytes.

Transforming growth factor beta (TGF-beta) is a cytokine with immunoregulatory properties that acts negatively on T lymphocyte proliferation. However, with the EL 4-6.1 variant of the murine thymoma EL 4 activated with phorbol ester and/or interleukin-1 (IL-1), we recently found that it up-regulates interleukin-2-receptor (IL-2R) expression. Since EL 4-6.1 cells share phenotypic and functional characteristics with the immature thymic subset lacking CD4 and CD8 accessory molecules (DN), we investigated the effect of TGF-beta 1 on the IL-2R 55kD alpha chain expression and proliferation of activated DN cells and especially in DN cells that do not express CD3. We observed that TGF-beta 1 was able to increase both the percentage of CD3-DN cells expressing IL-2R alpha chains and the expression of IL-2R alpha chain in these cells. This stimulatory effect of TGF-beta 1 was distal from early transduction events. In addition, TGF-beta 1 was found to modulate CD3-DN cell proliferation. During differentiation in the thymus, CD3-DN cells transiently express the IL-2R alpha chain of the IL-2R and these IL-2R+ CD3-DN cells are preprogrammed to down-regulate the IL-2R alpha chain and up-regulate the CD4 and CD8 accessory molecule. We thus also tested the effect of TGF-beta 1 on IL-2R alpha chain expression in these in vitro differentiating CD3-DN cells. We found that TGF-beta 1 neither significantly affected IL-2R expression nor changed CD4 or CD8 expression. Hence, in CD3-DN cells, the effect of TGF-beta 1 on IL-2R expression seems to be restricted to proliferating cells.

Animals↗
PubMed1993

Generation of drug-resistant variants of human immunodeficiency virus type 1 by in vitro passage in increasing concentrations of 2',3'-dideoxycytidine and 2',3'-dideoxy-3'-thiacytidine.

We selected in vitro human immunodeficiency virus type 1 variants that are resistant to each of 2',3'-dideoxycytidine (ddC) and the racemic mixture of 2',3'-dideoxy-3'-thiacytidine (BCH-189). The median effective concentrations of ddC and BCH-189 obtained for the resistant viruses ranged between 10 and 50 times above those for parental wild-type strains, and extensive cross-resistance was observed against 2',3'-dideoxyinosine (ddI) but not 3'-azido-3'-deoxythymidine (AZT). Two dimer compounds, in which either AZT and ddI or AZT and BCH-189 were linked through phosphodiester linkages, did not permit the emergence of variants resistant to BCH-189, ddI, or AZT but were ineffective at inhibiting the replication of AZT-resistant viruses.

Acquired Immunodeficiency Syndrome↗
PubMed1993

The same mutation that encodes low-level human immunodeficiency virus type 1 resistance to 2',3'-dideoxyinosine and 2',3'-dideoxycytidine confers high-level resistance to the (-) enantiomer of 2',3'-dideoxy-3'-thiacytidine.

Variants of human immunodeficiency virus type 1 that display 500- to 1,000-fold resistance to the (-) enantiomer of 2'-deoxy-3'-thiacytidine and approximately 4- to 8-fold resistance to 2',3'-dideoxycytidine and 2',3'-dideoxyinosine have been generated through in vitro selection with the former compound. The polymerase regions of several of these resistant viruses shared a codon alteration at site 184 (ATG-->GTG; methionine-->valine), a mutation previously associated with low-level resistance to 2',3'-dideoxycytidine. The biological relevance of this mutation for the (-) enantiomer of 2'-deoxy-3'-thiacytidine was confirmed by site-directed mutagenesis with the HXB2-D clone of human immunodeficiency virus type 1.

Antiviral Agents↗
PubMed1993

Synthesis of DNA, rRNA, and protein by Rickettsia prowazekii growing in untreated or gamma interferon-treated mouse L929 cells.

The syntheses of DNA, rRNA, and protein by Rickettsia prowazekii growing in mouse fibroblastic L929 cells were measured at various times after the addition of gamma interferon (IFN-gamma) to correlate the inhibition of a site of macromolecular synthesis with the established IFN-gamma-induced inhibition of rickettsial growth. A method was developed to measure the syntheses of DNA, rRNA, and protein by R. prowazekii during a 2-h pulse-labeling period while the rickettsiae were growing within cultured host cells that had intact macromolecular synthesis. This method involved incubation of the rickettsia-infected cells with a radioactive precursor (H3 32PO4 or Tran35S-label), purification of the rickettsiae, purification of rickettsial nucleic acids, and analysis of rickettsial nucleic acids and proteins by electrophoresis and autoradiography. A key feature of the method involved the use of calculated specific activities from a densitometric analysis of gels and autoradiograms, a procedure that made the data independent of rickettsial recovery. Rickettsial DNA and rRNA syntheses were both inhibited 12 h after the addition of IFN-gamma to infected cultures, whereas the synthesis of rickettsial proteins was not inhibited at this time. In contrast, at 20 h after the addition of IFN-gamma, rickettsial DNA, rRNA, and protein syntheses were all inhibited.

Animals↗
PubMed1993

Eupenifeldin, a novel cytotoxic bistropolone from Eupenicillium brefeldianum.

Eupenifeldin was isolated from cultures of Eupenicillium brefeldianum ATCC 74184 by extraction and crystallization. The compound was identified as a pentacyclic bistropolone on the basis of spectral data and its complete structure was established by single-crystal X-ray analysis. The compound is cytotoxic against the HCT-116 cell line and has in vivo antitumor activity in the P388 leukemia model.

Animals↗
PubMed1993

Clinical correlates and molecular basis of HIV drug resistance.

It has been widely reported that zidovudine (ZDV)-resistant variants of human immunodeficiency virus type 1 (HIV-1) can be isolated from patients undergoing prolonged therapy with this drug. At the same time, treatment of HIV-infected individuals with ZDV and other forms of nucleotide therapy, including didanosine (ddI), have enabled patients to live longer than would otherwise be the case and to enjoy improved quality of life. HIV resistance to ZDV, ddI, and other nucleosides is attributable to a series of point mutations within the pol gene of HIV-1 that encodes the viral enzyme, reverse transcriptase (RT). This is not surprising as the virus is known to replicate at high rates in infected individuals; moreover the RT that mediates transcription of proviral DNA from viral genomic RNA is known to be highly error prone. Thus, mutants of HIV-1, which possess a drug-resistance phenotype and genotype, may be expected to emerge under the selective pressure of long-term antiviral chemotherapy. This article describes a novel mutation at site 184 within the pol gene that accounts for resistance against both ddI and zalcitibine (ddC). HIV drug resistance occurs most commonly in individuals with low CD4 cell counts who have progressed to more serious forms of disease. Moreover, viruses obtained from patients with AIDS generally display higher levels of resistance, relative to pretreatment isolates, than do viruses from patients with more-limited illness. Although observations of drug resistance can be correlated with disease progression and a weakened immune system, it is still unclear whether a cause-and-effect relationship exists. Because of the error-prone nature of viral RT and the fact that the HIV-1 genome can mutate efficiently, it can be anticipated that viral drug resistance may emerge for all forms of nucleotide therapy to be offered in the future. In addition, resistance may also become apparent with regard to drugs that block HIV replication by acting at sites within the viral replication cycle other than RT.

CD4-Positive T-Lymphocytes↗
PubMed1992

DNA-ditercalinium interactions: implications for recognition of damaged DNA.

Ditercalinium is unique among known DNA-binding chemotherapeutic agents. Ditercalinium treatment of Escherichia coli causes cell death by provoking malfunction of the (A)BC exinuclease excision DNA repair system. In this report, we describe the three-dimensional X-ray structure of a ditercalinium-[d(CGCG)]2 complex in detail with an analysis of both the structure and its implications. Ditercalinium bisintercalates in the DNA fragment; the positively-charged linker of the drug interacts with the major groove. The DNA retains an underwound, right-handed, double-helical conformation with a bend of around 15 degrees in the helical axis. One striking feature of the complex is the extensive interaction of ditercalinium with guanines in contrast to the near absence of interaction with cytosines. The terminal cytosines in particular are "unstacked" from the rest of the complex. A systematic comparison of the three-dimensional structure of the DNA-ditercalinium complex with those of triostin A and nogalamycin (chemotherapeutic agents that act by conventional mechanisms) allows us to suggest a general model for recognition by the (A)BC exinuclease excision repair system. It is commonly hypothesized that the same distorted conformation of DNA results from modification by each member of a diverse family of DNA-damaging agents. This specific conformation of DNA would then be recognized by the (A)BC exinuclease excision repair system. Alternatively, we propose that each of these damaging agents causes local instability of DNA (but not necessarily a common conformation) and that the (A)BC exinuclease excision repair system recognizes excessive or unusual deformability of damaged DNA in comparison to normal DNA.

Carbazoles↗
PubMed1992

Identification and characterization of a 30K glycoprotein of herpes simplex virus type 2.

A herpes simplex virus type 2 (HSV-2) type-specific monoclonal antibody (MAb), CH-A9, precipitated a glycoprotein with an M(r) of approximately 30,000 (g30K) from extracts of HSV-2-infected BHK cells labelled with [3H]leucine, [14C]fructose or [3H]glucosamine. The M(r) of this glycoprotein is lower than those of other HSV glycoproteins. Immunoassays of BHK cells infected with HSV-1-HSV-2 intertypic recombinants localized the gene encoding the target antigen of MAb CH-A9 to the unique long (UL) region at map units 0.490 to 0.564. Tunicamycin effectively inhibits N-linked glycosylation of g30K, which suggests that g30K may be modified by addition of N-linked oligosaccharides and that the amino acid sequence may contain Asn-X-Ser or Asn-X-Thr. The g30K was also purified on an immunoadsorbent column consisting of MAb CH-A9 linked to Sepharose 4B and was shown to be an HSV-2 type-specific antigen by indirect ELISA. The glycoprotein could induce HSV-2 type-specific neutralizing antibody in BALB/c mice. This evidence suggests that g30K may be a novel glycoprotein of HSV-2.

Animals↗
PubMed1992

In vitro selection of variants of human immunodeficiency virus type 1 resistant to 3'-azido-3'-deoxythymidine and 2',3'-dideoxyinosine.

Drug-resistant variants of human immunodeficiency virus type 1 (HIV-1) have been isolated by in vitro selection. MT-4 cells were infected with either a laboratory strain (HIV-IIIB) or a clinical isolate (no. 187) of HIV-1 and maintained in medium containing subeffective concentrations of the drugs 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI). By gradually increasing the drug concentration in the culture medium during propagation of the virus on fresh MT-4 cells, we were able to isolate variants of HIV-IIIB and clinical isolate 187 which showed up to 100-fold increases in resistance to the drugs. The drug resistance phenotypes remained stable after propagation of the variants in the absence of drug pressure for over 2 months. However, variants resistant to one drug showed little or no cross-resistance to the other, suggesting that the genetic bases for resistance to the compounds differed. Genotypic analysis of these nucleoside-resistant variants by polymerase chain reaction (PCR) with primer pairs previously shown to correspond to mutations responsible for resistance to AZT was also carried out. A heterogeneity of genotypes was observed, with known mutations at pol codons 70 and 215 occurring in most of the AZT-resistant variants generated from either HIV-IIIB or clinical strain 187. However, mutations in codons 67 and 219 were less frequently detected, and none of these changes were observed in each of four variants resistant to ddI. Cloning and sequencing studies of the reverse transcriptase coding region of two of the isolates were also performed and confirmed the PCR data that had been obtained. In addition to previously described mutation sites responsible for resistance to AZT, an HIV-IIIB-resistant variant was shown to be mutated at positions 108 (Val----Ala) and 135 (Ile----Thr), while a resistant variant of strain 187 was mutated at positions 50 (Ile----Val) and 135 (Ile----Val).

Base Sequence↗
PubMed1992

Novel mutation in the human immunodeficiency virus type 1 reverse transcriptase gene that encodes cross-resistance to 2',3'-dideoxyinosine and 2',3'-dideoxycytidine.

We have used the technique of in vitro selection to generate variants of human immunodeficiency virus type 1 (HIV-1) that are resistant to 2',3'-dideoxyinosine (ddI) and cross-resistant to 2',3'-dideoxycytidine (ddC). The complete reverse transcriptase (RT)-coding regions, plus portions of flanking sequences, of viruses possessing a ddI-resistant phenotype were cloned and sequenced by polymerase chain reaction (PCR)-based methods. We observed that several of these viruses possessed mutations at amino acid sites 184 (Met-->Val; ATG-->GTG) and 294 (Pro-->Ser; CCA-->TCA). These mutations were introduced in the pol gene of infectious, cloned HXB2-D DNA by site-directed mutagenesis. Viral replication assays confirmed the importance of site 184 with regard to resistance to ddI. The recombinant viruses thus generated displayed more than fivefold-greater resistance to ddI than parental HXB2-D did. Moreover, more than fivefold-greater resistance to ddC was also documented; however, the recombinant viruses continued to be inhibited by zidovudine (AZT). No resistance to ddI, ddC, or AZT was introduced by inclusion of mutation site 294 in the pol gene of HXB2-D. PCR analysis performed on viral samples obtained from patients receiving long-term ddI therapy confirmed the presence of mutation site 184 in five of seven cases tested. In three of these five positive cases, the wild-type codon was also detected, indicating that mixtures of viral quasispecies were apparently present. Viruses possessing a ddI resistance phenotype were isolated from both subjects whose viruses contained only the mutated rather than wild-type codon at position 184 as well as from a third individual, whose viruses appeared to be mostly of the mutated variety.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Two-site pan-species monoclonal antibody ELISA for detection of blood stage malaria antigen.

A two-site pan-species monoclonal antibody sandwich ELISA (MAb-MAb ELISA) was developed to detect both Plasmodium vivax and P. falciparum antigens in whole blood impregnated on filter paper. In this assay, the plates were coated with pan-species MAb 3F9 and another pan-species MAb M26-32 conjugated with alkaline phosphatase was used for detection of bound antigen. The sensitivity of this assay was 5, 10 and 10 parasites per 10(6) erythrocytes for cultured P. falciparum, patient-derived P. vivax and P. falciparum, respectively. The coincidence rates for this assay were 93% (92/99) with healthy individuals and 93% (42/45) with microscopically confirmed vivax malaria cases. After two weeks treatment, 77.7% (14/18) of vivax malaria were still positive by this assay but with diminished level of reactivities [corrected].

Animals↗
PubMed1992

Generation of nucleoside-resistant variants of HIV-1 by in vitro selection in the presence of AZT or DDI but no by combinations.

Several laboratories have shown that AZT-resistant variants of HIV-1 can be isolated from patients who have received prolonged therapy with this drug. Our laboratory has now been able to generate HIV-1 variants resistant to both AZT and ddI, in tissue culture, by using step-wise increases in the concentrations of each of these compounds over a 10-week period. This work has been performed by culturing wild-type clinical strains of HIV-1 as well as the HIV-3b laboratory strain of this virus under such conditions. The ID50 values obtained for the resistant viruses thus generated vary between 50-100 times above those of the parental wild-type strains in each case. Furthermore, we have identified several new mutation sites in the HIV-1 pol gene that are responsible for the observed resistance to AZT and ddI. We have not succeeded, however, in generating drug-resistant strains of HIV-1, under conditions in which several compounds or anti-viral agents were simultaneously present during the in vitro selection process. Combinations of drugs which failed to yield drug-resistant variants included AZT plus ddI, AZT plus alpha-interferon, and ddI plus alpha-interferon. These findings indicate that HIV drug resistance is less likely to occur in tissue culture when combinations of drugs are used, and provide rationale for the development of combination clinical trials for treatment of HIV-associated disease.

Amino Acid Sequence↗