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Biomedical subjects

D Sun

Publications and source records attributed to D Sun.

At least 145 records · Page 8Linked to original sources
PubMed1999

MRP4: A previously unidentified factor in resistance to nucleoside-based antiviral drugs.

Dideoxynucleosides, which are potent inhibitors of HIV reverse transcriptase and other viral DNA polymerases, are a common component of highly active anti-retroviral therapy (HAART) (ref. 1). Six reverse transcriptase inhibitors have been approved for human use: azidothymidine; 2'3'-dideoxycytidine; 2'3'-dideoxyinosine; 2', 3'-didehydro-3'deoxythymidine; 2',3'-dideoxy-3'-thiacytidine; and 4-[2-amino-6-(cyclopropylamino)-9H-purin-9-yl]-2-cyclopentene-1-++ +metha nol. Although drug-resistant HIV strains resulting from genetic mutation have emerged in patients treated with HAART (ref. 1), some patients show signs of drug resistance in the absence of drug-resistant viruses. In our study of alternative or additional mechanisms of resistance operating during antiviral therapy, overexpression and amplification of the MRP4 gene correlated with ATP-dependent efflux of PMEA (9-(2-phosphonylmethoxyethyl)adenine) and azidothymidine monophosphate from cells and, thus, with resistance to these drugs. Overexpression of MRP4 mRNA and MRP4 protein severely impaired the antiviral efficacy of PMEA, azidothymidine and other nucleoside analogs. Increased resistance to PMEA and amplification of the MRP4 gene correlated with enhanced drug efflux; transfer of chromosome 13 containing the amplified MRP4 gene conferred resistance to PMEA. MRP4 is the first transporter, to our knowledge, directly linked to the efflux of nucleoside monophosphate analogs from mammalian cells.

Adenine↗
PubMed1999

A human breast cancer model for the study of telomerase inhibitors based on a new biotinylated-primer extension assay.

Telomerase is an RNA-dependent polymerase that synthesizes telomeric DNA (TTAGGG)n repeats. The overall goal of our work was to establish human cancer models that can be used to design clinical trials with telomerase inhibitors. The objectives of this study were (1) to set up a human breast cancer system that allows evaluation of the effects of telomerase inhibitors in cultured cells using a non-amplified telomerase assay and (2) to test this system using two drugs (cisplatin and TMPyP4) that affect the telomerase expression in breast cancer cells in culture. We first compared the telomerase activity in a variety of human breast cancer cell lines to that of other tumour types using a new biotinylated-primer extension assay. Our method, based on a non-amplified primer extension assay shows the direct incorporation of 32P-labelled nucleotides induced by telomerase on human telomeric primers. The 32P-dGTP labelled telomerase-extended 5'-biotinylated (TTAGGG)3 primer can subsequently be separated using streptavidin-coated magnetic beads. As compared to other non-amplified method, we showed that this procedure improved the characterization and the quantification of the banding pattern resulting from telomerase extension by reducing the radioactive background. Using this method, we observed that telomerase activity varies markedly in a panel of 39 human cancer cell lines. For example, MCF7 breast cancer cells in culture showed intermediate telomerase activity corresponding to 33.8+/-3.4% of that of the HeLa cells (reference cell line). Similarly, the telomere length varied with each cell line (average: 6.24+/-6.16). No correlation between the level of telomerase and telomere length was observed, suggesting that a high processivity is not required to maintain telomeres and that, in some cell lines, another mechanism of telomere elongation can maintain telomere length. From this study, we selected MCF7 and MX1 models that showed reproducible telomerase activity and a relatively limited telomere length for the testing of potential telomere-telomerase interacting agents. Using cisplatin and a new porphyrin-derived compound TMPyP4, we showed that our model was able to detect a down-regulation of the telomerase activity in MCF7 cells in culture and in a human MX1 tumour xenografts. Based on these results, a breast cancer model for evaluating telomerase and telomere interactive agents is proposed.

Animals↗
PubMed1999

Subclassifying atypical squamous cells in Thin-Prep cervical cytology correlates with detection of high-risk human papillomavirus DNA.

Recent studies have proposed subclassifying ASCUS into "favor reactive" (ASFR), "not otherwise specified" (ASNOS), and "favor squamous intraepithelial lesion (SIL)" (ASFS). This study explored the reproducibility of these diagnoses with Thin-Prep cytology and their association with high-risk human papillomavirus DNA (HRHPV). Three pathologists and 1 cytotechnologist with 2 to 25 years of experience reviewed 144 Thin-Prep (Cytyc, Boxborough, MA) specimens previously diagnosed as normal, ASFR, ASNOS, ASFS, and SIL. Interobserver reproducibility was computed with the kappa statistic. The original laboratory diagnosis was compared with the presence of HRHPV types. Interobserver reproducibility for a normal or SIL diagnosis was very good (kappa = .68 and .63). Reproducibility for ASFR, ASNOS, and ASFS ranged from poor to fair (kappa = .21, .19, and .32). In a weighted analysis, kappa values for ASFR/ASNOS and ASFS/SIL were .36 and .62, respectively. HRHPV-positivity for preparations originally diagnosed as N, ASFR, ASNOS, ASFS, and SIL were 5.7%, 8.8%, 17.4%, 47.8%, and 54.5%, respectively. The difference in index of HRHPV for either N or ASFR and ASFS or SIL was significant (P < .001). Reproducibility for ASCUS is generally poor, but better reproducibility is obtained by combining ASFS with SIL and, to a lesser degree, ASNOS with ASFR. ASFS and SIL confer a similar index of HRHPV and merit similar management. ASFR may be managed with cytologic follow-up; but this may depend upon the individual laboratory. HPV testing, in conjunction with cytologic and biopsy follow-up, appears useful for estimating the significance of ASCUS subgroups in laboratory practice.

DNA, Viral↗
PubMed1999

LSIL biopsies after HSIL smears. Correlation with high-risk HPV and greater risk of HSIL on follow-up.

Can the risk associated with a high-grade cervical smear be disregarded when followed by a low-grade biopsy? We examined the distribution of human papillomavirus (HPV) types in such cases to see whether they segregated preferentially with low-risk or high-risk viruses and compared the distribution with that reported in the literature for women with high-grade squamous intraepithelial lesions (HSILs) and low-grade squamous intraepithelial lesions (LSILs). We identified 48 cases of HSIL smears with corresponding LSIL biopsy specimens. Biopsy specimens were tested and typed for HPV by polymerase chain reaction amplification with consensus primers followed by restriction fragment length polymorphism analysis, and HPVs were scored as low-risk or high-risk types. Thirty-seven cases scored positive for HPV DNA: 2 for low-risk HPV types, 17 for high-risk types, and 18 for types of unknown oncogenicity. The prevalence of high-risk HPV was significantly higher than that of low-risk HPV. There was a higher rate of high-risk HPV than that seen in historic unselected LSIL cases. Cases of HSIL cytology/LSIL histology represent a group distinct from unselected LSILs by virtue of their higher prevalence of high-risk HPV types and, therefore, warrant closer clinical follow-up.

Biopsy↗
PubMed1999

Regulatory T cells in experimental allergic encephalomyelitis. I. Frequency and specificity analysis in normal and immune rats of a T cell subset that inhibits disease.

We have shown previously that administration of myelin basic protein (MBP)-reactive T cells to naive Lewis rats induces not only autoimmune encephalomyelitis (EAE) but also a near total resistance to subsequent disease. By isolating the effector cells that are responsible for the resistance, we demonstrated that disease protection paralleled with increased numbers of a CD8+ regulatory T cell (RTC) subset and that co-injection of this RTC subset with encephalitogenic T cells aborted the pathogenic activity of the latter cells. Here, we show that a radio-sensitive splenic population of RTC also exists in naive rats that can be recruited and activated to inhibit the onset of secondary episodes of adoptive EAE. In co-transfer experiments, this protective RTC subpopulation can be isolated to neutralize the pathogenic activity of stimulatory MBP-reactive T cells in vivo. We show that the frequency of RTC with specificity for MBP-reactive T cells in naive rats is two orders of magnitude higher than the frequency of MBP-specific precursors, the activity of RTC increases substantially with age and RTC frequencies increase as a consequence of immunization with MBP-reactive cells lines. In specificity studies, we show that RTC isolated from naive rats and RTC from animals primed with one MBP-reactive cell line show cross-reactive responses to a variety of different MBP-reactive T cell lines. However, following repeated stimulation with a given MBP line, these RTC display a more limited, clonotypic response to the selecting line and assume a uniform CD8 phenotype. Finally, functional studies with RTC indicate that proliferative and lytic specificities do not necessarily correlate and that activated rat RTC are especially lytic for a Fas-sensitive murine cell line.

Adoptive Transfer↗
PubMed1999

The presence of a heterotrimeric G protein and its role in signal transduction of extracellular calmodulin in pollen germination and tube growth

The role of heterotrimeric G proteins in pollen germination, tube growth, and signal transduction of extracellular calmodulin (CaM) was examined in lily pollen. Two kinds of antibodies raised against animal Gzalpha, one against an internal sequence and the other against its N terminus, cross-reacted with the same 41-kD protein from lily pollen plasma membrane. This 41-kD protein was also specifically ADP ribosylated by pertussis toxin. Microinjection of the membrane-impermeable G protein agonist GTP-gamma-S into a pollen tube increased its growth rate, whereas microinjection of the membrane-impermeable G protein antagonist GDP-beta-S and the anti-Galpha antibody decreased pollen tube growth. The membrane-permeable G protein agonist cholera toxin stimulated pollen germination and tube growth. Anti-CaM antiserum inhibited pollen germination and tube growth, and this inhibitory effect was completely reversed by cholera toxin. The membrane-permeable heterotrimeric G protein antagonist pertussis toxin completely stopped pollen germination and tube growth. Purified CaM, when added directly to the medium of plasma membrane vesicles, significantly activated GTPase activity in plasma membrane vesicles, and this increase in GTPase activity was completely inhibited by pertussis toxin and the nonhydrolyzable GTP analogs GTP-gamma-S and guanylyl-5'-imidodiphosphate. The GTPase activity in plasma membrane vesicles was also stimulated by cholera toxin. These data suggest that heterotrimeric G proteins may be present in the pollen system where they may be involved in the signal transduction of extracellular CaM and in pollen germination and tube growth.

Journal Article↗
PubMed1999

Role in cell permeability of an essential two-component system in Staphylococcus aureus.

A temperature-sensitive lethal mutant of Staphylococcus aureus was found to harbor a mutation in the uncharacterized two-component histidine kinase (HK)-response regulator (RR) pair encoded by yycFG; orthologues of yycFG could be identified in the genomes of Bacillus subtilis and other gram-positive bacteria. Sequence analysis of the mutant revealed a point mutation resulting in a nonconservative change (Glu to Lys) in the regulator domain of the RR at position 63. To confirm that this signal transduction system was essential, a disrupted copy of either the RR (yycF) or the HK (yycG) was constructed with a set of suicide vectors and used to generate tandem duplications in the chromosome. Resolution of the duplications, leaving an insertion in either the yycF or the yycG coding region, was achieved only in the presence of an additional wild-type copy of the two open reading frames. Phenotypic characterization of the conditional lethal mutant showed that at permissive growth conditions, the mutant was hypersusceptible to macrolide and lincosamide antibiotics, even in the presence of the ermB resistance determinant. Other mutant phenotypes, including hypersensitivity to unsaturated long-chain fatty acids and suppression of the conditional lethal phenotype by high sucrose and NaCl concentrations, suggest that the role of the two-component system includes the proper regulation of bacterial cell wall or membrane composition. The effects of this point mutation are strongly bactericidal at the nonpermissive temperature, indicating that this pathway provides an excellent target for the identification of novel antibiotics.

Amino Acid Sequence↗
PubMed1999

Localization of periplasmic redox proteins of Alcaligenes faecalis by a modified general method for fractionating gram-negative bacteria.

A lysozyme-osmotic shock method is described for fractionation of Alcaligenes faecalis which uses glucose to adjust osmotic strength and multiple osmotic shocks. During phenylethylamine-dependent growth, aromatic amine dehydrogenase, azurin, and a single cytochrome c were localized in the periplasm. Their induction patterns are different from those for the related quinoprotein methylamine dehydrogenase and its associated redox proteins.

Alcaligenes↗
PubMed1999

Differential regulation of COX-2 expression in the kidney by lipopolysaccharide: role of CD14.

Induction of the inducible cyclooxygenase isoform COX-2 is likely to be an important mechanism for increased prostaglandin production in renal inflammation. We examined the effect of lipopolysaccharide (LPS) on regional renal COX-2 expression in the rat. In the inner medulla, LPS injection (4 mg/kg ip) induced a twofold and 2.5-fold increase in the levels of COX-2 mRNA and COX-2 protein, respectively. In contrast, COX-2 expression in the renal cortex was not significantly altered. COX-2 promoter transgenic mice were created using the 2.7-kb flanking region of the rat COX-2 gene. In these animals, LPS injection induced reporter gene expression predominately in the inner medulla. The LPS receptor CD14, usually regarded as a monocyte/macrophage-specific marker, was found to be abundantly expressed in the inner medulla and in dissected inner medullary collecting duct (IMCD) cells, suggesting that it may mediate medullary COX-2 induction. CD14 was present only at low levels in cortex and cortical segments, including glomeruli. In cultured cells, it was abundant in mouse IMCD (mIMCD-K2) cells and renal medullary interstitial cells, but largely undetectable in mesangial cells and M1 cells, a cell line derived from mouse cortical collecting ducts. In the mIMCD-K2 cell line, LPS significantly induced COX-2 mRNA expression, with concomitant induction of CD14. LPS-stimulated COX-2 expression was reduced by the addition of an anti-CD14 monoclonal antibody to the culture medium. These results demonstrate that LPS selectively stimulates COX-2 expression in the renal inner medulla through a CD14-dependent mechanism.

Animals↗
PubMed1999

Na+-K+-2Cl- cotransporter in immature cortical neurons: A role in intracellular Cl- regulation.

Na+-K+-2Cl- cotransporter has been suggested to contribute to active intracellular Cl- accumulation in neurons at both early developmental and adult stages. In this report, we extensively characterized the Na+-K+-2Cl- cotransporter in primary culture of cortical neurons that were dissected from cerebral cortex of rat fetus at embryonic day 17. The Na+-K+-2Cl- cotransporter was expressed abundantly in soma and dendritic processes of cortical neurons evaluated by immunocytochemical staining. Western blot analysis revealed that an approximately 145-kDa cotransporter protein was present in cerebral cortex at the early postnatal (P0-P9) and adult stages. There was a time-dependent upregulation of the cotransporter activity in cortical neurons during the early postnatal development. A substantial level of bumetanide-sensitive K+ influx was detected in neurons cultured for 4-8 days in vitro (DIV 4-8). The cotransporter activity was increased significantly at DIV 12 and maintained at a steady level throughout DIV 12-14. Bumetanide-sensitive K+ influx was abolished completely in the absence of either extracellular Na+ or Cl-. Opening of gamma-aminobutyric acid (GABA)-activated Cl- channel or depletion of intracellular Cl- significantly stimulated the cotransporter activity. Moreover, the cotransporter activity was elevated significantly by activation of N-methyl-D-aspartate ionotropic glutamate receptor via a Ca2+-dependent mechanism. These results imply that the inwardly directed Na+-K+-2Cl- cotransporter is important in active accumulation of intracellular Cl- and may be responsible for GABA-mediated excitatory effect in immature cortical neurons.

Animals↗
PubMed1999

The natural history of the systemic inflammatory response syndrome and the evaluation of SIRS criteria as a predictor of severity in patients hospitalized through emergency services.

Based on the concept of the systemic inflammatory response syndrome (SIRS), a one-year retrospective study was carried out among a total of 2389 patients transported to the emergency room by ambulance. With respect to 351 patients who had all data necessary for evaluating SIRS criteria in 369 hospitalized patients, 200 met SIRS criteria within 24 hours of admission (24h-SIRS). The mortality rate for 24h-SIRS patients was significantly higher than that of non-SIRS patients. The mortality rate for 24h-SIRS patients increased sequentially as more SIRS criteria were met. In 235 patients who met SIRS criteria during hospitalization (overall-SIRS), 108 had sepsis. Of these, 60 developed severe sepsis, and 50 developed septic shock. The mortality rate for patients who had 3 or more consecutive days of SIRS was significantly higher than that for those with less than 3 consecutive SIRS days. Among 153 patients who had all data necessary for APACHEIII scoring within 24 hours of admission, the mortality rate for SIRS patients whose APACHEIII score was 50 or higher was 40.7%, significantly higher than that of other patients. In conclusion, SIRS criteria were demonstrated to be useful as indicators of severity and for predicting outcome in patients hospitalized through emergency services. Patients who met the following criteria were found to be a high-risk population among hospitalized emergency patients with SIRS: (1) Those who had three or more consecutive days of SIRS. (2) Those whose APACHEIII score was 50 or higher.

Adolescent↗
PubMed1999

An experimental study of electro-acupuncture on auditory impairment caused by kanamycin in guinea pigs.

Frequency following response (FFR) and auditory brain stem evoked potential response (ABR) were used to determine the auditory acuity in evaluating the effect of electro-acupuncture treatment of kanamycin-induced auditory impairment in guinea pigs. The succinate dehydrogenase (SDH) activity and morphological changes of the inner ear receptors were examined under the light and scanning electron microscope in cochlear spread preparations. The results showed that 1) electro-acupuncture was effective but no significant differences were found among the stimulating wave forms; 2) Tinggong (SI 19), Yifeng (SJ 17), Shenshu (UB 23), Sanyinjiao (Sp 6), Zhubin (K 9) and Waiguan (SJ 5) are all effective acupoints, especially the combination of Tinggong (SI 19), Sanyinjiao (Sp 6) and Zhubin (K 9) acupoints; 3) improvement in the cochlear function and excitability of the cortical and lower auditory center and increase of the mitochondrial SDH activity and energy supply in hair cells might contribute to the mechanism of the treatment.

Animals↗
PubMed1999

The quinobenzoxazines: relationship between DNA binding and biological activity.

The quinobenzoxazine compounds, derived from antibacterial quinolones, is active in vitro and in vivo against murine and human tumors. In this contribution, we show that the relative DNA binding affinity of the quinobenzoxazine compounds correlates with their cytotoxicity, their ability to inhibit gyrase-DNA complex formation, and the decatenation of kinetoplast DNA by human topoisomerase II. DNA binding studies with the descarboxy-A-62176 analogue indicate that the beta-keto acid moiety of the quinobenzoxazine compounds plays an important role in their interaction with DNA.

Animals↗
PubMed1999

[Vaginal hysterectomy for large uterus].

OBJECTIVE: The technique and experience of vaginal hysterectomy for large uterus of 25 cases was reported. METHODS: Vaginal hysterectomy for 25 cases of uterine fibroids and adenomyosis were performed. The uterine sizes enlarged as large as 6-16 weeks of pregnancy, mobile, no adhesion by estimation. The technique used for reducing the uterine bulk included bisection, myomectomy, morcellation and coring. The most common way to reducing the uterine bulk was bisection combined with myomectomy and morcellation. RESULTS: Twenty four cases of vaginal hysterectomy were performed successfully. The average weight of uterus for the 25 cases was 360 g(range from 155 g to 755 g), the mean time for the operation was 95 minutes(range from 60 to 150 minutes), the average quantity of blood loss was 150 ml (range from 50 to 400 ml), the average admission duration of postoperation was 5.3 days(range from 3 to 8 days). CONCLUSION: Vaginal hysterectomy is a safe and effective option for removing enlarged uterus. The successful operation was related to the uterine sizes, mobile, operator' skill and experiences.

Adult↗
PubMed1999

The impact of codon 54 variation in intestinal fatty acid binding protein gene on the pathogenesis of diabetes mellitus in Chinese.

OBJECTIVE: To investigate whether or not the intestinal fatty acid binding protein gene (FABP2)-Ala54Thr variation is related to non-insulin dependent diabetes mellitus (NIDDM), obesity, dyslipidemia and glucose stimulated insulin secretion (GSIS) in Chinese. METHODS: The FABP2-Ala54Thr variation was detected by PCR/Hhal digestion in 231 Chinese subjects (116 with normal glucose tolerance (NGT), 54 with impaired glucose tolerance (IGT) and 61 with NIDDM). Plasma glucose, insulin and C-peptide levels before and after 75 g glucose load as well as fasting lipid profile were determined. RESULTS: (1) The Ala54 and Thr54 allele frequencies in Chinese were 0.71 and 0.29 respectively; (2) The FABP2-Ala54Thr variation was neither associated with fasting and post-challenged plasma glucose levels nor with NIDDM; (3) This variation was neither associated with fasting lipid profile nor with obesity; (4) The IGT subjects with genotype Thr54(+) (Thr54 homozygotes and heterozygotes) had lower fasting, 2-hour and total C-peptide levels and smaller AUC representing lesser C-peptide secretion after glucose challenge than those with genotype Thr54(-) (Ala54 homozygotes) (P = 0.04, 0.03, 0.01 and 0.01 respectively). The serum insulin levels changed in the same tendency. CONCLUSIONS: The glucose stimulated insulin secretion (GSIS) reserve of islet beta-cells is more limited in subjects with FABP2-Thr54(+) genotype than in those with FABP2-Thr54(-) genotype. It suggests that FABP2-codon 54 variation might contribute to the insufficient insulin secretion in the development of NIDDM in Chinese.

Alanine↗
PubMed1999

[In situ observation of apoptosis and proliferation in breast cancer and its precancerous lesions].

OBJECTIVE: To investigate the effects of apoptosis and proliferation on malignant transformation of breast precancerous lesions by in situ observation. METHODS: The density and distribution of apoptotic cells were examined in 31 cases with breast cancer, 20 with dysplasia and 20 with hyperplasia, using TUNEL (TdT-mediated dUTP nick end labeling) technique and immunohistochemical method respectively. Eight normal breast tissues were used as the control. RESULTS: In normal breast tissue, apoptotic cells were located at the lacteals and gland follicles, facing the cavity. The proliferating cells were located at the basal layer of mucosa. These characteristics of distribution were lost in breast cancer and precancerous lesions. The density of apoptotic cells and proliferating cells of dysplasia was significantly higher than that in breast cancer and normal mucosa, The density of apoptotic cells and proliferating cells in breast cancer was significantly higher than that in normal mucosa. The ratio of apoptotic to proliferating cells in hyperplasia and dysplasia was 1:1.3-1.4, 1:1.1-1.2 respectively, and 1:2.3-2.4 in breast cancer. CONCLUSION: Abnormality in cell apoptosis and proliferation may play important role in the genesis of breast cancer.

Adult↗
PubMed1999

[Clinical and experimental study on zhixie buye mixture in treating infantile diarrhea complicated with dehydration].

OBJECTIVE: To observe the therapeutic effect of Zhixie Buye mixture (ZXBYM) on infantile diarrhea patient with mild and middle degree of dehydration. METHODS: The controlled observation method was adopted in the clinical and experimental study. RESULTS: There was no significant difference in the treatment of dehydration between the ZXBYM group and ORS group (P > 0.05), while it was more effective for ZXBYM to eliminate the symptoms such as vomiting and abdominal distension (P < 0.01). In addition, it was confirmed that the ion concentration of sodium, potassium, chlorine, glucose and osmotic pressure in the mixture was similar to those in ORS. Animal experiment showed that it has the effect of rat intestine peristalsis suppression and water absorption promotion in colon for ZXBYM. And there was statistical difference in antidiarrhea effect between the mixture and ORS (P < 0.01). CONCLUSION: The ZXBYM is an effective mixture in the treatment of diarrhea with dehydration. It has a good prospect of development and application.

Animals↗