Biomedical subjects

L Chen

Publications and source records attributed to L Chen.

At least 235 records · Page 13Linked to original sources

PubMed2001

Repeated formaldehyde effects in an animal model for multiple chemical sensitivity.

Chemical intolerance is a phenomenon observed in multiple chemical sensitivity (MCS) syndrome, an ill-defined disorder in humans attributed to exposure to volatile organic compounds. Amplification of symptoms in individuals with MCS resembles the phenomenon of psychostimulant- and stress-induced sensitization in rodents. We have recently tested in rats the hypothesis that repeated chemical exposure produces sensitization of central nervous system (CNS) circuitry. A rat model of MCS in our laboratory has employed several endpoints of CNS function after repeated formaldehyde (Form) exposure (1 h/day x 5 days/week x 4 weeks). Repeated Form exposure produced behavioral sensitization to later cocaine injection, suggesting altered dopaminergic sensitivity in mesolimbic pathways. Rats given repeated Form also demonstrated increased fear conditioning to odor paired with footshock, implicating amplification of neural circuitry guiding fear responding to a conditioned odor cue. Recent studies examining the effects of repeated Form on locomotor activity during each daily exposure showed a decrease in rearing activity after 12-15 days of Form exposure compared to air-exposed controls. EEG recordings taken 1 week after withdrawal from daily Form revealed altered sleep architecture. Some of the differences in sleep disappeared after subsequent brief (15 min) challenge with Form the next day. Overall, the findings indicate that repeated low-level chemical exposure produces behavioral changes that may be akin to those observed in individuals with MCS, such as greater sensitivity to chemicals manifest as increased anxiety upon chemical exposure and altered sleep and/or fatigue. Study of the underlying CNS changes will provide a basis for mechanistically based animal models for MCS.

Animals↗

PubMed2001

Cariogenic actinomyces identified with a beta-glucosidase-dependent green color reaction to Gardenia jasminoides extract.

The oral bacteria Actinomyces naeslundii and Actinomyces viscosus are known to contribute to the initiation and progression of human dental caries, especially root caries. We report that both A. naeslundii and A. viscosus react with a component in the Gardenia jasminoides extract to produce a distinct green product. This green color reaction was found to be dependent on the bacterial beta-glucosidase. The reaction is specific for cariogenic actinomyces, and it can detect as few as 10(4) cells of A. naeslundii and A. viscosus per ml.

Actinomyces↗

PubMed2001

Kinetic analysis of the steps of the polyomavirus lytic cycle.

Kinetic studies of the accumulation of early and late transcripts, early and late proteins, genomes, and live virus, during the lytic cycle of murine polyomavirus wild-type A2, were carried out in synchronized NIH 3T3 cells released from G(0) by the addition of serum after infection. This first-time simultaneous analysis of all parameters of the virus life cycle led to new insights concerning the transcriptional control at the early-to-late transition. During the early phase, early transcripts were synthesized at very low levels, detectable only by reverse transcription-PCR, from 6 h postinfection (hpi). Large T protein could be detected by 8 hpi (while infected cells were in the G(1) phase). The level of expression of the middle T and small T proteins was lower than that of large T at all times, due, at least in part, to a splicing preference for the large-T 5' splice site at nucleotide 411. A large increase in the level of both early and late transcripts coincided closely with the detection in mid-S phase of viral genome amplification. Thereafter, both classes of transcripts continued to further accumulate up to the end of the experiments (48 hpi). In addition, during the late phase, "giant" multigenomic transcripts were synthesized from the early as well as the late promoter. Thus, a major type of transcriptional control appears to be applied similarly to the transcription of both early and late genes. This view differs from that in the literature, which highlights the enhancement of late transcription and the repression of early transcription. However, despite this parallel transcriptional control, additional regulations are applied which result in higher levels of late compared to early transcripts, as previously described. In the accompanying article, a key role for middle T and/or small T in this late-phase enhancement of early and late transcription is demonstrated (16). Other novel findings, e.g., the synthesis of a very abundant short early promoter proximal RNA, are also described.

3T3 Cells↗

PubMed2001

Role of middle T-small T in the lytic cycle of polyomavirus: control of the early-to-late transcriptional switch and viral DNA replication.

A comparative analysis of the lytic cycle of wild-type polyomavirus and middle T and small T defective mutants was carried out in the A2 genetic background. The results contrast with those obtained in comparisons between the hr-t type and their middle-T small-T-producing partners as previously described (20). The A2-derived mutants were found to share the maturation defect previously described for the hr-t mutants. However, their defect in DNA replication was more acute, resulting in a 5- to 100-fold decrease in the accumulation of viral genomes. Furthermore, their gene expression pattern was affected. A2-derived mutants displayed an early defect resulting in a 4- to 16-h delay in the expression of large T, and an alteration of the early-to-late transcriptional switch. In wild-type A2 infection, this switch is characterized by a large increase in the accumulation of early transcripts followed by late transcripts after the appearance of middle T and small T proteins and the onset of viral DNA replication (L. Chen and M. M. Fluck, J. Virol. 75: 8368-8379, 2001). In the mutant infection, increases in both classes of transcripts were delayed and reduced, but the effect on early transcripts was more pronounced. As has been described previously for the hr-t mutants (E. Goldman, J. Hattori, and T. Benjamin, Cell 13:505-513, 1979), the magnitude of these defects depended upon experimental conditions. Experiments using cytosine beta-arabinofuranoside to reduce genome amplification suggest that the effect of middle T-small T on the transcriptional switch is not solely mediated by the effect of these protein(s) on increasing the number of templates. These data provide the first direct demonstration of an effect of middle T and/or small T in the viral transcription pattern during viral infection. The results agree with previous results obtained with plasmid reporters and with our understanding that the downstream targets of the middle T signaling pathway include three transcription factors that have binding sites in the enhancer domain that play a key regulatory role in the expression of the viral genes.

Antigens, Polyomavirus Transforming↗

PubMed2001

Lowest number of brachial plexus nerve roots required for maintaining normal limb function--an experimental study.

Rat models with different division of nerve roots of the brachial plexus were designed to assess whether single or two nerve roots of the contralateral normal brachial plexus can be divided as donor nerves in treatment of brachial plexus root avulsion. Two hundred and fifty-two SD rats were randomly divided into 13 groups, i.e. five groups with single nerve root division, seven groups with two nerve roots division, and one group with three nerve roots division. Post-operative evaluation by electroneurophysiology and muscular histology was performed at two weeks and two months. Results showed that in groups with single nerve root division, the experimental limb function did not show an apparent damage; in groups with non-neighbouring two nerve roots cutting, it did show a harmful repercussions on the limb at two weeks, but most of them recovered to normal at two months; in groups with neighbouring two nerve roots division and with non-neighbouring three nerve root cutting, the limb function showed a persistent damage. In conclusion, division of nerve root alone or even non- neighbouring two nerve roots of the brachial plexus did not affect the forelimb function of the rat permanently.

Animals↗

PubMed2001

Endothelin-1 and nitric oxide synthase in short rebound reaction to short exposure to inhaled nitric oxide.

On withdrawal of inhalation of nitric oxide (INO) administered after lung injury, pulmonary artery pressure (PAP) and arterial oxygen tension (Pa(O(2))) may deteriorate more than before INO (rebound response). In this study, we investigated the possible roles of endothelin (ET)-1 and nitric oxide (NO) synthase (NOS) activity in the short rebound reaction to short-term inhalation of NO. Twenty-six anesthetized mechanically ventilated piglets were given endotoxin infusion. Twelve animals then received INO (30 parts per million) for two 30-min periods. Nine controls were not given NO. Measurements were made of blood gases and hemodynamic parameters, lung tissue ET-1 expression and NOS activity, and plasma ET-1 concentration. INO decreased PAP and increased Pa(O(2)), but INO withdrawal caused a short rebound reaction with an increase in PAP. Lung tissue expression and plasma concentration of ET-1 increased during INO, and plasma ET-1 increased further after its withdrawal. Activity of constitutive NOS decreased during INO, whereas that of inducible NOS was unchanged. Upregulation of ET-1 and downregulation of NOS activity may have influenced the short rebound reaction to short-term INO.

Administration, Inhalation↗

PubMed2001

Molecular mechanisms by which iron induces nitric oxide synthesis in cultured proximal tubule cells.

Nitric oxide (NO) levels are increased after exposure of cultured proximal tubule cells (PTC) to non-haem iron, potentially contributing to PTC injury in disease states associated with increased iron exposure, including proteinuric renal disease. The mechanisms underlying this observed increase were investigated. After 3 h exposure to 400 microM nitrilotriacetate (NTA)-Fe, inducible nitric oxide synthase (iNOS) mRNA expression was significantly increased, with a corresponding increase in iNOS protein after 12 h. The nuclear binding activity of NFkappaB with 400 microM NTA-Fe was increased, and pyrrolidine dithiocarbamate (PDTC), an antioxidant inhibitor of NFkappaB, prevented both activation of NFkappaB and NO production in response to NTA-Fe. Inhibition of protein tyrosine kinase reduced iNOS mRNA, iNOS protein levels and NO production in response to NTA-Fe. The effect of tyrosine kinase inhibition on NFkappaB activation was variable, with herbimycin but not genistein having an inhibitory effect. Activation of either protein kinase A or C increased iNOS mRNA and protein levels, and NO production in response to NTA-Fe, whereas only the protein kinase C activator phorbol dibutyrate (PDBu) had a stimulatory effect on NFkappaB activation. The protein kinase A activator forskolin did not alter iron-induced activation of NFkappaB. These data suggest that the observed increase in NO production by PTC in response to iron is due to increased transcription of iNOS. The transcriptional regulation of this response is complex and involves NFkappaB, protein tyrosine kinase and the protein kinases A and C.

Animals↗

PubMed2001

Na,K-ATPase gene transfer mitigates an oxidant-induced decrease of active sodium transport in rat fetal ATII cells.

We investigated whether adenovirus-mediated transfer of genes encoding for subunits of the Na,K-ATPase increases transepithelial Na(+) transport in rat fetal distal lung epithelial (FDLE) monolayers and renders them more resistant to hydrogen peroxide injury. FDLE cells, isolated from rat fetuses at a gestational age of 19 to 20 d (22 d = term), were seeded on filters and infected with replication-incompetent human type 5 adenoviruses containing complementary DNAs encoding for rat Na,K-ATPase alpha(1) or beta(1) subunits (ad alpha(1) and ad beta(1), respectively). Once confluent monolayers were formed, the filters were mounted in Ussing chambers and short circuit currents (I(SC)) were measured. Increased levels of alpha(1) or beta(1) subunit proteins after infection with ad alpha(1) and ad beta(1), respectively, were confirmed by Western blot analysis. Baseline I(SC) increased after transfection with 2 plaque-forming units (pfu) of ad beta(1) from 5.1 +/- 0.3 to 6.1 +/- 0.3 microA/cm(2) (mean +/- SEM; P < 0.05). Permeabilization of the apical membrane with amphotericin B caused a large increase in I(SC); the ouabain-sensitive component of the amphotericin B-elicited I(SC) (ouab(max)) was increased from 4.0 +/- 0.2 (n = 69) in controls to 4.8 +/- 0.2 (n = 15), 5.9 +/- 0.3 (n = 53), 6.9 +/- 0.4 (n = 25), 7.7 +/- 0.9 (n = 16) in monolayers infected with 1, 2, 11, and 22 pfu of ad beta(1), respectively; transfection with ad alpha(1) had no effect on any measured variables. Further, transfection with ad beta(1) in comparison to noninfected monolayers resulted in higher baseline and ouab(max) I(SC) after injury with 500 microM H(2)O(2). We conclude that overexpression of the beta(1) subunit of the Na,K-ATPase may help maintain normal levels of vectorial Na(+) transport across ATII cell monolayers in pathologic conditions.

Amphotericin B↗

PubMed2001

The regulation of T cell homeostasis and autoimmunity by T cell-derived LIGHT.

Costimulatory molecules on antigen-presenting cells (APCs) play an important role in T cell activation and expansion. However, little is known about the surface molecules involved in direct T-T cell interaction required for their activation and expansion. LIGHT, a newly discovered TNF superfamily member (TNFSF14), is expressed on activated T cells and immature dendritic cells. Here we demonstrate that blockade of LIGHT activity can reduce anti-CD3-mediated proliferation of purified T cells, suggesting that T cell-T cell interaction is essential for this proliferation. To test the in vivo activity of T cell-derived LIGHT in immune homeostasis and function, transgenic (Tg) mice expressing LIGHT in the T cell lineage were generated. LIGHT Tg mice have a significantly enlarged T cell compartment and a hyperactivated peripheral T cell population. LIGHT Tg mice spontaneously develop severe autoimmune disease manifested by splenomegaly, lymphadenopathy, glomerulonephritis, elevated autoantibodies, and severe infiltration of various peripheral tissues. Furthermore, the blockade of LIGHT activity ameliorates the severity of T cell-mediated diseases. Collectively, these findings establish a crucial role for this T cell-derived costimulatory ligand in T cell activation and expansion; moreover, the dysregulation of T cell-derived LIGHT leads to altered T cell homeostasis and autoimmune disease.

Animals↗

PubMed2001

Characterization of a polyclonal antibody to human pituitary tumor transforming gene 1 (PTTG1) protein.

Pituitary tumor transforming gene 1 (PTTG1), recently cloned from human testis, is a potent oncogene that is expressed in most tumors. However, assessment of its potential value as a prognostic marker is dependent on the development of a suitable antibody. We have developed a rabbit polyclonal antibody, SK601, that is highly specific for the PTTG1 gene product using recombinant PTTG1 protein (24 kD) containing an N-terminal His(6) tag as the immunogen. The antiserum is capable of detecting recombinant PTTG1 protein in ELISA assays at a titer of 1:100,000. Use of the antibody as the probe in Western blotting analyses revealed a single band with the anticipated relative molecular weights of 52 kD from E. coli expressing the GST-PTTG1 recombinant protein, and 56 kD from COS-7 cells transfected with the PTTG1-GFP chimeric construct. A single band with a relative molecular weight of 28 kD was observed in extract of COS-7 cells transfected with PTTG1 cDNA. The antiserum immunoprecipitated a protein of relative molecular weight of 56 kD from the extracts of COS-7 cells transfected with the PTTG1-GFP chimeric construct. Immunohistochemical analysis of COS-7 cells transfected with this construct confirmed that the antibody detected and was specific for expressing the PTTG1-GFP recombinant protein. Screening of various normal human tissues (testis, ovary, and breast) by immunohistochemistry indicated that these tissues did not exhibit staining with the exception of testis, a tissue that had previously been shown to express PTTG1 mRNA. In contrast all of the tumor tissues (testicular tumor, ovarian tumor, and breast tumor) that were assessed exhibited intense staining. The results suggest that antiserum SK601 is highly specific for the PTTG1 protein and therefore should prove useful in further analysis of the expression and interactions of this protein, including its potential application as an immunohistochemical marker of human tumors.

Animals↗

PubMed2001

Expression of the human beta(3)-adrenergic receptor gene in SK-N-MC cells is under the control of a distal enhancer.

Mechanisms of transcriptional regulation of the human beta(3)-adrenergic receptor were studied using SK-N-MC cells, a human neuroblastoma cell line that expresses beta(3)- and beta(1)-adrenergic receptors endogenously. Deletions spanning different portions of a 7-kb 5'-flanking region of the human beta(3)-adrenergic receptor gene were linked to a luciferase reporter and transfected in SK-N-MC, CV-1, and HeLa cells. Maximal luciferase activity was observed when a 200-bp region located between -6.5 and -6.3 kb from the translation start site was present. This region functioned only in SK-N-MC cells. Electrophoretic mobility shift assays of nuclear extracts from SK-N-MC, CV-1, and HeLa cells using double stranded oligonucleotides spanning different portions of the 200-bp region as probes and transient transfection studies revealed the existence of three cis-acting regulatory elements: A) -6.468 kb-AGGTGGACT--6.458 kb, B) -6.448 kb-GCCTCTCTGGGGAGCAGCTTCTCC-6.428 kb, and C) -6.405 kb-20 repeats of CCTT-6.385 kb. These elements act together to achieve full transcriptional activity. Mutational analysis, antibody supershift, and electrophoretic mobility shift assay competition experiments indicated that element A binds the transcription factor Sp1, element B binds protein(s) present only in nuclear extracts from SK-N-MC cells and brown adipose tissue, and element C binds protein(s) present in both SK-N-MC and HeLa cells. In addition, element C exhibits characteristics of an S1 nuclease-hypersensitive site. These data indicate that cell-specific positive cis-regulatory elements located 6.5 kb upstream from the translation start site may play an important role in transcriptional regulation of the human beta(3)-adrenergic receptor. These data also suggest that brown adipose tissue-specific transcription factor(s) may be involved in the tissue-specific expression of the beta(3)-adrenergic receptor gene.

Adipose Tissue, Brown↗

PubMed2001

Bone loss in men with prostate cancer treated with gonadotropin-releasing hormone agonists.

Prostate cancer is the most common visceral malignancy in men. As the tumor is testosterone dependent, a frequent treatment modality involves therapy with GnRH agonists (GnRH-a) resulting in hypogonadism. Because testosterone is essential for the maintenance of bone mass in men, we postulated that GnRH-a therapy would negatively impact skeletal integrity. We compared bone mineral density (BMD), biochemical markers of bone turnover, and body composition in 60 men with prostate cancer (19 men receiving GnRH-a therapy and 41 eugonadal men) and BMD in 197 community-living healthy controls of similar age. BMD was assessed by dual energy x-ray absorptiometry and ultrasound. Biochemical markers of bone turnover, included markers of bone resorption (urinary N-telopeptide) and bone formation markers (bone-specific alkaline phosphatase and osteocalcin). Body composition (total body fat and lean body mass) was assessed by dual energy x-ray absorptiometry. Significantly lower BMD was found at the lateral spine (0.69 +/- 0.17 vs. 0.83 +/- 0.20 g/cm(2); P < 0.01), total hip (0.94 +/- 0.14 vs. 1.05 +/- 0.16 g/cm(2); P < 0.05), and forearm (0.67 +/- 0.11 vs. 0.78 +/- 0.07 g/cm(2); P < 0.01) in men receiving GnRH-a compared with the eugonadal men with prostate cancer. Significant differences were also seen at the total body, finger, and calcaneus (all P < 0.01). BMD values in eugonadal men with prostate cancer and healthy controls were similar. Markers of bone resorption (urinary N-telopeptide) and bone formation (bone-specific alkaline phosphatase) were elevated in men receiving GnRH-a therapy compared with those in eugonadal men with prostate cancer. Men receiving GnRH-a also had a higher percent total body fat (29 +/- 5% vs. 25 +/- 5%; P < 0.01) and lower percent lean body weight (71 +/- 5% vs. 75 +/- 5%; P < 0.01) compared with eugonadal men with prostate cancer. In conclusion, men with prostate cancer receiving androgen deprivation therapy have a significant decrease in bone mass and increase in bone turnover, thus placing them at increased risk of fracture.

Aged↗

PubMed2001

Inhibin and p27 interact to regulate gonadal tumorigenesis.

Tumor suppressors function as antiproliferative signaling proteins, and defects in these genes lead to uncontrolled cell proliferation and cancer. For example, absence of the tumor suppressor p27(Kip1), a cyclin-dependent kinase inhibitor (CKI), results in increased body size, hyperplasia of several organs including the testes, and cancer in mice. Similarly, lack of inhibins, alpha/beta heterodimeric members of the transforming growth factor-beta (TGFbeta) superfamily, causes testicular and ovarian tumors of the granulosa/Sertoli cell lineage beginning at 4 weeks of age and adrenal tumors in gonadectomized mice. Neither the cell cycle alterations in the absence of inhibin nor the cause of the increased testis size in the p27 knockout mice is known. To study the molecular (cell cycle) changes that result from absence of inhibins, we analyzed the regulation of cell cycle proteins in gonadal tumors derived from inhibin alpha knockout mice (Inha(-/-)). Northern blot analyses demonstrate that cyclin-dependent kinase 4 (Cdk4) and cyclin D2 mRNA levels are elevated, and immunohistochemistry shows that p27 protein levels are decreased in both ovarian and testicular tumors from Inha(-/-) mice. These findings suggest that increased Cdk4/cyclin D2 (positive) activity and decreased p27 (negative) activity is causal for gonadal tumor formation. To test this hypothesis, we generated double mutant mice lacking both p27 and inhibin alpha to determine whether the tumor suppressors p27 and inhibin have additive suppressor activity in the gonads. Like Inha(-/-) mice, p27(-/-)Inha(-/-) mice demonstrate elevated serum activin levels, ovarian and testicular tumors, and a resultant lethal cachexia-like syndrome. However, whereas 95% of the Inha(-/-) female mice die by 18 weeks of age, 100% of the p27(-/-)Inha(-/-) female mice are dead by 8 weeks. Similarly, 95% of the Inha(-/-) single mutant males die by 13 weeks while 100% of the p27(-/-)Inha(-/-) male mice die by 10 weeks. Moreover, tumor foci in p27(-/-)Inha(-/-) mice can be observed as early as 2 weeks of age in males and as early as 4 weeks in females. These findings demonstrate that absence of both inhibin and p27 in mice causes earlier development of ovarian and testicular tumors and earlier death compared with absence of inhibin alone.

Activins↗

PubMed2001

Effects of exercise training on myocardial fatty acid metabolism in rats with depressed cardiac function induced by transient ischemia.

The effects of exercise training on metabolic and functional recovery after myocardial transient ischemia were investigated in a rat model. Male Wistar Kyoto rats were subjected either to a 30-min left coronary artery occlusion followed by reperfusion or to a sham operation. At 4 weeks after operation, the rats were randomly assigned either to sedentary conditions or to exercise training for 6 weeks. In the ischemic rats, pinhole SPECT (single photon emission computed tomography) imaging with thallium-201 (201Tl) and 123I-(rho-iodophenyl)-3-R,S-methylpentadecanoic acid (BMIPP) showed a reduction of both myocardial perfusion and fatty acid metabolism in the risk zone of the left ventricle (LV). The LV was dilated and the ejection fraction was decreased after ischemic injury. The severity score showed a significant decrease on both 201Tl and BMIPP (201Tl, from 19.9+/-2.7 to 17.0+/-2.2, p<0.05; BMIPP, from 21.5+/-2.4 to 18.6+/-1.9, p<0.05) after exercise training in the ischemic trained rats, but did not change significantly in their sedentary counterparts. Plasma levels of free fatty acids normalized in the ischemic trained rats, but elevated in the ischemic sedentary rats (0.53+/-0.05 vs 0.73+/-0.06 mmol/L, p<0.05). Furthermore, the trained rats had a significant increase in LV stroke volume (0.25+/-0.02 vs 0.21+/-0.01 ml/beat, p<0.05) and adaptive cardiac hypertrophy. These findings demonstrate that adaptive improvements in myocardial perfusion, fatty-acid metabolism and LV function were induced by exercise training after transient ischemia.

Animals↗

PubMed2001

1,N(2)-propanodeoxyguanosine adduct formation in aortic DNA following inhalation of acrolein.

Recent reports indicate that many of the cytotoxic and health-threatening components of environmental tobacco smoke (ETS) reside in the vapor phase of the smoke. We have reported previously that inhalation of 1,3-butadiene, a prominent vapor phase component of ETS, accelerates arteriosclerotic plaque development in cockerels. In this study we asked whether inhaled acrolein, a reactive aldehyde that is also a prominent vapor-phase component of ETS, damages artery-wall DNA and accelerates plaque development. Cockerels inhaled 0, 1, or 10 ppm acrolein mixed with HEPA-filtered air for 6 hr. Half were killed immediately (day 1 group) for detection of the stable, premutagenic 1,N(2)-propanodeoxyguanosine acrolein adduct (AdG3) in aortic DNA via a (32)P-postlabeling/HPLC method, and half were killed after 10 days (day 10 group) for indirect assessment of adduct repair. In the day 1 group, acrolein-DNA adducts were 5 times higher in the 1 and 10 ppm groups than in HEPA-filtered air controls. However, in the day 10 group, adduct levels in the 1 and 10 ppm acrolein groups were reduced to the control adduct level. For the plaque studies, cockerels inhaled 1 ppm acrolein (6 hr/day, 8 weeks), mixed with the same HEPA-filtered air inhaled by controls. Plaque development was measured blind by computerized morphometry. Unlike butadiene inhalation, acrolein inhalation did not accelerate plaque development. Thus, even though repeated exposure to acrolein alone has no effect on plaque size under the exposure conditions described here, a single, brief inhalation exposure to acrolein elicits repairable DNA damage to the artery wall. These results suggest that frequent exposure to ETS may lead to persistent artery-wall DNA damage and thus provide sites on which other ETS plaque accelerants can act.

Acrolein↗

PubMed2001

Human papillomavirus 16 E6 is associated with the nuclear matrix of esophageal carcinoma cells.

AIM: To explore the etiologic role of HPV infection in esophageal carcinoma, and the association of HPV-16 E6 with the nuclear matrix of carcinoma cells. METHODS: Two esophageal carcinoma cell lines,EC/CUHK1 and EC/CUHK2, were tested for HPV-16 E6 subgenetic fragment by polymerase chain reaction amplification of virus DNA associated nuclear matrix. RT-PCR and immunocytochemistry were also used to visualize the expression of E6 subgene in the cells. RESULTS: The HPV-16 E6 subgenetic fragment was found to be present in nuclear matrix-associated DNA, E6 oncoprotein localized in the nucleus where it is tightly associated with nuclear matrix after sequential extraction in EC/CUHK2 cells. It was not detected, however, in EC/CUHK1 cells. CONCLUSION: The interaction between HPV-16 E6 and nuclear matrix may contribute to the virus induced carcinogenesis in esophageal carcinoma.

Antigens, Nuclear↗

PubMed2001

Phosphorylation of hepatic stimulator substance on mitogen-activated protein kinase in BEL-7402 hepatoma cells.

OBJECTIVE: To obtain more insight into information of signal transduction of EGF-receptor-mediated pathway response to the stimulation of hepatic stimulator substance (HSS). METHODS: HSS was extracted from weanling rat liver and partially purified. Bioactivity of HSS was confirmed with its ability to proliferate hepatoma cell in vitro. Meanwhile, a rat recombinant HSS vector was constructed and expressed in BL-21 E. Coli. Mitogen-activated protein kinase (MAPK) activation marked by phosphorylation at Thr202/Tyr204 was determined by Western blot. RESULTS: The molecular weight of the biochemically purified HSS was found identically to that of the recombined HSS as expressed in the prokaryotic cells. After the treatment of HSS, cellular MAPK phosphorylation was initiated obviously at 15 min and maintained to 30 min. In comparison with EGF, MAPK phosphorylation as stimulated by HSS appeared less intensive and later time-kinetics as well. The HSS induction on cellular MAPK phosphorylation was gradually inhibited by PD98059, a specific inhibitor of MAPK kinase (MEK). A complete blockade was seen at 100 micromol/L of PD98059. CONCLUSIONS: The involvement of HSS on MAPK activation implies that this liver-specific growth factor might take part in, either individually or as combined with other growth factors, the regulation of TPK signaling cascade during hepatocyte proliferation.

Animals↗

PubMed2001

Loss of heterozygosity at 19q13.3 is associated with locally aggressive neuroblastoma.

A genome-wide allelic analysis of neuroblastoma (NB) revealed a previously undescribed increased incidence of loss of heterozygosity (LOH) on chromosome arm 19q13 primarily affecting stages 3 and 4N disease. Further allelic analysis of chromosome 19q13 in a cohort of 116 NB patients using 17 polymorphic microsatellite markers identified the shortest common region of loss between D19S606 and D19S112 at 19q13.3. In some cases, clonal LOH at 19q13 was acquired during the course of disease, and deleted clones remained after cytotoxic therapy. In multivariant analysis, 19q13 LOH was associated with overall survival in local-regional International Neuroblastoma Staging System stages 1, 2, and 3 patients and was specifically present in tumors at the site of recurrence.

Chromosomes, Human, Pair 19↗