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Biomedical subjects

M Yan

Publications and source records attributed to M Yan.

At least 91 records · Page 5Linked to original sources
PubMed1997

Immuno and functional characterization of CFTR in submandibular and pancreatic acinar and duct cells.

Cystic fibrosis results from defective Cl- channel activity mediated by the cystic fibrosis transmembrane conductance regulator (CFTR) gene product. In the gastrointestinal tract this is manifested in abnormal salivary secretion and pancreatic insufficiency. This is generally attributed to defective Cl- transport by the ductal system of the glands. We provide the first immunocytochemical and functional evidence for expression of CFTR protein and Cl- current in rat and mouse submandibular gland (SMG) and pancreatic acinar cells, a site proximal to the ductal system of these secretory glands. Monoclonal and polyclonal antibodies recognizing COOH-terminal epitopes of CFTR show that duct and acinar cells from the two glands express CFTR in the luminal membrane. Specificity of the polyclonal antibody was verified by absence of staining in duct and acinar cells of the SMG of cf-/cf- and delta F/delta F mice. Identification of CFTR in acinar cells was aided by demonstrating coexpression of CFTR and type 3 inositol 1,4,5-trisphosphate receptors in the luminal pole of acini and absence of type 3 inositol 1,4,5-trisphosphate receptors in ducts. Electrophysiological characterization in single SMG duct and acinar cells shows the presence of a protein kinase A-activated, voltage- and time-independent, ohmic Cl- current and absence of repolarization-dependent tail currents, all of which are kinetic properties of the CFTR-dependent Cl- channel. In addition, the channel was activated by the nonhydrolyzable ATP analog 5'-adenylylimidodiphosphate and the benzimidazalone NS-004. Channels activated by all activators were inhibited by glibenclamide and a known inhibitory antiserum [anti-CFTR-(505-511)]. Combined immunologic, functional, and pharmacological evidence allows us to conclude that acinar cells of the SMG and pancreas express functional CFTR-dependent Cl- channels. Because this site is proximal to the duct, modification of activity of this channel in acinar cells is likely to contribute to abnormal salivary secretion and pancreatic insufficiency typical of cystic fibrosis.

Animals↗
PubMed1997

Diagnosis of lysosomal storage disorders: evaluation of lysosome-associated membrane protein LAMP-1 as a diagnostic marker.

Early diagnosis of lysosomal storage disorders (LSDs), before the onset of irreversible pathologies, will be a key factor in the development of effective therapies for many of these disorders. Newborn screening offers a potential mechanism for the early detection of these disorders. From studies of both normal and LSD-affected human skin fibroblasts we identified the lysosome-associated membrane protein LAMP-1 as a potential diagnostic marker. We have developed a sensitive method for the quantification of this protein with a time-resolved fluorescence immunoassay. A soluble form of LAMP-1 was observed in plasma samples, and determination of 152 unaffected individuals gave a median value of 303 micrograms/L with the 5th and 95th percentile at 175 and 448 micrograms/L respectively. Plasma samples from 320 LSD-affected individuals representing 25 different disorders were assayed. We observed that 17 of the 25 disorder groups tested had > 88% of individuals above the 95th percentile of the control population, with 12 groups having 100% above the 95th percentile. Overall, 72% of patients had LAMP-1 concentrations above the 95th percentile of the unpartitioned control population. We suggest that LAMP-1 may be a useful marker in newborn screening for LSDs.

Adolescent↗
PubMed1997

[Evaluation of brain evoked potentials in the detection of subclinical hepatic encephalopathy in cirrhotics].

Brainstem auditory evoked potentials (BAEP), visual evoked potentials (VEP) and short latency somatosensory evoked potentials (SSEP) were examined in 30 nonalcoholic liver cirrhotics without clinically detectable hepatic encephalopathy and 30 healthy controls. In the cirrhotics, all peak latencies of the three kinds of evoked potentials, the interpeak latencies (IPLs) I-V, III-V of BAEP and the IPLs N13-N20, N13-P25 of SSEP were significantly prolonged compared with the controls, respectively. The amplitudes of P100, N125 of VEP were significantly lowered in the cirrhotics than those of the controls. Abnormal BAEP test and abnormal SSEP test results were found in both 60% of the cirrhotics, while VEP tests showed abnormalities in only 36.7%. In total, abnormal evoked potential test of one or more kinds were found in 90% of the cirrhotics. It is concluded that in cirrhotic patients, before the appearance of clinically encephalopathy, there were already brain evoked potential abnormalities or brain function changes. Our results argue in favor of the three kinds of evoked potential as the combined investigation for the sensitive and objective diagnosis of subclinical hepatic encephalopathy in patients with nonalcoholic cirrhosis.

Adult↗
PubMed1997

[The preventing function of garlic on experimental oral precancer and its effect on natural killer cells, T-lymphocytes and interleukin-2].

In order to study the effect and mechanism of garlic on preventing oral precancer, we divided randomly 32 Wistar rats into two groups. The garlic group was painted with garlic solution on the hard palatal mucosae. The control group was applied with distilled water that is equal in quantity. Then, chemical carinogen 4-nitroquinoline 1-oxide (4NQO) was painted on the same sites for both groups, three times weekly. Eight rats were randomly killed in the 10th, 13th week. The hard palatal mucosae were examined with light microscope. Meanwhile, lymphocytes were isolated from the rat spleens. The activation of natural killer (NK) cells and T-lymphocytes, and level of interleukin-2 were determined by radioimmunoassay. The results revealed that garlic effectively prevented oral precancer induced by 4NQO. This effect may be related to the following factors that garlic can improve the activation of NK cells, the function of T-lymphocytes, and the level of IL-2.

Animals↗
PubMed1996

Nucleotide sequence of the Kaposi sarcoma-associated herpesvirus (HHV8).

The genome of the Kaposi sarcoma-associated herpesvirus (KSHV or HHV8) was mapped with cosmid and phage genomic libraries from the BC-1 cell line. Its nucleotide sequence was determined except for a 3-kb region at the right end of the genome that was refractory to cloning. The BC-1 KSHV genome consists of a 140.5-kb-long unique coding region flanked by multiple G + C-rich 801-bp terminal repeat sequences. A genomic duplication that apparently arose in the parental tumor is present in this cell culture-derived strain. At least 81 ORFs, including 66 with homology to herpesvirus saimiri ORFs, and 5 internal repeat regions are present in the long unique region. The virus encodes homologs to complement-binding proteins, three cytokines (two macrophage inflammatory proteins and interleukin 6), dihydrofolate reductase, bcl-2, interferon regulatory factors, interleukin 8 receptor, neural cell adhesion molecule-like adhesin, and a D-type cyclin, as well as viral structural and metabolic proteins. Terminal repeat analysis of virus DNA from a KS lesion suggests a monoclonal expansion of KSHV in the KS tumor.

Base Sequence↗
PubMed1996

Mammalian mitogen-activated protein kinase pathways are regulated through formation of specific kinase-activator complexes.

Mammalian cells contain at least three signaling systems which are structurally related to the mitogen-activated protein kinase (MAPK) pathway. Growth factors acting through Ras primarily stimulate the Raf/MEK/MAPK cascade of protein kinases. In contrast, many stress-related signals such as heat shock, inflammatory cytokines, and hyperosmolarity induce the MEKK/SEK(MKK4)/SAPK(JNK) and/or the MKK3 or MKK6/p38(hog) pathways. Physiological agonists of these pathway types are either qualitatively or quantitatively distinct, suggesting few common proximal signaling elements, although past studies performed in vitro, or in cells using transient over-expression, reveal interaction between the components of all three pathways. These studies suggest a high degree of cross-talk apparently not seen in vivo. We have examined the possible molecular basis of the differing agonist profiles of these three MAPK pathways. We report preferential association between MAP kinases and their activators in eukaryotic cells. Furthermore, using the yeast 2-hybrid system, we show that association between these components can occur independent of additional eukaryotic proteins. We show that SAPK(JNK) or p38(hog) activation is specifically impaired by co-expression of cognate dominant negative MAP kinase kinase mutants, demonstrating functional specificity at this level. Further divergence and insulation of the stress pathways occurs proximal to the MAPK kinases since activation of the MAPK kinase kinase MEKK results in SAPK(JNK) activation but does not cause p38(hog) phosphorylation. Therefore, in intact cells, the three MAPK pathways may be independently regulated and their components show specificity in their interaction with cognate cascade members. The degree of intermolecular specificity suggests that mammalian MAPK signaling pathways may remain distinct without the need for specific scaffolding proteins to sequester components of individual pathways.

Animals↗
PubMed1996

Serologic association between human papillomavirus type 16 infection and esophageal cancer in Shaanxi Province, China.

BACKGROUND: The existence of large geographic variations in the prevalence of esophageal cancer in some countries, such as China, indicates that environmental risk factors may be important in the development of this disease. Some studies have implicated genital-mucosal strains of human papillomaviruses (HPVs) in the etiology of this cancer. PURPOSE: We conducted a case-control study in Shaanxi Province, China, an area with a population at high risk for esophageal cancer, to assess the association of this disease with infection by HPV type 16 (HPV16), the most common cancer-associated genital-mucosal HPV type. METHODS: Ninety individuals with esophageal cancer and 121 cancer-free control subjects were identified among the patients in two hospitals in Xi'an, Shaanxi Province. The control subjects were matched to the case patients on the basis of age and sex. Blood specimens were drawn from all study subjects, and serum was isolated by routine methods. The presence of HPV16 antibodies in serum samples was determined by use of an enzyme-linked immunosorbent assay (ELISA) that used baculovirus-derived HPV16 virus-like particles as the antigen. A similar ELISA that used bovine papillomavirus type 1 (BPV1) virus-like particles as the antigen controlled for the specificity of HPV16 seroreactivity. Data from the HPV16 and the BPV1 assays were normalized with respect to results obtained in each assay with a control serum of known HPV16 seroreactivity. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to examine the association between HPV16 seroreactivity and esophageal cancer. Reported P values are two-sided. RESULTS: The mean seroreactivity to HPV16 virus-like particles was significantly higher for the cancer patients than for the control subjects (mean value +/- standard deviation = 0.85 +/- 0.22 versus 0.74 +/- 0.18; P<.0001). When the cancer patients and control subjects were compared by sex and age groups, the differences in mean seroreactivity remained statistically significant. The difference in mean seroreactivity to BPV1 virus-like particles between cancer patients and control subjects was not statistically significant (0.81 +/- 0.28 versus 0.88 +/- 0.32; P = .12); this result was not altered when sex and age groups were compared. By use of a cutoff point for HPV16 seropositivity that was established in studies of cervical neoplasia, 24% of the cancer patients were seropositive compared with 7% of the control subjects, yielding a sex- and age-adjusted OR of 4.5 (95% CI = 1.8-11.9). In general, the OR for esophageal cancer increased with increasing HPV16 seroreactivity. CONCLUSIONS AND IMPLICATIONS: HPV16 infection may be a risk factor for esophageal cancer. Further studies of the association between HPV16 infection and the incidence of esophageal cancer are needed.

Antibodies, Viral↗
PubMed1996

Identification of the gltX gene encoding glutamyl-tRNA synthetase from Methanobacterium thermoautotrophicum.

The gltX gene encoding glutamyl-tRNA synthetase from Methanobacterium thermoautotrophicum has been cloned, sequenced, and identified. The gene is located immediately downstream of idsA in an operon containing at least three additional ORFs. The deduced protein sequence from gltX contains conserved regions (HIGH and KMSKS) indicative of a class I aminoacyl-tRNA synthetase.

Amino Acid Sequence↗
PubMed1996

Induction of mitogen-activated protein kinase phosphatase 1 by the stress-activated protein kinase signaling pathway but not by extracellular signal-regulated kinase in fibroblasts.

The intracellular mechanisms involved in the activation of extracellular signal-regulated kinase (ERK) are relatively well understood. However, the intracellular signaling pathways which regulate the termination of ERK activity remain to be elucidated. Mitogen-activated protein kinase phosphatase 1 (MKP-1) has been shown to dephosphorylate and inactivate ERK in vitro and in vivo. In the present study, we show in NIH3T3 fibroblasts that activation of the stress-activated protein kinase (SAPK) pathway by either specific extracellular stress stimuli or via induction of MEKK, an upstream kinase of SAPK, results in MKP-1 gene expression. In contrast, selective stimulation of the ERK pathway by 12-O-tetradecanoylphorbol-13-acetate or following expression of constitutively active MEK, the upstream dual specificity kinase of ERK did not induce the transcription of MKP-1. Hence, these findings demonstrate the existence of cross-talk between the ERK and SAPK signaling cascades since activation of SAPK induced the expression of MKP-1 that can inactivate ERK. This mechanism may modulate the cellular response to stimuli which employ the SAPK signal transduction pathway.

3T3 Cells↗
PubMed1996

Raf, but not MEK or ERK, is sufficient for differentiation of hippocampal neuronal cells.

To elucidate signal transduction pathways leading to neuronal differentiation, we have investigated a conditionally immortalized cell line from rat hippocampal neurons (H19-7) that express a temperature sensitive simian virus 40 large T antigen. Treatment of H19-7 cells with the differentiating agent basic fibroblast growth factor at 39 degrees C, the nonpermissive temperature for T function, resulted in the activation of c-Raf-1, MEK, and mitogen-activated protein (MAP) kinases (ERK1 and -2). To evaluate the role of Raf-1 in neuronal cell differentiation, we stably transfected H19-7 cells with v-raf or an oncogenic human Raf-1-estrogen receptor fusion gene (deltaRaf-1:ER). deltaRaf-1:ER transfectants in the presence of estradiol for 1 to 2 days expressed a differentiation phenotype only at the nonpermissive temperature. However, extended exposure of the deltaRaf-1:ER transfectants to estradiol or stable expression of the v-raf construct yielded cells that extended processes at the permissive as well as the nonpermissive temperature, suggesting that cells expressing the large T antigen are capable of responding to the Raf differentiation signal. deltaRaf-1:ER, MEK, and MAP kinase activities in the deltaRaf-1:ER cells were elevated constitutively for up to 36 h of estradiol treatment at the permissive temperature. At the nonpermissive temperature, MEK and ERKs were activated to a significantly lesser extent, suggesting that prolonged MAP kinase activation may not be sufficient for differentiation. To test this possibility, H19-7 cells were transfected or microinjected with constitutively activated MEK. The results indicate that prolonged activation of MEK or MAP kinases (ERK1 and -2) is not sufficient for differentiation of H19-7 neuronal cells and raise the possibility that an alternative signaling pathway is required for differentiation of H19-7 cells by Raf.

Animals↗
PubMed1996

A three-step pathway of transcription initiation leading to promoter clearance at an activation RNA polymerase II promoter.

The progress of transcription bubbles during inhibition in vitro was followed in order to learn how RNA polymerase II begins transcription at the activated adenovirus E4 promoter. The issues addressed include the multiple roles of ATP, the potential effect of polymerase C-terminal domain phosphorylation, and the ability of polymerase to clear the promoter for reinitiation. The results lead to a three-step model for the transition from closed complex to elongation complex, two steps of which use ATP independently. In the first step, studied previously, ATP is hydrolyzed to open the DNA strands over the start site. In a second step, apparently independent of ATP, transcription bubbles move into the initial transcribed region where RNA synthesis can stall. In the third step, transcripts can be made as polymerase is released from these stalled positions with the assistance of an ATP-dependent process, likely phosphorylation of the polymerase C-terminal domain. After this third step, the promoter becomes cleared, allowing for the reinitiation of transcription.

Adenosine Triphosphate↗
PubMed1996

[A study on cAMP and Ca-CaM of the epithelium in human normal and cataractous lenses].

OBJECTIVE: The study was designed to study the relationship between calcium-calmodulin (Ca-CaM) and cyclic adenosine monophosphate (cAMP) systems and their possible cataractogenic effects. METHODS: We measured cAMP, Ca and Ca-CaM in the epithelium of human normal and cataractous lenses by enzymic and radioimmunoassay methods. RESULTS: The levels of Ca-CaM in the epithelium of senile cataractous lens were higher, while of cAMP were lower than that in the normal lens. CONCLUSION: The results imply that the low level of cAMP might cause the high level of calcium in the lens which is possibly a cataractogenic factor. The dangerous role of calcium is amplified by the elevation of Ca-CaM.

Adolescent↗
PubMed1995

Abortive initiation and first bond formation at an activated adenovirus E4 promoter.

Abortive initiation at the adenovirus E4 promoter was studied by following the production of RNA formed from the initiating nucleotides UpA and CTP. Formation of a specific short RNA via a reaction with appropriate alpha-amanitin sensitivity required promoter, activator, and ATP. In the absence of any of these, an alpha-amanitin-resistant reaction led to lower levels of a product of unknown origin. The alpha-amanitin-sensitive reaction required open promoter complexes, as assayed directly by permanganate probing. This reaction was not blocked by the inhibition of polymerase C-terminal domain kinase activity or by the lack of DNA supercoiling. Thus, formation of the initial bond of the mRNA appears to require activator and ATP to open the DNA but not phosphorylation of the polymerase C-terminal domain. In addition, the abortive initiation reaction was strongly suppressed when all elongation substrates were present, suggesting that cycling to produce high amounts of abortive product is strongly disfavored during productive initiation at this promoter.

Adenosine Triphosphate↗