Biomedical subjects

M Li

Publications and source records attributed to M Li.

At least 379 records · Page 21Linked to original sources

PubMed2000

Pituitary adenylate cyclase-activating polypeptide precursor is processed solely by prohormone convertase 4 in the gonads.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is abundant not only in the brain, but also in the testis. Immunohistochemical studies have shown that PACAP-LI in rat testis is expressed stage specifically in spermatids. This suggests that testicular PACAP participates in the regulatory mechanism of spermatogenesis. Additionally, the ovary contains a relatively small amount of PACAP, conceivably involved in the regulation of folliculogenesis. PACAP is synthesized as a preprohormone and is processed by prohormone convertases, such as PC1, PC2, and PC4. PC4 is expressed only in the testis and ovary, where neither PC1 nor PC2 is expressed. However, whether PC4 is the sole endoprotease for the PACAP precursor in the gonads remains unknown. Recent studies using PC4-transgenic mice revealed that male PC4-null mice exhibited severely impaired fertility, although spermatogenesis appeared to be normal. The female PC4-null mice exhibited delayed folliculogenesis in the ovaries. To examine whether PC4 is the sole processing enzyme for the PACAP precursor in the gonads, we analyzed testicular and ovarian extracts from the PC4-null and wild-type mice for PACAP (PACAP38 and PACAP27) and its messenger RNA using reverse phase HPLC combined with specific RIAs and ribonuclease protection assay, respectively. For RIAs, three different polyclonal antisera with different recognition sites were used to identify PACAP38, PACAP27, and its precursor. Neither the testis nor the ovary from the PC4-null mice expressed PACAP38 or PACAP27, but the levels of PACAP transcripts in the testis and ovary of homozygous PC4-deficient mice were considerably elevated compared with those of the wild-type and heterozygous animals. The findings indicate that PC4 is the sole processing enzyme for the precursor of PACAP in the testis and ovary of mice. The possibility that the absence of bioactive PACAP in the testis and ovary of PC4-null mice caused severely impaired fertility in the males and delayed folliculogenesis in females warrants investigation.

Animals↗

PubMed2000

GNAS1 mutation and Cbfa1 misexpression in a child with severe congenital platelike osteoma cutis.

We evaluated a 7-year-old girl with severe platelike osteoma cutis (POC), a variant of progressive osseous heteroplasia (POH). The child had congenital heterotopic ossification of dermis and subcutaneous fat that progressed to involve deep skeletal muscles of the face, scalp, and eyes. Although involvement of skeletal muscle is a prominent feature of POH, heterotopic ossification has not been observed in the head, face, or extraocular muscles. The cutaneous ossification in this patient was suggestive of Albright hereditary osteodystrophy (AHO); however, none of the other characteristic features of AHO were expressed. Inactivating mutations of the GNAS1 gene, which encodes the alpha-subunit of the stimulatory G protein of adenylyl cyclase, is the cause of AHO. Mutational analysis of GNAS1 using genomic DNA of peripheral blood and of lesional and nonlesional tissue from our patient revealed a heterozygous 4-base pair (bp) deletion in exon 7, identical to mutations that have been found in some AHO patients. This 4-bp deletion in GNAS1 predicts a protein reading frameshift leading to 13 incorrect amino acids followed by a premature stop codon. To investigate pathways of osteogenesis by which GNAS1 may mediate its effects, we examined the expression of the obligate osteogenic transcription factor Cbfa1/RUNX2 in lesional and uninvolved dermal fibroblasts from our patient and discovered expression of bone-specific Cbfa1 messenger RNA (mRNA) in both cell types. These findings document severe heterotopic ossification in the absence of AHO features caused by an inactivating GNAS1 mutation and establish the GNAS1 gene as the leading candidate gene for POH.

Amino Acid Sequence↗

PubMed2000

Successful use of argon plasma coagulation and tranilast to treat granulation tissue obstructing the airway after tracheal anastomosis.

We successfully used argon plasma coagulation (APC) and tranilast to treat granulation tissue that had formed on an end-to-end tracheal anastomosis. APC had several advantages over laser in the management of exuberant granulation tissue, and we considered tranilast to be effective in our patient. To our knowledge, this is the first description of the use of APC in the management of anastomotic granulation tissue in the trachea or bronchus.

Adult↗

PubMed2000

Dihydrocercosporin singlet oxygen production and subcellular localization: a possible defense against cercosporin phototoxicity in Cercospora.

Fungi in the genus Cercospora produce cercosporin, a potent singlet oxygen (1O2)-generating photosensitizer that plays a critical role in the ability of these fungi to parasitize plants. Although plants, mice, bacteria and many fungi are sensitive to cercosporin, Cercospora species are resistant to its toxicity. The cellular resistance of these fungi to cercosporin has been correlated with fungal cell surface reducing ability and the ability to maintain cercosporin in a chemically reduced state. As a model for reduced cercosporin we employed a reduced, acetylated derivative (hexaacetyl-dihydrocercosporin, HAC) that we tested for 1O2 production in a range of solvents. We found that as a 1O2 photosensitizer, HAC was only moderately effective in organic solvents (phi SO = 0.14-0.18) and very poor in water (phi SO = 0.02-0.04). By contrast, the 1O2 quantum yield of cercosporin itself was unaffected by solvent (phi SO = 0.84-0.97). To investigate the localization of reduced cercosporin in fungal cells, we developed a fluorescence assay using laser scanning confocal microscopy. This assay showed a uniform green fluorescence, indicative of reduced cercosporin, in the cytoplasm of hyphal cells treated with cercosporin. We hypothesize that the main protection mechanism against cercosporin phototoxicity in the fungus consists of transformation of cercosporin to a reduced state and localization of this reduced form in the aqueous compartment of the cell, thus decreasing intracellular 1O2 production to levels that can be tolerated by the fungus. In addition, we have, for the first time, directly detected 1O2 phosphorescence from fungal culture, either stained with the photosensitizer rose bengal or actively synthesizing cercosporin, demonstrating 1O2 production in vivo and from cercosporin in culture.

Animals↗

PubMed2000

Drug discrimination under two concurrent fixed-interval fixed-interval schedules.

Pigeons were trained to discriminate 5.0 mg/kg pentobarbital from saline under a two-key concurrent fixed-interval (FI) 100-s FI 200-s schedule of food presentation, and later tinder a concurrent FI 40-s FI 80-s schedule, in which the FI component with the shorter time requirement reinforced responding on one key after drug administration (pentobarbital-biased key) and on the other key after saline administration (saline-biased key). After responding stabilized under the concurrent FI 100-s FI 200-s schedule, pigeons earned an average of 66% (after pentobarbital) to 68% (after saline) of their reinforcers for responding under the FI 100-s component of the concurrent schedule. These birds made an average of 70% of their responses on both the pentobarbital-biased key after the training dose of pentobarbital and the saline-biased key after saline. After responding stabilized under the concurrent FI 40-s FI 80-s schedule, pigeons earned an average of 67% of their reinforcers for responding under the FI 40 component after both saline and the training dose of pentobarbital. These birds made an average of 75% of their responses on the pentobarbital-biased key after the training dose of pentobarbital, but only 55% of their responses on the saline-biased key after saline. In test sessions preceded by doses of pentobarbital, chlordiazepoxide, ethanol, phencyclidine, or methamphetamine, the dose-response curves were similar under these two concurrent schedules. Pentobarbital, chlordiazepoxide, and ethanol produced dose-dependent increases in responding on the pentobarbital-biased key as the doses increased. For some birds, at the highest doses of these drugs, the dose-response curve turned over. Increasing doses of phencyclidine produced increased responding on the pentobarbital-biased key in some, but not all, birds. After methamphetamine, responding was largely confined to the saline-biased key. These data show that pigeons can perform drug discriminations under concurrent schedules in which the reinforcement frequency under the schedule components differs only by a factor of two, and that when other drugs are substituted for the training drugs they produce dose-response curves similar to the curves produced by these drugs under other concurrent interval schedules.

Animals↗

PubMed2000

Cloning of a T-type Ca2+ channel isoform in insulin-secreting cells.

The T-type Ca2+ channel is an important determinant of electrical activity and of Ca2+ influx in rat and human pancreatic beta-cells. We have identified and sequenced a cDNA encoding a T-type Ca2+ channel alpha1-subunit derived from INS-1, the rat insulin-secreting cell line. The sequence of the cDNA indicates a protein composed of 2,288 amino acids that shares 96.3% identity to alpha1G, the neuronal T-type Ca2+ channel subunit. The transmembrane domains of the protein are highly conserved, but the isoform contains three distinct regions and 10 single amino acid substitutions in other regions. Sequencing rat genomic DNA revealed that the alpha1-subunit we cloned is an alternative splice isoform of alpha1G. By using specific primers and reverse transcription-polymerase chain reaction, we demonstrated that both splice variants are expressed in rat islets. The isoform deduced from INS-1 was also expressed in brain, neonatal heart, and kidney. Functional expression of this alpha1G isoform in Xenopus oocytes generated low voltage-activated Ba2+ currents. These results provide the molecular biological basis for studies of function of T-type Ca2+ channels in beta-cells, which is where these channels may play critical roles in diabetes.

Alternative Splicing↗

PubMed2000

Recombinant human betacellulin promotes the neogenesis of beta-cells and ameliorates glucose intolerance in mice with diabetes induced by selective alloxan perfusion.

Betacellulin (BTC), a member of the epidermal growth factor family, is expressed predominantly in the human pancreas and induces the differentiation of a pancreatic acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC has a physiologically important role in the endocrine pancreas. In this study, we examined the in vivo effect of recombinant human BTC (rhBTC) on glucose intolerance and pancreatic morphology using a new mouse model with glucose intolerance induced by selective alloxan perfusion. RhBTC (1 microg/g body wt) or saline was injected subcutaneously every day from the day after alloxan treatment. The intraperitoneal glucose tolerance test revealed no difference between rhBTC-treated and rhBTC-untreated glucose-intolerant mice at 2-4 weeks. However, glucose tolerance was significantly improved and body weight was significantly increased in rhBTC-treated mice compared with untreated mice at 8 weeks. Islet-like cell clusters, consisting mainly of beta-cells, were increased in the pancreas and were localized in contact with the ductal lining cells and sometimes with acinar cells. In conclusion, administration of rhBTC improved glucose tolerance in this mouse model by increasing beta-cell volume, primarily through accelerated neogenesis from ductal lining cells.

Animals↗

PubMed2000

Epstein-Barr virus RNA detection and glandular differentiation in nasopharyngeal carcinoma: report of 2 cases.

Most tumors arising in the nasopharynx are either squamous cell carcinoma or so-called undifferentiated carcinoma of the nasopharyngeal type. Primary adenocarcinomas of the nasopharynx are rare, and glandular differentiation in undifferentiated carcinoma of the nasopharyngeal type has not been reported to date. We report 2 cases of undifferentiated carcinoma of the nasopharyngeal type that show distinct glandular differentiation by light microscopy, histochemistry, immunohistochemistry, and ultrastructure. Both tumors showed equal positivity for Epstein-Barr virus latent membrane protein and in situ hybridization for Epstein-Barr virus genome in the undifferentiated areas of the tumor and those featuring glandular differentiation.

Adenocarcinoma↗

PubMed2000

Identification of the novel K15 gene at the rightmost end of the Kaposi's sarcoma-associated herpesvirus genome.

Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a distinct open reading frame called K15 at a position equivalent to the gene encoding LMP2A of Epstein-Barr virus (EBV). K15 isolates from body cavity-based lymphoma (BCBL) cells exhibited a dramatic sequence variation and a complex splicing pattern. However, all K15 alleles are organized similarly with the potential SH2 and SH3 binding motifs in their cytoplasmic regions. Northern blot analysis showed that K15 was weakly expressed in latently infected BCBL-1 cells, and the level of its expression was significantly induced by tetradecanoyl phorbol acetate stimulation. K15 encoded 40- to 55-kDa proteins, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and was localized at the cytoplasm and plasma membrane. To demonstrate the signal-transducing activity of the K15 protein, we constructed a chimeric protein in which the cytoplasmic tail of the human CD8alpha polypeptide was replaced with that of KSHV K15. While the CD8-K15 chimera was not capable of eliciting cellular signal transduction upon stimulation with an anti-CD8 antibody, it significantly inhibited B-cell receptor signaling, as evidenced by a suppression of tyrosine phosphorylation and intracellular calcium mobilization. This inhibition required the putative SH2 or SH3 binding motif in the cytoplasmic region of K15. Biochemical study of CD8-K15 chimeras showed that the cytoplasmic region of K15 was constitutively tyrosine phosphorylated and that the tyrosine residue within the putative SH2 binding motif of K15 was a primary site of phosphorylation. These results demonstrate that KSHV K15 resembles LMP2A in genomic location, splicing pattern, and protein structure and by the presence of functional signal-transducing motifs in the cytoplasmic region. Thus, KSHV K15 is likely a distant evolutionary relative of EBV LMP2A.

Alleles↗

PubMed2000

Functional domains and expression of truncated atrial natriuretic peptide receptor-A: the carboxyl-terminal regions direct the receptor internalization and sequestration in COS-7 cells.

The objective of this study was to determine the role of cytoplasmic (protein kinase-like homology and guanylyl cyclase catalytic) domains of atrial natriuretic peptide (ANP) receptor-A (Npra) in postbinding events and metabolic turnover of ligand-receptor complexes. Using deletion mutagenesis, the specific regions in the intracellular domains of Npra relevant to the receptor function, namely ligand-binding, cGMP production, and internalization and sequestration of ligand-receptor complexes, have been determined in transiently expressing COS-7 cells. Deletion of 12 aa (aa) at the carboxyl-terminal end of receptor (Delta1045-Npra) affected neither ligand-binding efficiency nor cGMP production. However, deletion of 120 to 170 aa residues (Delta937-Npra, Delta916-Npra, Delta902-Npra, and Delta887-Npra) decreased ligand binding by 16 to 20% and cGMP production by 50 to 90%. Further deletion of 422 aa and 569 aa (Delta635-Npra and Delta488-Npra) reduced ligand binding efficiency by 40% and 90%, respectively. The deletion of 12 aa (Delta1045-Npra) did not affect the internalization of Npra; however, deletions up to 170 aa (Delta937-Npra, Delta916-Npra, Delta887-Npra) reduced the internalization of ligand-receptor complexes by 60%. Cells expressing either full-length (wild-type) Npra or 120 aa deleted receptor (Delta937-Npra) released 40 to 45% (125)I-ANP radioactivity into culture medium, but only 10 to 15% radioactivity was released from the cells that expressed Delta635-Npra. Furthermore, 35 to 40% (125)I-ANP radioactivity was detected into the intracellular compartments of cells that expressed the wild-type Npra, and only 5 to 10% (125)I-ANP radioactivity was observed in cells expressing the Delta635-Npra (-422 aa) or Delta488-Npra (-569 aa) mutant receptors. These results show that specific regions within the intracellular domains of Npra determine the extent of ligand-binding efficiency, cGMP production, endocytosis, and intracellular sequestration of ligand-receptor complexes in cDNA expressing COS-7 cells.

Animals↗

PubMed2000

[Exclusive mapping on polydactyly with markers on chromosome 7 and 2 in a Chinese kindred].

OBJECTIVE: This study was aimed at mapping polydactyly related genes in a Chinese kindred. METHODS: Linkage analysis was performed using 7 markers on chromosome 7 and 1 marker on chromosome 2. RESULTS: Pairwise linkage analysis showed no linkage between the markers and polydactyly related gene in the kindred. CONCLUSION: The polydactyly related gene in this kindred may be located on a new locus.

Chromosome Mapping↗

PubMed2000

A mibefradil metabolite is a potent intracellular blocker of L-type Ca(2+) currents in pancreatic beta-cells.

It has been shown that mibefradil (Ro 40-5967) exerts a selective inhibitory effect on T-type Ca(2+) currents, although at higher concentrations it can antagonize high voltage-activated Ca(2+) currents. The action of mibefradil on Ca(2+) channels is use- and steady-state-dependent and the binding site of mibefradil on L-type Ca(2+) channels is different from that of dihydropyridines. By using conventional whole-cell and perforated patch-clamp techniques, we showed that mibefradil has an inhibitory effect on both T- and L-type Ca(2+) currents in insulin-secreting cells. However, the effect on L-type Ca(2+) currents was time-dependent and poorly reversible in perforated patch-clamp experiments. By using mass spectrometry, we demonstrated that mibefradil accumulates inside cells, and furthermore, a metabolite of mibefradil was detected. Intracellular application of this metabolite selectively blocked the L-type Ca(2+) current, whereas mibefradil exerted no effect. This study demonstrates that mibefradil permeates into cells and is hydrolyzed to a metabolite that blocks L-type Ca(2+) channels specifically by acting at the inner side of the channel.

Calcium Channel Blockers↗

PubMed2000

Aspirin induction of apoptosis in esophageal cancer: a potential for chemoprevention.

The potential use of non-steroidal anti-inflammatory drugs (NSAIDs) in the prevention of gastrointestinal cancers has been highlighted recently. However, it is not known whether NSAIDs could also be useful for preventing esophageal cancer, although regular users of these drugs appear to have a decreased incidence of esophageal cancer. Therefore, we examined the effect of aspirin on growth and apoptosis in 10 esophageal cancer cell lines as well as the expression and modulation of its target enzymes, cyclooxygenases (COXs), and their product prostaglandin E2. Growth inhibition of these cells by aspirin was dose- and time-dependent and associated with the induction of apoptosis. COX-1 and COX-2 were expressed in 7 of the 10 cell lines. Bile acids could induce COX-2 expression in six of eight cell lines tested, which was correlated with prostaglandin E2 production, and aspirin could inhibit COX-2 enzymatic activity even after bile acid stimulation but was unable to change the COX-2 protein level in these cell lines. Down-regulation of bcl-2 by aspirin was found in the two cell lines tested. These results suggest that induction of apoptosis by aspirin may be a mechanism by which it can intervene in esophageal carcinogenesis and may be indicative of the potential of NSAIDs as chemopreventive agents in esophageal cancer.

Actins↗

PubMed2000

The value of serum GPDA-F for the diagnosis of primary hepatocellular carcinoma.

OBJECTIVE: To investigate the role of glycylproline dipeptidyl aminopeptidase isoenzymes in the diagnosis of primary hepatocellular carcinoma (PHC). METHODS: We developed a stage gradient polyacrylamide gel electrophoresis system to separate serum GPDA isoenzymes. Total GPDA activities, alpha-fetoprotein, the sizes of tumors and alanine aminotransferase (ALT) activities were also measured simultaneously and the correlation between GPDA-F and these indices was analyzed. RESULTS: Serum GPDA was separated into two bands, namely fast band (GPDA-F) and slow band (GPDA-S). GPDA-F was negative in all healthy persons as well as in the patients with benign liver filling defects, while it was positive in 85.3% cases of PHC. Liver cirrhosis, chronic hepatitis, extrahepatic carcinoma and metastatic liver carcinoma had low positive rates. GPDA-F was positively correlated with serum total GPDA activities, but had no correlation with AFP and size of the tumors. There was the correlation between GPDA-F and ALT in benign liver diseases, but no correlation between GPDA-F and ALT in PHC. Serial measurements of serum GPDA-F showed that GPDA-F was persistently positive in PHC but might change into negative in benign liver diseases. Dynamic determination of GPDA-F might be helpful to differentiate true positive of PHC from false positive of benign liver diseases. CONCLUSION: GPDA-F is a new serum marker of PHC. Measurement of serum GPDA-F is of value for the diagnosis of PHC, especially for those at early stage or with negative AFP.

Alanine Transaminase↗

PubMed2000

The effect of zhu yu hua tan tang on intracranial pressure in case of acute cerebral hemorrhage.

The therapeutic effect of Zhu Yu Hua Tan Tang ([symbol: see text] Decoction for removing blood stasis and resolving phlegm) plus general and modern treatment on acute cerebral hemorrhage in 44 cases, and the pressure-lowering effect of Zhu Yu Hua Tan Tang and mannitol in 20 cases of acute cerebral hemorrhage in the basilar region, and in rabbits with experimental encephaledema induced by injection of olive oil into the common carotid artery were observed in this study. The results showed that Zhu Yu Hua Tan Tang plus general treatment can better improved the conscious state of the patients than western medicine, and it can make the intracranial pressure drop clinically by 40.44%. The intracranial pressure-lowering effect of Zhu Yu Hua Tan Tang, though slow, is smooth and long-lasting without any rebound phenomenon, as compared to those of mannitol. Similar results were found in animal experiments, but the intracranial pressure-lowering amplitude of Zhu Yu Hua Tan Tang was stronger than that of mannitol. The difference between the clinical and experimental results needs to be further studied.

Aged↗

PubMed2000

[Study of CFTR gene mutation in Chinese CUAVD patients].

OBJECTIVE: To analyze the frequency and hot spots of cystic fibrosis transmembrane conductance regulator(CFTR) gene in Chinese congenital unilateral absence of the vas deferens (CUAVD) patients. METHODS: The mutation of CFTR exons 2, 3, 4, 5, 6a, 8, 10, 11, 12, 13, 15A, 17b, 19A, 20, 21, and 23 were detected. PCR-single strand conformation polymorphism(SSCP) and direct sequencing were performed on 15 cases of Chinese CUAVD. RESULTS: One case exhibited an abnormal shift SSCP band in exon 17b of CFTR gene and subsequent DNA sequencing showed C to A transversion at position 3295 that led to a predicted change of Leusine(codon 1055, CTT) to Isoleucine(codon ATT). CONCLUSION: CFTR mutation could be detected in Chinese CUAVD patients. The missense mutation, Leu1055Ile, was identified as a novel CFTR mutation. It is necessary that Chinese CUAVD patients and their wives should be screened for CFTR gene prior to intracytoplasmic sperm injection.

Cystic Fibrosis Transmembrane Conductance Regulato↗

PubMed2000

[Cultivation optimization of prolyl endopeptidase and high cell density fermentation].

Engineered E. coli BL21/pGEM-PEP could constitutively express recombinant prolyl endopeptidase from Aeromonas punctata, which was extremely effected by culture conditions, so fermentation conditions were optimized to obtain it's high expression level. Firstly, the stability of BL21/pGEM-PEP was investigated, then, culture temperature, pH, time, medium were optimized in shake flaskd. The L9(3(4)) orthogonal experiment confirmed that shaker speed, pH, culture temperature, culture time had high degree statistical meaning. Based on these data, high cell density fermentation of E. coli BL21/pGEM-PEP on NBS BioFlo 3000 5 L fermentor was achieved, after 20 h cultivation, the final density(dwt) was 60OD600(22.5 g/L), the expressed PEP was about 28% of total cellular protein, the yield was 3.15 g per litter broth.

Culture Media↗

PubMed2000

Down-regulation of benzodiazepine binding to alpha 5 subunit-containing gamma-aminobutyric Acid(A) receptors in tolerant rat brain indicates particular involvement of the hippocampal CA1 region.

Chronic benzodiazepine treatment can produce tolerance and changes in gamma-aminobutyric acid (GABA)(A) receptors. To study the effect of treatment on a selected population of receptors, assays were performed using [(3)H]RY-80, which is selective for GABA(A) receptors with an alpha 5 subunit. Rats were given a flurazepam treatment known to produce tolerance and down-regulation of benzodiazepine binding, or a diazepam treatment shown to produce tolerance but not receptor down-regulation. Quantitative receptor autoradiography using sagittal brain sections bound with [(3)H]RY-80 showed binding in areas known to express alpha 5 mRNA. Brains from flurazepam-treated rats showed significantly decreased 1 nM [(3)H]RY-80 binding in hippocampal formation (e.g., 32% decrease in CA1) and superior colliculus, but not other areas. Using 5 nM [(3)H]RY-80 showed similar decreases in hippocampus. A corresponding 29% decrease in B(max) but no change in K(d) was found with a filtration binding assay using hippocampal homogenates. Down-regulation of [(3)H]RY-80 binding had returned to control by 2 days after withdrawing flurazepam treatment. The magnitude of down-regulation of [(3)H]RY-80 binding suggested that GABA(A) receptors with an alpha 5 subunit may play a prominent role in the adaptive responses associated with benzodiazepine tolerance. Chronic diazepam treatment also resulted in decreased [(3)H]RY-80 binding. However, the regional selectivity was even more pronounced than in flurazepam-treated rats, and only the hippocampal CA1 region showed decreased binding (27%). This localized down-regulation persisted for several days after the end of diazepam treatment. These data indicate that synapses in the hippocampal CA1 region are particularly involved in the adaptive response to chronic benzodiazepine treatments.

Alkynes↗