Search PubMed⌕ Search

Biomedical subjects

L M Chen

Publications and source records attributed to L M Chen.

At least 91 records · Page 5Linked to original sources
PubMed1995

Cellular localization of tissue kallikrein and kallistatin mRNAs in human kidney.

The renal kallikrein-kinin system has been implicated in the regulation of blood pressure and sodium/water excretion. The activity of renal kallikrein is controlled by a number of factors in vivo. Kallistatin is a newly identified serine proteinase inhibitor (serpin) which binds to tissue kallikrein and inhibits its enzymatic activity in vitro. To understand the role of kallistatin in modulating tissue kallikrein's function in vivo, we examined the anatomical relationship between human tissue kallikrein and kallistatin in the kidney by in situ hybridization histochemistry. Tissue kallikrein and kallistatin gene transcripts were identified using digoxigenin-labeled riboprobes at the cellular level. Antisense and sense riboprobes corresponding to the 3' region of the human kallikrein and kallistatin mRNAs were synthesized by in vitro transcription and used for hybridization. Using an antisense kallikrein riboprobe, sites of kallikrein synthesis were localized in the distal tubules, collecting ducts and Henle's loops of the kidney. To a lesser degree, juxtaglomerular cells were also stained. Kallistatin mRNA was found at the same sites where kallikrein mRNA was localized. The most intense signals of both kallikrein and kallistatin were seen in the distal tubules and collecting ducts. Hybridization was specific for the target mRNA since sense kallikrein or kallistatin riboprobe did not bind to the sections. Immunoreactive human renal kallikrein and kallistatin levels were measured in the kidney and urine by immunoassays using specific antibodies. Co-localization of kallikrein and kallistatin mRNA in the kidney suggests a potential role of kallistatin in regulating tissue kallikrein's function.

Carrier Proteins↗
PubMed1994

Molecular cloning and characterization of a ras-related gene of ran/tc4/spi1 subfamily in Giardia lamblia.

The significance of Ras-like proteins in the protozoa is relatively unexplored. In this report, a gene encoding a Ras-like nuclear (Ran) protein was identified in Giardia lamblia by a polymerase chain reaction-based cloning strategy. The sequence analyses suggest that the gene was intronless, and had short 5'-untranslated leader sequences in the corresponding mRNA up to -2, -4, or -29 bases upstream of the first initiation codon. The full-length cDNA sequence predicted a protein comprising 226 amino acids, in which the highly conserved functional motifs of the Ras superfamily were all preserved. This protein showed 52% identity to human TC4 and 50% identity to yeast Spi1 proteins, suggesting that it is closely related to the Ran proteins, and it was therefore designated gRan. gRan produced from recombinant Escherichia coli exhibited GTP binding activity by an overlay assay. In good agreement with the predicted size of gRan, a 27-kDa protein was identified in a lysate of G. lamblia by Western blotting using antiserum raised against recombinant gRan. The protein was further localized in both nuclei of G. lamblia by immunofluorescence staining. Recombinant gRan exhibited low affinity for GTP with a Kd value of 16.8 microM. The affinity was enhanced to a Kd value of 2.2 microM in the presence of 10 mM Mg2+. The intrinsic GTPase activity of gRan was observed only in the presence of 10 mM Mg2+ and had an estimated Km of 5.6 microM and a Kcat of 0.33/h. These observations demonstrate the presence of Ras-like proteins in the most primitive eukaryotic cells, G. lamblia, and infer that the Ran protein may play a functional role in the nuclei of this organism.

Animals↗
PubMed1994

The interrelationship between impaired glucose tolerance and other risk factors for cardiovascular disease: is it a predictor for cardiovascular disease?

Glucose intolerance and cardiovascular risk factors were assessed among 215 subjects aged 27-86 years in Ho-long, Taiwan. An oral glucose tolerance test (OGTT) was normal in 107 subjects, impaired in 41 subjects, and abnormal in 67 subjects (as noninsulin dependent diabetes mellitus, NIDDM). Body fatness, blood pressure, carbohydrate and lipid metabolic factors were examined. One-way ANCOVA was used to compare age- and/or BMI-adjusted differences among these groups. Male subjects with NIDDM and impaired glucose tolerance (IGT) had higher systolic (SBP) and diastolic (DBP) blood pressures, plasma insulin (fasting and 1-hour after OGTT), serum triglycerides (TG), heavier body weight, and larger BMI than normal. IGT subjects were also found to have higher TG, insulin (2-hour) and larger insulin area under curve (IAUC) after OGTT than NIDDM. In females, subjects with NIDDM had higher TG, insulin (fasting and 1-hour after OGTT), and larger IAUC than normal. Moreover, female IGT subjects were also found to have significantly higher SBP, DBP, insulin (1-and 2-hour after OGTT), and larger IAUC than both normal and NIDDM subjects. These results suggest that adverse cardiovascular risk factors are present not only in NIDDM subjects, but also in IGT subjects.

Adult↗
PubMed1994

Molecular cloning and characterization of a novel kallikrein transcript in colon and its distribution in human tissues.

1. We have cloned and characterized a new species of kallikrein cDNA from a human colon cDNA library. The new kallikrein cDNA clone contains a part of intron 2 of the tissue kallikrein gene which is spliced to the remaining exon sequences. It does not contain exons 1 and 2. 2. An in-frame open reading frame is present in the new kallikrein cDNA allowing translation of a 216-amino acid product. The intron-containing kallikrein transcript was detected in salivary glands, pancreas, kidney, colon, prostate gland, testis, spleen, and lung by reverse-transcription/polymerase chain reaction followed by Southern blot analysis using an intron-containing kallikrein-specific oligonucleotide probe. 3. The results indicate that the new species of kallikrein may be processed by alternative splicing or arises from a different transcription initiation site.

Amino Acid Sequence↗
PubMed1993

Kallistatin: a novel human serine proteinase inhibitor. Molecular cloning, tissue distribution, and expression in Escherichia coli.

We have recently purified a novel human serine proteinase inhibitor (serpin), designated as kallistatin, which binds to tissue kallikrein and inhibits kallikrein's kininogenase and amidolytic activities. In the present studies, we have cloned a full-length cDNA encoding kallistatin from human liver RNA by the polymerase chain reaction. The cDNA is 1284 base pairs in length and encodes 427 amino acid residues, including a 26-residue signal peptide and a 401-residue mature peptide. The translated amino acid sequence of kallistatin matches with the protein sequence and shares 44-46% sequence identity with human alpha 1-antichymotrypsin, protein C inhibitor, corticosteroid-binding globulin, alpha 1-antitrypsin, thyroxin-binding globulin, and rat kallikrein-binding protein. Kallistatin is a new member of the serpin superfamily with a unique reactive site P1-P1' of Phe-Ser. Four potential glycosylation sites are found in the translated amino acid sequence of kallistatin. In a Southern blot analysis following reverse transcription and polymerase chain reaction, kallistatin was found to be expressed in human liver, stomach, pancreas, kidney, aorta, testes, prostate, artery, atrium, ventricle, lung, renal proximal tubular cell, and a colonic carcinoma cell line T84. A genomic Southern blot using the full-length kallistatin cDNA probe revealed simple banding patterns suggesting the gene encoding kallistatin is single-copied. The kallistatin cDNA encoding the mature peptide was expressed in Escherichia coli. The recombinant kallistatin forms an SDS-stable complex with 125I-human tissue kallikrein and has a molecular mass of 40 kDa. The cloning of human kallistatin cDNA established the identity of the novel kallikrein inhibitor and its expression in a functional form in E. coli provides means for studying its structure-function relationship through protein engineering.

Amino Acid Sequence↗
PubMed1993

Prediction of chemotherapy response in human leukemia using in vitro chemosensitivity test.

We performed the dye exclusion assay (DEA) and the MTT dye reduction assay to determine the drug sensitivity of acute leukemia using short-term microplate cultures. The in vitro results were compared with the clinical response of 31 patients with acute leukemia treated by combination chemotherapy. The true-positive rates of the DEA and MTT assays were 86.7 and 91.3%, respectively; the true-negative rates were 33.3 and 77.8%, respectively; and the predictive accuracy was 62.5 and 87.5%, respectively. The DEA and MTT assays gave comparable results in drug sensitivity testing of leukemic blast cells. Our data suggest that MTT assay is the more suitable for assessing chemosensitivity in acute leukemia.

Adult↗
PubMed1993

The reappraisal of the association of glycosylated hemoglobin A1c (HbA1c) and blood pressure: a hypertension and diabetes study in a Taiwan rural area.

The association of age, body weight, body mass index (BMI) and glycosylated hemoglobin A1c (HbA1c) to blood pressure in the general population of the Ho-long area in Taiwan was examined. A total of 1341 subjects, 746 men and 595 women, aged 40-90 years, were studied. Sex-specific multiple regression analyses were carried out on variables both univariately and significantly associated with blood pressure. Results show HbA1c to be significantly associated with SBP in both males and females before age, weight and/or BMI are adjusted (p < 0.05). After controlling for these confounders, the strength of SBP-HbA1c association was slightly reduced in males, but remained significant. In contrast, there was no significance in females. Moreover, no significant relationship in DBP-HbA1c was found in either sex group. These findings suggest that although a positive correlation was found between HbA1c and SBP, HbA1c or glucose status appeared to be a less important contributing factor in hypertension than age or body weight.

Age Factors↗
PubMed1993

Heel replantation and subsequent analysis of gait.

A case of sharp, complete amputation of the heel with successful replantation is presented. The bones, periosteum, tendons, and neurovascular bundle were repaired. Seventeen months after injury, both motor and sensory recovery were satisfactory. The objective study of gait analysis also revealed an essentially normal gait pattern of the injured foot. These results suggest that in a suitable candidate, the procedure of choice for amputation of the heel is microsurgical replantation.

Adult↗
PubMed1993

Effect of nicardipine treatment on carbohydrate and lipoprotein metabolism in patients with hypertension.

This study evaluated the effect of nicardipine, calcium channel blocker, monotherapy on blood pressure and metabolic changes. Various aspect of carbohydrate and lipoprotein metabolism were studied before and after 12 weeks of nicardipine treatment in 23 patients with mild to moderate essential hypertension. Nicardipine was well tolerated and induced a significant decrease (p < 0.001) in both systolic and diastolic blood pressure without any changes in heart rate. Plasma levels of fasting glucose, insulin, C-peptide and hemoglobin A1c, hepatic extraction of insulin were similar following nicardipine treatment. Plasma glucose and insulin responses to an oral glucose challenge did not change in association with nicardipine therapy. Although high density lipoprotein (HDL) cholesterol concentration increased slightly, it did not reach statistical significance. Total cholesterol and low density lipoprotein (LDL) cholesterol levels also increased insignificantly. LDL triglyceride and very low density lipoprotein (VLDL) triglyceride concentrations were higher marginally, which resulted in slightly but insignificantly increase in total triglyceride concentration in association with nicardipine monotherapy for 12 weeks. In conclusion, treatment of patients with mild to moderate hypertension with nicardipine led to lower blood pressure effectively while had no significant influence on carbohydrate and lipoprotein metabolism.

Adult↗
PubMed1993

Regulation of kininogen gene expression and localization in the lung after monocrotaline-induced pulmonary hypertension in rats.

Pyrrolizidine monocrotaline (MCT) from plant seed produces pulmonary endothelial cell injury, pulmonary hypertension, and inflammation in rats, providing a useful animal model for studying progressive pulmonary vascular disease. Kininogen is the precursor of proinflammatory kinins and may also exert anti-inflammatory actions by inhibiting cysteine proteinases. Given the potential roles of kininogen in vascular injury and inflammation, we have investigated the regulation of kininogen gene expression in the MCT-induced pulmonary hypertensive rat model. Sprague-Dawley rats, in groups of six, were given a single subcutaneous injection of monocrotatine (60 mg/kg body wt) and sacrificed 10 and 20 days later. Northern blot hybridization using a kininogen cDNA probe showed kininogen gene expression in the liver, lung, and kidney. MCT treatment induced a time-dependent increase in kininogen mRNA levels, whereas it reduced rat alpha 1-antitrypsin and kallikrein-binding protein mRNA levels in the liver. Similarly, kininogen mRNA levels were low in the normal lung and were increased 7.5- and 13.7-fold, respectively, after MCT injection for 10 and 20 days. Immunoreactive kininogen levels in perfused liver and lung extracts of rats receiving MCT injection increased up to 20-fold, as measured by a T-kininogen radioimmunoassay. Western blot analyses showed that a 68-kilodalton immunoreactive kininogen increased in the serum and lung extracts of MCT-treated rats compared to those in the control rats. In control rats, immunostaining for kininogen in the lung was most marked in venous endothelial cells and alveolar macrophages. After MCT treatment, staining for kininogen increased dramatically throughout the lung tissues, often covering the epithelial surfaces of alveoli and bronchi. The present studies have shown that the toxin MCT altered the synthesis and distribution of pulmonary kininogen and suggest that the kininogen/kinin system may be associated with the pulmonary vascular injury, remodeling, and inflammation seen in this animal model.

Animals↗
PubMed1992

Differential regulation of kininogen gene expression by estrogen and progesterone in vivo.

Kininogens which have multifunctional domains, serve as the precursors of potent vasoactive kinin peptides and also function as cysteine proteinase inhibitors. Given its potential role in blood pressure homeostasis and inflammation, we have examined the regulation of rat kininogen gene expression by sex hormones in vivo. Our studies indicate a differential regulation of kininogen gene expression in rat liver by estrogen and progesterone. Northern and dot blot analysis using a rat low molecular weight kininogen cDNA probe show that kininogen mRNA levels in the liver of female rats are 4-fold higher than those in male rats. Ovariectomy results in a reduction of kininogen transcripts in the liver, while estradiol replacement of the ovariectomized rats increases kininogen mRNA levels. Similarly, Northern blot analysis using a kallikrein cDNA probe shows that estradiol treatment induces an increase of kallikrein gene expression in the kidney of the same animals. In contrast, progesterone treatment of the ovariectomized rats results in an increase in renal kallikrein mRNA levels while it reduces kininogen gene expression as compared to vehicle-treated ovariectomized animals. Immunoreactive kininogen levels in the serum, analyzed by a direct radioimmunoassay and Western blot, are increased by estradiol but slightly decreased by progesterone treatment. Western blot of serum proteins on a two-dimensional polyacrylamide gel reveals that in estradiol-treated ovariectomized rats, the levels of several 68,000 Da kininogens varying in charge are markedly higher than those in ovariectomized rats. The results indicate that estrogen is one of the determinants in regulating low molecular weight kininogen gene expression in vivo. The impact of estrogen-regulated kininogen expression on cardiovascular function awaits further investigation.

Animals↗
PubMed1992

Expression of kallikrein-binding protein and alpha 1-antitrypsin genes in response to sex hormones, growth, inflammation and hypertension.

We have recently purified rat kallikrein-binding protein (RKBP) and alpha 1-antitrypsin (alpha 1-AT) to homogeneity and isolated, sequenced cDNAs encoding these potential regulators of tissue kallikreins. Characterization of the cDNA and the gene has established the identity of the kallikrein-binding protein as a new member of the serpin (serine proteinase inhibitor) superfamily. Using the cDNA probes in Northern blot hybridization, we found a differential regulation of RKBP and alpha 1-AT gene expression in the liver. Ovariectomy results in a 67% reduction of RKBP mRNA levels but a 30% increase of alpha 1-AT mRNA levels. Estradiol or progesterone treatment of the ovariectomized rats increases RKBP transcripts by 2.5- and 6.5-fold, respectively, but reduces alpha 1-AT mRNA level by 30% and 45%, respectively. In contrast to kininogen expression, both RKBP and alpha 1-AT mRNA levels in the liver are at the lowest at birth and rapidly increase during growth and development. Rats injected with endotoxin from 4 to 24 h show a time-dependent decrease of RKBP mRNA levels while the same treatment induces alpha 1-AT gene expression. RKBP mRNA levels in the normotensive Wistar Kyoto (WKY) rats are higher than those in the spontaneously hypertensive rats (SHR) while there are no differences of alpha 1-AT mRNA levels between SHR and WKY.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute-Phase Reaction↗
PubMed1992

Identification of a nodD-like gene in Frankia by direct complementation of a Rhizobium nodD-mutant.

Clones from a Frankia At4 gene bank were pooled into groups and mass conjugated into a nodD mutant of Rhizobium leguminosarum bv. viciae by triparental matings. When peas were inoculated with the pooled transconjugants, nodulation was observed. A plasmid, pAt2GX containing Frankia DNA, was isolated from bacteria recovered from these nodules. This plasmid was shown to complement a nodD mutant of R. leguminosarum bv. viciae. Thus pAt2GX contains a Frankia gene that is functionally equivalent to nodD of R. leguminosarum bv. viciae.

Bacterial Proteins↗
PubMed1992

Myeloblastic leukemoid reaction in paroxysmal nocturnal hemoglobinuria associated with myelodysplasia.

Paroxysmal nocturnal hemoglobinuria (PNH) has been observed to evolve into myelofibrosis and acute myeloid leukemia. Myeloblastic leukemoid reaction has not been described in PNH. We described a patient with PNH with myelodysplasia and septicemia. The marrow aspirates showed a picture of myeloblastosis which subsided when sepsis was controlled. The myeloblastic leukemoid reaction in our patient related to overwhelming sepsis, splenectomy and overt hemolysis.

Bone Marrow↗
PubMed1992

[Evaluation of marrow neutrophilic reserve in normal subjects and aplastic anemia patients].

The widespread use of potent marrow suppressing drugs in malignant diseases has gained increasing importance in the functional assessment of neutrophil system. A peripheral neutrophil count of 500 cells/cumm is considered to be a good indicator of risk of bacterial infection. However, despite extreme neutropenia, infection is rare in chronic benign neutropenia patients. It may be presumed that adequate marrow reserve delivers mature neutrophil during bacterial infection in these patients. Currently available methods such as determination of blood neutrophil concentration, differential counts, estimation of bone marrow cellularity provide a relatively crude estimates of the functional capacity of the neutrophil system. The use of endotoxin, etiocholanolone and steroids have been introduced to determine the marrow neutrophil reserve with greater accuracy. In this study, intravenous hydrocortisone 100 mg was used to evaluate marrow neutrophilic reserve in normal and aplastic anemia subjects. The results showed significant statistical differences (P less than 0.001) between controls and aplastic subjects in 2, 3, 4 and 5th hour groups after steroid injection. The sensitivity and specificity occurring 3 and 4 hours after steroid injection were 78.6% and 100%, respectively. The normal marrow neutrophilic reserve showed more than 1198/cumm of increment neutrophilic counts 4 hours after steroid injection. No complication was found. The present finding suggests that steroid may be useful as an accurate evaluator of marrow neutrophilic reserve.

Adult↗
PubMed1991

Regulation of proline utilization in enteric bacteria: cloning and characterization of the Klebsiella put control region.

Enteric bacteria can grow on proline as the sole nitrogen and carbon source. Expression of the proline utilization (put) operon in Klebsiella strains and Escherichia coli is responsive to nitrogen regulation. In contrast, Salmonella typhimurium cannot activate put operon expression when growing in medium with glucose as a carbon source and proline as the sole nitrogen source. To compare nitrogen regulatory sites in the control regions of the put operons in these three closely related genera, we cloned the Klebsiella put operon onto a plasmid. The putA and putP genes were localized on the plasmid by transposon mutagenesis. The DNA sequence of the put control region was determined and compared with those of the put control regions from S. typhimurium and E. coli. The overall size and organization of the put control region were very similar in all three bacteria. However, no obvious ntr regulatory sites were found in this region, and transcription of the put genes started at the same sites during growth with limiting or excess nitrogen. These results strongly suggested that the Klebsiella put operon may not be directly regulated by the ntr system.

Base Sequence↗