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Biomedical subjects

L M Chen

Publications and source records attributed to L M Chen.

At least 73 records · Page 4Linked to original sources
PubMed1997

Adenovirus-mediated delivery of human kallistatin gene reduces blood pressure of spontaneously hypertensive rats.

Human kallistatin, or human tissue kallikrein-binding protein (HKBP), is a serine proteinase inhibitor (serpin). Transgenic mice overexpressing rat kallikrein-binding protein are hypotensive. To elucidate therapeutic potentials of kallistatin in hypertension, the human kallistatin gene in an adenoviral vector was directly introduced into spontaneously hypertensive rats (SHR) through portal vein injection. The kallistatin cDNA construct (RSV-cHKBP) under the promoter control of Rous sarcoma virus 3' long terminal repeat (LTR) was incorporated into adenovirus (Ad.RSV-cHKBP). Recombinant kallistatin in 293 cells transfected with RSV-cHKBP or Ad.RSV-cHKBP was measured by ELISA and by its complex formation with tissue kallikrein. A single intraportal vein injection of Ad.RSV-cHKBP at a dose of 8 x 10(10) pfu results in a significant reduction of blood pressure of SHR for 4 weeks. Human kallistatin mRNA was detected in the liver, spleen, kidney, aorta, and lung of rats receiving gene delivery. Immunoreactive human kallistatin in rat serum was detected at the highest level 1 day post injection and at lesser amounts in rat tissues. This study shows that adenovirus harboring Ad.RSV-cHKBP produces functional kallistatin, and adenovirus-mediated transfer of the human kallistatin gene reduces blood pressures of SHR. The results suggest that kallistatin may function as a vasodilator in vivo and provide important information for a potential gene therapy approach to hypertension.

Adenoviridae↗
PubMed1997

Beneficial effects of kallikrein-binding protein in transgenic mice during endotoxic shock.

Rat kallikrein-binding protein (RKBP) is a negative acute phase protein. The potential role of RKBP in inflammation was evaluated in transgenic mice overexpressing the RKBP gene under the control of the mouse metallothionein metal-responsive promoter. Bacterial endotoxic lipopolysaccharide (LPS) was injected intraperitoneally into mice at a dose of 600 microg/25 g body weight. The death toll was recorded every 12 hours for 3 days. The survival rate of transgenic male mice (n=78) was 33.3% while that of control male mice (n=54) was 9.3% 3 days post LPS injection. In comparison, the survival rate of transgenic female mice (n=59) was 55.9% while that of control female mice (n=65) was 30.8%. Recombinant RKBP levels in the circulation of these mice increased by 3-fold after LPS treatment. The results show that RKBP transgenic mice have a higher survival rate than their non-transgenic control littermates after endotoxin shock and female mice are more resistant to lethality induced by endotoxin shock than male mice in both transgenic and control groups. These findings suggest that kallikrein-binding protein has a protective effect during acute phase inflammation.

Animals↗
PubMed1997

A retrospective review of a large cohort of patients undergoing the process of withholding or withdrawal of life support.

OBJECTIVES: To determine the proportion of patients who died as a result of the withdrawal or withholding of life support (WD/WHLS) in the intensive care units (ICUs) of three teaching hospitals and to describe the process involved by determining: a) why the decision was made to withdraw support (WDLS); b) when WDLS took place; and c) how the WDLS process was conducted. DESIGN: Retrospective cohort study. SETTING: Three university-affiliated ICUs. PATIENTS: Four hundred nineteen patients who died in one of three academic, tertiary care ICUs over a 1-yr period. INTERVENTIONS: Retrospective chart review. Data collected included age, gender, admitting diagnoses, comorbid disease, Acute Physiology and Chronic Health Evaluation II score, and mode of death (brain death, death due to withholding of life support, death due to WDLS, or death despite ongoing therapy). For those patients dying due to WDLS, the reason for WDLS, person initiating discussion, timing of WDLS, degree of organ dysfunction, order of withdrawal of life support modalities, and the use of sedatives and analgesics were recorded. MEASUREMENTS AND MAIN RESULTS: Seventy percent of patients died by WD/WHLS and 8.4% were brain dead. Patients undergoing WD/WHLS were older and had a longer length of stay than patients dying from other causes. Poor prognosis was the most common reason given for WDLS, reflected by significant organ dysfunction at the time of WDLS. Future quality of life was a less frequently cited reason. Most patients undergoing WDLS did so early in their ICU stay, although time to WDLS appeared to reflect diagnostic group. Few meetings occurred before WDLS and death occurred soon after initiating WDLS. There was a preference of withdrawing mechanical ventilation last and large amounts of morphine (mean 21 +/- 33 [SD] mg/hr) and benzodiazepines (mean 8.6 +/- 11 mg/hr) were used. Little variability in practice was apparent among the studied ICUs. CONCLUSIONS: Similar to other studies, WD/WHLS was the most common cause of death in academic ICUs and poor patient prognosis was considered the most important factor in deciding on WDLS. However, in contrast to other studies, future quality of life was not as frequently cited a reason for WDLS and larger amounts of morphine were used during WDLS. These discrepancies suggest areas for potential future research.

APACHE↗
PubMed1997

Underreporting and misclassification of maternal mortality in Taiwan.

BACKGROUND: Underreporting and misclassification of maternal deaths are universal. The purposes of this study were to quantify the level of underestimation of maternal mortality and to analyze possible factors that contribute to underreporting and misclassification of maternal mortality. METHODS: An interview census of all registered deaths that occurred during 1984-1988 in women of reproductive age was undertaken in Taiwan. Pregnancy-related deaths were screened from all collected questionnaires and death certificates by the researchers. The screened pregnancy-related deaths were then reviewed and evaluated by obstetrician-gynecologists; a cause of each death was assigned. RESULTS: For the five years, on average, the rate of underreporting of maternal mortality is 58.38% and the correct/confirmed rate of classification is 53.28%. Underreported and misclassified maternal deaths are more likely for women aged 20-24, with stillbirth and fetal death, care sought for non-obstetric reasons, care received in private hospitals and clinics, occurrence in the home, certification by non-obstetrician-gynecologists and court doctors, and death from non-obstetric causes. CONCLUSIONS: This study shows the limitations of official vital registration and concludes that dependence on death certificates alone to identify maternal deaths is incomplete and incorrect.

Adult↗
PubMed1996

Requirement of CDC42 for Salmonella-induced cytoskeletal and nuclear responses.

The bacterial pathogen Salmonella typhimurium triggers host cell signaling pathways that lead to cytoskeletal and nuclear responses required for pathogenesis. Here, the role of the small guanosine triphosphate (GTP)-binding protein CDC42Hs in these responses was examined. Expression of a dominant interfering mutant of CDC42 (CDC42HsN17) prevented S. typhimurium-induced cytoskeletal reorganization and subsequent macropinocytosis and bacterial internalization into host cells. Cells expressing constitutively active CDC42 (CDC42HsV12) internalized an S. typhimurium mutant unable to trigger host cell responses. Furthermore, expression of CDC42HsN17 prevented S. typhimurium-induced JNK kinase activation. These results indicate that CDC42 is required for bacterial invasion and induction of nuclear responses in host cells.

Animals↗
PubMed1996

Tissue kallikrein-binding protein reduces blood pressure in transgenic mice.

The kallikrein-kinin system participates in blood pressure regulation. One of the kallikrein-kinin system components, kallikrein-binding protein, binds to tissue kallikrein and inhibits its activity in vitro. To investigate potential roles of rat kallikrein-binding protein (RKBP) in vivo, we have developed transgenic mice that express an RKBP gene under the control of the mouse metallothionein metal-responsive promoter. Expression of the transgene, RKBP, was detected in the liver, kidney, lung, heart, pancreas, salivary glands, spleen, brain, testis, and adrenal gland at the mRNA and protein levels. Systolic blood pressures of homozygous transgenic mice were 88.5 +/- 0.8 mm Hg (mean +/- S.E., n = 19, P < 0.001) for one line and 88.8 +/- 1.6 mm Hg (mean +/- S.E., n = 19, P < 0.001) for another, as compared with 100.5 +/- 0.8 mm Hg (mean +/- S.E., n = 18) for control mice. Direct blood pressure measurements of these transgenic mice through an arterial cannula showed similar reductions of blood pressure. Intravenous injection of purified RKBP into mice via a catheter produced a dose-dependent reduction of the mean arterial blood pressure. Our findings suggest that RKBP may function as a vasodilator in vivo, independent of regulating the activity of tissue kallikrein.

Animals↗
PubMed1996

The cytosolic subunit p67phox contains an NADPH-binding site that participates in catalysis by the leukocyte NADPH oxidase.

The NADPH-dependent respiratory burst oxidase of human neutrophils catalyzes the reduction of oxygen to superoxide using NADPH as the electron donor and is essential for normal host defenses. To gain insight into the function of the various oxidase subunits that are required for the full expression of catalytic activity, we studied the interactions between the 2',3'-dialdehyde derivative of NADPH (NADPH dialdehyde) and neutrophil cytosol. NADPH dialdehyde treatment of cytosol resulted in the loss of the ability of the cytosol to participate in cell-free oxidase activation; this inactivation was blocked by NADPH but not by NAD, NADP, or GTP. Partial purification of neutrophil cytosol yielded a single peak which could restore the activity lost in cytosol treated with NADPH dialdehyde. This peak contained p67phox but not p47phox or Rac2. Purified recombinant p67phox was similarly able to restore the activity lost in NADPH dialdehyde-treated cytosol and bound [32P]NADPH dialdehyde in a specific fashion. The activity of recombinant p67phox in cell-free oxidase assays was lost on treatment with NADPH dialdehyde. Together, these data suggest p67phox contains the catalytic NADPH-binding site of the leukocyte NADPH oxidase.

Aldehydes↗
PubMed1996

Systemic and portal vein delivery of human kallikrein gene reduces blood pressure in hypertensive rats.

There is an inverse correlation between systemic blood pressure and urinary kallikrein levels in humans and hypertensive animal models, suggesting that the tissue kallikrein-kinin system plays an important role in blood pressure regulation. In this study, we explored the potential of human kallikrein gene delivery on blood pressure reduction in spontaneously hypertensive rats (SHR). The human tissue kallikrein gene or cDNA was placed under the control of following promoters: the metallothionein gene metal response-element (MRE-pHK), albumin gene (ALB-pHK), Rous sarcoma virus 3' long terminal repeat (LTR) (RSV-cHK), and cytomegalovirus (CMV-cHK). A single injection of these kallikrein DNAs results in a significant reduction of blood pressure in SHR, which lasts for 5-6 weeks. Systemic delivery of CMV-cHK, RSV-cHK, and MRE-pHK has a greater effect on blood pressure reduction than ALB-pHK, whereas intraportal vein gene delivery of ALB-pHK is more effective than the other kallikrein DNA constructs. The degree of blood pressure reduction depends on the amount of administered DNA and the age of the animals. Reduction of blood pressure was observed in adult, but not young, SHR. The expression of human tissue kallikrein in rats was identified by an ELISA that is specific for human tissue kallikrein. No antibodies to either human tissue kallikrein or its DNA were detected in rat sera after somatic gene delivery. These results show that somatic gene delivery of human tissue kallikrein causes a lowering effect of systolic blood pressure in genetically hypertensive rats and provide valuable information for kallikrein gene therapy in the treatment of hypertension.

Animals↗
PubMed1996

Kallistatin, a novel human tissue kallikrein inhibitor: levels in body fluids, blood cells, and tissues in health and disease.

Kallistatin, a human serine proteinase inhibitor, is a newly identified tissue kallikrein inhibitor. It binds strongly to tissue kallikrein but weakly to other serine proteinases such as chymotrypsin and elastase. The tissue distribution and changes in kallistatin levels in human diseases were characterized by using specific monoclonal and polyclonal antibodies against kallistatin. Kallistatin antigen levels in blood cells, fluids, and tissues measured with a specific enzyme-linked immunosorbent assay showed displacement curves that were parallel with those in purified kallistatin, indicating their immunologic identity. Expression of kallistatin mRNA in platelets, neutrophils, lymphocytes, monocytes, endothelial cells, hepatocytes, and colon and prostate carcinoma cells was identified by reverse transcription-polymerase chain reaction followed by Southern blot analysis. Plasma kallistatin concentration was 22.1 +/- 3.5 micrograms/ml in 30 normal subjects and 21.1 +/- 3.8 micrograms/ml in 5 patients with C1 inhibitor deficiency. A significantly reduced kallistatin level (7.2 +/- 2.5 micrograms/ml, p < 0.001) was seen in plasma samples from 9 patients with liver disease and 10 patients with sepsis (7.7 +/- 3.5 micrograms/ml, p < 0 .001). Further, kallistatin levels in 10 women taking oral contraceptives (19.8 +/- 3.8 micrograms/ml) and 21 pregnant women (14.9 +/- 3.3 microg/ml) were significantly lower than those seen in healthy individuals. These data suggest that kallistatin is found in plasma, is produced mostly in the liver, and can be consumed during sepsis. Its consumption in sepsis may indicate a protective role to prevent blood pressure lowering.

Antibodies, Monoclonal↗
PubMed1996

Salmonella spp. are cytotoxic for cultured macrophages.

We have shown by a variety of microscopical and biochemical techniques that Salmonella spp. are cytotoxic for cultured J774A.1 and bone marrow-derived murine macrophages. The cytotoxicity is initially manifested by inhibition of membrane ruffling and macropinocytosis in infected macrophages, and is followed by cell death. Macrophages killed by Salmonella spp. exhibited features of apoptosis such as condensation and fragmentation of chromatin, membrane blebbing, and the presence of cytoplasmic nucleosomes and apoptotic bodies. Cytotoxicity does not require bacterial internalization as cytochalasin D, a drug that prevents bacterial uptake, did not prevent Salmonella-induced macrophage cell death. However, the cytotoxic effects are strictly dependent upon the expression of the invasion-associated Type III protein-secretion system encoded at centisome 63 of the Salmonella chromosome. Wild-type Salmonella typhimurium grown under conditions that do not allow optical expression of this system or strains of Salmonella carrying mutations in genes that encode components of this protein-secretion system were devoid of macrophage cytotoxicity. In addition, mutations in invJ, spaO, sipB, sipC and sipD, which encode proteins that are secreted via this secretion apparatus and are required for bacterial entry into non-phagocytic cells, also abolished the toxicity. In contrast, mutations in sipA and sptP, which encode secreted proteins that are not required for bacterial invasion, had no effect on macrophage cytotoxicity. These results indicate a close correlation between the mechanisms of bacterial internalization into non-phagocytic cells and those that lead to macrophage cytotoxicity. Host-adapted serotypes of Salmonella such as S. typhi, S. gallinarum and S. dublin were also toxic for murine macrophages, indicating that this virulence property is probably present in most Salmonella spp. and is not associated with the mechanisms responsible for host range.

Animals↗
PubMed1996

Expression and cellular localization of tissue kallikrein-kinin system in human adrenal gland.

The tissue kallikrein-kinin system has been implicated in regulating blood pressure and electrolyte homeostasis. To understand the function of this system, we identified the expression and cellular localization of its components including tissue kallikrein, kallistatin, kininogen, and bradykinin B1 and B2 receptors in human adrenal gland. Reverse transcription-polymerase chain reaction followed by Southern blot analysis showed that these five components of this system were all expressed in human adrenal gland. In situ hybridization histochemistry with respective digoxigenin-labeled antisense riboprobes revealed localization of kallikrein transcript throughout the adrenal cortex and medulla except the zona glomerulosa, whereas kallistatin mRNA was only localized in the zona fasciculata. Low-molecular-weight kininogen and B2 receptor mRNAs were colocalized in the zona glomerulosa and zona fasciculata and also in the zona reticularis and chromaffin cells but to a lesser degree. The B1 receptor mRNA was stained in the zona fasciculata and medulla. These results show the expression and differential colocalization of the components of the tissue kallikrein-kinin system and reveal the potential action sites of this system in the adrenal gland.

Adrenal Glands↗
PubMed1996

High-salt diet upregulates kininogen and downregulates tissue kallikrein expression in Dahl-SS and SHR rats.

Tissue kallikrein cleaves low-molecular-weight (low-M(r)) kininogen to produce the vasoactive kinin peptide. It has been suggested that hypertensive patients with low urinary kallikrein excretion may have a defect in sodium handling. In this study, we examined the effect of a high-salt diet on the expression of tissue kallikrein and kininogen genes in Dahl salt-sensitive rats (Dahl-SS), spontaneously hypertensive rats (SHR), and normotensive Sprague-Dawley rats (SD) by Northern and Western blot analysis and radioimmunoassay. Control and experimental groups received normal and high-salt diets containing 0.4% and 8% NaCl, respectively, for 6 wk. High-salt diet induced a significant time-dependent increase of blood pressure in both strains of hypertensive rats and a slight but significant increase of blood pressure in normotensive SD rats. Hepatic kininogen mRNA levels of both Dahl-SS and SHR on a high-salt diet increased 2.4-fold and 2.0-fold, respectively, while alpha 1-antitrypsin mRNA levels were not changed in rats receiving high-salt diet. Immunoreactive total kininogen and low-M(r) kininogen (58 kDa) levels in sera increased in response to high-salt diet in both strains of hypertensive rats. In SD rats, the low-M(r) kininogen level in sera was unaltered, whereas total kininogen increased in response to high-salt diet. Tissue kallikrein mRNAs in the kidney and salivary glands of Dahl-SS, SHR, and SD rats were reduced, whereas beta-actin mRNA was not altered by high-salt diet. Similarly, immunoreactive intrarenal kallikrein levels were reduced in these rats in response to high-salt diet. These studies show that increases in blood pressure after salt loading in Dahl-SS and SHR are accompanied by increases in low-M(r) kininogen. Tissue kallikrein gene expression in hypertensive Dahl-SS and SHR and normotensive SD rats is suppressed after salt loading. These findings show that reduced renal kallikrein expression and increased kininogen expression is regulated at the transcriptional level during salt loading.

Animals↗
PubMed1996

Localization of the prostatic apex for radiotherapy planning: a comparison of two techniques.

Autopsy and pathology studies have shown that the caudal portion of the prostate gland harbors tumour in 64-75% of specimens examined. Accurate localization of the prostatic apex may be important in improving local control with external beam radiation therapy. We compared the location of the apex obtained with CT based treatment planning versus localization using retrograde urethrography in 32 consecutive patients. The prostatic apex, localized by CT and retrograde urethrography, was compared relative to the ischial tuberosities and the symphysis pubis. Discordance between the location of the prostatic apex as defined on CT scan and retrograde urethrography was found in 50% of patients evaluated. There was 31% discordance between the location of the prostatic apex as defined on CT and retrograde urethrography when data were analysed with the location of the prostatic apex 1 cm above the narrowing on urethrography, a definition others have suggested. The urethrogram defined prostatic apex, as compared with the CT definition, necessitated the treatment of more of the surrounding normal tissues in 31% of our cases, with four-field techniques. Comparison of dose-volume histograms of the bladder, rectum and penis irradiated for target volumes defined by CT versus retrograde urethrography showed that more penis was irradiated in some patients with the urethrogram defined prostatic apex; irradiation of the base of the penis could be relatively avoided by using a six-field treatment plan instead of the standard four-field box. There is discordance between the CT and urethrogram defined prostatic apex. Dose-volume histogram information suggests that differences in apex localization can significantly affect doses to normal adjacent prostatic tissues. Combining CT localization with the urethrogram localization of the prostatic apex optimizes radiotherapy planning and dose delivery.

Humans↗
PubMed1996

Examination of pericardial effusions by cytology and immunocytochemistry.

BACKGROUND: Cytologic examination of pericardial effusions (PE) has been uncommonly reported in the literature. Fewer reports have studied the role of immunocytochemistry in diagnosis of PE. We described our experience, according to cytologic examination and immunocyto-chemical staining of PE. METHODS: Over a four-year period, 50 PE specimens from 36 patients were examined cytologically by Wright-Giemsa stain and/or Papanicolaou stain, at Tri-Service General Hospital. Immunocytochemical stainings were further performed to ensure the cell nature. RESULTS: Eighteen of these patients had benign etiologies; the others suffered from malignant diseases. Sixteen cases, including eight benign and eight malignant, received pericardial biopsy for comparison. Commonly encountered cells in PE with any kind of etiology included neutrophils, lymphocytes macrophages and mesothelial cells. Cancer cells were additionally seen in malignant PE, but not in all cases with cancers. Furthermore, reactive mesothelial cells and macrophages were sometimes not easily distinguished from cancer cells, and reactive lymphocytes frequently mimicked hematopoietic malignancies. Further, the nature of small round tumor cells can hardly be ensured by morphologic examination only. Immunocytochemical studies were successfully used to solve these dilemmas in some suspicious cases. CONCLUSIONS: This study showed that cytology together with clinical information and immunocytochemistry can achieve excellent sensitivity and specificity in identification of malignant PE. The use of immunocytochemistry can even ensure the nature of certain cancers such as small cell carcinoma and B cell lymphoma.

Adult↗
PubMed1995

Molecular cloning, purification and in situ localization of human colon kallikrein.

We have cloned and characterized a full-length cDNA encoding tissue kallikrein from a human colon carcinoma cell line (T84). The nucleic acid sequence of the colon kallikrein cDNA is identical to that of renal/pancreatic or tissue kallikrein cDNA. Reverse-transcription PCR followed by Southern-blot analysis using specific oligonucleotide probes showed expression of tissue kallikrein in human colon, pancreas and kidney. Tissue kallikrein mRNA was localized in glandular epithelial cells (goblet cells) in colon by in situ hybridization histochemistry. Human colon kallikrein was purified to apparent homogeneity by DEAE-Sepharose Cl-6B, aprotinin-affinity, and HQ/M perfusion chromatography. The purified colon kallikrein migrated as a broad, 40-45 kDa band in SDS/PAGE and was recognized by antibodies to human tissue kallikrein. The linear displacement curves for the colon kallikrein in an RIA were parallel with the human tissue kallikrein standard curve, indicating their immunological identity. The N-terminal sequence of the purified colon kallikrein matches completely with that of purified urinary or tissue kallikrein. These results indicate that human colon kallikrein is transcribed from the tissue kallikrein gene.

Amino Acid Sequence↗