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Biomedical subjects

B Han

Publications and source records attributed to B Han.

At least 73 records · Page 4Linked to original sources
PubMed1999

Quantification of iNOS mRNA with reverse transcription polymerase chain reaction directly from cell lysates.

Inducible nitric oxide synthase (iNOS) is a member of a family of primary inflammatory response genes. Quantitative measurement of iNOS mRNA levels is important for the study of gene expression of this enzyme during the process of inflammation. We report here a method for quantitative measurement of iNOS mRNA levels with rtPCR directly from cells lysed with a single step phenol/chloroform/ether extraction. Using a mouse macrophage cell line, J774.2, which expresses iNOS mRNA upon LPS + IFN-gamma treatment as the model, the effects of the extraction on iNOS mRNA recovery and cytosolic RNase removal have been studied. The cells are lysed and RNases denatured and removed by phenol/chloroform extraction. Trace amounts of the phenol partitioned in the samples are then removed by ether extraction. After the extraction, the samples can be used directly for reverse transcription and PCR without further purification of RNA. The recovery of specific mRNA is not affected by the extraction procedure and externally added iNOS cRNA shows no degradation by the extracted cell lysates. Measurement of iNOS mRNA with this procedure is linear using serially double-diluted cells in the range from 94 to 6000 cells. The efficiencies of rtPCR of iNOS wild-type and deletion cRNAs are also compared in our study. By controlling the molecular size of the deletion construct to within 10% of that of the wild type and maintaining PCR cycling below 25 cycles, the rtPCR efficiencies of iNOS wild type and deletion are identical. The detection of rtPCR products is enhanced by hybridization with specific probes. Under these conditions, iNOS mRNA concentration can directly be calculated from the internal standard in each tube without a standard curve. We conclude that our procedure provides an accurate method for quantitative measurement of iNOS mRNA from limited amount of cells without complete RNA isolation.

Animals↗
PubMed1999

Requirement for the homeobox gene Hb9 in the consolidation of motor neuron identity.

The homeobox gene Hb9, like its close relative MNR2, is expressed selectively by motor neurons (MNs) in the developing vertebrate CNS. In embryonic chick spinal cord, the ectopic expression of MNR2 or Hb9 is sufficient to trigger MN differentiation and to repress the differentiation of an adjacent population of V2 interneurons. Here, we provide genetic evidence that Hb9 has an essential role in MN differentiation. In mice lacking Hb9 function, MNs are generated on schedule and in normal numbers but transiently acquire molecular features of V2 interneurons. The aberrant specification of MN identity is associated with defects in the migration of MNs, the emergence of the subtype identities of MNs, and the projection of motor axons. These findings show that HB9 has an essential function in consolidating the identity of postmitotic MNs.

Animals↗
PubMed1999

The linoleic acid metabolite, 13-HpODE augments the phosphorylation of EGF receptor and SHP-2 leading to their increased association.

In previous studies with Syrian hamster embryo fibroblasts, we found that a specific lipoxygenase metabolite of linoleic acid, 13(S)-hydroperoxyoctadecadienoic acid (HpODE), enhanced epidermal growth factor (EGF) signal transduction in a tumor suppressor gene plus phenotype (supB+); with a diminished response to 13(S)-HpODE in a tumor suppressor gene minus phenotype (supB-). This differential response was attributed to differences in the rate of EGF receptor (EGFR) dephosphorylation. To further define the molecular basis for these observations, in this report we examine the interaction of phosphorylated EGFR with the SH2 domain-containing protein tyrosine phosphatase, SHP-2, a positive modulator of EGF dependent cell growth. SHP-2 associated with phosphorylated EGFR to a greater extent in supB+ cells when compared to supB-. This differential association could not be accounted for by differences between suppressor gene phenotypes in SHP-2 protein level or mutations in the molecular sequence. The addition of 13(S)-HpODE stimulated a concentration-dependent increase in EGF-dependent phosphorylation of SHP-2 and its association with EGFR. A more dramatic response was observed in the supB+ cells. Differences in SHP-2 interaction with EGFR may account, in part, for phenotypic differences in the growth rates and responsiveness to EGF between the supB+ and supB- cells. EGFR-SHP-2 association appears to play an important role in the regulation of EGFR signal transduction.

Animals↗
PubMed1999

Supraphysiologic concentrations of cerulein induce apoptosis in the rat pancreatic acinar cell line AR4-2J.

Little is known as yet about the role of apoptosis in pancreatic damage. This study evaluated the effects of supraphysiologic concentrations of the cholecystokinin (CCK) analog, cerulein, which causes cell damage in vitro and acute pancreatitis in vivo, on cell proliferation and DNA fragmentation in the rat pancreatic acinar cell line AR4-2J. Cerulein inhibited the cell proliferation of AR4-2J time- and dose-dependently to approximately 60% of the control level at 10(-6) M after 72 h. DNA fragmentation, as assessed by both electrophoresis and enzyme-linked immunosorbent assay (ELISA), occurred at cerulein concentrations > or = 10(-8) M. The maximal DNA fragmentation as measured by ELISA was reached after 24 h. Cerulein at concentrations > or = 10(-9) M induced wild-type p53. Glutathione (1 mM) diminished the effects of cerulein on both cell proliferation and DNA fragmentation, whereas spermine (100 microM), which partially attenuated DNA fragmentation, did not have an effect on cell proliferation. The CCK-A-receptor antagonist loxiglumide completely abolished the effect of cerulein on DNA fragmentation. The serine-protease inhibitor FUT-175 (10 microM), the cysteine-protease inhibitor NCO-700 (5 mM), and ethylene glycol tetraacetic acid (EGTA; 500 microM) all had no effects on the changes in cell proliferation and DNA fragmentation induced by cerulein. The data suggest that supraphysiologic concentrations of cerulein rapidly induce apoptosis in AR4-2J cells and only later inhibit cell proliferation. These effects are mediated by CCK-A receptors. Cerulein-induced apoptosis may involve the induction of wild-type p53 or glutathione depletion or both.

Animals↗
PubMed1999

Cholecystokinin induction of mob-1 chemokine expression in pancreatic acinar cells requires NF-kappaB activation.

Inflammatory mediators are involved in the early phase of acute pancreatitis, but the cellular mechanisms responsible for their generation within pancreatic cells are unknown. We examined the role of nuclear factor-kappaB (NF-kappaB) in cholecystokinin octapeptide (CCK-8)-induced mob-1 chemokine expression in pancreatic acinar cells in vitro. Supraphysiological, but not physiological, concentrations of CCK-8 increased inhibitory kappaB (IkappaB-alpha) degradation, NF-kappaB activation, and mob-1 gene expression in isolated pancreatic acinar cells. CCK-8-induced IkappaB-alpha degradation was maximal within 1 h. Expression of mob-1 was maximal within 2 h. Neither bombesin nor carbachol significantly increased mob-1 mRNA or induced IkappaB-alpha degradation. Thus the concentration, time, and secretagogue dependence of mob-1 gene expression and IkappaB-alpha degradation were similar. Inhibition of NF-kappaB with pharmacological agents or by adenovirus-mediated expression of the inhibitory protein IkappaB-alpha also inhibited mob-1 gene expression. These data indicate that the NF-kappaB signaling pathway is required for CCK-8-mediated induction of mob-1 chemokine expression in pancreatic acinar cells. This supports the hypothesis that NF-kappaB signaling is of central importance in the initiation of acute pancreatitis.

Adenoviridae↗
PubMed1999

Family caregiving for patients with stroke. Review and analysis.

BACKGROUND: The literature on family caregiving for stroke patients is reviewed with the goals of (1) evaluating the effects of stroke caregiving on caregivers' well-being, (2) outlining deficiencies and methodological limitations of current research, and (3) outlining policy and practice implications of current studies. SUMMARY OF REVIEW: A total of 20 published stroke caregiving research articles were included in this review. Across studies, the effects of stroke caregiving on caregivers' well-being and the significant predictors of caregivers' depression were analyzed. Current evidence suggests that stroke caregivers have elevated levels of depression at both the acute stroke phase and the chronic stroke phase. However, major gaps are apparent in this literature, with few studies addressing such areas as caregiver physical health, ethnicity, and caregiver interventions. CONCLUSIONS: Given the increasing prevalence of stroke as well as the increasing pressures on families to provide care, more research is needed to guide policy and practice in this understudied topic.

Caregivers↗
PubMed1999

Induction of apoptosis by Coprinus disseminatus mycelial culture broth extract in human cervical carcinoma cells.

Extract of Coprinus disseminatus (pers. Fr.) (C. disseminatus) culture broth (EDCB) inhibits proliferation and induces apoptosis in the human cervical carcinoma cells at 5 microg/ml. To determine whether the cell death induced by the EDCB recruits caspases or not, one of the exclusive pathways in cell death, we examined caspase-3 activity in this cell death process. The activity of caspase-3 was remarkably increased when the cell was treated with EDCB, and this activity was nullified by Z-VAD-FMK, a well known caspase-3 inhibitor. From these results, we would expect the EDCB to contain substances with the ability to induce apoptosis in the human cervical carcinoma cells. The extent of the EDCB induced apoptosis is cell line-dependent.

Amino Acid Chloromethyl Ketones↗
PubMed1999

Age- and growth hormone-induced alterations in renal sulfate transport.

The effects of growth hormone (GH) treatment on renal sodium sulfate cotransport (NaSi-1) were studied in adult (9-10 months) and old (22-23 months) male Fischer 344 rats. All animals received twice-daily s.c. injections of recombinant human GH (hGH; 4 mg/kg) for up to 6 days. Animals were sacrificed by exsanguination on days 0, 1, 2, 3, 4, 5, and 6. Kidneys were removed, and kidney cortex was trimmed immediately and used for RNA and membrane preparations. Plasma hGH concentrations were significantly lower in old rats during the hGH treatment (P <.05). Insulin-like growth factor-I (IGF-I) levels were significantly increased and remained stable after day 2 of hGH treatment in both age groups (P <.05). There was no significant difference in plasma IGF-I levels between age groups. Plasma IGF-I binding protein 3 (IGFBP-3) concentrations were significantly higher in 9- to 10-month-old rats compared with that in 22- to 23-month-old animals (P <.001). There were no significant differences in plasma IGFBP-3 concentrations between days of hGH treatment. The NaSi-1 mRNA levels were significantly lower in 22- to 23-month-old rats compared with that in 9- to 10-month-old animals (P <.001). The NaSi-1 mRNA levels were significantly increased on days 2 and 3 of hGH treatment (P <.05) and then gradually decreased to the control value. The NaSi-1 protein levels in old animals (22-23 months) were also significantly lower than that of 9- to 10-month-old animals and were significantly increased from day 2 of hGH treatment, reaching a maximum level on day 3 or 4 and then returning to the baseline level in both age groups. From these results, it was concluded that 1) NaSi-1 mRNA and protein levels are lower in old animals and increase in both adult and aged rats after hGH treatment, 2) plasma IGF-I levels are similar in adult and aged rats and increase after hGH treatment, and 3) plasma IGFBP-3 levels are lower in old rats and remain unchanged after hGH treatment.

Aging↗
PubMed1999

Aluminum uptake and effects on transferrin mediated iron uptake in primary cultures of rat neurons, astrocytes and oligodendrocytes.

Transferrin (Tf) is known primarily for its role in the transport and cellular uptake of iron (Fe). Tf is also the major serum binding protein for Al. In this study, primary rat oligodendrocyte, neuron and astrocyte cultures were found to differ in Tf mediated Fe and Al uptake and in the effect of Al-Tf on Fe-Tf uptake during 4 h incubation periods. When incubated with Al-Tf (1.25 microM), oligodendrocytes displayed a 3- to 4-fold increase (p=.0002) in Al, neurons demonstrated a much smaller (p=.06) increase, and no increase was seen for astrocytes. When incubated with equimolar Al citrate or Al chloride, no increase in cellular Al was seen in any of the three cell types. Oligodendrocytes, astrocytes and neurons all demonstrated greater 59Fe uptake from Fe-Tf than Fe chloride. This uptake could be inhibited by excess Fe-Tf in oligodendrocytes and neurons, but not astrocytes. A small but significant inhibition of 59Fe uptake from Fe-Tf was seen after addition of Al-Tf to the incubation medium of oligodendrocytes, but not neurons or astrocytes. Oligodendrocytes may be particularly vulnerable to the accumulation of excess intracellular Al, and to interference of Al with Fe uptake. Such effects could contribute to Al-induced neurotoxicity if they result in altered myelin formation or maintenance.

Aluminum↗
PubMed1999

[The development of a RF electrical pole catheter for heart ablation].

This article describes a RF electrical pole catheter for heart ablation. There is a device in the catheter, that can adjust the bending of the catheter tip on which there are a pair of electrical poles that is able to record and map the electrical potential of the heart, and to transfer 50 W electrical energy for focus ablution of heart disease.

Cardiac Surgical Procedures↗
PubMed1998

Brain-wave recognition of sentences.

Electrical and magnetic brain waves of two subjects were recorded for the purpose of recognizing which one of 12 sentences or seven words auditorily presented was processed. The analysis consisted of averaging over trials to create prototypes and test samples, to each of which a Fourier transform was applied, followed by filtering and an inverse transformation to the time domain. The filters used were optimal predictive filters, selected for each subject. A still further improvement was obtained by taking differences between recordings of two electrodes to obtain bipolar pairs that then were used for the same analysis. Recognition rates, based on a least-squares criterion, varied, but the best were above 90%. The first words of prototypes of sentences also were cut and pasted to test, at least partially, the invariance of a word's brain wave in different sentence contexts. The best result was above 80% correct recognition. Test samples made up only of individual trials also were analyzed. The best result was 134 correct of 288 (47%), which is promising, given that the expected recognition number by chance is just 24 (or 8.3%). The work reported in this paper extends our earlier work on brain-wave recognition of words only. The recognition rates reported here further strengthen the case that recordings of electric brain waves of words or sentences, together with extensive mathematical and statistical analysis, can be the basis of new developments in our understanding of brain processing of language.

Adult↗
PubMed1998

Repin-induced neurotoxicity in rodents.

Russian knapweed is a perennial weed found in many parts of the world, including southern California. Chronic ingestion of this plant by horses has been reported to cause equine nigropallidal encephalomalacia (ENE), which is associated with a movement disorder simulating Parkinson's disease (PD). Repin, a principal ingredient purified from Russian knapweed, is a sesquiterpene lactone containing an alpha-methylenebutyrolactone moiety and epoxides and is a highly reactive electrophile that can readily undergo conjugation with various biological nucleophiles, such as proteins, DNA, and glutathione (GSH). We show in this study that repin is highly toxic to C57BL/6J mice and Sprague-Dawley rats and acutely induces uncoordinated locomotion associated with postural tremors, hypothermia, and inability to respond to sonic and tactile stimuli. We also show that repin intoxication reduces striatal and hippocampal GSH and increases total striatal dopamine (DA) levels in mice. Striatal microdialysis in rats, however, has demonstrated a significant reduction of extracellular DA levels. These findings, coupled with the absence of any demonstrable change in striatal DOPAC levels, suggest that repin acts by inhibiting DA release, a hypothesis that is further supported by our demonstration that, in cultured PC12 cells, repin inhibits the release of DA without affecting its uptake. We believe, therefore, that inhibition of DA release represents one of the earliest pathogenetic events in ENE, leading eventually to striatal extracellular DA denervation, oxidative stress, and degeneration of nigrostriatal pathways. Since the neurotoxic effects of repin appear to be mediated via oxidative stress, and since repin is a natural product isolated from a plant in our environment that can cause a movement disorder associated with degeneration of nigrostriatal pathways, clarification of the mechanism of repin neurotoxicity may provide new insights into our understanding of the pathogenesis of PD.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1998

Estimation of target hazard quotients and potential health risks for metals by consumption of seafood in Taiwan.

The purpose of this paper is to describe the impact of metal pollution on the main seafood and assess the potential health risk from consuming the contaminated seafood in Taiwan. The results of geometric mean (GM) metal concentrations in various seafood showed that the copper, zinc, and arsenic concentrations in oysters were significantly (p < 0.001) higher than those in the other seafood by about 1,057, 74.3, and 56.2 times, respectively. The green color found in the oysters was due to high GM copper and zinc concentrations of 909 (ranging from 113-2,805) and 1,293 (ranging from 303-3,593) microg/g dry wt, respectively. In addition, using a maximum consumption rate of 139 g/day of oysters for individuals, calculations yield target hazard quotients (daily intake/reference dose) of below 1 for cadmium and mercury and high values of 1.61, 9. 33, and 1.77 for inorganic arsenic, copper, and zinc in adults, respectively. The various lifetime cancer risks for inorganic arsenic (maximum exposed individuals risk ranging from 9.93 x 10(-6) to 3.11 x 10(-4)) might be caused by consuming different seafood in Taiwan. The highest risk estimate for inorganic arsenic was 5.10 x 10(-4) for consumption of oysters by Machu Islands residents. The long-term exposure of metals through consumption of oysters, especially for some high-risk groups, could be dangerous. Taking inorganic arsenic for example, a 10(-6) upper limit on lifetime risk as the health protection standard would require maximum oyster residue levels of approximately 0.0076-0.056 microg/g wet wt, for consumption rates of 139-18.6 g/d. In the light of known risks to public health, the government should issue an immediate warning to the public to refrain from eating all seafood harvested from the Taiwan coastal areas, especially the Hsiangshan area and the Machu Islands.

Environmental Exposure↗
PubMed1998

Detection and quantitation of a sodium-dependent sulfate cotransporter (NaSi-1) by sandwich-type enzyme-linked immunosorbent assay.

The sodium-dependent sulfate transporter (NaSi-1) DNA has been recently identified from rat kidney cortex. The objective of this study was to develop a quantitative assay for the NaSi-1 transporter protein. The NaSi-1 antigen was prepared by fusion protein techniques following analysis of the primary sequence for antigenicity. Polyclonal and monoclonal antibodies against the NaSi-1 antigen were raised in rabbits and mice, respectively. The specificity of the raised antibodies was examined by Western analysis using brush-border membrane (BBM) and basolateral membrane (BLM) purified from rat kidney cortex. Both NaSi-1 polyclonal and monoclonal antibodies detected a 69-kDa protein in the BBM. Using the purified monoclonal antibody as the capture antibody and the polyclonal antibody as the detecting antibody, a simple and sensitive sandwich-type enzyme-linked immunosorbent assay was developed to quantitate NaSi-1 transporter protein levels in tissue. The specificity of the assay was examined using BBM, BLM and NaSi-1-transfected Madin-Darby canine kidney cells. The assay was capable of detecting NaSi-1 at levels as low as 6.58 fmol. The concentration of NaSi-1 transporter protein in crude membrane isolated from rat kidney cortex was 0.094+/-0.014 fmol/ microg protein (mean+/-SD of three preparations).

Animals↗
PubMed1998

The use of a chimera HIV-1/HIV-2 envelope protein for immunodiagnosis of HIV infection: its expression and purification in E. coli by use of a translation initiation site within HIV-1 env gene.

A chimera HIV-1/HIV-2 envelope sequence composed of multiple conserved immunodominant epitopes of HIV-1 envelope protein (HIV-1 IIIB: env482-518 + env548-675) and the HIV-2 gp36 immunodominant epitope (env592-603), was constructed and directly over-expressed in E. coli by using a prokaryotic translation initiation sequence contained within the gene of HIV-1 envelope. The recombinant product was purified and applied in antibody-screening assay. The purified chimera antigen reacted with all the thirty-eight HIV-1 positive serum samples, the two HIV-2 serum samples, and had no cross-reaction with all the eighty-eight normal healthy serum sample. The results indicated that this recombinant chimera HIV-1/HIV-2 envelope protein could be useful for diagnostic purposes of HIV infection.

AIDS Serodiagnosis↗
PubMed1998

Inhibition of cathepsin B does not affect the intracellular activation of trypsinogen by cerulein hyperstimulation in isolated rat pancreatic acinar cells.

Activation of trypsinogen is thought to trigger the autodigestive process in acute pancreatitis. The lysosomal enzyme cathepsin B was suggested to cause the activation of trypsinogen because it is known that cathepsin B is able to activate trypsinogen in special circumstances and that lysosomal and digestive enzymes are colocalized within intracellular vacuoles in the early stage of pancreatitis. As yet this hypothesis has been difficult to prove because activated trypsin is difficult to quantify in pancreatitis by conventional enzymatic measurements. We therefore employed an ELISA for trypsin activating peptide (TAP), which is a small peptide cleaved during the activation of trypsinogen and can be determined reliably. Supraphysiological concentrations of cerulein (1 nM-1 microM) resulted in a marked increase in TAP in freshly isolated pancreatic acinar cells, indicating activation of trypsinogen. This activation as determined by the TAP increase was significantly reduced by the serine protease inhibitor Fut-175 but not by the cathepsin B inhibitors E-64 and NCO-700. The concentrations of NCO-700 and E-64 abolished the cathepsin B activity of pancreatic acinar cells but did not significantly reduce the trypsin activity (after enterokinase preincubation); correspondingly the concentrations of Fut-175 used abolished the trypsin activity but did not reduce the cathepsin B activity. The results indicate that an autoactivation of trypsin rather than an activation of trypsinogen by cathepsin B triggers trypsin activation by supramaximal cerulein concentrations.

Animals↗