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Biomedical subjects

J Han

Publications and source records attributed to J Han.

At least 271 records · Page 15Linked to original sources
PubMed1997

Applications of pressure differential scanning calorimetry in the study of pharmaceutical hydrates. I. Carbamazepine dihydrate.

The dehydration of carbamazepine dihydrate (C(15)H(12)N(2)O.2H(2)O) was studied by both conventional differential scanning calorimetry (DSC) and by pressure differential scanning calorimetry. Variable temperature powder X-ray diffractometry (VTXRD) and thermogravimetric analysis were used as complementary techniques. By performing DSC at elevated pressures, the dehydration and vaporization endotherms were separated and it was possible to determine the enthalpy of dehydration. Over the range of 100-600 psi, the enthalpy of dehydration was unaffected by pressure. However, the solid-state of the anhydrous phase formed was influenced by the DSC conditions. At ambient pressure, dehydration resulted in the formation of the gamma-form of anhydrous carbamazepine while at elevated pressures, the anhydrous beta-form appeared which converted to the gamma-form at higher temperatures. At high pressures, the water liberated on dehydration is not immediately removed and its presence appears to be responsible for the formation of beta-carbamazepine. VTXRD permitted in situ study of dehydration and this confirmed the DSC results. A specially fabricated sample holder permitted VTXRD studies at elevated pressures.

Journal Article↗
PubMed1997

Characterization of the structure and function of the fourth member of p38 group mitogen-activated protein kinases, p38delta.

We have cloned and characterized a new member of the p38 group of mitogen-activated protein kinases here termed p38delta. Sequence comparisons revealed that p38delta is approximately 60% identical to the other three p38 isoforms but only 40-45% to the other mitogen-activated protein kinase family members. It contains the TGY dual phosphorylation site present in all p38 group members and is activated by a group of extracellular stimuli including cytokines and environmental stresses that also activate the other three known p38 isoforms. However, unlike the other p38 isoforms, the kinase activity of p38delta is not blocked by the pyridinyl imidazole, 4-(4-fluorophenyl)-2-2(4-hydroxyphenyl)-5-(4-pyridyl)-imidazole (identicalto SB202190). p38delta can be activated by MKK3 and MKK6, known activators of the other isoforms. Nonetheless, in-gel kinase assays provide evidence for additional activators. The data presented herein show that p38delta has many properties that are similar to those of other p38 group members. Nonetheless important differences exist among the four members of the p38 group of enzymes, and thus each may have highly specific, individual contributions to biologic events involving activation of the p38 pathways.

Amino Acid Sequence↗
PubMed1997

Evidence for autocrine inhibition of gonadotropin-releasing hormone (GnRH) gene transcription by GnRH in hypothalamic GT1-1 neuronal cells.

To examine whether an ultrashort feedback mechanism of gonadotropin-releasing hormone (GnRH) operates at the level of gene transcription, we studied the effects of GnRH analogs on GnRH promoter activity and GnRH mRNA level in hypothalamic GT1-1 neuronal cells. Treatment of GT1-1 cells with buserelin, a GnRH agonist, or native GnRH for 24 h significantly decreased GnRH promoter activity and its mRNA level, whereas that with GnRH antagonists, antide or [D-Phe2,D-Ala6]-GnRH, showed no effect. The inhibitory effects of buserelin on GnRH gene transcription and GnRH mRNA level were dose-related, and a significant inhibition was observed in cells treated with buserelin at concentrations higher than 0.1 microM. Time-course experiments showed that significant decreases in GnRH promoter-driven luciferase activity and GnRH mRNA level were observed within 12 h and sustained up to 48 h. Moreover, treatment with GnRH agonist for 12 h significantly decreased the transcription rate of the mouse GnRH gene, as revealed by nuclear run-on transcription assay. The promoter analysis with the 5'-deletional constructs demonstrated that cis-acting elements important for GnRH autoregulation by GnRH agonist reside within -854 bp upstream from the transcription start site. These data clearly demonstrate that GnRH can exert autocrine regulation at the level of GnRH gene transcription.

Animals↗
PubMed1997

Diversity among clinical isolates of Helicobacter pylori in Korea.

Eleven strains of Helicobacter pylori were isolated from biopsy specimens obtained at the A-San Medical Center from December, 1995 to February, 1996. Every H. pylori positive patient was diagnosed to have chronic, erosive, or mild superficial gastritis. To determine the diversity in clinical isolates, the following were studied: total protein profile, plasmid profile, presence of cagA, and variation in DNA sequence. Protein profiles of nine isolates were similar to each other while two isolates had a 35 kDa protein which did not appear in others. The presence of cagA was detected with PCR in seven isolates (63%). Among eleven isolates, seven (63%) carried plasmids. Each isolate showed a big diversity with a PCR-based randomly amplified polymorphic DNA method. Even though all H. pylori isolates used in this study were isolated from gastritis patients at the same hospital, their molecular and biological characteristics were quite different from another showing a big diversity in H. pylori.

Antigens, Bacterial↗
PubMed1997

Native and modified low density lipoproteins increase the functional expression of the macrophage class B scavenger receptor, CD36.

The uptake of oxidized low density lipoprotein (OxLDL) by macrophages is a key event implicated in the initiation and development of atherosclerotic lesions. Two macrophage surface receptors, CD36 (a class B scavenger receptor) and the macrophage scavenger receptor (a class A scavenger receptor), have been identified as the major receptors that bind and internalize OxLDL. Expression of CD36 in monocyte/macrophages in tissue culture is dependent both on the differentiation state as well as exposure to soluble mediators (cytokines and growth factors). The regulatory mechanisms of this receptor in vivo are undetermined as is the role of lipoproteins themselves in modulating CD36 expression. We studied the effect of lipoproteins, native LDL and modified LDL (acetylated LDL (AcLDL) and OxLDL) on the expression of CD36 in J774 cells, a murine macrophage cell line. Exposure to lipoproteins resulted in a marked induction of CD36 mRNA expression (4-8-fold). Time course studies showed that maximum induction was observed 2 h after treatment with AcLDL and at 4 h with LDL and OxLDL. Increased expression of CD36 mRNA persisted for 24 h with each treatment group. Induction of CD36 mRNA expression was paralleled by an increase in CD36 protein as determined by Western blot with the greatest induction by OxLDL (4-fold). In the presence of actinomycin D, treatment of macrophages with LDL, AcLDL, or OxLDL did not affect CD36 mRNA stability, implying that CD36 mRNA was transcriptionally regulated by lipoproteins. To determine the mechanism(s) by which lipoproteins increased expression of CD36 we evaluated the effects of lipoprotein components on CD36 mRNA expression. ApoB 100 increased CD36 mRNA expression significantly, whereas phospholipid/cholesterol liposomes had less effect. Incubation of macrophages with bovine serum albumin or HDL reduced expression of CD36 mRNA in a dose-dependent manner. Finally, to evaluate the in vivo relevance of the induction of CD36 mRNA expression by lipoproteins, peritoneal macrophages were isolated from mice following intraperitoneal injection of lipoproteins. Macrophage expression of CD36 mRNA was significantly increased by LDL, AcLDL, or OxLDL in relation to mice infused with phosphate-buffered saline, with OxLDL causing the greatest induction (8-fold). This is the first demonstration that exposure to free and esterified lipids augments functional expression of the class B scavenger receptor, CD36. These data imply that lipoproteins can further contribute to foam cell development in atherosclerosis by up-regulating a major OxLDL receptor.

Animals↗
PubMed1997

A cell binding domain from the alpha3 chain of type IV collagen inhibits proliferation of melanoma cells.

Our previous studies have shown that a peptide corresponding to the residue sequence 185-203 of the NC1 domain of the alpha3 chain of basement membrane collagen (type IV) inhibits the activation of polymorphonuclear leukocytes. Peptides from the same region of the alpha1, alpha2, alpha4, and alpha5(IV) chains did not exhibit this property. Because of the intimate relationship between metastasizing neoplastic cells and vascular as well as epithelial basement membranes, we measured the cell adhesion-promoting activity of peptides from the NC1 domain of type IV collagen and their effect on proliferation of human melanoma cells. We found that peptide alpha3(IV)185-203 (CNYYSNSYSFWLASLNPER) not only promotes adhesion of human melanoma cells but also inhibits their proliferation. Adhesion increased by 50-60% over control. Melanoma cell proliferation was inhibited by 40% when cells were grown in a medium containing 5 microg/ml peptide for 5 days. Studies showed that replacement of serine in position 189 or 191 by alanine resulted in significantly reduced adhesion. Similarly, serine replacement resulted in reduced ability to inhibit proliferation. Our data suggest that a region of the NC1 domain of the alpha3(IV) chain, contained within the sequence 185-203, not only specifically promotes adhesion but also inhibits proliferation of melanoma cells. These properties appear to be dependent on the presence of the triplet sequence -SNS- (residues 189-191), which is unique to the alpha3 chain and may represent an important functional epitope.

Amino Acid Sequence↗
PubMed1997

Structure-function studies of p38 mitogen-activated protein kinase. Loop 12 influences substrate specificity and autophosphorylation, but not upstream kinase selection.

Several mitogen-activated protein kinase (MAPK) cascades have been identified in eukaryotic cells. The activation of MAPKs is carried out by distinct MAPK kinases (MEKs or MKKs), and individual MAPKs have different substrate preferences. Here we have examined how amino acid sequences encompassing the dual phosphorylation motif located in the loop 12 linker (L12) between kinase subdomains VII and VIII and the length and amino acid sequence of L12 influence autophosphorylation, substrate specificity, and upstream kinase selectivity for the MAPK p38. Conversion of L12 of p38 to an "ERK-like" structure was accomplished in several ways: (i) by replacing glycine with glutamate in the dual phosphorylation site, (ii) by placing a six-amino acid sequence present in L12 of ERK (but absent in p38) into p38, and (iii) by mutations of amino acid residues in loop 12. Two predominant effects were noted: (i) the Xaa residue in the dual phosphorylation motif Thr-Xaa-Tyr as well as the length of L12 influence p38 substrate specificity, and (ii) the length of L12 plays a major role in controlling autophosphorylation. In contrast, these modifications do not result in any change in the selection of p38 by individual MAPK kinases.

Amino Acid Sequence↗
PubMed1997

Characterization of a novel mammalian RGS protein that binds to Galpha proteins and inhibits pheromone signaling in yeast.

Genetic studies of molecules that negatively regulate G-coupled receptor functions have led to the identification of a large gene family with an evolutionarily conserved domain, termed the RGS domain. It is now understood that RGS proteins serve as GTPase-activating proteins for subfamilies of the heterotrimeric G-proteins. We have isolated from mouse pituitary a full-length cDNA clone encoding a novel member of the RGS protein family, termed RGS16, as well as the full-length cDNA of mRGS5 and mRGS2. Tissue distribution analysis shows that the novel RGS16 is predominantly expressed in liver and pituitary, and that RGS5 is preferentially expressed in heart and skeletal muscle. In contrast, RGS2 is widely expressed. Genetic analysis using the pheromone response halo assay and FUS1 gene induction assay show that overexpression of the RGS16 gene dramatically inhibits yeast response to alpha-factor, whereas neither RGS2 nor RGS5 has any discernible effect on pheromone sensitivity, pointing to a possible functional diversity among RGS proteins. In vitro binding assays reveal that RGS5 and RGS16 bind to Galphai and Galphao subunits of heterotrimeric G-proteins, but not to Galphas. Based on mutational analysis of the conserved residues in the RGS domain, we suggest that the G-protein binding and GTPase-activating protein activity may involve distinct functional structures of the RGS proteins, indicating that RGS proteins may exert a dual function in the attenuation of signaling via G-coupled receptors.

Amino Acid Sequence↗
PubMed1997

Activation of the transcription factor MEF2C by the MAP kinase p38 in inflammation.

For cells of the innate immune system to mount a host defence response to infection, they must recognize products of microbial pathogens such as lipopolysaccharide (LPS), the endotoxin secreted by Gram-negative bacteria. These cellular responses require intracellular signalling pathways, such as the four MAP kinase (MAPK) pathways. In mammalian cells the MAPK p38 is thought to play an important role in the regulation of cellular responses during infection through its effects on the expression of proinflammatory molecules. One means of understanding the role of p38 in these responses is to identify proteins with functions regulated by p38-catalysed phosphorylation. Here we demonstrate a link between the p38 pathway and a member of the myocyte-enhancer factor 2 (MEF2) group of transcription factors. We found that in monocytic cells, LPS increases the transactivation activity of MEF2C through p38-catalysed phosphorylation. One consequence of MEF2C activation is increased c-jun gene transcription. Our results show that p38 may influence host defence and inflammation by maintaining the balance of c-Jun protein consumed during infection.

Binding Sites↗
PubMed1997

The structure of mitogen-activated protein kinase p38 at 2.1-A resolution.

The structure of mitogen-activated protein (MAP) kinase p38 has been solved at 2.1-A to an R factor of 21.0%, making p38 the second low activity MAP kinase solved to date. Although p38 is topologically similar to the MAP kinase ERK2, the phosphorylation Lip (a regulatory loop near the active site) adopts a different fold in p38. The peptide substrate binding site and the ATP binding site are also different from those of ERK2. The results explain why MAP kinases are specific for different activating enzymes, substrates, and inhibitors. A model presented for substrate and activator interactions has implications for the evolution of protein kinase cascades.

Amino Acid Sequence↗
PubMed1997

NIK is a new Ste20-related kinase that binds NCK and MEKK1 and activates the SAPK/JNK cascade via a conserved regulatory domain.

Nck, an adaptor protein composed of one SH2 and three SH3 domains, is a common target for a variety of cell surface receptors. We have identified a novel mammalian serine/threonine kinase that interacts with the SH3 domains of Nck, termed Nck Interacting Kinase (NIK). This kinase is most homologous to the Sterile 20 (Ste20) family of protein kinases. Of the members of this family, GCK and MSST1 are most similar to NIK in that they bind neither Cdc42 nor Rac and contain an N-terminal kinase domain with a putative C-terminal regulatory domain. Transient overexpression of NIK specifically activates the stress-activated protein kinase (SAPK) pathway. Both the kinase domain and C-terminal regulatory region of NIK are required for full activation of SAPK. NIK likely functions upstream of MEKK1 to activate this pathway; a dominant-negative MEK kinase 1 (MEKK1) blocks activation of SAPK by NIK. MEKK1 and NIK also associate in cells and this interaction is mediated by regulatory domains on both proteins. Two other members of this kinase family, GCK and HPK1, contain C-terminal regulatory domains with homology to that of NIK. These findings indicate that the C-terminal domain of these proteins encodes a new protein domain family and suggests that this domain couples these kinases to the SAPK pathway, possibly by interacting with MEKK1 or related kinases.

Adaptor Proteins, Signal Transducing↗
PubMed1997

Identification and characterization of a predominant isoform of human MKK3.

We have obtained a novel cDNA species of MKK3, termed MKK3b. MKK3b cDNA differs from its original form, MKK3, at the 5'-end, encoding 29 extra amino acids in the N-terminus. Analysis of MKK3 genomic DNA structure revealed that the MKK3b-unique 5'-end sequence is derived from an exon different from that of MKK3, and that they share identical sequences thereafter. This suggests that the two cDNA forms of MKK3 are either generated by differential splicing of the same gene or derived from differential promoter utilization. Northern blotting analysis showed that MKK3b mRNA is much more abundant than MKK3. Functional characterization based on the activation of p38 revealed that MKK3b is more efficient than MKK3 in mediating downstream signalling events.

Amino Acid Sequence↗
PubMed1997

Apoptosis signaling pathway in T cells is composed of ICE/Ced-3 family proteases and MAP kinase kinase 6b.

Fas/APO-1(CD95) ligation activates programmed cell death, a cellular process that plays an important role in the maturation of the host immune response. We show that activation of a specific MAP kinase kinase (MKK), MKK6b, is necessary and sufficient for Fas-induced apoptosis of Jurkat T cells. MKK6b activation occurs downstream of an interleukin-1 converting enzyme-like (ICE-like) protease(s), while execution of the apoptotic pathway by MKK6b requires both ICE- and CPP32-like proteases. Surprisingly, the p38 MAP kinase protein, a known substrate of MKK6b, does not participate in Fas/MKK6b-mediated apoptosis. These findings indicate a divergence of the MKK6b signaling pathways, one of which activates p38 and leads to regulation of gene expression, and one of which activates the ICE/Ced-3 family of proteases and leads to cell death. These studies represent a demonstration of an apoptotic pathway that is comprised of both the ICE/Ced-3 family of proteases and MAP kinase kinase 6.

Amino Acid Sequence↗
PubMed1997

Inhibitory effects of topical application of low doses of curcumin on 12-O-tetradecanoylphorbol-13-acetate-induced tumor promotion and oxidized DNA bases in mouse epidermis.

The effects of topical applications of very low doses of curcumin (the major yellow pigment in turmeric and the Indian food curry) on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced oxidation of DNA bases in the epidermis and on tumor promotion in mouse skin were investigated. CD-1 mice were treated topically with 200 nmol of 7,12-dimethylbenz[a]anthracene followed one week later by 5 nmol of TPA alone or together with 1, 10, 100 or 3000 nmol of curcumin twice a week for 20 weeks. Curcumin-mediated effects on TPA-induced formation of the oxidized DNA base 5-hydroxymethyl-2'-deoxyuridine (HMdU) and tumor formation were determined. All dose levels of curcumin inhibited the mean values of TPA-induced HMdU formation in epidermal DNA (62-77% inhibition), but only the two highest doses of curcumin strongly inhibited TPA-induced tumor promotion (62-79% inhibition of tumors per mouse and tumor volume per mouse). In a second experiment, topical application of 20 or 100 nmol (but not 10 nmol) of curcumin together with 5 nmol TPA twice a week for 18 weeks markedly inhibited TPA-induced tumor promotion. Curcumin had a strong inhibitory effect on DNA and RNA synthesis (IC50 = 0.5-1 microM) in cultured HeLa cells, but there was little or no effect on protein synthesis.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1997

Vascular and neural anatomy of the thenar area of the hand: its surgical applications.

Thirty fresh cadaver hands were injected with a silicone rubber compound (Microfil) and dissected to examine the vascular and neural supplies of the thenar area. In 10 specimens, a selective injection technique was used to determine the extent of skin territory nourished by the superficial palmar branch of the radial artery. The purpose of this study was to clarify the anatomic basis for clinical applications of vascularized free- or island-flap transfers from the thenar area. The superficial palmar branch of the radial artery was found in all hands; the average diameter of the branch measured at its bifurcation site was 1.4 mm (0.8 to 3.0 mm). The constant area nourished by the superficial palmar branch was an area approximately 4 x 3 cm located over the proximal parts of the abductor pollicis brevis and opponens pollicis muscles. These results indicate that a fasciocutaneous flap from the radial thenar region can be raised based on the superficial palmar branch and consistently transferred as a free flap. In 63 percent of the hands dissected, the superficial palmar branch was connected to other arteries in the palm, suggesting that the flap can be transferred as a reverse-pedicle island flap in such hands. Innervation of the flap was provided chiefly by a branch of the superficial radial nerve. The radial aspect of the thenar eminence can provide a new and useful donor source for an innervated and vascularized free- or island-flap transfer for reconstruction of various skin defects of the volar side of the fingers.

Arteries↗
PubMed1997

Lck regulates Vav activation of members of the Rho family of GTPases.

Vav is a member of a family of oncogene proteins that share an approximately 250-amino-acid motif called a Dbl homology domain. Paradoxically, Dbl itself and other proteins containing a Dbl domain catalyze GTP-GDP exchange for Rho family proteins, whereas Vav has been reported to catalyze GTP-GDP exchange for Ras proteins. We present Saccharomyces cerevisiae genetic data, in vitro biochemical data, and animal cell biological data indicating that Vav is a guanine nucleotide exchange factor for Rho-related proteins, but in similar genetic and biochemical experiments we fail to find evidence that Vav is a guanine nucleotide exchange factor for Ras. Further, we present data indicating that the Lck kinase activates the guanine nucleotide exchange factor and transforming activity of Vav.

Animals↗
PubMed1997

Modulation of ATP-sensitive K+ channels in rabbit ventricular myocytes by adenosine A1 receptor activation.

The objective of the present study was to characterize the role of adenosine in the regulation of ATP-sensitive K (KATP) channel activity in isolated rabbit ventricular myocytes using the patch-clamp technique. In an outside-out patch exposed to guanosine 5'-triphosphate and ATP at the intracellular surface, external adenosine stimulated KATP channel activity. In an inside-out patch exposed to external adenosine, ATP reduced KATP channel activity and guanosine 5'-triphosphate stimulated KATP channel activity. Guanosine 5'-O-(3-thiotriphosphate) resulted in a gradual increase of KATP channel activity even in the absence of adenosine. When myocytes were preincubated with pertussis toxin or 8-cyclopentyl-1,3-dipropylxanthine, adenosine A1 receptor activation failed to activate the KATP channel. Analysis of the open and closed time distributions showed that adenosine A1 receptor activation increased burst duration and decreased interburst duration. In a dose-response relationship for ATP, adenosine A1 receptor activation shifted the half-maximal inhibition of the KATP channel from 70 to 241 microM.

Adenosine↗