Biomedical subjects

H Lin

Publications and source records attributed to H Lin.

At least 271 records · Page 15Linked to original sources

PubMed1998

Synthesis and biological activity of binuclear platinum complexes containing two monofunctional cis-[Pt(NH3)2Cl]+ units bridged by 4,4'-dipyridyl selenides or sulfides.

The synthesis of four binuclear platinum complexes of general formula ¿cis-[Pt(NH3)2Cl]2(L)¿ (NO3)2 [L is 4,4'-dipyridyl sulfide (compound I), 4,4'-dipyridyl selenide (compound II), bis(3-methyl-4-pyridyl) sulfide (compound III) or bis(3-methyl-4-pyridyl) selenide (compound IV)] has been achieved. These compounds have been characterized by elemental analysis, IR, 1H-NMR, 13C-NMR and 195Pt-NMR spectroscopy. The above dinuclear platinum complexes have been assayed for antitumor activity in vitro against the cisplatin-sensitive L1210 (the mice leukemia) and cisplatin-insensitive HCT8 (the human coloadenocarcinoma) cell lines. The compounds show IC50 values comparable to or higher than cisplatin against L1210 cell line; however, they have lower IC50 values than cisplatin against the cisplatin-insensitive HCT8 cell line. The complexes containing S exhibit a cytotoxicity against the two cancer cell lines superior to the Se analogues. DNA binding studies indicate the compound I possibly interacts with DNA nonintercalatively. The mode of DNA binding of the dinuclear Pt(II) complexes bridged by 4,4'-dipyridyl selenides or sulfides may be different to that of the aliphatic diaminebridged dinuclear Pt(II) complexes reported previously.

Animals↗

PubMed1998

Estrogen receptors and glucocorticoid receptors in human well-differentiated thyroid cancer.

Previously, we and others have found that the prognosis of patients with well differentiated thyroid cancer is related to patient age and gender. In this study, we examined estrogen and glucocorticoid receptors in thyroid cancers and adjacent non-neoplastic thyroid tissues from 31 Chinese patients. Our results indicate that the level of estrogen and glucocorticoid receptors in thyroid cancer tissues was similar to that in non-neoplastic thyroid tissues. Significant higher estrogen and glucocorticoid receptor levels were found in thyroid cancers from men than women. There was no difference in estrogen receptor levels from thyroid cancer in women less than or older than 45. However, the level of glucocorticoid receptor in thyroid cancer from women less than 45 years old was significantly higher than that of women older than 45. Estrogen or glucocorticoid receptor levels did not correlate with tumor invasiveness. The pathobiological role of glucocorticoid receptors in thyroid cancer is not known but may be involved in changes in circadian rhythm of thyroid tumors.

Journal Article↗

PubMed1997

MMAC1/PTEN mutations in primary tumor specimens and tumor cell lines.

A candidate tumor suppressor gene, MMAC1/PTEN, located in human chromosome band 10q23, was recently identified based on sequence alterations observed in several glioma, breast, prostate, and kidney tumor specimens or cell lines. To further investigate the mutational profile of this gene in human cancers, we examined a large set of human tumor specimens and cancer cell lines of many types for 10q23 allelic losses and MMAC1 sequence alterations. Loss of heterozygosity (LOH) at the MMAC1 locus was observed in approximately one-half of the samples examined, consistent with the high frequency of 10q allelic loss reported for many cancers. Of 124 tumor specimens exhibiting LOH that have been screened for MMAC1 alterations to date, we have detected variants in 13 (approximately 10%) of these primary tumors; the highest frequency of variants was found in glioblastoma specimens (approximately 23%). Novel alterations identified in this gene include a missense variant in a melanoma sample and a splicing variant and a nonsense mutation in pediatric glioblastomas. Of 76 tumor cell lines prescreened for probable LOH, microsequence alterations of MMAC1 were detected in 12 (approximately 16%) of the lines, including those derived from astrocytoma, leukemia, and melanoma tumors, as well as bladder, breast, lung, prostate, submaxillary gland, and testis carcinomas. In addition, in this set of tumor cell lines, we detected 11 (approximately 14%) homozygous deletions that eliminated coding portions of MMAC1, a class of abnormality not detected by our methods in primary tumors. These data support the occurrence of inactivating MMAC1 alterations in multiple human cancer types. In addition, we report the discovery of a putative pseudogene of MMAC1 localized on chromosome 9.

Brain Neoplasms↗

PubMed1997

Growth suppression of glioma cells by PTEN requires a functional phosphatase catalytic domain.

Deletions of all or part of chromosome 10 are the most common genetic alterations in high-grade gliomas. The PTEN gene (also called MMAC1 and TEP1) maps to chromosome region 10q23 and has been implicated as a target of alteration in gliomas and also in other cancers such as those of the breast, prostate, and kidney. Here we sought to provide a functional test of its candidacy as a growth suppressor in glioma cells. We used a combination of Northern blot analysis, protein truncation assays, and sequence analysis to determine the types and frequency of PTEN mutations in glioma cell lines so that we could define appropriate recipients to assess the growth suppressive function of PTEN by gene transfer. Introduction of wild-type PTEN into glioma cells containing endogenous mutant alleles caused growth suppression, but was without effect in cells containing endogenous wild-type PTEN. The ectopic expression of PTEN alleles, which carried mutations found in primary tumors and have been shown or are expected to inactivate its phosphatase activity, caused little growth suppression. These data strongly suggest that PTEN is a protein phosphatase that exhibits functional and specific growth-suppressing activity.

Brain Neoplasms↗

PubMed1997

Electromagnetic field exposure induces rapid, transitory heat shock factor activation in human cells.

Stimulation of human promyelocytic HL60 cells by a 60Hz magnetic field at normal growth temperatures results in heat shock factor 1 activation and heat shock element binding, a sequence of events that mediates the stress-induced transcription of the stress gene HSP70 and increased synthesis of the stress response protein hsp70kD. Thus, the events mediating the electromagnetic field-stimulated stress response appear to be similar to those reported for other physiological stresses (e.g., hyperthermia, heavy metals, oxidative stress) and could well be the general mechanism of interaction of electromagnetic fields with cells.

Binding Sites↗

PubMed1997

Spectrosomes and fusomes anchor mitotic spindles during asymmetric germ cell divisions and facilitate the formation of a polarized microtubule array for oocyte specification in Drosophila.

In the Drosophila ovary, membrane skeletal proteins such as the adducin-like Hts protein(s), spectrin, and ankyrin are found in the spectrosome, an organelle in germline stem cells (GSC) and their differentiated daughter cells (cystoblasts). These proteins are also components of the fusome, a cytoplasmic structure that spans the cystoblast's progeny that develop to form a germline cyst consisting of 15 nurse cells and an oocyte. Spectrosomes and fusomes are associated with one pole of spindles during mitosis and are implicated in cyst formation and oocyte differentiation. Here we show that the asymmetric behavior of the spectrosome persists throughout the cell cycle of GSC. Eliminating the spectrosome by the htsl mutation leads to randomized spindle orientation, suggesting that the spectrosome anchors the spindle to ensure the asymmetry of GSC division; eliminating the fusome in developing cysts results in defective spindles and randomized spindle orientation as well as asynchronous and reduced cystocyte divisions. These observations suggest that fusomes are required for the proper formation and asymmetric orientation of mitotic spindles. Moreover, they reinforce the notion that fusomes are required for the four synchronous divisions of the cystoblast leading to cyst formation. In htsl cysts which lack fusomes and fail to incorporate a hts gene product(s) into ring canals following cyst formation, polarized microtubule networks do not form, the dynamics of cytoplasmic dynein is disrupted, and oskar and orb RNAs fail to be transported to the future oocyte. These observations support the proposed role of fusomes and ring canals in organizing a polarized microtubule-based transport system for RNA localization that leads to oocyte differentiation.

Animals↗

PubMed1997

Cryogenic frozen solution matrixes for analysis of DNA by time-of-flight mass spectrometry.

We present an alternative matrix system for mass analysis of high molecular weight biomolecules by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). Single-stranded DNA oligomers are embedded in thin films of frozen solutions containing photoabsorbing substituted phenols. Ultraviolet laser desorption from these cryogenically cooled matrixes generates internally colder ions than from the equivalent room-temperature crystalline matrixes containing no solvent, as evidenced by a substantial reduction in the extent of fragmentation in the corresponding mass spectra of oligonucleotides. Furthermore, the homogeneity of the frozen solution samples allows analyte signals to be obtained uniformly throughout the sample. Sensitivity and mass resolution approaching that obtained using the most effective room-temperature crystalline matrix for DNA (3-hydroxypicolinic acid) are consistently and reproducibly obtained in both positive and negative ion mass spectra. As fragmentation is a principle factor limiting the maximum attainable mass resolution in MALDI-TOF mass analysis of DNA, this improvement in generating intact DNA ions is advantageous.

DNA↗

PubMed1997

Transcriptional targeting of replication-defective adenovirus transgene expression to smooth muscle cells in vivo.

Gene transfer using replication-defective adenoviruses (RDAd) holds promise for the treatment of vascular proliferative disorders, but is potentially limited by the capacity of these viruses to infect multiple cell lineages. We have generated an RDAd vector, designated AdSM22-lacZ, which encodes the bacterial lacZ reporter gene under the transcriptional control of the smooth muscle cell (SMC)-specific SM22alpha promoter. Here, we show that in vitro AdSM22-lacZ programs expression of the lacZ reporter gene in primary rat aortic SMCs and immortalized A7r5 SMCs, but not in primary human umbilical vein endothelial cells (HUVECs) or NIH 3T3 cells. Consistent with these results, after intraarterial administration of AdSM22-lacZ to control and balloon-injured rat carotid arteries, beta-galactosidase activity was detected within SMCs of the tunica media and neointima, but not within endothelial or adventitial cells. Moreover, intravenous administration of AdSM22-lacZ did not result in lacZ gene expression in the liver or lungs. Finally, we have shown that direct injection of AdSM22-lacZ into SMC-containing tissues such as the ureter and bladder results in high-level transgene expression in visceral SMCs. Taken together, these results demonstrate that transgene expression after infection with an RDAd vector can be regulated in an SMC lineage-restricted fashion by using a transcriptional cassette containing the SMC-specific SM22alpha promoter. The demonstration of an efficient gene delivery system targeted specifically to SMCs provides a novel means to restrict expression of recombinant gene products to vascular or visceral SMCs in vivo.

Adenoviridae↗

PubMed1997

Influence of the positions of cis double bonds in the sn-2-acyl chain of phosphatidylethanolamine on the bilayer's melting behavior.

In an attempt to examine the effects of different numbers and positions of cis double bonds in the sn-2-acyl chain of phosphatidylethanolamine (PE) on the bilayer's melting behavior, 21 molecular species of PE were first semisynthesized, and their Tm and DeltaH values were subsequently determined by high resolution differential scanning calorimetry. In the plot of Tm versus the number of the cis double bond, some characteristic profiles were observed for the various series of PEs. For instance, if the cis double bond was first introduced into the sn-2-acyl chain of C(20):C(20)PE at the Delta5-position, the Tm was observed to reduce drastically. Subsequent stepwise additions of up to five cis double bonds at the methylene-interrupted positions toward the methyl end resulted in a progressive yet smaller decrease in Tm. If, on the other hand, the cis double bonds were introduced sequentially at the Delta11-, Delta11,14-, and Delta11,14,17-positions along the sn-2-acyl chain of C(20):C(20)PE, the Tm profile in the Tm versus the number of the cis double bond showed a down-and-up trend. Most interestingly, for positional isomers of C(20):C(20:3Delta5,8,11)PE, C(20):C(20:3Delta8,11,14)PE, and C(20):C(20:3Delta11,14,17)PE, an inverted bell-shaped Tm profile was detected in the plot of Tm against the position of the omega-carbon for these isomers. Similar Tm profiles were also observed for C(18):C(20)PE, C(20):C(18)PE, and their unsaturated derivatives. This work thus demonstrated that both the positions and the numbers of cis double bonds in the sn-2 acyl chain could exert noticeable influence on the gel-to-liquid crystalline phase transition behavior of the lipid bilayer. Finally, a molecular model was presented, with which the behavior of the gel-to-liquid crystalline phase transition observed for lipid bilayers composed of various sn-1-saturated/sn-2-unsaturated lipids can be rationalized.

Acylation↗

PubMed1997

Activated alleles of yeast SLN1 increase Mcm1-dependent reporter gene expression and diminish signaling through the Hog1 osmosensing pathway.

Two-component signal transduction systems involving histidine autophosphorylation and phosphotransfer to an aspartate residue on a receiver molecule have only recently been discovered in eukaryotes, although they are well studied in prokaryotes. The Sln1 protein of Saccharomyces cerevisiae is a two-component regulator involved in osmotolerance. Phosphorylation of Sln1p leads to inhibition of the Hog1 mitogen-activated protein kinase osmosensing pathway. We have discovered a second function of Sln1p by identifying recessive activated alleles (designated nrp2) that regulate the essential transcription factor Mcm1. nrp2 alleles cause a 5-fold increase in the activity of an Mcm1-dependent reporter, whereas deletion of SLN1 causes a 10-fold decrease in reporter activity and a corresponding decrease in expression of Mcm1-dependent genes. In addition to activating Mcm1p, nrp2 mutants exhibit reduced phosphorylation of Hog1p and increased osmosensitivity suggesting that nrp2 mutations shift the Sln1p equilibrium toward the phosphorylated state. Two nrp2 mutations map to conserved residues in the receiver domain (P1148S and P1196L) and correspond to residues implicated in bacterial receivers to control receiver phosphorylation state. Thus, it appears that increased Sln1p phosphorylation both stimulates Mcm1p activity and diminishes signaling through the Hog1 osmosensing pathway.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1997

Long-lifetime metal-ligand pH probe.

We describe the synthesis and fluorescence spectral characterization of a pH-sensitive metal-ligand complex, [Ru(deabpy)(bpy)2]2., where deabpy is 4,4'-diethylaminomethyl-2,2'-bipyridine. This metal-ligand complex (MLC) was found to display pH-dependent intensities, emission spectra, and decay times, with the changes centered near the physiological useful pH value of 7.5. The apparent pKa values were not found to be dependent on ionic strength. The compound was found to be useful for lifetime-based sensing by phase-modulation fluorometry. Global analysis of the intensity decays over a range of pH values revealed two decay times of 235 and 380 ns, associated with the protonated and unprotonated forms, respectively. Because of its long decay time, optical pH measurements could be accomplished by phase-modulation fluorometry with a conveniently low modulation frequency of 700 kHz. The lifetime data were obtained with either a amplitude-modulated laser or with an amplitude-modulated blue-light-emitting diode. This pH-sensitive complex also displays a modest spectral shift with change in pH, allowing its use as a wavelength-ratiometric MLC probe. One can imagine lifetime sensors for a variety of blood cations and point-of-care assays based on long-lifetime metal-ligand complexes and simple solid-state light sources and detectors.

2,2'-Dipyridyl↗

PubMed1997

Formation of the male pronuclear lamina in Drosophila melanogaster.

Upon fertilization, a sperm nucleus reorganizes to become a male pronucleus. This reorganization includes breakdown and reformation of the nuclear envelope of the male pronucleus. In this study, we used a maternally encoded nuclear lamina protein, YA, in parallel with another lamina protein, lamin Dm, as probes to study the formation of the male pronuclear lamina in Drosophila melanogaster. Ectopically expressed YA is present in the nuclear envelopes of spermatocytes, but not in mature sperm, similar to endogenous lamin Dm. This suggests that the nuclear envelope of Drosophila sperm differs from that of somatic cells. Upon fertilization, YA and lamin Dm are recruited to the periphery of the male-derived nucleus before or during the early stages of migration by the male pronucleus. Using a paternal effect mutation, snky, we found that recruitment of lamina proteins to the male pronucleus requires, and probably accompanies, reorganization of the sperm nucleus. In order to identify factors that affect the recruitment of nuclear lamina proteins to the male pronucleus, we examined the subcellular localization of YA and lamin Dm in mutant embryos defective for the function of either the male pronucleus (mh, K81, and pal or both pronuclei (gnu, png, and plu). None of these mutations affect the recruitment of YA or lamin Dm to the male pronuclear envelope, suggesting that the mutations affect processes independent of, or after, reorganization of the nuclear envelope. Double mutant analyses between Ya and gnu suggest that YA plays a role in the nuclear envelope permissive for rounds of DNA replication.

Animals↗

PubMed1997

Downregulation of RAR alpha in mice by antisense transgene leads to a compensatory increase in RAR beta and RAR gamma and development of lymphoma.

Retinoic acid receptors (RARs) alpha, beta, and gamma contain retinoic acid response elements (RAREs) in their promoter regions and respond to their own activation, thus forming an autoregulatory loop. We generated transgenic mice that expressed an antisense construct of the RAR alpha. Homozygous transgenic mice demonstrated 30% to 80% reduction in RAR alpha protein expression in various tissues. Unlike RAR alpha null mice generated by knockout, our antisense mice demonstrated significant compensatory increases in the expression of RAR beta and RAR gamma proteins. Coarse fur, male sterility, and low body weight were other abnormalities observed in these mice. Most importantly, lymphoma developed in 44% of our homozygous transgenic mice at an early stage of life. These data suggest that RAR alpha is necessary for appropriate response of the RAR beta and RAR gamma genes to physiologic changes and deregulation of the RAR alpha in transgenic mice, which resulted in upregulation of RAR beta and RAR gamma, can be associated with lymphomagenesis. Thus, the data support the hypothesis that a balance among the RARs is necessary for appropriate response to various homeostatic needs.

Animals↗

PubMed1997

Purification and characterization of the small subunit of phage T4 terminase, gp16, required for DNA packaging.

Phage T4 terminase is an enzyme that binds to the portal protein of proheads and cuts and packages concatemeric DNA. The T4 terminase is composed of two subunits, gene products (gp) 16 and 17. The role of the small subunit, gp16, in T4 DNA packaging is not well characterized. We developed a new purification procedure to obtain large quantities of purified gp16 from an overexpression vector. The pure protein is found in two molecular weight forms, due to specific C-terminal truncation, displays in vitro packaging activity, and binds but does not hydrolyze ATP. gp16 forms specific oligomers, rings, and side-by-side double rings, as judged by native polyacrylamide gel electrophoresis and scanning transmission electron microscopy measurements. The single ring contains about eight monomers, and the rings have a diameter of about 8 nm with a central hole of about 2 nm. A DNA-binding helix-turn-helix motif close to the N terminus of gp16 is predicted. The oligomers do not bind to DNA, but following denaturation and renaturation in the presence of DNA, binding can be demonstrated by gel shift and filter binding assays. gp16 binds to double-stranded DNA but not single-stranded DNA, and appears to bind preferentially to a gene 16-containing DNA sequence.

Adenosine Triphosphatases↗

PubMed1997

Simultaneous determination of purine bases, ribonucleosides and ribonucleotides by capillary electrophoresis-electrochemistry with a copper electrode.

Simultaneous determination of purine bases, ribonucleosides and ribonucleotides was achieved by coupling capillary electrophoresis (CE) with wall-jet amperometric detection. A 200 microm diameter copper disk electrode was applied at working potential, +0.65 V vs. saturated calomel electrode. The current response of high sensitivity and stability was obtained in strong basic solutions which were suitable for satisfactory CE separations. The calibration curve was linear over 2-3 orders of magnitude and the limits of detection for adenine, guanine, xanthine, uric acid, adenosine, guanosine, adenosine-5'- monophosphate and guanosine-5'-monophosphate were below 9 fmol (SIN=3). The use of this method for the separation and detection of compounds present in human plasma samples was reported.

Calibration↗

PubMed1997

The enhanced tumorigenic activity of a mutant epidermal growth factor receptor common in human cancers is mediated by threshold levels of constitutive tyrosine phosphorylation and unattenuated signaling.

Deregulation of signaling by the epidermal growth factor receptor (EGFR) is common in human malignancy progression. One mutant EGFR (variously named DeltaEGFR, de2-7 EGFR, or EGFRvIII), which occurs frequently in human cancers, lacks a portion of the extracellular ligand-binding domain due to genomic deletions that eliminate exons 2 to 7 and confers a dramatic enhancement of brain tumor cell tumorigenicity in vivo. In order to dissect the molecular mechanisms of this activity, we analyzed location, autophosphorylation, and attenuation of the mutant receptors. The mutant receptors were expressed on the cell surface and constitutively autophosphorylated at a significantly decreased level compared with wild-type EGFR activated by ligand treatment. Unlike wild-type EGFR, the constitutively active DeltaEGFR were not down-regulated, suggesting that the altered conformation of the mutant did not result in exposure of receptor sequence motifs required for endocytosis and lysosomal sorting. Mutational analysis showed that the enhanced tumorigenicity was dependent on intrinsic tyrosine kinase activity and was mediated through the carboxyl terminus. In contrast with wild-type receptor, mutation of any major tyrosine autophosphorylation site abolished these activities suggesting that the biological functions of DeltaEGFR are due to low constitutive activation with mitogenic effects amplified by failure to attenuate signaling by receptor down-regulation.

Amino Acid Sequence↗

PubMed1997

Deletion mapping of chromosome 4 in head and neck squamous cell carcinoma.

Genomic deletions involving chromosome 4 have recently been implicated in several human cancers. To identify and characterize genetic events associated with the development of head and neck squamous cell carcinoma (HNSCC), a fine mapping of allelic losses associated with chromosome 4 was performed on DNA isolated from 27 matched primary tumor specimens and normal tissues. Loss of heterozygosity (LOH) of at least one chromosome 4 polymorphic allele was seen in the majority of tumors (92%). Allelic deletions were confined to short arm loci in four tumors and to the long arm loci in 12 tumors, suggesting the presence of two regions of common deletion. One region of frequent deletion was centered at D4S405 on 4p and included the loci D4S1546 to D4S428 in approximately 41% of the tumors. The common region of deletion on 4q was more complex and extended from D4S1571 to D4S1573. Frequent genetic alterations were observed within this region (4q25) and one marker, D4S407, exhibited a high frequency of LOH (>75%). These results indicate that alterations of chromosome 4 regions are associated with HNSCC tumorigenesis and further localizes the regions that may harbor tumor suppressor genes.

Alleles↗

PubMed1997

Electron microscopic localization of lacZ expression in the proximal convoluted tubular cells of the kidney in transgenic mice carrying chimeric erythropoietin/lacZ gene constructs.

Regulated expression of the erythropoietin (EPO) gene in the adult kidney plays a key role in the regulation of erythropoiesis. However, uncertainty exists regarding the type of kidney cells involved in EPO gene expression. We previously showed by light microscopy that the lacZ reporter gene is expressed and inducible by hypoxia/anemia in the proximal convoluted tubular (PCT) cells of the kidneys of transgenic mice carrying the 5'-lacZ construct, in which the lacZ gene was placed downstream of a 7.0-kb mouse EPO gene segment containing 6.5 kb of the 5'-flanking sequence. We, report here the light and transmission electron microscopic examination of lacZ expression in the kidneys of transgenic mice carrying the 5'-lacZ construct and two additional constructs carrying the 6.5-kb 5'-flanking sequence with the body of the gene alone, or along with the 1.2-kb 3'-flanking sequence. The electron microscopic analyses unequivocally demonstrated that lacZ under the regulatory control of the 6.5-kb 5'-flanking sequence with or without the body of the gene and the 1.2-kb 3'-flanking sequence was expressed predominantly in the proximal convoluted tubular cells of the kidney following hypoxia induction.

Anemia↗