Search PubMed⌕ Search

Biomedical subjects

J J Lin

Publications and source records attributed to J J Lin.

At least 181 records · Page 10Linked to original sources
PubMed1988

Rapid purification of mammalian cardiac troponin T and its isoform switching in rat hearts during development.

A rapid purification of troponin T from adult hearts of various species has been developed. The purification procedure included 60 degrees C treatment of the high salt extract, ammonium sulfate fractionation, and DEAE-cellulose column chromatography. The troponin T purified from the bovine left ventricle contained two isoforms, which differed in both apparent molecular mass and isoelectric point. Both isoforms were able to bind to F-actin filaments only in the presence of tropomyosin. Monoclonal antibody JLT12 against rabbit skeletal troponin T cross-reacted with both isoforms of bovine cardiac troponin T. There was no detectable difference in the relative amount of these two isoforms among different portions (atria, right and left ventricles) of the bovine heart. The purified protein was used as an antigen to immunize mice, and a mouse antiserum with high titer and specificity to both isoforms was subsequently obtained. This antiserum also cross-reacted with cardiac troponin T from chicken, rabbit, and rat. The antibodies were further used to probe cardiac development in rats by Western blotting and immunoprecipitation. The results clearly showed that there was a switch of troponin T isoforms between hearts from 20-day-old rat embryos and hearts from 14-day-old rats. Immunoprecipitation of the in vitro translation products of poly(A)+ RNA isolated from day 5 rat hearts revealed the presence of two isoforms of troponin T, suggesting that two mRNAs coding for these two isoforms existed in the heart cells. It is of interest to not that some profound changes in the morphology and function of cardiac muscle have also been detected at this time of development. Troponin T isoform switching thus may well represent an important marker for cardiac development and function.

Actins↗
PubMed1988

A trans-acting gene is required for the phenotypic expression of a tyrosinase gene in Streptomyces.

The melanin locus (melC) from Streptomyces antibioticus was previously shown to be composed of two open reading frames (ORFs), melC1 and melC2. The melC2 ORF codes for the polypeptide chain of tyrosinase (apotyrosinase). The function of melC1 is not known except that insertional mutation within it abolishes the tyrosinase activity. Here, we show that in Streptomyces lividans TK64 harboring melC1 mutated and melC2 intact (melC1- melC2+) plasmids, while there was no tyrosinase activity, melC transcript was synthesized and apotyrosinase could be detected. The apotyrosinase could be activated to a limited degree by incubation with copper ions, or by mixing the mycelial extract from a culture harboring a melC1- melC2+ (pPF950) plasmid with that from a culture containing a melC1+ melC2- (pSA1) plasmid. Complementation analysis showed that melC1 acted in trans on the tyrosinase gene expression. Together, these results suggest that melC1 encodes or regulates a copper-transfer protein serving an in vivo copper-donor function in the biosynthesis of active tyrosinase.

Catechol Oxidase↗
PubMed1988

Theophylline in rats during maintenance phase of post-ischemic acute renal failure.

We have shown previously that theophylline increases both renal plasma flow (RPF) and glomerular filtration rate (GFR) during the initiation phase of post-ischemic acute renal failure (ARF) in rats. The purpose of the present experiments was to determine the effects of theophylline during the maintenance phase of ARF, five days after initiation. Clearance techniques were used to measure renal function in a control group of pentobarbital anesthetized rats (group C) and in three experimental groups, five days after subjecting the left kidney to a thirty-minute period of complete ischemia. Group SS received saline during both the ischemic episode and the clearance measurements; group ST received saline during ischemia and theophylline, acutely, during the clearance measurements; group TS received theophylline during ischemia and saline during the clearance measurements. In comparison with the values for the control group (group C), RPF and GFR of the post-ischemic left kidneys of group SS were approximately half normal. In groups ST and TS, RPF and GFR of the left kidneys were higher than in group SS. Collectively, these results demonstrate that pretreatment with theophylline during the initiation phase of ischemia-induced ARF leads to increased RPF and GFR during the maintenance phase, and that acute theophylline treatment during the maintenance phase acutely increases RPF and GFR. Since increases in GFR were associated with increases in RPF, and since theophylline is an adenosine receptor antagonist, these results are consistent with the hypothesis that adenosine-mediated hemodynamic changes play a pathogenic role in ischemia-induced ARF in rats.

Acute Kidney Injury↗
PubMed1988

Motility-dependence of the heterogenous staining of culture cells by a monoclonal anti-tropomyosin antibody.

A monoclonal antibody (CG1) which recognizes tropomyosin isoforms 1 and 3 of chicken embryo fibroblasts was used to detect what is a motility-dependent change in the availability of the antigenic determinant in tropomyosin molecules along microfilaments. Immunofluorescence microscopy with this antibody revealed a heterogenous staining pattern among chicken embryo fibroblasts cells such that a population (17%) of cells showed only background staining. Stress fibers in about half the population of the cells stained weakly with this antibody, while the stress fibers in another population of cells (35%) showed very strong staining. After glycerination or cytochalasin B treatment, all of the cells became positive in reaction to CG1 antibody, suggesting that the antigenic determinant was present in every cell. On the other hand, all of the cells after brief nonionic detergent treatment became negative to CG1 antibody. The CG1 staining pattern was not significantly changed in cells at different stages after release from colcemid blockage, nor was a brief treatment of cells with buffer containing 2 M urea, mild trypsin, chymotrypsin, or V.8 protease effective in changing the reactivity. However, most of the cells with a morphology typical of movement, and all of the contracted, glycerinated cells were strongly positive to CG1 antibody. These results suggest that the unmasking of the CG1 determinant may be motility-dependent. Immunoblot analysis showed that forced modification on the cysteine residue of tropomyosin molecules, caused either by performic acid oxidation or by disulfide cross-linking with the chemical 5,5'-dithiobis (2-nitrobenzoate), results in drastic changes in the reactivity of the different isoforms to CG1 antibody. These results indicate that the cysteine residue is involved in the CG1 determinant. The motility-dependent unmasking of this determinant may suggest an important role for nonmuscle tropomyosin in regulating cell motility.

Actin Cytoskeleton↗
PubMed1988

Differential localization of tropomyosin isoforms in cultured nonmuscle cells.

We have previously shown that chicken embryo fibroblast (CEF) cells and human bladder carcinoma (EJ) cells contain multiple isoforms of tropomyosin, identified as a, b, 1, 2, and 3 in CEF cells and 1, 2, 3, 4, and 5 in human EJ cells by one-dimensional SDS-PAGE (Lin, J. J.-C., D. M. Helfman, S. H. Hughes, and C.-S. Chou. 1985. J. Cell Biol. 100: 692-703; and Lin, J. J.-C., S. Yamashiro-Matsumura, and F. Matsumura. 1984. Cancer Cells 1:57-65). Both isoform 3 (TM-3) of CEF and isoforms 4,5 (TM-4,-5) of human EJ cells are the minor isoforms found respectively in normal chicken and human cells. They have a lower apparent molecular mass and show a weaker affinity to actin filaments when compared to the higher molecular mass isoforms. Using individual tropomyosin isoforms immobilized on nitrocellulose papers and sequential absorption of polyclonal antiserum on these papers, we have prepared antibodies specific to CEF TM-3 and to CEF TM-1,-2. In addition, two of our antitropomyosin mAbs, CG beta 6 and CG3, have now been demonstrated by Western blots, immunoprecipitation, and two-dimensional gel analysis to have specificities to human EJ TM-3 and TM-5, respectively. By using these isoform-specific reagents, we are able to compare the intracellular localizations of the lower and higher molecular mass isoforms in both CEF and human EJ cells. We have found that both lower and higher molecular mass isoforms of tropomyosin are localized along stress fibers of cells, as one would expect. However, the lower molecular mass isoforms are also distributed in regions near ruffling membranes. Further evidence for this different localization of different tropomyosin isoforms comes from double-label immunofluorescence microscopy on the same CEF cells with affinity-purified antibody against TM-3, and monoclonal CG beta 6 antibody against TM-a, -b, -1, and -2 of CEF tropomyosin. The presence of the lower molecular mass isoform of tropomyosin in ruffling membranes may indicate a novel way for the nonmuscle cell to control the stability and organization of microfilaments, and to regulate the cell motility.

Animals↗
PubMed1988

Studies on cardiac myofibrillogenesis with antibodies to titin, actin, tropomyosin, and myosin.

Cardiac myofibrillogenesis was examined in cultured chick cardiac cells by immunofluorescence using antibodies against titin, actin, tropomyosin, and myosin. Primitive cardiomyocytes initially contained stress fiber-like structures (SFLS) that stained positively for alpha actin and/or muscle tropomyosin. In some cases the staining for muscle tropomyosin and alpha actin was disproportionate; this suggests that the synthesis and/or assembly of these two isoforms into the SFLS may not be stoichiometric. The alpha actin containing SFLS in these myocytes could be classified as either central or peripheral; central SFLS showed developing sarcomeric titin while peripheral SFLS had weak titin fluorescence and a more uniform stain distribution. Sarcomeric patterns of titin and myosin were present at multiple sites on these structures. A pair of titin staining bands was clearly associated with each developing A band even at the two or three sarcomere stage, although occasional examples of a titin band being associated with a half sarcomere were noted. The appearance of sarcomeric titin patterns coincided or preceded sarcomere periodicity of either alpha actin or muscle tropomyosin. The early appearance of titin in myofibrillogenesis suggests it may have a role in filament alignment during sarcomere assembly.

Actins↗
PubMed1988

Monoclonal antibodies against caldesmon, a Ca++/calmodulin- and actin-binding protein of smooth muscle and nonmuscle cells.

Monoclonal antibodies, C2, C9, C18, C21 and C23, against chicken gizzard caldesmon have been prepared and characterized. These antibodies reacted with gizzard caldesmon (150 KDa) by enzyme-linked immunosorbent assay and protein immunoblotting. Immunofluorescence microscopy with these antibodies on cultured gizzard cells showed strong stress fiber and membrane ruffle stainings. Surprisingly, in addition to these cytoplasmic staining patterns, the C23 antibody also stained nuclei in these cells. Preabsorption of C23 with purified caldesmon abolished the staining of stress fibers and membrane ruffles as well as the staining of nuclei, suggesting that a common epitope existed in both gizzard caldesmon and the nuclear protein. Western blot analysis on the cell extract of chicken embryo fibroblast (CEF) showed that antibodies C2, C9, C18 and C21 recognized a nonmuscle caldesmon (66 KDa), whereas C23 reacted with a protein (60 KDa) different from nonmuscle caldesmon. Antibody C21 also crossreacted with a nonmuscle caldesmon (80 KDa) in normal rat kidney (NRK) cells, with a nonmuscle caldesmon (78 KDa) in human cells, and with a nonmuscle caldesmon (72 KDa) in gerbil fibroma cells. This antibody had broad-species specificity. Immunofluorescent staining of CEF cells with antibodies C2, C9, C18 and C21 showed some stress fibers and ruffles, but mostly diffuse staining. Antibody C23 crossreacted with 62 KDa and 55 KDa proteins in NRK cells, 63 KDa and 55 KDa proteins in gerbil fibroma cells and 66 KDa and 56 KDa proteins in human bladder carcinoma cells. These polypeptides were identified as nuclear lamins A and C by an anti-lamin antibody in immunoblots and two-dimensional gel analysis. Like the nuclear lamins, the C23 antigens also underwent a reversible disassembly during mitosis, as detected by double-label immunofluorescence with C23 antibody and a polyclonal anti-tubulin antibody. Tropomyosin-enriched microfilaments isolated from fibroblastic and epithelial types of NRK cells by monoclonal anti-tropomyosin antibody contained an 80 KDa protein, which had the heat-resistant property of caldesmon. The polyclonal antiserum generated against this 80 KDa protein showed a crossreactivity with purified gizzard caldesmon and vice versa. The amount of this nonmuscle caldesmon associated with the microfilaments of Kirstein virus-transformed NRK cells was greatly decreased.

Actin Cytoskeleton↗
PubMed1988

Ethylene effect on extensin and peroxidase distribution in the subapical region of pea epicotyls.

In dark grown pea (Pisum sativum) seedlings ethylene causes the triple response in which elongation growth is inhibited, radial growth is promoted, and orientation of shoots to gravity is altered. The distribution of extensin and peroxidase activity in pea epicotyls upon ethylene treatment was studied by tissue printing on nitrocellulose paper. It was found that the localization of extensin and peroxidase activity changes after 72 and 96 hours of ethylene treatment. In untreated plants, peroxidase activity is detected only in the vascular bundles. Nonetheless, after 72 and 96 hours of ethylene treatment peroxidase activity is hardly detected in the vascular system but present in the epidermal and cortical cells. Extensin increases in the epidermal and cortical cells upon ethylene treatment but it also appears in the vascular system when peroxidase activity is no longer detected.

Journal Article↗
PubMed1987

The extent of amino-terminal heterogeneity in rabbit fast skeletal muscle troponin T.

The extent and nature of fast troponin T (TnT) heterogeneity has been assessed in rabbit skeletal muscle. Previous studies identified two major fast TnT species (TnT1f and TnT2f), in the fast white muscle erector spinae, differing in their N-terminal cyanogen bromide (CNBr) fragments. Here a monoclonal antibody that recognizes a conserved region of TnT was used to characterize two additional TnT species (TnT3f and TnT4f) in the epaxial and limb musculature and a minor species (TnTcf) in craniofacial muscles. A combination of CNBr peptide mapping, immunoblotting and specific labelling of the N-terminus shows that these TnT species also differ in their N-terminal region. This observation is consistent with cDNA studies that predicted the N-terminal region is hypervariable. One additional species, a variant of TnT2f present in the tongue, was identified by two-dimensional gel electrophoresis. The limited number of TnT variants indicates that the full potential for heterogeneity inferred from the cDNA studies is not realized. This conclusion is supported by immunoblot analysis with a monoclonal antibody that recognizes an epitope in the hypervariable N-terminal region which is present in all variants of TnT1f and TnT2f but absent from the lower molecular weight species TnT3f and TnT4f.

Amino Acid Sequence↗
PubMed1987

Phytic Acid Metabolism in Lily (Lilium longiflorum Thunb.) Pollen.

The accumulation of phytic acid during development of lily (Lilium longiflorum Thunb.) pollen and its degradation during germination have been studied. A substantial amount of phytic acid accumulates in lily pollen by 5 days before anthesis, and little change occurs during subsequent maturation. Mature lily pollen contains 7 to 8 micrograms phytic acid per milligram pollen. Considerable degradation of phytic acid occurs by 15 minutes of incubation in glucose culture medium, and very little is left by 3 hours. No partially phosphorylated myo-inositol accumulates during germination. The breakdown of phytic acid proceeds at a constant rate during this time period. The rate is calculated to be 0.037 microgram phytic acid/milligram pollen/minute. Two phytases are detected in germinated lily pollen extract using high performance liquid chromatography with an anion exchange column (diethylaminoethyl-5PW). The results suggest that one of the phytases is already present in mature ungerminated lily pollen and the other one is newly synthesized during germination from a long-lived, pre-existing mRNA.

Journal Article↗
PubMed1987

The effect of dipyridamole on the initiation phase of postischemic acute renal failure in rats.

Several previous observations support the hypothesis that increased adenosine production and release mediate, at least in part, the reductions in renal blood flow and glomerular filtration rate in ischemic acute renal failure (ARF). If this hypothesis is correct, dipyridamole should potentiate these changes, since it blocks cellular adenosine uptake, thereby increasing the concentration and potentiating the effects of extracellular adenosine. Moreover, theophylline should block the effects of dipyridamole, since it is an adenosine receptor antagonist. These predictions were tested in three groups of anesthetized rats. All rats were subjected to 30 min of left renal artery occlusion; 30 min after relieving the occlusion, a 45-min clearance period was begun. The control group was given saline i.v.; the two experimental groups received either dipyridamole (24 micrograms X min-1 X kg-1) or dipyridamole plus theophylline i.v. (111 mumol/kg as a prime, 1.1 mumol X min-1 X kg-1 as an infusion). In the control group, the previously ischemic left kidneys exhibited decreased clearances of para-aminohippurate and inulin (CPAH and CIn), filtration fraction (FF), and urine/plasma inulin concentration (U/PIn), and increased urine flow (V), Na excretion (UNaV), and fractional Na excretion (FENa) in comparison with the contralateral right kidney. Dipyridamole pretreatment did not affect the right kidney, but it intensified the reductions in left kidney CPAH, CIn, and FF. Theophylline blocked all these effects of dipyridamole on the left kidney, and increased renal plasma flow (CPAH/PAH extraction), despite a decrease in systemic arterial blood pressure. These results are further support for the hypothesis that adenosine mediates, at least in part, the hemodynamic changes in postischemic ARF in rats.

Acute Kidney Injury↗
PubMed1986

The use of interstitial radiation therapy in the treatment of persistent, localized, and unresectable cancer in children.

Two children with cancer that persisted after multiple exploratory laparotomies, external beam radiation therapy, and multidrug chemotherapy had gold 198 (198Au) seeds implanted into their localized but unresectable tumor. Both children are alive, are receiving no therapy, and are disease-free more than 2 years later. These two cases indicate the value of interstitial implant therapy in the treatment of some children with cancer.

Abdominal Neoplasms↗
PubMed1986

Is the nuclear envelope a 'generator' of membrane? Developmental sequences in cytomembrane elaboration.

Early diplotene oocytes from Necturus maculosus ranging from approximately 0.2 to 0.5 mm in diameter were examined by electron microscopy. In the smallest oocytes of this range, the cytoplasm is largely devoid of membranes, but contains primarily ribosomes and mitochondria. In slightly larger oocytes, smooth-surfaced cytomembranes first appear in the perinuclear cytoplasm. At this time, the outer layer of the germinal vesicle nuclear envelope (GVNE) shows frequent connections with long membranous lamellae that extend for considerable, but variable distances into the juxtanuclear ooplasm. The number of smooth membranous lamellae increases tremendously as the oocytes increase in diameter. In such oocytes as well, frequent continuities are observed between the outer membrane of the GVNE and many of the cytoplasmic membranes. Eventually, as the ooplasm becomes populated with extensive numbers of membranous lamellae, instances of continuity between the membranous lamellae and nuclear envelope now become sparse and eventually non-existent. The frequent connections observed between membranous lamellae and the outer membrane of the GVNE during a circumscribed interval of diplotene strongly implicate the GVNE in the generation of extensive amounts of cytoplasmic membrane. The ooplasm of larger oocytes in the size range indicated contain numerous Golgi complexes and large quantities of annulate lamellae most of which are positioned in the peripheral or subcortical ooplasm, as well as extensive quantities of smooth membranes of the endoplasmic reticulum and lipid droplets.

Animals↗
PubMed1986

The influence of repeated carotid plaque hemorrhages on the production of cerebrovascular symptoms.

Single carotid intraplaque hemorrhage has been related to the occurrence of cerebrovascular ischemic symptoms. We were unable to reproduce these findings; therefore, our patient population was reinvestigated to ascertain the importance of repeated plaque hemorrhages and their possible relationship to the production of neurologic symptoms. Eighty-five consecutive patients underwent 95 carotid endarterectomies. Plaques were separated into three groups: those from patients with lateralizing symptoms, nonlateralizing symptoms, and no symptoms. All the plaques were inspected microscopically for evidence of hemorrhage, specifically, repeated hemorrhages. Repeated hemorrhages were found in 29 of 44 plaques (66%) in the lateralizing symptom group, 6 of 19 (32%) in the nonlateralizing symptom group, and 12 of 32 (37%) in the no symptom group. We conclude that repeated rather than single intraplaque hemorrhage is the critical factor related to the production of specific cerebrovascular ischemic symptoms.

Aged↗
PubMed1986

Assembly of different isoforms of actin and tropomyosin into the skeletal tropomyosin-enriched microfilaments during differentiation of muscle cells in vitro.

We have used a monoclonal antibody (CL2) directed against striated muscle isoforms of tropomyosin to selectively isolate a class of microfilaments (skeletal tropomyosin-enriched microfilaments) from differentiating muscle cells. This class of microfilaments differed from the one (tropomyosin-enriched microfilaments) isolated from the same cells by a monoclonal antibody (LCK16) recognizing all isoforms of muscle and nonmuscle tropomyosin. In myoblasts, the skeletal tropomyosin-enriched microfilaments had a higher content of alpha-actin and phosphorylated isoforms of tropomyosin as compared with the tropomyosin-enriched microfilaments. Moreover, besides muscle isoforms of actin and tropomyosin, significant amounts of nonmuscle isoforms of actin and tropomyosin were found in the skeletal tropomyosin-enriched microfilaments of myoblasts and myotubes. These results suggest that different isoforms of actin and tropomyosin can assemble into the same set of microfilaments, presumably pre-existing microfilaments, to form the skeletal tropomyosin-enriched microfilaments, which will eventually become the thin filaments of myofibrils. Therefore, the skeletal tropomyosin-enriched microfilaments detected here may represent an intermediate class of microfilaments formed during thin filament maturation. Electron microscopic studies of the isolated microfilaments from myoblasts and myotubes showed periodic localization of tropomyosin molecules along the microfilaments. The tropomyosin periodicity in the microfilaments of myoblasts and myotubes was 35 and 37 nm, respectively, whereas the nonmuscle tropomyosin along chicken embryo fibroblast microfilaments had a 34-nm repeat.

Actin Cytoskeleton↗