Biomedical subjects

M Murakami

Publications and source records attributed to M Murakami.

At least 397 records · Page 22Linked to original sources

PubMed1998

The functions of five distinct mammalian phospholipase A2S in regulating arachidonic acid release. Type IIa and type V secretory phospholipase A2S are functionally redundant and act in concert with cytosolic phospholipase A2.

We examined the relative contributions of five distinct mammalian phospholipase A2 (PLA2) enzymes (cytosolic PLA2 (cPLA2; type IV), secretory PLA2s (sPLA2s; types IIA, V, and IIC), and Ca2+-independent PLA2 (iPLA2; type VI)) to arachidonic acid (AA) metabolism by overexpressing them in human embryonic kidney 293 fibroblasts and Chinese hamster ovary cells. Analyses using these transfectants revealed that cPLA2 was a prerequisite for both the calcium ionophore-stimulated immediate and the interleukin (IL)-1- and serum-induced delayed phases of AA release. Type IIA sPLA2 (sPLA2-IIA) mediated delayed AA release and, when expressed in larger amounts, also participated in immediate AA release. sPLA2-V, but not sPLA2-IIC, behaved in a manner similar to sPLA2-IIA. Both sPLA2s-IIA and -V, but not sPLA2-IIC, were heparin-binding PLA2s that exhibited significant affinity for cell-surface proteoglycans, and site-directed mutations in residues responsible for their membrane association or catalytic activity markedly reduced their ability to release AA from activated cells. Pharmacological studies using selective inhibitors as well as co-expression experiments supported the proposal that cPLA2 is crucial for these sPLA2s to act properly. The AA-releasing effects of these sPLA2s were independent of the expression of the M-type sPLA2 receptor. Both cPLA2, sPLA2s-IIA, and -V were able to supply AA to downstream cyclooxygenase-2 for IL-1-induced prostaglandin E2 biosynthesis. iPLA2 increased the spontaneous release of fatty acids, and this was further augmented by serum but not by IL-1. Finally, iPLA2-derived AA was not metabolized to prostaglandin E2. These observations provide evidence for the functional cross-talk or segregation of distinct PLA2s in mammalian cells in regulating AA metabolism and phospholipid turnover.

Animals↗

PubMed1998

Fas-induced arachidonic acid release is mediated by Ca2+-independent phospholipase A2 but not cytosolic phospholipase A2, which undergoes proteolytic inactivation.

Fas-mediated apoptosis of human leukemic U937 cells was accompanied by increased arachidonic acid (AA) and oleic acid release from membrane glycerophospholipids, indicating phospholipase A2 (PLA2) activation. During apoptosis, type IV cytosolic PLA2 (cPLA2), a PLA2 isozyme with an apparent molecular mass of 110 kDa critical for stimulus-coupled AA release, was converted to a 78-kDa fragment with concomitant loss of catalytic activity. Cleavage of cPLA2 correlated with increased caspase-3-like protease activity in apoptotic cells and was abrogated by a caspase-3 inhibitor. A mutant cPLA2 protein in which Asp522 was replaced by Asn, which aligns with the consensus sequence of the caspase-3 cleavage site (DXXD downward arrowX), was resistant to apo-ptosis-associated proteolysis. Moreover, a COOH-terminal deletion mutant of cPLA2 truncated at Asp522 comigrated with the 78-kDa fragment and exhibited no enzymatic activity. Thus, caspase-3-mediated cPLA2 cleavage eventually leads to destruction of a catalytic triad essential for cPLA2 activity, thereby terminating its AA-releasing function. In contrast, the activity of type VI Ca2+-independent PLA2 (iPLA2), a PLA2 isozyme implicated in phospholipid remodeling, remained intact during apoptosis. Inhibitors of iPLA2, but neither cPLA2 nor secretory PLA2 inhibitors, suppressed AA release markedly and, importantly, delayed cell death induced by Fas. Therefore, we conclude that iPLA2-mediated fatty acid release is facilitated in Fas-stimulated cells and plays a modifying although not essential role in the apoptotic cell death process.

Antibodies↗

PubMed1998

Regulation of cyclooxygenase-2 and endogenous cytokine expression by bacterial lipopolysaccharide that acts in synergy with c-kit ligand and Fc epsilon receptor I crosslinking in cultured mast cells.

Emerging evidence has suggested the pivotal role of mast cells in a host defense against bacterial infection. In this paper, we report that bacterial lipopolysaccharide (LPS) is a potent enhancer of the cytokine- and IgE-dependent delayed responses of IL-3-dependent mouse bone marrow-derived cultured mast cells (BMMC). LPS, although showing minimal effects, significantly augmented the c-kit ligand (KL)- or IgE-dependent expression of cyclooxygenase (COX)-2 and the attendant delayed PGD2 generation, with IL-10 and IL-4 acting as potentiating and inhibitory cytokines, respectively. The COX-2-inducing activity of LPS was mimicked by exogenous IL-1 beta. Assessment of endogenous cytokine induction revealed that IL-1 beta expression was stimulated by either LPS or exogenous IL-1 beta. IL-6 expression occurred in parallel with COX-2 expression. IL-10 expression, which lagged behind COX-2 expression, depended on exogenous IL-10, but not on LPS and IL-1 beta. Thus, LPS and IL-1 beta exhibited similar biological activities in terms of COX-2 and endogenous cytokine expression. However, adding an antibody against the type I IL-1 receptor to BMMC, which abrogated the effects of IL-1 beta, failed to neutralize the effects of LPS. These results suggest that LPS activates BMMC through the signal transduction pathway shared with exogenous IL-1 beta, rather than exerting its action indirectly via the production of endogenous IL-1 beta.

Animals↗

PubMed1998

Assessment of left ventricular function using serum cardiac troponin I measurements following myocardial infarction.

The prognosis and extent of injury to the myocardium have previously been assessed by increased serum creatine kinase (CK) MB levels. We report findings from 39 consecutive, acute myocardial infarction (AMI) patients presenting 4.5 h (range, 0.7-12.1 h) after the onset of chest pain. We compared CK MB mass (upper reference limit, 5.0 ng/ml) and cardiac troponin I (cTnI; upper reference limit, 0.8 ng/ml) (Stratus II, Dade International) in serial serum specimens obtained over 36 h after chest pain from AMI patients; within 6 h after onset of chest pain. While the appearance of the kinetics of CK MB and cTnI were similar during the initial 24 h following the onset of chest pain, cTnI was increased significantly (p < 0.05) over CK MB after 9 to 12 h. Half-life determinations (mean+/-S.D.) in 22 of the 39 AMI patients demonstrated a significantly (p < 0.01) shorter half-life in non-Q-wave infarcts [t1/2 6.8 h (+/-5.6)] vs. Q-wave infarcts [t1/2 20.4 h (+/-10.7)]. Further serial time versus marker (mean+/-S.D.) results were significantly correlated (p < 0.001, r = 0.66). Sixteen of twenty patients assessed by echocardiography had an abnormal left ventricular ejection fraction (LVEF); mean 37.6 (S.D. 15.2)%, ranging from 15.4 to 67.6%. LVEF was significantly and inversely correlated to peak CK MB (r = .50, p = 0.03), as well as to peak cTnI (r = 0.46, p = 0.04). Based on these findings, cTnI shows excellent promise as a useful marker of infarct size, for the assessment of left ventricular function, and may potentially replace CK MB as the cardiac-specific marker for AMI detection.

Biomarkers↗

PubMed1998

Modulation of actomyosin ATPase by goniodomin A differs in types of cardiac myosin.

Goniodomin A causes the conformational change of actin to modify actomyosin ATPase activity [Furukawa, K.-I., Sakai, K., Watanabe, S., Maruyama, K., Murakami, M., Yamaguchi, K., Ohizumi, Y., 1993. Goniodomin A induces modulation of actomyosin ATPase activity mediated through conformational change of actin. J. Biol. Chem. 268, 26026-26031]. Goniodomin A inhibited the ATPase activities of atrial myofibrils, myosin B and reconstituted actomyosin in a concentration-dependent manner. Interestingly, these ATPase activities of ventricular muscle were enhanced by goniodomin A (3 x 10(-8)-3 x 10(-7) M), but were decreased when the concentration was further raised. The stimulatory effect of goniodomin A was significantly inhibited by troponin tropomyosin complex. These results suggest that goniodomin A affects actin to modify cardiac actomyosin ATPase activity, and that this modulation differs in types of cardiac myosin.

Animals↗

PubMed1998

Cyclic AMP potentiates substance P-induced amylase secretion by augmenting the effect of calcium in the rat parotid acinar cells.

Potentiation of amylase secretion by the combination of isoproterenol and substance P was examined in perfused rat parotid acinar cells. Combined additions of substance P and isoproterenol evoked biphasic changes in amylase secretion, an initial large peak and the following sustained plateau: the magnitudes of the both responses were higher than the sum of the responses induced by each agonist alone. Isoproterenol also increased the maximum response and the apparent affinity (EC50) for substance P to evoke the initial peak response; the EC50 values were about 20 and 0.8 nM, respectively, in the absence and the presence of isoproterenol. On the other hand, 1 nM substance P was sufficient for evoking the maximum potentiation of the sustained plateau response. Substance P did not change the EC50 for isoproterenol. The effect of isoproterenol was mimicked with dibutyryl cyclic AMP and agonists that increase parotid cyclic AMP. Omission of Ca2+ or addition of 5 mM nickel chloride almost completely abolished the potentiation of the sustained plateau, but little decreased that of the initial peak. Depletion of Ca2+ in InsP3-sensitive intracellular stores with thapsigargin, on the other hand, decreased the initial peak response, but not the sustained plateau, to substance P. The potentiation was also observed between isoproterenol and Ca2+ ionophores. Switching to the solutions containing higher concentrations of Ca2+ during the continuous stimulation with isoproterenol or IBMX evoked a large, but transient, response of amylase secretion. Time course of changes in amylase secretion induced by isoproterenol and substance P in combination was very similar to that of substance P, but not of isoproterenol. Isoproterenol did not enhance the effect of substance P on [Ca2+]i. These results show that the potentiation is mainly, if not totally, caused by cyclic AMP-induced enhancement of the potency and the efficacy in the pathway regulated by Ca2+.

1-Methyl-3-isobutylxanthine↗

PubMed1998

Primary culture of cells from hyperfunctioning thyroid adenoma with an activating mutation of G alphas.

We analyzed cultured cells from hyperfunctioning thyroid adenoma and its surrounding thyroid tissue from a Japanese woman and determined the nucleotide sequences of genes encoding the alpha subunit of the stimulatory G-protein 1 (G alphas) and thyrotropin (TSH) receptor in its tumor tissue. Primary culture of cells from hyperfunctioning thyroid adenoma and its surrounding thyroid tissue revealed that cAMP production was constitutively activated while intracellular Ca2+ concentration was suppressed both at the basal level and in the response to TSH stimulation in the cells from tumor tissue compared with those from non-tumor tissue. Nucleotide sequence analysis demonstrated the somatic missense mutation at codon 201 (CGT(Arg)-CAT(His)) of G alphas gene in tumor tissue but not in its surrounding tissue. No mutation was observed in the transmembrane region of TSH receptor. These results suggest that cAMP regulatory cascade is constitutively activated while phospholipase C-Ca2+ signaling cascade is suppressed in hyperfunctioning thyroid adenoma with an activating mutation of G alphas gene in the present case.

Adenoma↗

PubMed1998

Neoadjuvant concurrent chemoradiotherapy followed by definitive high-dose radiotherapy or surgery for operable thoracic esophageal carcinoma.

PURPOSE: A prospective clinical trial was undertaken to investigate the feasibility of concurrent chemoradiotherapy for esophageal carcinomas. MATERIALS AND METHODS: Between June 1989 and May 1996, forty patients with operable squamous cell carcinoma of the thoracic esophagus (Stage 0 to III: UICC 1987), ages 45 to 78 years (mean: 64), were enrolled in a study of neoadjuvant concurrent chemoradiotherapy followed by definitive high-dose radiotherapy (CRT group) or surgery (CRT-S group). Neoadjuvant chemoradiotherapy consisted of 44 Gy in 40 fractions for 4 weeks (2.2 Gy/2 Fr/day) through 10-MVX rays, with 2 courses of cisplatin (80-100 mg/body, mean: 60 mg/m2, Day 1, bolus injection) and 5-fluorouracil (500-1000 mg/body/day, mean: 400 mg/m2, Days 1-4, continuous infusion). After completion of neoadjuvant chemoradiotherapy, an intermediate clinical response was assessed by barium swallow, esophagoscopy with/without biopsy, EUS in most cases, thoracic and upper abdominal CT scan, and cervical US. Definitive chemoradiotherapy was performed in patients when regression of more than 75% was evident (CRT Group), and esophageal resection was indicated in those who remained at less than 75% (CRT-S Group). In CRT Group, a cumulative dose of 60-70 Gy for Tis, T1 and 65-75 Gy for T2-T4 tumor with high-dose-rate intraluminal brachytherapy and a total of 3 courses of chemotherapy were planned. In CRT-S Group, intraoperative radiotherapy for abdominal lymphatic system and postoperative supraclavicular irradiation were added. RESULTS: At the time of intermediate assessment, complete response (CR) was observed in 16 patients, a partial response (PR) in 22, and no change (NC) in 2. Thirty responding patients (CR, 16; PR, 14) entered the CRT Group, and 10 nonresponding patients (PR, 8; NC, 2) were followed by surgery (CRT-S Group). Radiotherapy was completed satisfactorily, but chemotherapy was suspended in 26 patients (65%) because of acute toxicity. Clinical CR rate at the completion of treatment showed 90% in CRT Group, and pathologic CR rate 10% in CRT-S Group. The overall median survival was 45 months, survival at 1, 2, and 3 years being 100%, 72%, and 56%, respectively. Local-regional failure was observed in 7 patients (all in CRT Group), distant failure in 6 (3 in CRT Group, 3 in CRT-S Group) and local-regional with distant failure in 1 (CRT Group). Four patients with local-regional recurrence in the CRT Group were salvaged by surgery. Overall survival at 2 and 3 years for CRT vs. CRT-S Group was 72%, 64% vs. 75%, 38%, respectively. No treatment-related mortality was observed. The rate of the 'esophagus conservation' was 65% (Stage 0: 1 of 1, 100%; Stage I: 11 of 12, 92%; Stage II: 8 of 17, 47%; Stage III: 6 of 10, 60%). CONCLUSION: Our results demonstrated that almost all early disease (Stage 0-I) and about half of advanced disease (Stage II-III) could be conserved, their esophagus treated by the multidisciplinary approach centering on high-dose radiotherapy and concurrent chemotherapy.

Aged↗

PubMed1998

Age-dependent appearance of NK1.1+ T cells in the livers of beta2-microglobulin knockout and SJL mice.

NK1.1+ T cells, a specialized set of T cells that recognize CD1, are reportedly absent in young beta2-microglobulin-deficient (beta2m-knockout (KO)) and SJL mice. In this study, we show that a significant number of NK1.1+ T cells exist in the livers of older beta2m-KO and SJL mice, and that the number of liver NK1.1+ T cells increases as the animals age. The surface phenotypes of liver NK1.1+ T cells from beta2m-KO and SJL mice were similar to NK1.1+ T cells from C57BL/6 mice, except that the bulk of these cells were CD4- CD8-. After anti-CD3 injection in vivo, the cells promptly expressed IL-4 mRNA just as NK1.1+ T cells did in normal mice. Using L cells expressing CD1, liver NK1.1+ T cells from both beta2m-KO and SJL mice were stimulated to proliferate, although to a lesser degree than were such cells from C57BL/6 mice. Our studies show that some NK1.1+ T cells accumulate in the livers of older beta2m-KO and SJL mice, and that they appear to have functional properties similar to "normal" NK1.1+ T cells.

Age Factors↗

PubMed1998

Segregated coupling of phospholipases A2, cyclooxygenases, and terminal prostanoid synthases in different phases of prostanoid biosynthesis in rat peritoneal macrophages.

We examined herein the functional linkage of enzymes regulating the initial, intermediate, and terminal steps of PG biosynthesis to provide PGs in rat peritoneal macrophages stimulated with LPS and/or A23187. Quiescent cells stimulated with A23187 produced thromboxane B2 (TXB2) in marked preference to PGE2 within 30 to 60 min (constitutive immediate response), which was mediated by preexisting cytosolic phospholipase A2 (cPLA2), cyclooxygenase-1 (COX-1), and TX synthase. Cells treated with LPS predominantly produced PGE2 during culture for 3 to 24 h (delayed response), where cPLA2 and secretory PLA2 functioned cooperatively with inducible COX-2, which was, in turn, coupled with inducible PGE2 synthase. Cells primed for 12 h with LPS and stimulated for 30 min with A23187 produced PGE2 in marked preference to TXB2 (induced immediate response), in which three inducible enzymes, cPLA2, COX-2, and PGE2 synthase, were functionally linked. Preferred coupling of the two inducible enzymes, COX-2 and PGE2 synthase, was further confirmed by the ability of LPS-treated cells to convert exogenous arachidonic acid to PGE2 optimally at a time when both enzymes were simultaneously induced. These results suggest that distinct PG biosynthetic enzymes display segregated functional coupling following different transmembrane stimulation events even when enzymes that catalyze similar reactions in vitro coexist in the same cells.

Animals↗

PubMed1998

Cytosolic phospholipase A2 is required for cytokine-induced expression of type IIA secretory phospholipase A2 that mediates optimal cyclooxygenase-2-dependent delayed prostaglandin E2 generation in rat 3Y1 fibroblasts.

Activation of rat fibroblastic 3Y1 cells with interleukin-1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF alpha) induced delayed prostaglandin (PG) E2 generation over 6-48 h, which occurred in parallel with de novo induction of type IIA secretory phospholipase A2 (sPLA2) and cyclooxygenase (COX)-2, without accompanied by changes in the constitutive expression of type IV cytosolic PLA2 (cPLA2) and COX-1. Types V and IIC sPLA2s were barely detectable in these cells. Studies using an anti-type IIA sPLA2 antibody, sPLA2 inhibitors, and a type IIA sPLA2-specific antisense oligonucleotide revealed that IL-1 beta/TNF alpha-induced delayed PGE2 generation by these cells was largely dependent on inducible type IIA sPLA2, which was functionally linked to inducible COX-2. Delayed PGE2 generation was also suppressed markedly by the cPLA2 inhibitor arachidonoyl trifluoromethyl ketone (AACOCF3), which attenuated induction of type IIA sPLA2, but not COX-2, expression. AACOCF3 inhibited the initial phase of cytokine-stimulated arachidonic acid release, and supplementing AACOCF3-treated cells with exogenous arachidonic acid partially restored type IIA sPLA2 expression. These results suggest that certain metabolites produced by the cPLA2-dependent pathway are crucial for the subsequent induction of type IIA sPLA2 expression and attendant delayed PGE2 generation. Some lipoxygenase-derived products might be involved in this event, since IL-1 beta/TNF alpha-induced type IIA sPLA2 induction and PGE2 generation were reduced markedly by lipoxygenase, but not COX, inhibitors. In contrast, Ca2+ ionophore-stimulated immediate PGE2 generation was regulated predominantly by the constitutive enzymes cPLA2 and COX-1, even when type IIA sPLA2 and COX-2 were maximally induced after IL-1 beta/TNF alpha treatment, revealing functional segregation of the constitutive and inducible PG biosynthetic enzymes.

Animals↗

PubMed1998

Expression and co-stimulatory function of B7-2 on murine CD4+ T cells.

Co-stimulatory signals mediated by the interaction of B7-1/B7-2 with CD28 are important for the activation of CD4+ T cells stimulated with antigen on antigen-presenting cells. There are controversies about the expression and function of B7-1/B7-2 on CD4+ T cells. The aim of this study was to analyze the expression of B7-1/B7-2 on naive and memory CD4+ T cells and the co-stimulatory function in the activation of naive CD4+ T cells stimulated by TCR ligation. Present results indicate that memory CD4+ T cells express B7-2 molecules on their surface, whereas naive CD4+ T cells do not. Neither memory nor naive CD4+ T cells expressed B7-1 molecule on their surface, although B7-1 mRNA was faintly expressed in memory T cells. B7-2 molecules expressed on memory T cells co-stimulated CD4+ naive T cells stimulated with plate-coated anti-CD3 to produce IL-2. Naive CD4+ T cells were shown to express B7-2 after co-stimulation with B7-2 and TCR ligation, because the naive T cells stimulated with anti-CD3 and B7-2CHO expressed B7-2 on their surface, although it remained to be studied whether the co-stimulation with B7-2 directly induced B7-2 expression on naive T cells. Our present results indicate that memory CD4+ T cells play some role in the activation of naive CD4+ T cells through the co-stimulation with B7-2 molecules.

Animals↗

PubMed1998

Effects of centrifugation and lipid removal on the cryopreservation of in vitro produced bovine embryos at the eight-cell stage.

The effects of intracellular lipid polarization and lipid removal treatments on the postthawed in vitro development of frozen bovine embryos at the 8-cell stage were studied. As the first step, bovine presumptive zygotes were centrifuged at 16,000 g for 20 min for the cytoplasmic lipid polarization and their lipid layers were removed by micromanipulation in order to examine the influence of these treatments on the developmental capacity of bovine zygotes. As the second step, bovine embryos developed to the 8-cell stage following centrifugation treatment at various forces (8000, 12,000, and 16,000 g) or lipid removal treatment at the 1-cell stage were frozen in 1.8 M ethylene glycol + 0.05 M trehalose supplemented with 5% polyvinylpyrrolidone in a one-step procedure. There were no significant differences among the control (nontreatment), lipid-polarized, and lipid-removed groups with respect to the developmental capacity of fresh nonfrozen zygotes (experiment 1). The rates of survival and development to the blastocyst of frozen-thawed 8-cell embryos increased slightly with increasing force of centrifugation (experiment 2). The rate of development into blastocysts of the frozen-thawed 8-cell embryos was significantly higher in the groups that underwent centrifugation (at 16,000 g for 20 min; P < 0.05) or lipid removal (P < 0.01) treatments than the control (intact) group. However, there were no significant differences among the groups with respect to the rate of development to the expanded/hatched blastocyst stage. In addition, the mean cell numbers of embryos developed into blastocysts (day 8) derived from frozen-thawed 8-cell embryos tended to be low in the centrifugation and lipid removal groups compared to the controls (experiment 3). These results suggest that although the centrifugation with or without lipid removal treatments has no detrimental effects on the developmental capacity of bovine zygotes, the freezing tolerance of bovine 8-cell embryos was not improved by these treatments.

Animals↗

PubMed1998

Expression of the alpha 1 and alpha 2 chains of type IV collagen in the infarct zone of rat myocardial infarction.

Healing processes affect ventricular remodeling, an important prognostic factor in acute myocardial infarction (AMI). Type IV collagen, a major component of the basement membrane, has been demonstrated to appear in the infarct zone. Six type IV alpha chains, alpha 1 to alpha 6 [alpha 1(IV) to alpha 6(IV)], have been identified. We examined the expression of the alpha 1(IV), alpha 2(IV), alpha 3(IV) and alpha 5(IV) in experimental rat AMI induced by permanent ligation of the left coronary artery in male Sprague-Dawley rats. Neonatal cardiac fibroblasts cultured to reach confluency were also used to examine the hypoxic induction of the expression of these chains. Northern blotting, in situ hybridization, and immunohistochemical staining were employed to examine the alpha chain expression. With the in situ hybridization, alpha 1(IV) and alpha 2(IV) mRNA signals were observed in spindle-shaped mesenchymal cells in the intercellular spaces, in the vascular endothelial cells and in some myocytes in the normal myocardium. Signals for both alpha 1(IV) and alpha 2(IV) mRNA appeared in the spindle-shaped mesenchymal cells located between the surviving myocytes in the infarct peripheral zone at 24 h after the ligation. These signals extended into the central zone and reached the center point of the infarct 5 days after the ligation. Neither alpha 3(IV) nor alpha 5(IV) mRNA signals were observed in the infarct zone. Northern blotting demonstrated increased expressions of alpha 1(IV) and alpha 2(IV) mRNA in the infarct zone compared to the sham-operated hearts. The expression of alpha 5(IV) mRNA was weak in the sham-operated hearts but was not detected in the infarct zone. alpha 3(IV) mRNA was not expressed in the infarct zones or sham-operated hearts. In the cultured cardiac fibroblasts, a hypoxic induction of the expression of both alpha 1(IV) and alpha 2(IV) mRNA was demonstrated by Northern hybridization. Immunohistochemistry revealed alpha 1(IV) and alpha 2(IV) staining in the infarct lesion. The findings demonstrate that among the type IV collagen alpha chains examined, alpha 1(IV) and alpha 2(IV) produced by mesenchymal cells appeared in the infarct zone, suggesting that type IV collagen composed by alpha 1 and alpha 2 chains contributes to the pathological course of myocardial infarction.

Animals↗

PubMed1998

Increase in the expression of biglycan mRNA expression Co-localized closely with that of type I collagen mRNA in the infarct zone after experimentally-induced myocardial infarction in rats.

Biglycan, a small dermatan sulphate proteoglycan, has been postulated to interact with other components of the extracellular matrix (ECM), specifically collagens. We hypothesized that biglycan messenger ribonucleic acid (mRNA) is increased in the myocardial infarct zone. Biglycan mRNA expression after acute myocardial infarction (AMI) in rats was determined with the use of Northern blotting and in situ hybridization, and its expression pattern was compared to that of type I collagen mRNA [alpha1(I) collagen]. The left coronary artery was ligated in male Sprague-Dawley rats, and the hearts were excised on days 2 and 7. The Northern blot analysis demonstrated that expression of biglycan mRNA in the infarct on days 2 and 7 were 4.0- and 6.8-fold higher, respectively, compared to the sham-operated hearts. The in situ hybridization revealed intense signals for both biglycan and alpha1(I) collagen mRNA on day 2 in the spindle-shaped mesenchymal cells located between the surviving myocytes in the infarct peripheral zone. On day 7, biglycan mRNA signals were observed in the interior of the infarct around the infarct granulation tissue, a distribution that was essentially the same as that of alpha1(I) collagen. These results demonstrated that the increases in the infarct biglycan mRNA expression produced by mesenchymal cells (presumably myofibroblasts and fibroblasts) was closely co-localized with that of type I collagen mRNA, indicating that biglycan contributes to the infarct healing processes.

Animals↗

PubMed1998

Histopathological assessment of localized proliferation in cases of Bowen's disease using immunostaining and a laser cytometer.

In order to evaluate the localized proliferative activity of intratumor cells in Bowen's disease using tissue sections, skin specimens from ten patients were compared with skin samples from seven normal individuals for their expression of proliferating cell nuclear antigen (PCNA), Ki-67 immunostaining and intranuclear DNA contents, quantitated with a laser cytometer (LCM). In normal epidermis, the largest proportion of PCNA- and Ki-67-positive cells was observed in the basal cell layer, with the amounts decreasing through the suprabasal cell layer towards the prickle cell layer. Examination by LCM also revealed the highest average fluorescence intensity of individual nuclei in the basal cell layer and, as with the immunohistological parameters, reducing towards the upper layer of the epidermis. In the Bowen's disease tissue sections, the largest proportion of PCNA- and Ki-67-positive cells was found in contact with the basement membrane (base of the tumor), with lower amounts in the center of the tumor nest and in the marginal epidermis. The average fluorescence intensities of individual nuclei were in line with these results. These results show that tumor cells distributed in Bowen's disease tumor nests have different proliferative activities depending on their location.

Bowen's Disease↗

PubMed1998

Genetic changes in primary colorectal cancer by comparative genomic hybridization.

Comparative genomic hybridization (CGH) is a powerful new technique for the molecular cytogenetic analysis of cancer. In this method, at first the cancer DNA and normal DNA are labeled with biotin and digoxigenin, respectively, and then the labeled DNAs are applied onto normal lymphocyte metaphase preparations in hybridization. After hybridization, they are stained with FITC and rhodamine, respectively, so chromosomal gains and losses in cancer are thus detected by using a green:red ratio. In this study, we analyzed the abnormal chromosomes in nine cases with human primary colon cancer. A gain in chromosomes 11p, 12q, 16p, 20p, and 20q were observed, while a loss of 18q and 22q were discovered. CGH may thus provide us with important information for analyzing the genes in colon cancer.

Chromosome Aberrations↗

PubMed1998

Northern analysis of type II iodothyronine deiodinase mRNA in rat Harderian gland.

It has been known that type II iodothyronine deiodinase activity is present in rat Harderian gland and the activity is significantly increased by isoproterenol administration. We have performed Northern analyses to study whether the transcript for type II iodothyronine deiodinase is expressed in rat Harderian gland and whether the isoproterenol stimulation of type II iodothyronine deiodinase activity in rat Harderian gland is due to the change in its mRNA level. Northern analyses have demonstrated that type II iodothyronine deiodinase mRNA, approximately 7.5 kb in size, is expressed in rat Harderian gland, and the mRNA levels as well as the deiodinase activities are greater in hypothyroid rats than those in euthyroid rats. Type II iodothyronine deiodinase mRNA levels and the deiodinase activities in Harderian gland were increased by isoproterenol administration, and the increase in the mRNA levels preceded that in the deiodinase activities. These results indicate that 7.5 kb transcript for type II iodothyronine deiodinase is expressed in rat Harderian gland and beta-adrenergic stimulation of type II iodothyronine deiodinase activity is due at least in part to the increase in its mRNA level.

Actins↗