Biomedical subjects

M Fujimura

Publications and source records attributed to M Fujimura.

At least 127 records · Page 7Linked to original sources

PubMed1999

Up-regulation of ICH-1L protein by thromboxane A2 antagonists enhances cisplatin-induced apoptosis in non-small-cell lung-cancer cell lines.

We evaluated the effect of thromboxane A2 (TXA2) blockade on cisplatin-induced apoptosis in non-small-cell lung cancer (NSCLC) cell lines. Cisplatin induced apoptosis in PC/9 and PC-9/CDDP in a dose-dependent manner. Treatment with specific TXA2 antagonist, calcium 5(Z)-1R,2S,3S,4S-7-[3-phenylsulfonylaminobicyclo[2,2,1]hept-2-yl]- 5-heptonoate hydrate (S-1452) and 5(Z-6-[(1R,2R,3R,4S)-3-(N-4-bromobenzenesulfonyl aminomethyl) bicyclo[2,2,1]heptane-2-yl]-hex-5-enoic acid (ONO-NT-126), enhanced the cisplatin-induced apoptosis in each cell line. Acetyl-L-aspartyl-glutamyl-valyl-aspart-1-aldehyde (Ac-DEVD-CHO) inhibited cisplatin-induced apoptosis and enhancement of the apoptosis by TXA2 blockade, but acetyl-L-tyrosyl-valyl-alanyl-aspart-1-aldehyde (Ac-YVAD-CHO) had no effect on the apoptosis. There was no difference in the interleukin-1beta-converting enzyme (ICE) protease protein expression in either cell line. Cysteine protease p32(CPP32) protein expression was lower in PC-9/CDDP but was not changed by S-1452, cisplatin, or cotreatment with cisplatin and S-1452. Ice and Ced-3 homolog (ICH-1L) expression was significantly lower in PC-9/CDDP and was up-regulated by S-1452 or ONO-NT-126. These data suggest that ICH-1L might play a critical role in cisplatin-induced apoptosis and that TXA2 blockade up-regulates ICH-1L protein expression. Overexpression of ICH-1L and treatment with cisplatin might result in an increase in apoptosis in NSCLC cell lines.

Apoptosis↗

PubMed1999

In vivo airway eosinophil accumulation does not enhance antigen- or propranolol-induced bronchoconstriction in guinea pigs.

BACKGROUND: Chronic airway eosinophil accumulation is characteristic of asthma. However, it remains unclear whether airway eosinophils enhance or reduce release of chemical mediators and/or action of the released mediators in the airways in vivo, because previous investigators have indicated that eosinophil-derived factors such as histaminase and arylsulfatase may alter the allergic reaction by metabolizing chemical mediators. Recently, we have developed a guinea pig model of propranolol-induced bronchoconstriction (PIB), which is mediated by lipid mediators such as thromboxane A2 (TxA2), cysteinyl leukotrienes (cLTs) and platelet activation factor (PAF). This study was conducted to explain the influence of airway eosinophil accumulation on antigen-induced bronchoconstriction and the following PIB, both of which are mediated by lipid mediators. METHODS: Guinea pigs were transnasally treated with 75 microg/kg of polymyxin-B or vehicle twice a week for a total of 3 weeks. Guinea pigs were anesthetized and treated with diphenhydramine hydrochloride, and then artificially ventilated 24 h after the last administration of polymyxin-B or vehicle followed by passive sensitization. Propranolol at a concentration of 10 mg/ml was inhaled 20 min after an aerosolized antigen challenge. RESULTS: The proportion of eosinophils in bronchoalveolar lavage fluid obtained 15 min after the propranolol inhalation was significantly increased in guinea pigs treated with polymyxin-B compared with the vehicle. The polymyxin-B treatment did not affect antigen-induced bronchoconstriction or the following PIB. CONCLUSIONS: We conclude that eosinophils accumulated in the airways by polymyxin-B does not affect release of chemical mediators induced by antigen or propranolol inhalation, or action of released mediators in vivo.

Administration, Inhalation↗

PubMed1999

Early decrease in apurinic/apyrimidinic endonuclease is followed by DNA fragmentation after cold injury-induced brain trauma in mice.

Apurinic/apyrimidinic endonuclease, a multifunctional protein in the DNA base excision repair pathway, plays a central role in repairing DNA damage caused by reactive oxygen species. We examined protein expression of apurinic/apyrimidinic endonuclease before and after cold injury-induced brain trauma in mice, where we have previously shown reactive oxygen species to participate. Immunohistochemistry showed the nuclear expression of apurinic/apyrimidinic endonuclease in the entire region of control brains. One hour after cold injury-induced brain trauma, nuclear immunoreactivity was predominantly decreased in the inner boundary of the lesion, whereas there was a slight increase in the outer boundary area. Four hours after cold injury-induced brain trauma, nuclear immunoreactivity was almost absent in the entire lesion, and remained so until 24 h. At this time, a marked increase in apurinic/apyrimidinic endonuclease immunoreactivity was seen in the outer boundary zone. Western blot analysis of the sample from the non-ischemic area showed a characteristic 37,000 mol. wt band, which decreased markedly 24 h after cold injury-induced brain trauma. A time-dependent increase in DNA fragmentation was also observed after cold injury-induced brain trauma. Our data provide the first evidence that apurinic/apyrimidinic endonuclease decreased rapidly in the lesion after cold injury-induced brain trauma, whereas it was significantly increased at the outer boundary zone. Although further examination is necessary to elucidate the direct relationship between apurinic/apyrimidinic endonuclease alteration and the pathogenesis of cold injury-induced brain trauma, our results suggest the possibility that an early decrease in apurinic/apyrimidinic endonuclease and failure of the DNA repair mechanism may contribute to DNA-damaged neuronal cell death after cold injury-induced brain trauma.

Animals↗

PubMed1999

Interleukin-8 inhalation directly provokes bronchoconstriction in guinea pigs.

BACKGROUND: Although it has been reported that the concentration of interleukin (IL)-8 in nasal lavage fluid and sputum and its production in bronchial epithelium were increased in asthmatic subjects, the direct effects of IL-8 on the airways in vivo is unclear. METHODS: We examined bronchoconstriction in response to IL-8 inhalation through an endotracheal cannula in anesthetized, artificially ventilated guinea pigs. RESULTS: Inhalation of IL-8 at concentrations of 1 and 10 microg/ml caused significant bronchoconstriction, as revealed by the elevation of pressure at the airway opening. Moreover, the bronchoconstriction induced by IL-8 was significantly inhibited by the antihistamines diphenhydramine and terfenadine, suggesting the involvement of histamine release in the IL-8-induced bronchoconstriction. No significant leukocyte infiltration was observed in the bronchoalveolar lavage fluid or histologic findings 25 min after the first IL-8 inhalation. CONCLUSIONS: IL-8 provokes bronchoconstriction without leukocyte accumulation in the airways, mediated in part by histamine release, in guinea pigs.

Administration, Inhalation↗

PubMed1999

Increased airway responsiveness to acetaldehyde in asthmatic subjects with alcohol-induced bronchoconstriction.

Bronchial responsiveness to acetaldehyde, a main factor in alcohol-induced bronchoconstriction, and methacholine were compared between 10 subjects with alcohol-induced bronchoconstriction and 16 asthmatic subjects without alcohol sensitivity. In the alcohol-sensitive group, the geometric mean (geometric SEM (GSEM)) of the provocative concentration of methacholine (PC20,meth) and acetaldehyde (PC20,acet) causing a 20% fall in forced expiratory volume in one second were 0.947 mg x mL(-1) (GSEM 0.139) and 21.0 mg x mL(-1) (GSEM 0.112), respectively, which were not significantly different from those in the nonalcohol-sensitive group, which were 0.634 mg x mL(-1) (GSEM 0.115) and 31.7 mg x mL(-1) (GSEM 0.077), respectively. The ratio of airway responsiveness to acetaldehyde relative to methacholine (log PC20,acet/PC20,meth) was 1.345+/-0.093 (mean+/-SEM) in the alcohol-sensitive group, which was significantly different from the value of 1.699+/-0.059 in the nonalcohol-sensitive group (p=0.0025). A significant correlation was observed between PC20,meth and PC20,acet in both the alcohol-sensitive group (r=-0.742, p=0.0115) and nonsensitive group (r=0.882, p<0.0001). In conclusion, the airways of asthmatic subjects with alcohol-induced bronchoconstriction have a selective hyperresponsiveness to acetaldehyde.

Acetaldehyde↗

PubMed1999

Motor activity of vascularly perfused rat duodenum. 1. Characteristics of spontaneous movement.

We developed an ex vivo model of arterially perfused rat duodenum to examine the motor activity of intestine. In this preparation, spontaneously occurring pressure waves with regular rhythm were observed. The oxygen consumption and motor activity of the intestine were compared at different arterial perfusion rates to determine the degree of oxygenation required to elicit spontaneous motility. Pressure waves with regular rhythm occurred at a frequency of 1 min-1 when the arterial perfusion was 3-5 mL min-1, and stopped when the perfusion rate fell below 2 mL min-1. Atropine and hexamethonium reduced the percentage motor index/10 min of pressure waves in a dose-dependent manner, and tetrodotoxin completely blocked motor activity. Acetylcholine stimulated motor activity, and this effect was not antagonized by TTX. These findings suggest that spontaneous contraction in the ex vivo perfused rat duodenum might be mediated by a cholinergic mechanism via muscarinic receptors on smooth muscle, but that noncholinergic mechanisms may also participate in this response.

Animals↗

PubMed1999

Motor activity of vascularly perfused rat duodenum. 2. Effects of VIP, PACAP27 and PACAP38.

We examined the mechanisms of effects of vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase activating polypeptide (PACAP) 27 and PACAP38 on spontaneously occurring pressure waves in ex vivo perfused rat duodenum. VIP and PACAPs dose-dependently reduced the percentage motor index of pressure waves; this reduction was not prevented by atropine, hexamethonium or tetrodotoxin (TTX). VIP and PACAPs abolished acetylcholine-induced stimulation of pressure waves, even in the presence of TTX. These findings suggest that VIP and PACAPs may exert direct inhibitory effects via VIP/PACAP receptors located on smooth muscle rather than via cholinergic receptors. The inhibitory effects of VIP and PACAPs were partially antagonized by the VIP receptor antagonists VIP(10-28), suggesting that VIP and PACAPs share common receptor sites on intestinal smooth muscle. The effects of VIP and PACAPs were completely antagonized by nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine (L-NA), suggesting that NO mediates the inhibitory effects of VIP and PACAPs on duodenal motility. Furthermore, single injection of L-NA stimulated spontaneously occurring pressure waves, while VIP(10-28) did not affect them. These findings suggest that VIP/PACAPs and NO strongly interact as an inhibitory mediator on duodenal motility, but that their modes of action in doing so may differ.

Animals↗

PubMed1999

Involvement of 5-HT3 and 5-HT4 receptors in the motor activity of isolated vascularly perfused rat duodenum.

The involvement of serotonin (5-HT) receptor subtypes in motor activity of the ex vivo vascularly perfused rat duodenum was investigated. Clusters of phasic contractions (CPCs), migrating in an oral to anal direction, were obtained without any stimulation. Drug effects were evaluated by changes in different components of the pressure waves, such as motor index (MI), frequency, amplitude and duration of the CPC. The effect of 5-HT depletion on motor activity was examined in animals treated with p-chlorophenylalanine (PCPA). The MI, frequency and duration of CPC were decreased by PCPA, but the amplitude was not affected, suggesting that endogenous 5-HT may play an important role in regulation of the motor activity of the rat intestine. The importance of the 5-HT receptor subtypes in the regulation of motor activity was examined. Neither the nonselective 5-HT1 and 5-HT2 receptor antagonist, methysergide, nor the 5-HT2 receptor antagonist, ketanserin, affected motor activity. However, the 5-HT3 receptor antagonists, granisetron and azasetron, decreased percentage MI, frequency, percentage amplitude and percentage duration of CPC. The 5-HT4 receptor antagonist, SB204070, exerted both excitatory and inhibitory actions, with a higher dose (10 nM) stimulating percentage MI, frequency, percentage amplitude and percentage duration, and a lower dose (0.1 nM or 1 nM) decreasing percentage MI and percentage duration of CPC. These results suggest that endogenous 5-HT regulates the motor activity of the rat duodenum through 5-HT3 and 5-HT4 receptors, with the former mediating the stimulatory influence and the latter mediating both stimulatory and inhibitory influences.

Animals↗

PubMed1999

Duration of tachyphylaxis in response to methacholine in healthy non-asthmatic subjects.

It is well recognized that bronchial responsiveness to methacholine is reduced after methacholine-induced bronchoconstriction in non-asthmatic subjects, but not in asthmatic subjects. However, it is unknown how long the methacholine tachyphylaxis lasts. The present study was conducted to elucidate duration of the methacholine tachyphylaxis in healthy non-asthmatic subjects. Measurements of methacholine responsiveness were repeated six times at intervals of 1 h and 1, 2, 3 and 7 days in eight healthy non-asthmatic young female subjects in whom methacholine concentrations causing a 20% fall in forced expiratory volume in 1 sec (FEV1; PC20-Meth) were 40 mg/mL or less. Geometric mean value (GSEM) of PC20-Meth was 10.6 (1.44) mg/mL at the first challenge, 63.4 (1.85) at the 1-h interval, 50.2 (1.67) at the 1-day interval, 49.7 (1.68) at the 2-day interval, 17.3 (1.51) at the 3-day interval and 13.1 (1.44) mg/mL at the 7-day interval. The values at intervals of 1 h to 3 days were significantly greater than the initial value. These results indicate that measurements of methacholine responsiveness should be separated by at least 7 days to avoid the tachyphylaxis when the tests are repeated in non-asthmatic subjects.

Adult↗

PubMed1999

Malignant mesothelioma presenting as pulmonary metastasis ahead of growth of primary tumour.

A 59-year-old woman was admitted to Houju Memorial Hospital, Ishikawa, Japan, because of cough and fever on 30 March 1997. A diagnosis of pneumonia was made and she was given antibiotics. Her symptoms improved but failed to resolve completely on antibiotic therapy. On 9 September 1997, she revisited the hospital because of bodyweight loss and malaise. There was no history of exposure to asbestos. The chest roentgenogram revealed infiltrative shadows with vague and indistinct margins suggesting inflammatory processes, which were more extensive than those investigated on her last visit. One month later, a giant tumour was detected rapidly growing from the mediastinum and open biopsy was performed. The histological examination confirmed that the tumour was a malignant mesothelioma and the intrapulmonary nodules were its metastases. This is a rare case of pulmonary metastasis being present for several months before an appearance of primary mesothelioma.

Female↗

PubMed1999

Difference in bronchoprotective effects of bronchodilators on postallergic propranolol-induced bronchoconstriction.

Administration of propranolol provokes bronchoconstriction only in asthmatic patients. It is unknown whether bronchodilator therapy can prevent the propranolol-induced bronchoconstriction (PIB). We previously reported an animal model of PIB in which bronchoconstriction is caused by propranolol when inhaled 20 minutes after an antigen provocation in passively sensitized guinea pigs. Our goal was to evaluate the bronchoprotective effects of bronchodilators on the PIB in our animal model. Propranolol was inhaled 20 minutes after an antigen challenge in passively sensitized, anesthetized, and artificially ventilated guinea pigs. Atropine (5 mg/kg) and equipotent doses of salbutamol (0.6 microgram/kg) and aminophylline (25 mg/kg), which were determined by the dose-response curves for inhibition of histamine-induced bronchoconstriction, were intravenously administered 5 minutes before the propranolol inhalation. Treatment of the animals with 25 mg/kg of aminophylline, but not with 0.6 microgram/kg of salbutamol or 5 mg/kg of atropine, significantly prevented the PIB. These results show that our animal model is an experimental model of PIB which is resistant to beta 2-agonists or anticholinergics and suggest that aminophylline may be useful to prevent and treat PIB resistant to beta 2-agonists.

Airway Resistance↗

PubMed1999

Comparison of bronchoalveolar lavage cell findings in complete-resolution pneumonia and delayed-resolution pneumonia.

BACKGROUND: In some patients with bacterial pneumonia, the resolution of chest radiograph shadows are delayed. There have been many clinical and pathological studies on delayed-resolution pneumonia (DR). However, there are no reports concerning inflammatory cell findings of bronchoalveolar lavage (BAL) fluid in patients with DR. We compared the BAL fluid cell findings in patients with DR with those in patients with complete-resolution pneumonia (CR). METHODS: The subjects included six patients whose chest radiograph shadows were completely resolved within 2 weeks after an appropriate antibiotic administration (CR), and nine patients whose chest radiograph shadows were unresolved more than 2 weeks after the treatment (DR). BAL was done 2-3 weeks after the antibiotic treatment in both groups. We compared differential counts and lymphocyte subsets in BAL fluid among patients with CR, patients with DR, and asymptomatic subjects. RESULTS: There were no significant differences in BAL fluid cell findings between CR groups and asymptomatic groups. On the other hand, the percentages of lymphocytes, neutrophils and eosinophils in DR group were significantly increased compared with those in CR and normal groups. There was no significant difference in the CD4+/CD8+ ratio of BAL lymphocytes among the three groups. CONCLUSIONS: It is suggested that infiltration of inflammatory cells in the lung exists in DR, despite the disappearance of inflammatory reaction in the peripheral blood.

Adult↗

PubMed1999

Early decrease of apurinic/apyrimidinic endonuclease expression after transient focal cerebral ischemia in mice.

The authors examined the protein expression of apurinic/apyrimidinic endonuclease (APE/Ref-1), a multifunctional protein in the DNA base excision repair pathway, before and after transient focal ischemia in mice. Immunohistochemistry showed the nuclear expression of APE/Ref-1 in the entire region of the control brains. Nuclear immunoreactivity was decreased as early as 5 minutes after 60 minutes of ischemia in the ischemic core, which was followed by a significant reduction of APE/Ref-1-positive cells in the entire middle cerebral artery territory. Western blot analysis of the sample from the nonischemic brain showed a characteristic 37-kDa band, which was reduced after ischemia. A significant amount of DNA fragmentation was observed at 24 hours, but not at 4 hours, after ischemia. The authors' data provide the first evidence that APE/Ref-1 rapidly decreases after transient focal ischemia, and that this reduction precedes the peak of DNA fragmentation in the brain regions that are destined to show necrosis and apoptosis. Although further examination is necessary to elucidate the direct relationship between the APE/Ref-1 decrease and ischemic necrosis and apoptosis, our results suggest the possibility that rapid decrease of APE/Ref-1 and the failure of the DNA repair mechanism may contribute to necrosis or apoptosis after transient focal ischemia.

Animals↗

PubMed1999

Inhibition of interleukin-1beta converting enzyme family proteases (caspases) reduces cold injury-induced brain trauma and DNA fragmentation in mice.

The authors examined the effect of z-VAD.FMK, an inhibitor that blocks caspase family proteases, on cold injury-induced brain trauma, in which apoptosis as well as necrosis is assumed to play a role. A vehicle alone or with z-VAD.FMK was administered into the cerebral ventricles of mice 15 minutes before and 24 and 48 hours after cold injury. At 24 hours after cold injury, infarction volumes in the z-VAD.FMK-treated animals were significantly smaller than infarction volumes in the vehicle-treated animals, and were further decreased at 72 hours (0.92 +/- 1.80 mm3, z-VAD.FMK-treated animals; 7.46 +/- 3.53 mm3, vehicle-treated animals; mean +/- SD, n = 7 to 8). The amount of DNA fragmentation was significantly decreased in the z-VAD.FMK-treated animals compared with the vehicle-treated animals, as shown by terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end labeling staining and DNA gel electrophoresis. By Western blot analysis, both the proform and activated form of interleukin-1beta converting enzyme (caspase 1) were detected in the control brain, and the activated form showed moderate reduction after cold injury-induced brain trauma. These results indicate that caspase inhibitors could reduce cold injury-induced brain trauma by preventing neuronal cell death by DNA damage. The caspase family proteases appear to contribute to the mechanisms of cell death in cold injury-induced brain trauma and to provide therapeutic targets for traumatic brain injury.

Amino Acid Chloromethyl Ketones↗

PubMed1999

Early appearance of activated matrix metalloproteinase-9 after focal cerebral ischemia in mice: a possible role in blood-brain barrier dysfunction.

During cerebral ischemia blood-brain barrier (BBB) disruption is a critical event leading to vasogenic edema and secondary brain injury. Gelatinases A and B are matrix metalloproteinases (MMP) able to open the BBB. The current study analyzes by zymography the early gelatinases expression and activation during permanent ischemia in mice (n = 15). ProMMP-9 expression was significantly (P < 0.001) increased in ischemic regions compared with corresponding contralateral regions after 2 hours of ischemia (mean 694.7 arbitrary units [AU], SD +/- 238.4 versus mean 107.6 AU, SD +/- 15.6) and remained elevated until 24 hours (mean 745.7 AU, SD +/- 157.4). Moreover, activated MMP-9 was observed 4 hours after the initiation of ischemia. At the same time as the appearance of activated MMP-9, we detected by the Evan's blue extravasation method a clear increase of BBB permeability. Tissue inhibitor of metalloproteinase-1 was not modified during permanent ischemia at any time. The ProMMP-2 was significantly (P < 0.05) increased only after 24 hours of permanent ischemia (mean 213.2 AU, SD +/- 60.6 versus mean 94.6 AU, SD +/- 13.3), and no activated form was observed. The appearance of activated MMP-9 after 4 hours of ischemia in correlation with BBB permeability alterations suggests that MMP-9 may play an active role in early vasogenic edema development after stroke.

Animals↗

PubMed1999

Copper-zinc superoxide dismutase prevents the early decrease of apurinic/apyrimidinic endonuclease and subsequent DNA fragmentation after transient focal cerebral ischemia in mice.

BACKGROUND AND PURPOSE: DNA damage and its repair mechanism are thought to be involved in ischemia/reperfusion injury in the brain. We have previously shown that apurinic/apyrimidinic endonuclease (APE/Ref-1), a multifunctional protein in the DNA base excision repair pathway, rapidly decreased after transient focal cerebral ischemia (FCI) before the peak of DNA fragmentation. To further investigate the role of reactive oxygen species in APE/Ref-1 expression in vivo, we examined the expression of APE/Ref-1 and DNA damage after FCI in wild-type and transgenic mice overexpressing copper-zinc superoxide dismutase. METHODS: Transgenic mice overexpressing copper-zinc superoxide dismutase and wild-type littermates were subjected to 60 minutes of transient FCI by intraluminal blockade of the middle cerebral artery. APE/Ref-1 protein expression was analyzed by immunohistochemistry and Western blot analysis. DNA damage was evaluated by gel electrophoresis and terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end-labeling (TUNEL). RESULTS: A similar level of APE/Ref-1 was detected in the control brains from both groups. APE/Ref-1 was significantly reduced 1 hour after transient FCI in both groups, whereas the transgenic mice had less reduction than that seen in wild-type mice 1 and 4 hours after FCI. DNA laddering was detected 24 hours after FCI and was decreased in transgenic mice. Double staining with APE/Ref-1 and TUNEL showed that the neurons that lost APE/Ref-1 immunoreactivity became TUNEL positive. CONCLUSIONS: These results suggest that reactive oxygen species contribute to the early decrease of APE/Ref-1 and thereby exacerbate DNA fragmentation after transient FCI in mice.

Animals↗

PubMed1999

Early decrease of XRCC1, a DNA base excision repair protein, may contribute to DNA fragmentation after transient focal cerebral ischemia in mice.

BACKGROUND AND PURPOSE: DNA damage and the DNA repair mechanism are known to be involved in ischemia/reperfusion injury in the brain. The x-ray repair cross-complementing group 1 (XRCC1) protein plays a central role in the DNA base excision repair pathway by interacting with DNA ligase III and DNA polymerase beta. The present study examined the protein expression of XRCC1 and DNA fragmentation before and after transient focal cerebral ischemia (FCI). METHODS: Adult male CD-1 mice were subjected to 60 minutes of FCI by intraluminal blockade of the middle cerebral artery. XRCC1 protein expression was analyzed by immunohistochemistry and Western blot analysis. DNA damage was evaluated by gel electrophoresis and terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end-labeling (TUNEL). The spatial relationship between XRCC1 expression and DNA damage was examined by double staining with XRCC1 and TUNEL after FCI. RESULTS: Immunohistochemistry showed the nuclear expression of XRCC1 in all regions of the control brains and that it was predominant in the hippocampus. The XRCC1 level was markedly reduced in the caudate putamen at 10 minutes, further decreased in the entire middle cerebral artery territory at 1 hour, and remained reduced until 4 and 24 hours after FCI. Western blot analysis of the normal control brain showed a characteristic band of 70 kDa, which decreased after FCI. A significant amount of DNA fragmentation was detected by DNA gel electrophoresis 24 hours but not 4 hours after FCI. Double staining showed that the neurons that lost XRCC1 immunoreactivity became TUNEL positive. CONCLUSIONS: These results suggest that the early decrease of XRCC1 and the failure of the DNA repair mechanism may contribute, at least in part, to DNA fragmentation after FCI.

Animals↗

PubMed1999

Reduction of apurinic/apyrimidinic endonuclease expression after transient global cerebral ischemia in rats: implication of the failure of DNA repair in neuronal apoptosis.

BACKGROUND AND PURPOSE: To clarify the relationship between apurinic/apyrimidinic endonuclease (APE/Ref-1), a multifunctional protein in the DNA base excision repair pathway, and delayed neuronal cell death associated with apoptosis, we examined the expression of APE/Ref-1 before and after transient global ischemia in rats. METHODS: Global ischemia was induced by bilateral common carotid artery occlusion and hypotension. Expression of the APE/Ref-1 protein was evaluated by Western blot and immunohistochemical analyses. Apoptosis after global ischemia was observed by DNA electrophoresis and terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end labeling (TUNEL) staining. RESULTS: Immunohistochemistry showed the nuclear expression of APE/Ref-1 in the control brains. Nuclear immunoreactivity of APE/Ref-1 was significantly decreased 2 days after 10 minutes of ischemia in the hippocampal CA1 subregion. Western blot analysis of a sample from the normal brains showed a characteristic 37-kDa band, which was reduced in the hippocampal CA1 subregion after ischemia. A significant amount of DNA fragmentation was observed at 3 days but not at 1 day after ischemia. Double staining with APE/Ref-1 and TUNEL clearly showed that the neurons that lost APE/Ref-1 immunoreactivity became TUNEL positive. CONCLUSIONS: Our data provide evidence that APE/Ref-1 decreased in hippocampal CA1 neurons after transient global ischemia and that this reduction precedes DNA fragmentation, which is destined to cause apoptosis. Our results suggest the possibility that a decrease of APE/Ref-1 activity and the failure of DNA repair may underlie the mechanism of apoptosis after transient focal ischemia.

Animals↗