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D K Das

Publications and source records attributed to D K Das.

At least 73 records · Page 4Linked to original sources
PubMed1999

Cardioprotection of red wine: role of polyphenolic antioxidants.

Epidemiological studies suggest that the consumption of wine, particularly of red wine, reduces the incidence of mortality and morbidity from coronary heart disease. This has given rise to what is now popularly termed the "French paradox". The cardioprotective effect has been attributed to antioxidants present in the polyphenol fraction of red wine. Grapes contain a variety of antioxidants, including resveratrol, catechin, epicatechin and proanthocyanidins. Of these, resveratrol is present mainly in grape skin while proanthocyanidin is present in the seeds. In this report, we provide evidence that red wine extract as well as resveratrol and proanthocyanidins are equally effective in reducing myocardial ischemic reperfusion injury, which suggests that these red wine polyphenolic antioxidants play a crucial role in cardioprotection.

Antioxidants↗
PubMed1999

Reactive oxygen species function as second messenger during ischemic preconditioning of heart.

Ischemic preconditioning has been shown to trigger a signaling pathway by potentiating tyrosine kinase phosphorylation leading to the activation of p38 MAP kinase and MAPKAP kinase 2. Recently, the nuclear transcription factor, NFkappaB, was found to play a role in the signaling process. Since NFkappaB is a target of oxygen free radicals, we hypothesized that reactive oxygen species might play a role in the signaling process. To test this hypothesis, isolated rat hearts were perfused in the absence or presence of either dimethyl thiourea (DMTU), a OH* radical scavenger, or SN 50 peptide, a NFkappaB blocker. Hearts were then subjected to ischemic preconditioning by four repeated episodes of 5 min ischemia each followed by 10 min reperfusion. All hearts were then made globally ischemic for 30 min followed by 2 h of reperfusion. The results of our study demonstrated enhanced tyrosine kinase phosphorylation during ischemic preconditioning which was blocked by DMTU. DMTU also inhibited preconditioning mediated increased phosphorylation of p38 MAP kinase and MAPKAP kinase 2 activity. However, DMTU had no effect on the translocation and activation of protein kinase C (PKC) resulting from preconditioning. Preconditioning reduced myocardial infarct size as expected. This cardioprotective effect of preconditioning was abolished by both DMTU and SN 50. Preconditioning resulted in the nuclear translocation and activation of NFkappaB. Increased NFkappaB binding was blocked by both DMTU and SN 50. The results of this study demonstrate that reactive oxygen species play a crucial role in signal transduction mediated by preconditioning. This signaling process appears to be potentiated by tyrosine kinase phosphorylation resulting in the activation of p38 MAP kinase and MAPKAP kinase 2 leading to the activation of NFkappaB suggesting a role of oxygen free radicals as second messenger. Free radical signaling seems to be independent of PKC although PKC is activated during preconditioning process suggesting the role of two separate signaling pathways in ischemic preconditioning.

Adaptation, Physiological↗
PubMed1998

Coordinated role of vasoactive intestinal peptide and nitric oxide in cardioprotection.

The present study sought to examine the interrelationship between nitric oxide (NO) and vasoactive intestinal peptide (VIP) in myocardial protection. Isolated rat hearts were perfused for 15 min with buffer only (Group I); 0.3 mM VIP (Group II); 3 mM L-arginine (a precursor of NO) (Group III); VIP and aminoguanidine (iNOS blocker) (Group IV); or L-arginine plus VIP 10-28 (VIP inhibitor) (Group V). Each heart was then made globally ischemic for 30 min followed by 2 h reperfusion. Both VIP and NO were found to provide cardioprotection during ischemia and reperfusion. However, the beneficial effects of VIP and NO were reduced by inhibition of NO and VIP, respectively, suggesting that cardioprotection by VIP is modulated by NO and vice versa. The results of this study suggested a coordinated regulation by cardioprotection by NO and VIP.

Animals↗
PubMed1998

An essential role of NFkappaB in tyrosine kinase signaling of p38 MAP kinase regulation of myocardial adaptation to ischemia.

We have recently demonstrated that myocardial adaptation to ischemia triggers a tyrosine kinase regulated signaling pathway leading to the translocation and activation of p38 MAP kinase and MAPKAP kinase 2. Since oxidative stress is developed during ischemic adaptation and since free radicals have recently been shown to function as an intracellular signaling agent leading to the activation of nuclear transcription factor, NFkappaB, we examined whether NFkappaB was involved in the ischemic adaptation process. Isolated perfused rat hearts were adapted to ischemic stress by repeated ischemia and reperfusion. Hearts were pretreated with genistein to block tyrosine kinase while SB 203580 was used to inhibit p38 MAP kinases. Ischemic adaptation was associated with the nuclear translocation and activation of NFkappaB which was significantly blocked by both genistein and SB 203580. The ischemically adapted hearts were more resistant to ischemic reperfusion injury as evidenced by better function recovery and less tissue injury during post-ischemic reperfusion. Ischemic adaptation developed oxidative stress which was reflected by increased malonaldehyde formation. A synthetic peptide containing a cell membrane-permeable motif and nuclear sequence, SN 50, which blocked nuclear translocation of NFkappaB during ischemic adaptation, significantly inhibited the beneficial effects of adaptation on functional recovery and tissue injury. In concert, SN 50 reduced the oxidative stress developed in the adapted myocardium. These results demonstrate that p38 MAP kinase might be upstream of NFkappaB which plays a role in ischemic preconditioning of heart.

Adaptation, Physiological↗
PubMed1998

Oxidative stress developed during the reperfusion of ischemic myocardium induces apoptosis.

Apoptosis or programmed cell death is a genetically controlled response for cells to commit suicide and, is associated with DNA fragmentation or laddering. The common inducers of apoptosis include oxygen free radicals/oxidative stress and Ca2+ which are also implicated in the pathogenesis of myocardial ischemic reperfusion injury. To examine whether ischemic reperfusion injury is mediated by apoptotic cell death, isolated perfused rat hearts were subjected to 15, 30 or 60 min of ischemia as well as 15 min of ischemia followed by 30, 60 or 120 min of reperfusion. At the end of each experiment, hearts were processed for the evaluation of apoptosis, DNA laddering. Apoptosis was studied by visualizing the apoptotic cardiomyocytes by direct fluorescence detection of digoxigenin-labeled genomic DNA using APOPTAG in situ apoptosis detection kit. DNA laddering was evaluated by subjecting the DNA obtained from the hearts to 1.8% agarose gel electrophoresis and photographed under UV illumination. The results of our study revealed apoptotic cells only in the 60 and 120 min reperfused hearts as demonstrated by the intense fluorescence of the immunostained digoxigenin-labeled genomic DNA when observed under fluorescence microscopy. None of the ischemic hearts showed any evidence of apoptosis. These results corroborated with the findings of DNA fragmentation which showed increased ladders of DNA bands in the same reperfused hearts representing integer multiples of the intenucleosomal DNA length (about 180 bp). The presence of apoptotic cells and DNA fragmentation in the myocardium were abolished by preperfusing the hearts in the presence of ebselen, which also removed the oxidative stress developed in the heart. Taken together, these results clearly demonstrate that oxidative stress developed in the ischemic reperfused myocardium induces apoptosis.

Animals↗
PubMed1998

High-performance liquid chromatographic peak identification of 2,4-dinitrophenylhydrazine derivatives of lipid peroxidation aldehydes by photodiode array detection.

Malonaldehyde (MDA), a product of lipid peroxidation, is a presumptive marker for the development of oxidative stress in tissues and plasmas. In this study we report the photodiode array detection of the 2,4-dinitrophenylhydrazine (DNPH) derivatives of MDA using HPLC. Oxidative stress was produced by injecting (i.p.) bacterial lipopolysaccharide (LPS) into rats at a dose of 100 micrograms/kg, or i.v. into rabbits (1 microgram/kg), or added to freshly drawn human blood (200 ng/ml). Blood was collected at several time points up to 5 h, centrifuged, and equal volumes of 20% TCA were used to precipitate proteins from the plasma. The supernatants were derivatized with DNPH, and the aldehyde-DNPHs were extracted with pentane. After evaporation, aliquots of 10 microliters in acetonitrile were injected onto a Beckman Ultrasphere C18 (3 microns) column, chromatographed with an acetonitrile-water-acetic acid gradient mobile phase and scanned using Waters 996 photodiode array detector. Peak identification and homogeneity was determined by comparing the experimental peaks and UV scans with those of authentic standards. A significant increase in the DNPH derivative of malonaldehyde (MDA-DNPH), but not of the other aldehyde-DNPH derivatives of formaldehyde (FDA), acetaldehyde (ADA), acetone and propionaldehyde (PDA) was seen over the first hour after LPS administration in anesthetized rats, while in conscious rabbits this trend lasted up to 3 h. The retention times as well as the UV scans of the derivatized aldehydes matched the authentic standards. Thus, photodiode array detection has proved valuable in establishing this HPLC method for estimating oxidative stress. This technique could accurately measure pmol amounts of MDA-DNPH indicating the usefulness of photodiode array detection method for estimating small changes in the oxidative stress.

Animals↗
PubMed1998

Ultrasound-guided fine-needle aspiration cytology diagnosis of gallbladder lesions: a study of 82 cases.

Carcinoma of the gallbladder (GB) is among the five most common forms of cancers and tops the list of gastrointestinal malignancies in females of the Delhi region. Lack of specific signs and symptoms prevents early detection of GB carcinoma. However, in recent years ultrasonographically (US)-guided fine-needle aspiration (FNA) cytology has been found to be a reliable procedure for its diagnosis. The present study was carried out during a period of 5 yr (1986-1990) in 64 female and 18 male patients to find out the diagnostic utility of US-guided FNA cytology in gallbladder lesions. Ultrasonography in these 82 cases revealed a mass in the gallbladder/GB area in 74 (90.2%), a mass in the GB/pancreas in 1 (1.2%), gallstones in 32 (39.0%), and miscellaneous gallbladder lesions in 4 (4.9%). The other findings included space-occupying lesions in liver in 18 (22.0%), portal lymphadenopathy in 12 (14.6%), and infiltration in other organs in 7 (8.5%). The initial cytodiagnosis was malignancy in 48 cases, inflammatory in 12, and inadequate in 22. Following review of the smears by one of the investigators (D.K.D.), the number of malignant cases remained 48 (58.5%). There were 10 (12.2%) inflammatory and 24 (29.3%) inadequate cases. Adenocarcinoma was the most common malignancy (83.3%), followed by squamous-cell and adenosquamous carcinoma (12.5%) and small round cell tumors (4.2%). The 10 inflammatory lesions showed slight (+) to excessive (+3) neutrophilic infiltration and included one case each of xanthogranulomatous cholecystitis and a necrotizing granulomatous lesion likely to be of tuberculous etiology.

Adenocarcinoma↗
PubMed1998

Detection of oxidative DNA damage to ischemic reperfused rat hearts by 8-hydroxydeoxyguanosine formation.

Reactive oxygen species that are generated in the ischemic heart upon reperfusion, play a significant role in the pathogenesis of reperfusion injury. Although DNA is a well known target for free radical attack, little attention has been paid to the injury of DNA molecules associated with ischemia and reperfusion. In this study, the formation of 8-hydroxydeoxyguanosine (8-OHDG), a product of hydroxyl radical (OH.)-DNA interaction, was monitored in the post-ischemic myocardium. A simple high performance liquid chromatography (HPLC), with uv detection, detected pmol levels of 8-OHDG in the pre-ischemic heart which increased steadily and progressively as a function of reperfusion time. A similar rise in 8-OHDG was noticed when isolated hearts were perfused with a OH. -generating system. Corroborating with the increased 8-OHDG formation, increased amount of creatine kinase was released from the coronary effluent indicating increased tissue injury. The formation of 8-OHDG was completely blocked when hearts were preperfused with oxygen-free-radical scavenger, 1,3-dimethyl-2-thiourea (DMTU) which also significantly reduced the appearance of CK in the coronary effluent, suggesting that oxidative DNA damage play a role in the pathophysiology of ischemic reperfusion injury.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1998

The role of heme propionate in controlling the redox potential of heme: square wave voltammetry of protoporphyrinato IX iron (III) in aqueous surfactant micelles.

The proton equilibrium in aqueous surfactant solutions of hemin, (PPIX)Fe(H2O)+2, involving the propionic acid groups is reported (PPIX = protoporphyrinato IX). The surfactant used are sodium dodecyl sulphate (SDS), triton X-100 (TX-100) and hexadecyl trimethylammonium bromide (CTAB). The pKa values, determined spectroscopically from the pH variations of the Soret absorption, are found at ca. 3.5 in SDS and TX-100 micelles. In the cationic surfactant (CTAB) the absorbance of hemin is independent of the hydrogen ion concentration in solution and this surfactant form a salt-link with the heme propionate group. The dependence of mid-point potential on pH indicates that the heme propionate undergoes a redox-linked changed in pKa from 3.2-3.5 in the ferric form to 4.4-4.5 in the ferrous form (27 degrees C, mu = 0.2 M). The change in the mid-point potential per unit change in pH. delta E/delta pH, is ca. -59 mV. Replacement of the water molecules in hemin by tetrahydrofuran, (PPIX)Fe(thf)+2, induces a considerable shift of pKa of the heme propionate: 6.2-6.8 in the ferric form and 7.5-7.8 in the ferrous form. The lower value of pKa in diaquo hemin is attributed to stabilisation of the propionate by a hydrogen bond with coordinated water molecules. All these results may be interpreted in terms of the involvement of a heme propionate group in redox linked uptake of protons and the influence of hydrogen bonding and salt-link formation on the pKa of the proton equilibrium.

Cetrimonium↗
PubMed1998

HPTLC analysis of sphingomylein, ceramide and sphingosine in ischemic/reperfused rat heart.

Since the sphingomylein-ceramide-sphingosine pathway, especially ceramide, has been shown to induce programmed cell death (apoptosis), and since apoptosis may be involved with ischemic/reperfused injury in the heart, it became desirable to quantitate the three components in ischemic/reperfused rat heart. One group of rat hearts (n = 6) was isolated and perfused with Krebs-Henseleit buffer using the Langendorff non-recirculating mode. The hearts were perfused for 10 min, made ischemic for 30 min and reperfused for 120 min. Hearts were collected and stored at - 70 degrees C before ischemia, after ischemia and after 30, 60 and 120 min of reperfusion. The hearts were homogenized, and lipids were extracted using the Folch method. The lipids were then chromatographed on Whatman silica gel 60 A high-performance thin-layered chromatography (HPTLC) plates. The plates were developed with iodine, photographed using Photoshop software and quantitated using NIH Imaging software. The results show a 50% decrease of sphingomylein during reperfusion with a corresponding increase in ceramide with sphingosine showing a smaller decrease as compared with the ceramide increase.

Animals↗
PubMed1998

Redistribution of phosphatidylethanolamine and phosphatidylserine precedes reperfusion-induced apoptosis.

Although cardiomyocyte death and infarction associated with ischemia-reperfusion are traditionally believed to be induced via necrosis, recent studies implicated apoptotic cell death in ischemic reperfused tissue. To examine whether myocardial ischemic reperfusion injury is mediated by apoptotic cell death, isolated perfused rat hearts were subjected to 15 and 30 min of ischemia as well as 15 min of ischemia followed by 30, 90, or 120 min of reperfusion. At the end of each experiment, hearts were processed for the evaluation of apoptosis and DNA laddering. Apoptosis was studied by visualizing the apoptotic cardiomyocytes by direct fluorescence detection of digoxigenin-labeled genomic DNA using APOPTAG in situ apoptosis detection kit. DNA laddering was evaluated by subjecting the DNA obtained from cardiomyocytes to 1.8% agarose gel electrophoresis and photographed under ultraviolet illumination. In addition, high-performance thin-layer chromatography (HPTLC) of aminophospholipids labeled with 2,4,6-trinitrobenzenesulfonate was performed to evaluate phospholipid topography in cardiomyocytes. The results of our study revealed apoptotic cells only in the 90- and 120-min reperfused hearts as demonstrated by the intense fluorescence of the immunostained digoxigenin-labeled genomic DNA when observed under fluorescence microscope. None of the ischemic hearts showed any evidence of apoptosis. These results corroborated with the findings of DNA fragmentation that showed increased ladders of DNA bands in the 120-min reperfused hearts, representing integer multiples of the internucleosomal DNA length (approximately 180 bp). Two-dimensional HPTLC of the phospholipids obtained from the cardiomyocytes and transbilayer organization of the phosphatidylethanolamine (PE) and phosphatidylserine (PS) in the myocytes indicated translocation of both PE and PS from the inner leaflet to the outer leaflet of the membrane as early as after 20 min of ischemia. These results demonstrate that the redistribution of PS and PE precedes the apototic cell death and DNA fragmentation associated with the reperfusion of ischemic myocardium, suggesting that ischemia may trigger the signal for apoptosis although it becomes evident during reperfusion.

Animals↗
PubMed1998

Ischemic preconditioning triggers tyrosine kinase signaling: a potential role for MAPKAP kinase 2.

Myocardial adaptation to ischemia has been shown to activate protein tyrosine kinase, potentiating activation of phospholipase D, which leads to the stimulation of mitogen-activated protein (MAP) kinases and MAP kinase-activated protein (MAPKAP) kinase 2. The present study sought to further examine the signal transduction pathway for the MAPKAP kinase 2 activation during ischemic adaptation. Isolated perfused rat hearts were adapted to ischemic stress by repeated ischemia and reperfusion. Hearts were pretreated with genistein to block tyrosine kinase, whereas SB-203580 was used to inhibit p38 MAP kinases. Western blot analysis demonstrated that p38 MAP kinase is phosphorylated during ischemic stress adaptation. Phosphorylation of p38 MAP kinase was blocked by genistein, suggesting that activation of p38 MAP kinase during ischemic adaptation is mediated by a tyrosine kinase signaling pathway. MAPKAP kinase 2 was estimated by following in vitro phosphorylation with recombinant human heat shock protein 27 as specific substrate for MAPKAP kinase 2. Again, both genistein and SB-203580 blocked the activation of MAPKAP kinase 2 during myocardial adaptation to ischemia. Immunofluorescence microscopy with anti-p38-antibody revealed that p38 MAP kinase is primarily localized in perinuclear regions. p38 MAP kinase moves to the nucleus after ischemic stress adaptation. After ischemia and reperfusion, cytoplasmic striations in the myocytes become obvious, indicating translocation of p38 MAP kinase from nucleus to cytoplasm. Corroborating these results, myocardial adaptation to ischemia improved the left ventricular functions and reduced myocardial infarction that were reversed by blocking either tyrosine kinase or p38 MAP kinase. These results demonstrate that myocardial adaptation to ischemia triggers a tyrosine kinase-regulated signaling pathway, leading to the translocation and activation of p38 MAP kinase and implicating a role for MAPKAP kinase 2.

Adaptation, Biological↗
PubMed1998

Preconditioning of rat heart with monophosphoryl lipid A: a role for nitric oxide.

Preconditioning with monophosphoryl lipid A (MLA) protects rabbit hearts from prolonged ischemic reperfusion injury by a mechanism involving inducible nitric oxide synthase (iNOS) activation. This study was undertaken to determine whether MLA also could precondition rat hearts in a similar manner. Rats were injected with two different doses of MLA (300 microg/kg or 450 microg/kg i.v.) or vehicle (control), and after 24 hr the animals were sacrificed for preparation of isolated perfused rat hearts. Hearts were then perfused by working mode, and then made ischemic for 30 min followed by 30 min of reperfusion. Another group of hearts were treated simultaneously with a nitric oxide (NO) blocker, L-nitro-arginine-methyl-ester (L-NAME) (10 mg/kg) and MLA (450 microg/kg). For arrhythmia studies, 12 hearts were used in each group (total, 48 hearts). Cardiac functions were examined in a separate group of 24 hearts (n = 6/group). MLA-treated hearts (either dose) were tolerant to ischemic reperfusion injury as evidenced by improved postischemic ventricular recovery [coronary flow (ml/min) 19.1 +/- 0.8 (300 microg/kg MLA), 22.6 +/- 1.0 (450 microg/kg MLA) vs. 15.9 +/- 0.7 (control); aortic flow (ml/min) 20.7 +/- 1.8 (300 microg/kg MLA), 25.8 +/- 1.4 (450 microg/kg MLA) vs. 11. 0 +/- 0.8 (control); left ventricular developed pressure (kPa) 13.3 +/- 0.6 (300 microg/kg MLA), 14.6 +/- 0.2 (450 microg/kg MLA) vs. 10. 3 +/- 0.7 (control)]. Incidences of ventricular fibrillation and ventricular tachycardia were decreased compared with the control group only in the 450 microg/kg dose of MLA-treated hearts (92% to 33%). Pretreatment of the hearts with L-NAME inhibited the preconditioning effect of MLA. To examine the induction of the iNOS expression, RNAs were extracted from the control and MLA-treated hearts (after 2, 4,6, 8, 12 and 24 hr of treatment) and Northern blot analyses were performed with a specific cDNA probe for iNOS. A single band of approximately 4.6 kb corresponding to iNOS mRNA was detected after 4 hr of MLA treatment, whereas the maximal iNOS expression was found between 6 and 8 hr of MLA treatment. The results of this study demonstrated that MLA induced the expression of iNOS and protected the myocardium from ischemic reperfusion injury which is blocked by an inhibitor of NO synthesis, which suggests a role of NO in MLA-mediated cardioprotection.

Adjuvants, Immunologic↗
PubMed1998

Ischemic preconditioning attenuates apoptotic cell death associated with ischemia/reperfusion.

Apoptosis or programmed cell death is a genetically controlled response for cells to commit suicide and is associated with DNA fragmentation or laddering. The common inducers of apoptosis include oxygen free radicals/oxidative stress and Ca2+ which are also implicated in the pathogenesis of myocardial ischemic reperfusion injury. To examine whether ischemic reperfusion injury is mediated by apoptotic cell death, isolated perfused rat hearts were subjected to 15, 30 or 60 min of ischemia as well as 15 min of ischemia followed by 30, 60, 90 or 120 min of reperfusion. At the end of each experiment, the heart was processed for the evaluation of apoptosis and DNA laddering. Apoptosis was studied by visualizing the apoptotic cardiomyocytes by direct fluorescence detection of digoxigenin-labeled genomic DNA using APOPTAG in situ apoptosis detection kit. DNA laddering was evaluated by subjecting the DNA obtained from the hearts to 1.8% agarose gel electrophoresis and photographed under UV illumination. The results of our study revealed apoptotic cells only in the 90 and 120 min reperfused hearts as demonstrated by the intense fluorescence of the immunostained digoxigenin-labeled genomic DNA when observed under fluorescence microscopy. None of the ischemic hearts showed any evidence of apoptosis. These results were corroborated with the findings of DNA fragmentation which showed increased ladders of DNA bands in the same reperfused hearts representing integer multiples of the internucleosomal DNA length (about 180 bp). The presence of apoptotic cells and DNA fragmentation in the myocardium were completely abolished by subjecting the myocardium to repeated short-term ischemia and reperfusion which also reduced the ischemic reperfusion injury as evidenced by better recovery of left ventricular performance in the preconditioned myocardium. The results of this study indicate that reperfusion of ischemic heart, but not ischemia, induces apoptotic cell death and DNA fragmentation which can be inhibited by myocardial adaptation to ischemia.

Adaptation, Physiological↗
PubMed1998

Induction of iNOS gene expression by monophosphoryl lipid A: a pharmacological approach for myocardial adaptation to ischemia.

Using the concept that exposing a cell to an adverse environment (stress) results in the stimulation of its endogenous defense system, hearts have been adapted to ischemia by exposing them to diverse stresses. Recently, 24-h pretreatment of monophosphoryl lipid A (MLA), a chemically modified derivative of endotoxin, was found to render the hearts more tolerant to ischemic reperfusion injury. Since nitric oxide has recently been implicated in myocardial preservation and since inducible nitric oxide synthetase (iNOS) was originally characterized in macrophages and shown to be maximally induced by bacterial lipopolysaccharides (endotoxin), we sought to determine whether MLA mediates its cardioprotective effects through the iNOS expression. For this, rats were injected with MLA (300 micrograms/kg) or vehicle (control), and after 24 h the animals were sacrificed and the isolated working hearts were made ischemic for 30 min followed by 30 min of reperfusion. MLA-treated hearts were found to be tolerant to ischemic reperfusion injury as evidenced by improved postischemic ventricular recovery. After 30 min of reperfusion, left ventricular developed pressure (LVDP) and its maximum first derivative (LVmaxdp/dt) were 13.3 +/- 0.3 kPa and 537 +/- 13 kPa/s, respectively, in the MLA-treated group, as compared with 10.2 +/- 0.4 kPa (p < 0.05) and 447 +/- 11 kPa/s (p < 0.05), respectively, for the control group. Aortic flow and coronary flow were 20.1 +/- 1.4 ml/min and 19.1 +/- 0.8 ml/min, respectively, in the MLA group, as compared with 9.5 +/- 0.8 ml/min (p < 0.05) and 15.9 +/- 0.7 ml/min (p < 0.05), respectively, for the untreated group. To examine the induction of the iNOS expression, RNAs were extracted from the control and MLA-treated hearts (after 2, 4, 6, 8, 12 and 24 h of treatment) and Northern blot analysis was performed using specific cDNA probe for iNOS. A single band of approximately 4.6 kb corresponding to iNOS mRNA was detected after 4 h of MLA treatment, while the maximal iNOS expression was found between 6-8 h of MLA treatment. The results of this study demonstrate that MLA induces the expression of iNOS and protects the myocardium from ischemic reperfusion injury.

Animals↗
PubMed1997

Glutathione peroxidase knockout mice are susceptible to myocardial ischemia reperfusion injury.

BACKGROUND: To test our hypothesis that intracellular antioxidant enzymes constitute a cellular defense against acute stress, we studied myocardial ischemia reperfusion injury in the setting of reduced level of glutathione peroxidase using GSHPx-1 gene knockout mice. METHODS: Knockout mice were developed by disrupting the coding sequence of GSHPx-1 gene after inserting a neomycin resistance gene derived from pMCIpol A into the EcoRI site located in exon 2. Isolated perfused hearts were prepared from two groups of mice-knockout and nontransgenic controls. A 4-0 silk was attached to the apex of the heart which in turn was attached to a force transducer. Hearts were perfused by the Langendorff mode, and after 20 minutes of stabilization subjected to 30 minutes of ischemia followed by 2 hours of reperfusion. The force developed by the heart (DF) and the first derivative of DF (dF/dt) were recorded. Creatine kinase (CK) release was measured in the perfusate and the infarct size was measured at the end of each experiment. RESULTS: For both GSHPx-1 knockout and nontransgenic control groups, DF and dF/dt were significantly lower during early postischemic reperfusion compared with baseline, but these values were significantly higher for the control group than the knockout mice throughout most of the reperfusion period. CK release from the heart increased during reperfusion for both groups, but this increase was significantly lower for the control group. The infarct size was also smaller for the control mice as compared with knockouts. CONCLUSIONS: The results indicate that the knockout mice are more susceptible to ischemia reperfusion injury, suggesting the importance of GSHPx-1 gene in myocardial protection from ischemic reperfusion injury.

Animals↗