Biomedical subjects

R Mukherjee

Publications and source records attributed to R Mukherjee.

At least 127 records · Page 7Linked to original sources

PubMed1996

Direct and interactive effects of cardioplegic arrest and protamine on myocyte contractility.

BACKGROUND: Cardioplegic arrest with rewarming and protamine administration have been implicated in causing transient left ventricular dysfunction perioperatively. However, whether interactive effects between cardioplegic arrest and rewarming with protamine occur with respect to myocyte contractile processes remains unclear. Accordingly, using an isolated myocyte model, the present study tested the hypothesis that simulated cardioplegic arrest with rewarming and protamine would have direct and interactive effects on myocyte contractile function. METHODS: Left ventricular isolated myocyte contractile function was examined using computer-aided videomicroscopy under normothermic conditions (37 degrees C, cell medium; n = 183) and after simulated hypothermic, hyperkalemic cardioplegic arrest with rewarming (4 degrees C, 24 mEq/L K+, 2 hours; then 37 degrees C, cell medium, 5 minutes; n = 268). Myocyte function was then examined in the presence of protamine (10 to 40 micrograms/mL) under normothermic conditions (n = 102) and after cardioplegic arrest with rewarming (n = 175). RESULTS: Myocyte contractile function decreased by 43% from baseline after simulated cardioplegic arrest with rewarming. Under normothermic conditions, protamine (20 micrograms/mL) reduced myocyte contractile function by 43.9% +/- 4.3%, whereas myocyte contractile function decreased by only 31.1% +/- 2.7% with protamine (20 micrograms/mL) after cardioplegic arrest with rewarming. Thus, the negative effects of protamine on myocyte contractility were attenuated after cardioplegic arrest when compared with normothermic conditions. CONCLUSIONS: The present study demonstrated that simulated cardioplegic arrest with rewarming and protamine have direct and interactive effects on myocyte contractile function, which are not additive or synergistic.

Animals↗

PubMed1996

Right and left ventricular geometry and myocyte contractile processes with dilated cardiomyopathy: myocyte growth and beta-adrenergic responsiveness.

OBJECTIVES: Comparison of the effects of supraventricular tachycardia-induced dilated cardiomyopathy on left and right ventricular isolated myocyte geometry and function. BACKGROUND: Chronic ventricular tachycardia and supraventricular tachycardia cause left ventricular dilation and dysfunction in humans. However, it is unknown whether supraventricular tachycardia-induced dilated cardiomyopathy is a homogenous process for both the left and right ventricles. METHODS: Dilated cardiomyopathy was induced by rapid atrial pacing (240 beats/min, 3 weeks) in 5 pigs. Five age- and weight-matched pigs served as controls. Ventricular mass was measured, myocyte dimensions were obtained, and isolated right and left ventricular myocyte contractile performance was evaluated at baseline and after beta-adrenergic receptor stimulation. RESULTS: With the development of dilated cardiomyopathy, there was no change in left ventricular mass. In contrast, right ventricular mass was increased, as was right ventricular myocyte cross-sectional area. In the control group, baseline right ventricular myocyte contractile function was increased compared to left ventricular myocytes. beta-adrenergic receptor stimulation increased myocyte contractile function in both left and right ventricular myocytes. With supraventricular tachycardia-induced cardiomyopathy, both left and right ventricular myocyte contractile function and beta-adrenergic responsiveness were reduced. CONCLUSIONS: This study demonstrated differences in left and right ventricular myocyte growth with supraventricular tachycardia-induced dilated cardiomyopathy and this differential growth response was associated with changes in contractile performance. Thus, in this model of cardiomyopathic disease, left and right ventricular growth and changes in contractile performance are not a homogenous process.

Adrenergic beta-Agonists↗

PubMed1996

Veno-occlusive disease of the small bowel. An entity in search of identity.

Veno-occlusive disease of the bowel is a rare entity, and, to our knowledge, only two studies have been reported in the literature; these reports describe 11 patients with veno-occlusive disease of the small and large bowels. We describe a patient who developed veno-occlusive disease of the small bowel, whose histologic findings were somewhat different from those previously reported.

Adult↗

PubMed1995

Direct effects of protamine sulfate on myocyte contractile processes. Cellular and molecular mechanisms.

BACKGROUND: Administration of the arginine-rich, highly charged protamine (PROT) molecule has been associated with episodes of acute left ventricular (LV) dysfunction. The objective of the present study was to test the hypothesis that PROT has direct effects on isolated LV myocyte contractile processes and sarcolemmal transduction systems. METHODS AND RESULTS: Exposure of porcine LV myocytes (n = 305) to 40 micrograms/mL PROT (reflecting a dose of 2.5 mg/kg) decreased basal contractile function and beta-adrenergic responsiveness. For example, myocyte percent shortening was 4.3 +/- 0.1% in control myocytes and decreased to 2.8 +/- 0.2% in the presence of 40 micrograms/mL PROT (P < .05). Myocyte percent shortening was 9.3 +/- 0.7% after beta-adrenergic receptor stimulation (isoproterenol; 25 nmol/L) and was significantly reduced in the presence of 40 micrograms/mL PROT (5.7 +/- 0.7%, P < .05). PROT reduced myocyte responsiveness to forskolin (100 mumol/L), which directly activates adenylate cyclase, by > 40% from forskolin. In addition, PROT abolished the inotropic effects of ouabain on myocyte contractile function. To determine contributory mechanisms for the effects of PROT on myocyte sarcolemmal systems, beta-receptor- and cardiac glycoside-binding characteristics were determined in sarcolemmal preparations. beta-receptor binding was 175 +/- 10 fmol/mg and was reduced to 140 +/- 6 fmol/mg in the presence of PROT (P < .05). Ouabain receptor binding was 7.1 pmol/mg and decreased to 2.6 +/- 0.4 pmol/mg in the presence of PROT. In addition, cAMP production after stimulation with isoproterenol and forskolin was significantly blunted in the presence of PROT. Variants of the PROT moelcule were constructed by specific amino acid substitutions and deletions, which provided a means to vary charge as well as structure. Substitution of arginine with lysine in the PROT peptide sequence ameliorated the negative effects on myocyte contractile processes; despite identical overall charge (21+). However, a PROT variant with an 18+ charge but different amino acid sequence induced significant negative effects on myocyte function and inotropic responsiveness. Thus, the effects of PROT on myocyte contractile processes are not due simply to the high positive charge of the molecule. To further establish that PROT can contribute to changes in LV function in the clinical setting, fluorescein-labeled PROT was circulated in antegradely perfused rabbit hearts. Microscopic examination revealed that PROT could traverse the vascular compartment of the myocardium and come in direct contact with the myocyte. CONCLUSIONS: The unique findings from the present study suggest that a fundamental contributory mechanisms for the changes in LV function observed after protamine administration may be the direct effect of unbound protamine on myocyte contractile processes.

Adrenergic beta-Agonists↗

PubMed1995

Angiotensin-converting enzyme inhibition and the progression of congestive cardiomyopathy. Effects on left ventricular and myocyte structure and function.

BACKGROUND: Clinical trials have demonstrated that angiotensin-converting enzyme inhibition (ACEI) improves survival in patients with long-term left ventricular (LV) dysfunction. However, it remained unclear from these clinical reports whether the beneficial effects of ACEI were due to direct improvements in LV myocardial structure and function. Accordingly, the overall objective of the present study was to examine the direct effects of ACEI on both LV and myocyte structure and function in the setting of cardiomyopathic disease. METHODS AND RESULTS: LV and isolated myocyte function and structure were examined in control dogs (n = 6), in dogs after the development of dilated cardiomyopathy caused by rapid ventricular pacing (RVP, 216 beats per minute, 4 weeks, n = 6), and in dogs with RVP and concomitant ACEI (RVP/ACEI, fosinopril 30 mg/kg BID, n = 6). LV ejection fraction fell with RVP compared with control values (35 +/- 3 versus 73 +/- 2%, P < .05) and was higher with RVP/ACEI compared with RVP values (41 +/- 4%, P = .048). LV end-diastolic volume increased with RVP compared with control values (78 +/- 7 versus 101 +/- 7 cm3, P < .05) and was lower with RVP/ACEI (82 +/- 3 cm3, P < .05). Isolated myocyte length increased with RVP (182 +- 1 versus 149 +/- 1 micron), and the velocity of shortening decreased (36 +/- 1 versus 57 +/- 1 micron/s) compared with control values (P < .05). With RVP/ACEI, myocyte length was reduced (169 +/- 1 micron) and velocity of shortening was increased (45 +/- 1 micron/s) compared with RVP values (P < .05). Myocyte velocity of shortening after beta-adrenergic receptor stimulation with 25 nmol/L isoproterenol was reduced with RVP compared with control values (142 +/- 5 versus 193 +/- 8 micron/s, P < .05) and significantly improved with RVP/ACEI (166 +/- 6 micron/s, P < .05). In the RVP group, beta-adrenergic receptor density fell 26%, and cAMP production with beta-adrenergic receptor stimulation was reduced 48% from control values. RVP/ACEI resulted in a normalization of beta-adrenergic receptor density and cAMP production. LV myosin heavy-chain content when normalized to dry weight of myocardium was unchanged with RVP (149 +/- 11 mg per gram dry weight of myocardium [gdwt]) and RVP/ACEI (150 +/- 4 mg/gdwt) compared with control values (165 +/- 4 mg/gdwt). LV collagen content decreased with RVP compared with control values (7.6 +/- 0.4 versus 9.6 +/- 0.8 mg per gram wet weight of myocardium [gwwt], P < .05) but was increased with RVP/ACEI (14.4 +/- 1.3 mg/gwwt, P < .05). CONCLUSIONS: Concomitant ACEI with chronic tachycardia reduced LV chamber dilation and improved myocyte contractile function and beta-adrenergic responsiveness. Contributory cellular and extracellular mechanisms for the beneficial effects of ACEI in this model of dilated cardiomyopathy included a normalization of beta-adrenergic receptor function and enhanced myocardial collagen support. The results from this study provide evidence that ACEI during the development of cardiomyopathic disease provided beneficial effects on LV myocyte contractile processes and myocardial structure.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1995

The human peroxisome proliferator-activated receptor (PPAR) subtype NUC1 represses the activation of hPPAR alpha and thyroid hormone receptors.

We have cloned two human peroxisome proliferator-activated receptor (PPAR) subtypes, hPPAR alpha and hNUC1. hPPAR alpha is activated by clofibric acid and other PPAR activators. hNUC1 is not activated by these compounds acting instead as a repressor of hPPAR alpha and human thyroid hormone receptor transcriptional activation. Repression is specific since hNUC1 does not significantly repress activation by the progesterone or retinoic acid receptors. We demonstrate co-operative binding of hNUC1 and hRXR alpha to a PPAR-responsive element and show that in the presence of hRXR alpha, the affinity of hNUC1 for the peroxisome proliferator is comparable to that of hPPAR alpha. Furthermore, repression of hPPAR alpha can be overcome by transfecting excess hPPAR alpha. We propose that hNUC1 represses the activity of hPPAR alpha by titrating out a factor required for activation. Our data further suggests convergence of thyroid hormone- and peroxisome-mediated fatty acid metabolism pathways. Overcoming hNUC1 repression could be a means of increasing the activity of these receptors.

Cell Line↗

PubMed1995

Direct effects of thrombin on myocyte contractile function.

Cardiopulmonary bypass activates the clotting cascade, resulting in elevated circulating levels of thrombin. In light of the fact that the function of a wide variety of cell types is modulated by thrombin, we hypothesized that thrombin may have a direct effect on myocyte function. Isolated left ventricular myocyte contractile function was measured from 6 adult dogs using videomicroscopy at baseline and after increasing concentrations of thrombin (1 to 10 U/mL). Indices of myocyte contractile function were reduced in a dose-dependent manner in the presence of increasing concentrations of thrombin. For example, myocyte percent shortening fell by 18% with 1 U/mL thrombin and by 43% with 2 U/mL thrombin. The addition of hirudin, a highly selective thrombin inhibitor, completely blocked the effects of thrombin on myocyte contractile function. beta-Adrenergic agonists are commonly used in the early post-cardiopulmonary bypass period. Accordingly, a final set of experiments examined the effects of thrombin on myocyte beta-adrenergic responsiveness using isoproterenol (25 nmol/L). In myocytes preincubated with 1 U/mL thrombin, myocyte beta-adrenergic responsiveness was significantly reduced. For example, in the presence of 1 U/mL thrombin, myocyte velocity of shortening fell by 25% from isoproterenol alone values. The results from the present study provide evidence that thrombin has a direct negative effect on steady-state contractile function and beta-adrenergic responsiveness in adult mammalian ventricular myocytes. These findings suggest that thrombin may be an additional contributory factor toward the transient left ventricular dysfunction that has been observed after cardiopulmonary bypass.

Animals↗

PubMed1995

Morphine inhibits the development of allogeneic immune responses in mouse lymph node.

Morphine and related opiates are often administered to relieve post-operative and chronic pain following transplantation surgery. Opiates have been shown to suppress a variety of immune parameters in both animal models and man. In the present study, we investigated whether morphine affects allogeneic immune responses by injecting C57BL/6 mice in the footpad with allogeneic spleen cells and examining changes in the draining popliteal lymph node (PLN). Morphine (administered as subcutaneous implants) had profound inhibitory effects on the development of alloreactivity manifested as a suppression of: (1) lymph node hyperplasia, (2) mixed lymphocyte reactivity (MLR) in PLN cells and (3) the number of CD4+ and Thy 1.2 lymphoid subsets. These inhibitory effects of morphine were abolished or dramatically reduced by co-administration of the opiate antagonist, naltrexone, indicating that suppression of allo-sensitization was opiate receptor mediated. In toto, these findings demonstrate that morphine administration interferes with the development of allogeneic immune response in mouse lymph node through an opiate receptor mediated mechanism.

Animals↗

PubMed1995

Addition of immunotherapy with Mycobacterium w vaccine to multi-drug therapy benefits multibacillary leprosy patients.

Immunotherapy with a vaccine consisting of autoclaved Mycobacterium w, was given in addition to standard chemotherapy (multidrug therapy (MDT)) to 93 multibacillary (MB) leprosy patients. One hundred and seven patients with similar types of disease served as controls and received MDT + placebo injections. The study was a double-blind randomised trial. On opening the codes, results obtained were in concordance with those in a single-blind trial which has been extensively reported. Bacteriological clearances were significantly more rapid in vaccinated patients (p < 0.03). Thirty-five LL or BL patients with a high bacterial index (BI) of 6 were completely cleared of acid-fast bacilli (AFB) after eight doses of vaccine. Only 8 patients in the control group became bacteriologically negative in the same time period. They all had BIs < 4. Associated with decreasing BI was accelerated clinical regression of lesions after vaccination and lepromin conversion rates of 100% for BB, 71% for BL and 70% for LL. A significant number of immunised patients showed histological improvement (p < 0.004). Thirty-six showed a complete disappearance of dermal granulomas and a picture of non-specific infiltration. The vaccine did not precipitate neuritis or deformities; episodes were noted in vaccinated patients as were incidences of Type 2 reaction. The overall improvement was reflected by a shorter duration of treatment and faster release of vaccinated patients.

Bacterial Vaccines↗

PubMed1995

Myocyte electrophysiological properties following the development of supraventricular tachycardia-induced cardiomyopathy.

Chronic supraventricular tachycardia (SVT) causes left ventricular (LV) dilatation and dysfunction, diminished myocyte contractile function, and abnormalities in sarcolemmal receptor systems. We hypothesized that changes in myocyte action potential characteristics and L-type Ca2+ channel (Ca2+ channel) function, which are major determinants of myocyte contractile processes, would occur with SVT cardiomyopathy. LV function and isolated myocyte contractile function were examined in 11 pigs with SVT cardiomyopathy (pace 240 bpm; 3 weeks) and 11 control pigs. With chronic SVT, LV fractional shortening fell and myocyte shortening velocity was reduced compared to controls (11 +/- 2 v 37 +/- 2%, P < 0.0001; and 32.5 +/- 1.2 v 55.7 +/- 1.6 microns/s, P < 0.0001, respectively). Isolated myocyte action potential upstroke velocity and amplitude were reduced with SVT cardiomyopathy compared to controls (92.8 +/- 4.8 v 129.5 +/- 3.1 V/s, P < 0.0001; and 98.2 +/- 2.2 v 110.3 +/- 1.3 mV, P < 0.0001, respectively). the duration of the myocyte action potential, defined as the time to 90% repolarization, was prolonged with SVT cardiomyopathy compared to controls (201.7 +/- 5.9 v 169.1 +/- 6.8 ms, P = 0.002). These specific abnormalities in the indices of myocyte contractile function and action potential characteristics which occurred with SVT cardiomyopathy were not normalized following beta-adrenergic receptor stimulation. In order to determine a potential mechanism for the changes in myocyte contractile function and action potential characteristics with SVT cardiomyopathy, Ca2+ channel function was examined in control and SVT myocytes. In SVT myocytes, peak L-type Ca2+ current (ICa) normalized to membrane capacitance and the Ca2+ channel inactivation time constant were reduced compared to controls (-2.30 +/- 0.24 v -3.79 +/- 0.28 pA/pF, P = 0.0001; and 104.0 +/- 10.8 v 199.9 +/- 27.4 ms, P = 0.005, respectively). The abnormalities in Ca2+ channel function with SVT cardiomyopathy persisted in myocytes with equivalent membrane capacitances and were not normalized with beta-adrenergic receptor stimulation. In conclusion, findings from the present study suggest that fundamental abnormalities in myocyte electrical events (action potential) and ionic flux (Ca2+ channel function) are contributory mechanisms for the depressed myocyte contractile function with SVT cardiomyopathy.

Action Potentials↗

PubMed1995

The direct effects of 3,5,3'-triiodo-L-thyronine (T3) on myocyte contractile processes. Insights into mechanisms of action.

Administration of 3,5,3'-triiodo-L-thyronine (T3) has recently been suggested to acutely improve left ventricular performance. However, the cellular and molecular mechanisms responsible for this improvement in left ventricular function with T3 remained unknown. Accordingly, the present study examined the direct effects of T3 administration on myocyte contractile function and the sarcolemmal systems that might potentially contribute to these effects. In isolated porcine left ventricular myocytes (n = 81), velocity of shortening increased in the presence of 80 pmol/L T3 compared with that in untreated myocytes (117.0 +/- 5.0 versus 77.3 +/- 3.3 microns/sec, p < 0.05). In a separate series of experiments (n = 29), myocyte velocity of shortening increased in the presence of both T3 and beta-adrenergic receptor stimulation (25 nmol/L isoproterenol) to greater than that with beta-adrenergic receptor stimulation alone (274.3 +/- 16.9 versus 203.7 +/- 16.2 microns/sec, p < 0.05). Cyclic adenosine monophosphate generation was next examined in isolated myocyte preparations (n = 9). In the presence of T3, no significant increase in cyclic-adenosine monophosphate generation was observed compared with that in untreated myocytes (39.1 +/- 8.3 versus 24.7 +/- 5.8 fmols/myocyte, p = 0.17). However, in the presence of both T3 and beta-adrenergic receptor stimulation, cyclic-adenosine monophosphate generation increased significantly to greater than that with beta-adrenergic receptor stimulation alone (224.4 +/- 61.1 versus 120.1 +/- 35.5 fmoles/myocyte, p < 0.05). Because cyclic-adenosine monophosphate modulates intracellular Ca2+ processes, L-type Ca+2 channel current (patch clamp methods; -picoamp/picofarad, n = 15) and peak intracellular Ca+2 levels (fura 2 ionic measurement, n = 47) were next measured. In the presence of T3, a shift in the activation voltage at peak L-type Ca+2 channel current was observed from baseline (5.5 +/- 1.4 versus 9.0 +/- 1.0 mV, p < 0.05). Furthermore, in the presence of both T3 and beta-adrenergic receptor stimulation, peak L-type Ca+2 channel current (8.9 +/- 0.7 versus 6.3 +/- 1.0 mV, p < 0.05) and peak intracellular Ca+2 levels (189.9 +/- 8.4 versus 171.7 +/- 8.3 nmol/L, p < 0.05) increased compared with values obtained with beta-adrenergic receptor stimulation alone. Important findings from the present study were twofold: (1) T3 improved myocyte contractile processes through a cyclic-adenosine monophosphate-independent mechanism and (2) T3 potentiated the effects of beta-adrenergic receptor stimulation transduction by increasing cyclic-adenosine monophosphate production, L-type Ca+2 channel current, and Ca+2 availability to the myocyte contractile apparatus.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1995

LV and myocyte structure and function after early recovery from tachycardia-induced cardiomyopathy.

Left ventricular (LV) function and mass were measured in six conscious dogs at weekly intervals during the progression of tachycardia-induced dilated cardiomyopathy (DCM) and during a 1-mo recovery period from DCM (post-DCM). LV end-diastolic volume and LV wall stress increased and LV ejection fraction decreased with each week of pacing. Despite the increased LV wall stress, LV mass did not change during the progression of tachycardia DCM. One week post-DCM resulted in an improved LV ejection fraction and normalization of neurohormonal profiles. However, 1 wk post-DCM was accompanied by a 26% increase in LV mass and persistent LV chamber dilation. Isolated myocyte function was examined and compared with that in six normal control dogs. Myocyte percent and myocyte velocity of shortening were 19 and 32% lower, respectively, in the post-DCM group compared with controls. Thus termination of the tachycardia subsequent to the development of DCM resulted in persistent LV chamber dilation and abnormalities in myocyte contractile function. The improved LV pump function with early recovery from tachycardia-induced DCM was mediated by LV hypertrophy and a subsequent reduction in LV wall stress rather than a normalization of LV geometry and myocyte contractile function.

Animals↗

PubMed1995

Effects of chronic supraventricular pacing tachycardia on relaxation rate in isolated cardiac muscle cells.

Chronic supraventricular pacing tachycardia (SVT) causes abnormalities in both ventricular and cellular relaxation. The mechanisms causing these abnormalities have not been fully determined. To examine two of the possible mechanisms, a decrease in restoring force or an impairment of the intrinsic myocardial relaxation process, cardiocytes were enzymatically isolated from the left ventricle of pigs subjected to left atrial pacing at 240 beats/min for 3 wk and normal control pigs. SVT caused a decrease in the extent of cardiocyte shortening and the velocity of cardiocyte lengthening. To determine whether the changes in the relaxation velocity merely reflected a concomitant decrease in the extent of cardiocyte shortening (and a resultant decrease in restoring forces) or, in addition, reflected impairment in intrinsic relaxation properties, the relation between cardiocyte relaxation velocity and cardiocyte shortening extent was examined. There was a direct relation between relaxation velocity and shortening extent in both control and SVT cardiocytes. However, SVT decreased the relaxation velocity at any common extent of shortening and decreased the slope of the direct relation (slope 5.91 in control vs. 3.51 s-1 in SVT, P < 0.05). Therefore, these data suggested that SVT caused a primary impairment in the intrinsic myocardial relaxation process independently of a decrease in restoring force.

Animals↗

PubMed1994

Identification of myelin basic proteins in circulating immune complexes associated with lepromatous leprosy.

Circulating immune complexes (CIC) were first measured in lepromatous patients (LL) by the 125I-C1q binding assay and the polyethylene glycol (PEG) precipitation test. High levels were found by both methods (95 and 90% of positives, respectively). LL-CIC were investigated for the presence of neural antigens. CIC were precipitated in 3.5% PEG, filtered through protein A-Sepharose affinity chromatography, eluted with glycine-HCl, pH 2.8, and washed with PBS; fractions after CIC dissociation were studied by SDS-PAGE and Western blotting. The LL-CIC PEG precipitates and the glycine-HCl eluates were positive in 76 and 71% respectively against anti-myelin basic proteins (MBP) monoclonal antibody, showing a single band at 15-25 kDa similar to the one obtained incubating MBP with anti-MBP. No reaction was detected with CIC-PBS fractions; strips were incubated with other anti-neural antibodies such as anti-glial fibrillary acidic proteins, anti-S-100, and anti-neurofilaments, without any reactivity. Our results demonstrate that LL-CIC contain MBP as an antigen; its significance could be related to the pathogenesis of leprosy since the liberation of MBP after Mycobacterium leprae nerve damage may elicit anti-MBP autoantibodies to myelin breakdown, which reacts with peripheral nerve MBP inducing CIC formation. This mechanism may be important in demyelination and destruction of nerve in leprosy.

Antigen-Antibody Complex↗

PubMed1994

Direct effects of acute administration of 3, 5, 3' triiodo-L-thyronine on myocyte function.

Recent studies have suggested that acute administration of 3, 5, 3' triiodo-L-thyronine (T3) may increase ventricular performance; however, the direct cellular effects of acute T3 administration are not understood. The objectives of this study were to determine (1) whether T3 acts directly on the cardiac muscle cell (myocyte) itself, and (2) whether T3 acts independently of the myocyte beta-adrenergic receptor. Accordingly, isolated myocyte function was examined using video-microscopy in normal porcine myocytes (n = 60) in the control state and in the presence of increasing T3 concentrations (10 to 500 pmol/L). T3 caused myocyte shortening extent, shortening velocity, and lengthening velocity to increase in a dose-dependent manner. For example, shortening velocity increased from 49.2 +/- 4.3 microns/s at baseline to 66.5 +/- 6.1 microns/s with 100 pmol/L T3 (p < 0.05). beta-Adrenergic stimulation with 25 nmol/L isoproterenol increased shortening velocity to 97.6 +/- 5.7 microns/s; isoproterenol with T3 increased shortening velocity further to 168.5 +/- 10.9 microns/s. Analysis of variance revealed that this increase with T3 was independent of and additive to the beta-adrenergic receptor system. In summary, T3 caused a dose-dependent increase in myocyte contractile performance, and these effects were independent of and additive to beta-adrenergic receptor stimulation. Thus, acute T3 administration may provide a novel modality to improve left ventricular contractile function independent of the beta-adrenergic receptor system.

Animals↗

PubMed1994

Effects of protamine on myocyte contractile function and beta-adrenergic responsiveness.

The use of protamine sulfate in patients has been associated with severe circulatory collapse and myocardial failure. However, the exact mechanisms responsible for these reactions to protamine remain unclear. Accordingly, we examined the effect of protamine on isolated myocyte contractile function. Indexes of isolated myocyte contractile function, percent shortening, and velocity of shortening were examined using videomicroscopy. Porcine cardiocytes (n = 75) were studied at baseline and in the presence of 80 micrograms/mL protamine. In addition, myocyte function was examined sequentially, first during treatment with 8 IU/mL heparin and then after the addition of a protamine dose sufficient to completely bind the heparin. The binding of heparin and protamine resulted in the formation of a heparin-protamine complex. The protamine concentration of 80 micrograms/mL is approximately equal to the serum concentration of protamine obtained in patients when administered in a dose of 5 mg/kg. In the presence of 80 micrograms/mL protamine, both percent shortening and velocity of shortening fell by more than 32% from baseline values (p < 0.05). The presence of either heparin alone or the heparin-protamine complex resulted in no change in baseline myocyte contractile measurements. Furthermore, to examine the effect of protamine on myocyte beta-adrenergic responsiveness a second series of experiments were performed. Myocyte contractile function was measured when 25 nmol/L isoproterenol was added to each of the protocols above. The presence of 80 micrograms/mL protamine resulted in a significant blunting of myocyte beta-adrenergic responsiveness. The presence of either heparin alone or the heparin-protamine complex resulted in no change in myocyte beta-adrenergic responsiveness.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Creatine kinase activity as an indicator of unopposed estrogen action in the mouse uterus associated with anti-progesterone treatment.

The brain isozyme of creatine kinase (CKB) is a major component of the estrogen-induced proteins in the rat uterus. Hormonal specificity of this response was studied in cotransfection assays using the rat CKB promoter linked to the bacterial chloramphenicol acetyltransferase gene. Response was specific for estrogen as 17 beta-estradiol in the presence of estrogen receptor dramatically stimulated the CKB promoter. This induction was completely blocked by the estrogen antagonist ICI 164,384. Nuclear receptors for progesterone, androgen, glucocorticoid and vitamin D did not significantly activate the CKB promoter in the presence of their respective ligands. Creatine kinase (CK) activity was analyzed in decidualized mouse uterus to assess estrogenic activity in vivo. Upon oil stimulation, uterine horns of day 4 pseudopregnant mice underwent a dramatic outgrowth in response to endogenous progesterone. This response was accompanied by a significant decrease in CK activity from a control value of 1.44 +/- 0.25 to 0.38 +/- 0.08 IU/mg protein (P < 0.001), indicating that the action of estrogen was suppressed. Treatment of females one day prior to oil-stimulation with progesterone receptor antagonists, RU486 (Mifepristone) or ZK299 (Onapristone), or with a monoclonal antibody to progesterone (DB3), abolished decidualization, and also restored the CK activity to the control value. These results suggest that CK can be used as a specific cellular marker to detect unopposed estrogen action in the mouse uterus associated with progesterone withdrawal or receptor blockade.

Animals↗

PubMed1994

Human and rat peroxisome proliferator activated receptors (PPARs) demonstrate similar tissue distribution but different responsiveness to PPAR activators.

We have isolated a human peroxisomal proliferator activated receptor (hPPAR) from a human liver cDNA library. Based on sequence analysis, we have determined that this cDNA encodes the human PPAR alpha. When assayed in a reconstituted hPPAR responsive transcription system in mammalian CV-1 cells, this receptor was shown to be transcriptionally activated by hypolipidemic agents like clofibric acid, and ETYA (5,8,11,14-eicosatetraynoic acid; a synthetic arachidonic acid homolog). When analyzed in CV-1 cells, the rat PPAR alpha was similarly transcriptionally regulated. However, when assayed in a human liver cell line (HepG2) we noticed that ETYA was a more efficient activator of hPPAR alpha than rPPAR alpha. Thus, factors other than the receptor are important in determining the cellular responsiveness to this class of compounds. Interestingly, WY-14,643, another peroxisome proliferator, was a much more potent activator of rPPAR alpha than human PPAR alpha when assayed in both cell lines. This may explain in part why certain fibrates are potent hepatocarcinogens in rodents. Northern analysis indicates that hPPAR alpha and rPPAR alpha are well expressed in heart, kidney and liver. We further demonstrate that hPPAR alpha and human retinoid X receptor alpha synergistically interact to bind and transactivate through a peroxisomal proliferator response element. Thus in a similar cell and promoter context the rat and human PPARs show a differential response to certain activators. Cumulatively these data suggest that differential ligand responsiveness does not provide a complete explanation for the different biological effects exhibited by hypolipidemic drugs when administered to humans and rats.

5,8,11,14-Eicosatetraynoic Acid↗