Permanently Hypoxic Cell Culture Yields Rat Bone Marrow Mesenchymal Cells with Higher Therapeutic Potential in the Treatment of Chronic Myocardial Infarction.

BACKGROUND: The mismatch between traditional in vitro cell culture conditions and targeted chronic hypoxic myocardial tissue could potentially hamper the therapeutic effects of implanted bone marrow mesenchymal stem cells (BMSCs). This study sought to address (i) the extent of change to BMSC biological characteristics in different in vitro culture conditions and (ii) the effectiveness of permanent hypoxic culture for cell therapy in treating chronic myocardial infarction (MI) in rats. METHODS: rat BMSCs were harvested and cultured in normoxic (21% O2, n=27) or hypoxic conditions (5% O2, n=27) until Passage 4 (P4). Cell growth tests, flow cytometry, and Bio-Plex assays were conducted to explore variations in the cell proliferation, phenotype, and cytokine expression, respectively. In the in vivo set-up, P3-BMSCs cultured in normoxia (n=6) or hypoxia (n=6) were intramyocardially injected into rat hearts that had previously experienced 1-month-old MI. The impact of cell therapy on cardiac segmental viability and hemodynamic performance was assessed 1 month later by 2-Deoxy-2[18F]fluoro-D-glucose (18F-FDG) positron emission tomography (PET) imaging and pressure-volume catheter, respectively. Additional histomorphological examinations were conducted to evaluate inflammation, fibrosis, and neovascularization. RESULTS: Hypoxic preconditioning significantly enhanced rat BMSC clonogenic potential and proliferation without altering the multipotency. Different profiles of inflammatory, fibrotic, and angiogenic cytokine secretion were also documented, with a marked correlation observed between in vitro and in vivo proangiogenic cytokine expression and tissue neovessels. Hypoxic-preconditioned cells presented a beneficial effect on the myocardial viability of infarct segments and intrinsic contractility. CONCLUSION: Hypoxic-preconditioned BMSCs were able to benefit myocardial perfusion and contractility, probably by modulating the inflammation and promoting angiogenesis.

TREM-1 Mediates Inflammatory Injury and Cardiac Remodeling Following Myocardial Infarction.

RATIONALE: Optimal outcome after myocardial infarction (MI) depends on a coordinated healing response in which both debris removal and repair of the myocardial extracellular matrix play a major role. However, adverse remodeling and excessive inflammation can promote heart failure, positioning leucocytes as central protagonists and potential therapeutic targets in tissue repair and wound healing after MI. OBJECTIVE: In this study, we examined the role of triggering receptor expressed on myeloid cells-1(TREM-1) in orchestrating the inflammatory response that follows MI. TREM-1, expressed by neutrophils and mature monocytes, is an amplifier of the innate immune response. METHODS AND RESULTS: After infarction, TREM-1 expression is upregulated in ischemic myocardium in mice and humans. Trem-1 genetic invalidation or pharmacological inhibition using a synthetic peptide (LR12) dampens myocardial inflammation, limits neutrophils recruitment and monocyte chemoattractant protein-1 production, thus reducing classical monocytes mobilization to the heart. It also improves left ventricular function and survival in mice (n=20-22 per group). During both permanent and transient myocardial ischemia, Trem-1 blockade also ameliorates cardiac function and limits ventricular remodeling as assessed by fluorodeoxyglucose-positron emission tomographic imaging and conductance catheter studies (n=9-18 per group). The soluble form of TREM-1 (sTREM-1), a marker of TREM-1 activation, is detectable in the plasma of patients having an acute MI (n=1015), and its concentration is an independent predictor of death. CONCLUSIONS: These data suggest that TREM-1 could constitute a new therapeutic target during acute MI.

Repairing chronic myocardial infarction with autologous mesenchymal stem cells engineered tissue in rat promotes angiogenesis and limits ventricular remodeling.

BACKGROUND: Tissue engineering scaffold constitutes a new strategy of myocardial repair. Here, we studied the contribution of a patch using autologous mesenchymal stem cells (MSCs) seeded on collagen-1 scaffold on the cardiac reconstruction in rat model of chronic myocardial infarction (MI). METHODS: Patches were cultured with controlled MSCs (growth, phenotype and potentiality). Twenty coronary ligated rats with tomoscingraphy (SPECT)-authenticated transmural chronic MI were referred into a control group (n = 10) and a treated group (n = 10) which beneficiated an epicardial MSC-patch engraftment. Contribution of MSC-patch was tested 1-mo after using non-invasive SPECT cardiac imaging, invasive hemodynamic assessment and immunohistochemistry. RESULTS: 3D-collagen environment affected the cell growth but not the cell phenotype and potentiality. MSC-patch integrates well the epicardial side of chronic MI scar. In treated rats, one-month SPECT data have documented an improvement of perfusion in MI segments compared to control (64 ± 4% vs 49 ± 3% p = 0.02) and a reduced infarction. Contractile parameter dp/dtmax and dp/dtmin were improved (p & 0.01). Histology showed an increase of ventricular wall thickness (1.75 ± 0.24 vs 1.35 ± 0.32 mm, p &0.05) and immunochemistry of the repaired tissue displayed enhanced angiogenesis and myofibroblast-like tissue. CONCLUSION: 3D-MSC-collagen epicardial patch engraftment contributes to reverse remodeling of chronic MI.

Clinical-grade mesenchymal stem cells derived from umbilical cord improve septic shock in pigs.

BACKGROUND: Septic shock is the leading cause of death in intensive care units. The pathophysiological complexity of this syndrome contributes to an absence of specific treatment. Several preclinical studies in murine models of septic shock have shown improvements to organ injury and survival after administration of mesenchymal stem cells (MSCs). To better mimic a clinical approach in humans, we investigated the impact of randomized controlled double-blind administration of clinical-grade umbilical cord-derived MSCs to a relevant pig model of septic shock. METHODS: Septic shock was induced by fecal peritonitis in 12 male domestic pigs. Animals were resuscitated by an experienced intensivist including fluid administration and vasopressors. Four hours after the induction of peritonitis, pigs were randomized to receive intravenous injection of thawed umbilical cord-derived MSCs (UCMSC) (1 × 106 UCMSCs/kg diluted in 75 mL hydroxyethyl starch (HES), (n = 6) or placebo (HES alone, n = 6). Researchers were double-blinded to the treatment administered. Hemodynamic parameters were continuously recorded. Gas exchange, acid-base status, organ function, and plasma cytokine concentrations were assessed at regular intervals until 24 h after the onset of peritonitis when animals were sacrificed under anesthesia. RESULTS: Peritonitis induced profound hypotension, hyperlactatemia, and multiple organ failure. These disorders were significantly attenuated when animals were treated with UCMSCs. In particular, cardiovascular failure was attenuated, as attested by a better mean arterial pressure and reduced lactatemia, despite lower norepinephrine requirements. As such, UCMSCs improved survival in this very severe model (60% survival vs. 0% at 24 h). CONCLUSION: UCMSCs administration is beneficial in this pig model of polymicrobial septic shock.

Intramyocardial Implantation of bone marrow-derived stem cells enhances perfusion in chronic myocardial infarction: dependency on initial perfusion depth and follow-up assessed by gated pinhole SPECT.

UNLABELLED: Cell therapy-induced changes in the perfusion of areas of myocardial infarction (MI) remain unclear. This study investigated whether an original pinhole SPECT technique could be applied to a rat MI model to analyze local improvement in myocardial perfusion relating to engraftment sites of bone marrow-derived stem cells (BMSCs). METHODS: Four-month-old MI rats were either untreated (n = 8) or treated (n = 10) by intramyocardial injection of (111)In-labeled BMSCs. Early distribution of (111)In-BMSCs within the MI target was evidenced by dual (111)In/(99m)Tc pinhole SPECT 48 h later. Myocardial perfusion was serially monitored by (99m)Tc-sestamibi pinhole gated SPECT up to 3 mo after transplantation. RESULTS: Forty-eight hours after transplantation, (111)In-BMSCs were observed in all treated rats and in 18 of their 32 underperfused MI segments (<70% sestamibi uptake before transplantation). During the subsequent 3-mo follow-up, the perfusion of MI segments worsened in untreated rats (absolute change in sestamibi uptake, -3% +/- 3%; P < 0.05) but improved in treated rats (+4% +/- 7%; P < 0.05). This perfusion improvement was unrelated to the initial detection of (111)In-BMSCs (+2% +/- 6% in segments with (111)In-BMSCs vs. +5% +/- 7% in those without; not statistically significant) but was strongly associated with less severe perfusion defects before transplantation (+6% +/- 6% in segments with 60%-70% sestamibi uptake [n = 19] vs. -1% +/- 6% in those with <60% uptake [n = 13]; P = 0.003). CONCLUSION: When BMSCs are injected within chronic MI, perfusion enhancement predominates in the MI areas showing a high enough residual perfusion before treatment but not in those of the initial cell engraftment, giving evidence of dependency on the perfusion and metabolic environment at implantation sites.

High Versus Low Blood-Pressure Target in Experimental Ischemic Prolonged Cardiac Arrest Treated with Extra Corporeal Life Support.

BACKGROUND: There is currently no recommendation for the mean arterial pressure target in the particular setting of Extracorporeal Cardiopulmonary Resuscitation (ECPR) in the first hours following cardiogenic shock complicated by cardiac arrest. This study aimed to assess the effects of two different levels of mean arterial pressure on macrocirculatory, microcirculatory, and metabolic functions. DESIGN: Randomized animal study. SETTING: University research laboratory. INTERVENTION: Ventricular fibrillation was induced in 14 male pigs by surgical ligature of the interventricular coronary artery. After 20 min of cardiopulmonary resuscitation, Extracorporeal Life Support (ECLS) was initiated to restore circulatory flow. Thereafter, animals were randomly allocated to a high mean arterial pressure group (High-MAP, 80-85 mm Hg) or to a standard mean arterial pressure group (Standard-MAP, 65-70 mm Hg). Assessments conducted at baseline, immediately following and 6 h after ECLS initiation were focused on lactate evolution, amount of infused fluid, and microcirculatory parameters. RESULTS: There was no significant difference between the two groups at the time of ECLS initiation and at 6 h with regard to lactate levels (High-MAP vs. Standard-MAP: 8.8 [6.7-12.9] vs. 9.6 [9.1-9.8] mmol·l, P = 0.779 and 8.9 [4.3-11.1] vs. 3.3 [2.4-11] mmol·l, P = 0.603). Infused fluid volume did not significantly differ between the two groups (4,000 [3,500-12,000] vs. 5,000 [2,500-18,000] mL, P = 0.977). There was also no significant difference between the two groups regarding renal and liver functions, and sublingual capillary microvascular flow index assessed by Sidestream Dark Field imaging. CONCLUSION: Compared with a standard mean arterial pressure regimen, targeting a high mean arterial pressure in the first hours of an experimental ECPR model did not result in any hemodynamic improvement nor in a decrease in the amount of infused fluid.

Moderate Hypothermia Improves Cardiac and Vascular Function in a Pig Model of Ischemic Cardiogenic Shock Treated With Veno-Arterial ECMO.

Cardiogenic shock (CS) patients treated with extracorporeal membrane oxygenation (ECMO) have severe cardiac failure, associated with ischemia-reperfusion. The use of moderate hypothermia during ischemia-reperfusion syndrome is supported by experimental data. We therefore studied the effects of moderate hypothermia on cardiac and vascular function in pig ischemic CS treated with veno-arterial extracorporeal membrane oxygenation (VA-ECMO). CS was induced in 12 anesthetized pigs by coronary ligation. After 1 h of CS, VA-ECMO was initiated and pigs were randomized to normothermia (38°C) or moderate hypothermia (34°C) during 8 h. Intrinsic cardiac function was measured using a left ventricular conductance catheter. At the end of the experiment, tissues were harvested for Western blotting. ECMO associated with norepinephrine infusion and volume resuscitation increased mean arterial pressure, mixed venous oxygen saturation as well as carotid, renal, and coronary blood flow without any differences between normothermia and hypothermia. Hypothermia was associated with less fluid and less norepinephrine infusion, lower lactate level, and higher urinary output. Vascular reactivity was superior in hypothermia comparatively to normothermia as expressed using norepinephrine dose-response curves. Pressure development during isovolumic contraction, left ventricular ejection fraction, and prerecruitable stroke work index were higher in the hypothermia group. There were no differences between normothermia and hypothermia with regard to carotid and mesenteric protein expression for iNOs, eNOS, and phospho AKt/AKt measured at the end of the experimentation. The incidence of surgical bleeding and coagulation disorders was the same in both groups. In conclusion, moderate and rapid hypothermia improves hemodynamics and cardiac and vascular function in a pig model of ischemic CS treated with ECMO.

Short-term heart retention and distribution of intramyocardial delivered mesenchymal cells within necrotic or intact myocardium.

Cell therapy with bone marrow mesenchymal stem cells (BMSCs) is a new strategy for treating ischemic heart failure, but data concerning the distribution and retention of transplanted cells remain poor. We investigated the short-term myocardial retention of BMSCs when these cells are directly injected within necrotic or intact myocardium. 111Indium-oxine-labeled autologous BMSCs were injected within either 1-month-old infarction (n = 6) or normal myocardium (n = 6) from rats. Serial in vivo pinhole scintigraphy was scheduled during 1 week in order to track the implanted cells. The myocardial retention of BMSCs was definitely higher in myocardial infarction than in normal myocardial area (estimated percent retention at 2 h: 63 +/- 3% vs. 25 +/- 4%, p < 0.001) and the estimated cardiac retention values were unchanged in both groups along the 7 days of follow-up. On heart sections at day 7, labeled BMSCs were still around the injection site and appeared confined to the scarred tissue corresponding either to the infarct area or to the myocardium damaged by needle insertion. BMSCs have a higher retention when they are injected in necrotic than in normal myocardial areas and these cells appear to stay around the injection site for at least a 7-day period.

Beneficial Effects of Norepinephrine Alone on Cardiovascular Function and Tissue Oxygenation in a Pig Model of Cardiogenic Shock.

INTRODUCTION: The present study was developed to investigate the effects of norepinephrine alone on hemodynamics and intrinsic cardiac function in a pig model of cardiogenic shock mimicking the clinical setting. METHODS: Cardiogenic shock was induced by 1-h ligation of the left anterior descending (LAD) artery followed by reperfusion. Pigs were monitored with a Swan-Ganz catheter, a transpulmonary thermodilution catheter, and a conductance catheter placed in the left ventricle for pressure-loop measurements. Measurements were performed before LAD occlusion, 1 h after LAD occlusion, and 4 h after myocardial reperfusion. RESULTS: Myocardial infarction and reperfusion was followed by cardiogenic shock characterized by a significant increase in heart rate and significant decreases in mean arterial pressure (MAP), mixed venous oxygen saturation (SVO2), left ventricular end-diastolic pressure (LVEDP), prerecruitable stroke work (PRSW), and cardiac power index (CPI). Lactate levels were significantly increased. The systemic vascular resistance index (SVRI) and global end-diastolic volume index (GEDVI) remained unchanged. When compared with the control group (n = 6), norepinephrine infusion (n = 6) was associated with no changes in heart rate, a significant increase in MAP, SVO2, left ventricular ejection fraction, pressure development during isovolumic contraction, SVRI, and CPI and a decrease in lactate level. Cardiac index tended to increase (P = 0.059), whereas PRSW did not change in the norepinephrine group. LVEDP and GEDVI remained unchanged. CONCLUSIONS: Norepinephrine alone is able to improve hemodynamics, cardiac function, and tissue oxygenation in a pig model of ischemic cardiogenic shock.

Pharmacological inhibition of the triggering receptor expressed on myeloid cells-1 limits reperfusion injury in a porcine model of myocardial infarction.

AIMS: Limitation of ischemia/reperfusion injury is a major therapeutic target after acute myocardial infarction (AMI). Toll-like receptors are implicated in the inflammatory response that occurs during reperfusion. The triggering receptor expressed on myeloid cells (TREM)-1 acts as an amplifier of the immune response triggered by toll-like receptor engagement. We hypothesized that administration of a TREM-1 inhibitory peptide (LR12) could limit reperfusion injury in a porcine model of AMI. METHODS AND RESULTS: AMI was induced in 15 adult minipigs by a closed-chest coronary artery occlusion-reperfusion technique. Animals were randomized to receive LR12 or vehicle before reperfusion (LR12 n = 7, vehicle n = 8), and were monitored during 18 h. AMI altered hemodynamics and cardiac function, as illustrated by a drop of mean arterial pressure, cardiac index, cardiac power index, ejection fraction, and real-time pressure-volume loop-derived parameters. TREM-1 inhibition by LR12 significantly improved these dysfunctions (P < 0.03) and limited infarct size, as assessed by lower creatine phosphokinase and troponin I concentrations (P < 0.005). Pulmonary, renal, and hepatic impairments occurred after AMI and were attenuated by LR12 administration as assessed by a better PaO2 to FiO2 ratio, a less positive fluid balance, and lower liver enzymes levels (P < 0.05). CONCLUSION: Inhibition of the TREM-1 pathway by a synthetic peptide limited myocardial reperfusion injury in a clinically relevant porcine model of AMI.

Effect of chronic left ventricular unloading on myocardial remodeling: Multimodal assessment of two heterotopic heart transplantation techniques.

BACKGROUND: Cardiac recovery is possible by means of mechanical unloading yet remains rare. Excessive unloading-associated myocardial atrophy and fibrosis may adversely affect the process of reverse remodeling. In this study, we sought to evaluate the effect of different intensities of chronic left ventricular (LV) unloading on myocardial remodeling. METHODS: Twenty-five isogenic Lewis rats underwent complete LV unloading (CU, n = 15) induced by heterotopic heart transplantation or partial LV unloading (PU, n = 10) by heterotopic heart-lung transplantation. Information obtained from serial echocardiography, 2-deoxy-2[(18)F]fluoro-d-glucose ((18)F-FDG)-positron emission tomography, and an LV pressure-volume catheter were used to evaluate the morphology, glucose metabolism, and hemodynamic performance of the orthotopic hearts and heterotopic transplants over 4 weeks. Cell size, collagen content, tissue cytokines (interleukin [IL]-1α, IL-2, IL-6, IL-10, tumor necrosis factor-α, and vascular endothelial growth factor), and matrix metalloproteinase-2 and -9 were also determined. The recorded parameters included LV end-systolic dimension, LV end-diastolic dimension, posterior wall thickness, diastolic interventricular septum thickness, LV fractional shortening, and LV ejection fraction. RESULTS: We demonstrated an LV load-dependent relationship using echo-based structural (left posterior wall thickness, diastolic interventricular septum thickness, and left ventricular end-diastolic dimension) and functional (LV fractional shortening and LV ejection fraction) parameters, as well as an (18)F-FDG uptake (all p < 0.05). This load-dependent relationship was also evidenced in measurements from the pressure-volume conductance catheter (stroke volume, stroke work, cardiac output, dP/dTmax, and -dP/dTmin; all p < 0.05). Significant myocardial atrophy and fibrosis were observed in unloaded hearts, whereas concentrations of cytokines and matrix metalloproteinases were comparable in both unloading conditions. CONCLUSIONS: Partial and complete unloading affected the remodeling of non-failing hearts in a rodent model to different extents on myocardial atrophy, fibrosis, glucose metabolism, and mechanical work. Cardiac atrophy is the prominent change after mechanical unloading, which exaggerates the proportion of total collagen that is responsible for diastolic dysfunction.

Sustained therapeutic perfusion outside transplanted sites in chronic myocardial infarction after stem cell transplantation.

This study aimed at comparing long-term variations in the perfusion of chronic myocardial infarction (MI) areas after local injections of autologous bone marrow stem cells (BMSCs). 14 coronary ligated rats with transmural chronic MI (4 months) were used: a control group (n = 7) versus a treated group (n = 7) in which (111)In labeled-BMSCs were directly engrafted on MI areas. By using (111)In/(99m)Tc SPECT and Sestamibi gated-SPECT,. left ventricle perfusion and function were monitored in all animals by serial (99m)Tc-Sestamibi pinhole gated-SPECT over a period of 6 months. Post-therapeutic myocardial perfusion improved as early as 48 h following injection in the 2 groups. This benefice was sustained during the 6-month follow-up in the non-engrafted MI-areas from treated rats (at 6-months: +10 ± 5 %), whereas the engrafted ones, as well as the MI areas from control rats, exhibited progressive deterioration over time (at 6-months: -9 ± 10 % and -5 ± 3 %, respectively). Perfusion enhancement of the chronic MI areas treated by BMSCs transplantation is: (1) marked in the following days, presumably because of an unspecific inflammatory reaction, and (2) sustained over the long term but only outside the sites of cell engraftment, suggesting a distant paracrine effect of transplanted cells.

Effects of a TREM-like transcript 1-derived peptide during hypodynamic septic shock in pigs.

The objective of this study was to determine the effects of a TREM (triggering receptor expressed on myeloid cells 1)-like transcript 1-derived peptide (LR12) administration during septic shock in pigs. Two hours after induction of a fecal peritonitis, anesthetized and mechanically ventilated adult male minipigs were randomized to receive LR12 (n = 6) or its vehicle alone (normal saline, n = 5). Two animals were operated and instrumented without the induction of peritonitis and served as controls (sham). Resuscitation was achieved using hydroxyethyl starch (up to 20 mL/kg) and norepinephrine infusion (up to 10 µg/kg per minute). Hemodynamic parameters were continuously recorded. Gas exchange, acid-base status, organ function, and plasma cytokines concentrations were evaluated at regular intervals until 24 h after the onset of peritonitis when animals were killed under anesthesia. Peritonitis induced profound hypotension, myocardial dysfunction, lactic acidosis, coagulation abnormalities, and multiple organ failure. These disorders were largely attenuated by LR12. In particular, cardiovascular failure was dampened as attested by a better mean arterial pressure, cardiac index, cardiac power index, and S(v)O(2), despite lower norepinephrine requirements. LR12, a TREM-like transcript 1-derived peptide, exhibits salutary properties during septic shock in adult minipigs.

Time-related changes in expression of collagen types I and III and of tenascin-C in rat bone mesenchymal stem cells under co-culture with ligament fibroblasts or uniaxial stretching.

Adult bone-marrow-derived mesenchymal stroma cells (BMSC) seem to be a potential cell source for tissue engineering of the ligament. The objective of this work was to study the time-related changes in mRNA expression and protein levels of collagen types I and III and of tenascin-C in BMSC under co-culture with fibroblasts or under a uniaxial cyclic condition. Rat BMSC harvested from the femur and tibial bone marrow were co-cultured with ligament fibroblasts or stimulated by cyclic 10% uniaxial stretching at 1 Hz. Image analysis showed significant cell loss in stretched BMSC, particularly in the directions close to the stretching direction. However, these BMSC displayed an equivalent growth rate to that of non-stretched cells. Real-time reverse transcription/polymerase chain reaction revealed that the mRNA expression of collagen types I and III and of tenascin-C by BMSC was significantly up-regulated by co-culture and cyclic stretching. Radioimmunoassay results confirmed the effects of these stimulations, showing increases in the level of these proteins. Thus, BMSC might be useful as a cell source for the tissue engineering of ligament.

Feasibility of in vivo dual-energy myocardial SPECT for monitoring the distribution of transplanted cells in relation to the infarction site.

PURPOSE: Cell therapy using bone marrow mesenchymal stem cells (BMSCs) shows promise in the treatment of myocardial infarction (MI) but accurate cell delivery within MI areas remains critical. In the present study, we tested the feasibility of in vivo pinhole SPECT imaging for monitoring the sites of intramyocardial implanted BMSCs in relation to targeted MI areas in rats. METHODS: BMSCs were labelled with (111)In-oxine and injected within the fibrotic areas of 3-month-old MI in ten rats. Two days later, dual (111)In/(99m)Tc-sestamibi pinhole SPECT was recorded for localisation of (111)In-BMSCs on a 15-segment left ventricular (LV) division. Additional (99m)Tc-sestamibi pinhole SPECT had been performed 1 month earlier and on the day before transplantation. In vitro counting on histological sections was used to validate the pinhole SPECT determination of (111)In-BMSC activity within LV segments. RESULTS: The underperfused MI area (segments with <70% uptake) was stable between the (99m)Tc-sestamibi SPECT study recorded at 1 month (4.6+/-1.9 segments) and at 1 day (4.7+/-2.3 segments) before transplantation. (111)In-BMSCs were detected by dual-energy SPECT in 56 segments: 33 (59%) were underperfused MI segments but 23 (41%) were not (14 adjacent and nine remote segments). Finally, (111)In-labelled BMSCs were not detected in 14 out of the 47 (30%) underperfused MI segments. CONCLUSION: When BMSCs are injected within MI areas in rats, sites of early cell retention do not always match the targeted MI areas. The dual-energy pinhole SPECT technique may be used for monitoring the sites of early retention of implanted BMSCs and the data obtained may have critical importance when analysing the effects of cardiac cell therapy.

Effect of a methionine-supplemented diet on the blood pressure of Wistar-Kyoto and spontaneously hypertensive rats.

The objectives of the present work were to evaluate the effect of a methionine-supplemented diet as a model of hyperhomocysteinaemia on the systolic blood pressure (BP) and vasomotor functions of aortic rings in Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). WKY and SHR rats, randomised into four groups, were fed a normal semisynthetic diet or a methionine (8 g/kg)-supplemented diet for 10 weeks. Systolic BP was measured non-invasively. At the end of the experiment, plasma homocysteine, methionine, cysteine and glutathione levels were determined. Vasoconstriction and vasodilatation of aortic rings were measured. The methionine-supplemented diet induced a significant increase in plasma homocysteine and methionine concentration in both WKY and SHR rats, an increase in plasma cysteine concentrations in WKY rats and an increase in the glutathione concentration in SHR. The systolic BP of WKY rats fed the methionine-supplemented diet increased significantly (P<0.01), whereas systolic BP was reduced in SHR. An enhanced aortic responsiveness to noradrenaline and a decreased relaxation induced by acetylcholine and bradykinin were observed in the WKY rats fed the methionine-enriched diet. In SHR, the bradykinin-induced relaxation was reduced, but the sodium nitroprusside response was increased. In conclusion, a methionine-enriched diet induced a moderate hyperhomocysteinaemia and an elevated systolic BP in WKY rats that was consistent with the observed endothelial dysfunction. In SHR, discrepancies between the decreased systolic BP and the vascular alterations suggest more complex interactions of the methionine-enriched diet on the systolic BP. Further investigations are needed to understand the paradoxical effect of a methionine-rich diet on systolic BP.