Efficient transduction of vascular smooth muscle cells with a translational AAV2.5 vector: a new perspective for in-stent restenosis gene therapy.

Coronary artery disease represents the leading cause of mortality in the developed world. Percutaneous coronary intervention involving stent placement remains disadvantaged by restenosis or thrombosis. Vascular gene therapy-based methods may be approached, but lack a vascular gene delivery vector. We report a safe and efficient long-term transduction of rat carotid vessels after balloon injury intervention with a translational optimized AAV2.5 vector. Compared with other known adeno-associated virus (AAV) serotypes, AAV2.5 demonstrated the highest transduction efficiency of human coronary artery vascular smooth muscle cells (VSMCs) in vitro. Local delivery of AAV2.5-driven transgenes in injured carotid arteries resulted in transduction as soon as day 2 after surgery and persisted for at least 30 days. In contrast to adenovirus 5 vector, inflammation was not detected in AAV2.5-transduced vessels. The functional effects of AAV2.5-mediated gene transfer on neointimal thickening were assessed using the sarco/endoplasmic reticulum Ca(2+) ATPase isoform 2a (SERCA2a) human gene, known to inhibit VSMC proliferation. At 30 days, human SERCA2a messenger RNA was detected in transduced arteries. Morphometric analysis revealed a significant decrease in neointimal hyperplasia in AAV2.5-SERCA2a-transduced arteries: 28.36±11.30 (n=8) vs 77.96±24.60 (n=10) µm(2), in AAV2.5-green fluorescent protein-infected, P<0.05. In conclusion, AAV2.5 vector can be considered as a promising safe and effective vector for vascular gene therapy.

SERCA2a gene transfer prevents intimal proliferation in an organ culture of human internal mammary artery.

Coronary restenosis, a major complication of percutaneous balloon angioplasty, results from neointimal proliferation of vascular smooth muscle cells (VSMCs). The sarco/endoplasmic reticulum calcium ATPase 2a isoform (SERCA2a), specific to contractile VSMCs, has been reported previously to be involved in the control of the Ca(2+)-signaling pathways governing proliferation and migration. Moreover, SERCA2a gene transfer was reported to inhibit in vitro VSMC proliferation and to prevent neointimal thickening in a rat carotid injury model. The aim of this study was to evaluate the potential therapeutic interest of SERCA2a gene transfer for prevention of in-stent restenosis using a ex vivo model of human left internal mammary artery (hIMA) intimal thickening. Left hIMAs, obtained at the time of aorto-coronary bypass surgeries, were subjected to balloon dilatation followed by infection for 30 min with adenoviruses encoding either human SERCA2 and green fluorescence protein (GFP) or control gene (ß-galactosidase, ß-gal) and GFP. Proliferation of subendothelial VSMCs and neointimal thickening were observed in balloon-injured hIMA maintained 14 days in organ culture under constant pressure and perfusion. SERCA2a gene transfer prevented vascular remodeling and significantly (P<0.01, n=5) reduced neointimal thickening in injured arteries (intima/media ratio was 0.07±0.01 vs 0.40±0.03 in ß-gal-infected arteries). These findings could have potential implications for treatment of pathological in-stent restenosis.

A calcium-sensitive promoter construct for gene therapy.

Targeting diseased cells is a challenging issue in both pharmacological and biological therapeutics. Gene therapy is emerging as a novel approach for treating rare diseases and for illnesses for which there is no other alternative. An important limitation of gene therapy has been the off-target effects and therefore efforts have been focused on increasing the specificity of gene transfer to the targeted organ. Here, we describe a promoter containing six nuclear factor of activated T cells (NFAT) consensus sequences, which is as efficient as the cytomegalovirus (CMV) promoter to drive expression in vascular smooth muscle cells both in vitro and in vivo. In contrast to the CMV promoter it is activated in a Ca(2+)-dependent manner after endoplasmic reticulum depletion and allows the transgene expression only in proliferative/diseased cells. Overexpression of sarco/endoplasmic reticulum (SR/ER) Ca(2+) ATPase 2a under the control of this NFAT promoter inhibits restenosis after angioplasty in rats. In conclusion, this promoter may be useful for gene therapy in vascular proliferative diseases and other diseases involving upregulation of the NFAT pathway.

Effect of cyclopiazonic acid, an inhibitor of the sarcoplasmic reticulum Ca-ATPase, on skeletal muscles from normal and mdx mice.

AIM: In this study, we investigated Ca2+ loading by the sarcoplasmic reticulum in skeletal muscle from mdx mice, an animal model of human Duchenne's muscular dystrophy, at two stages of development: 4 and 11 weeks. METHOD: Experiments were conducted on fast- (extensor digitorum longus, EDL) and slow- (soleus) twitch muscles expressing different isoforms of Ca2+-ATPase, which is responsible for the uptake of Ca2+ by the sarcoplasmic reticulum. RESULTS: In sarcoplasmic reticulum vesicles, the ATP-dependent activity and sensitivity to cyclopiazonic acid (CPA), an inhibitor of the sarcoplasmic reticulum Ca2+-ATPase, were similar in mdx and normal EDL muscle. Furthermore, in chemically-skinned fibres from both normal and mdx muscles, the presence of CPA induced a decrease in Ca2+ uptake by the sarcoplasmic reticulum. However, the sensitivity to CPA was lower in mdx EDL muscle than in normal muscle. In addition, in EDL muscle from 4-week-old mdx mice, the expression of the slow Ca2+-pump isoform (SERCA2a) was significantly increased, without any accompanying change in slow myosin expression. In contrast, the expression and function of the Ca2+-ATPase in mdx soleus muscles at 4- and 11-weeks of development did not differ from those in age-matched controls. CONCLUSION: These findings show that in dystrophic muscle, where the Ca2+ homeostasis was perturbed, the Ca2+ handling by the sarcoplasmic reticulum was altered in fast-twitch muscle, and this was associated with the expression of the slow isoform of SERCA. In these muscles, reduced Ca2+ uptake could then contribute to an elevated concentration of Ca2+ in the cytosol, and also to Ca2+ depletion of the sarcoplasmic reticulum.

Functional coupling between the caffeine/ryanodine-sensitive Ca2+ store and mitochondria in rat aortic smooth muscle cells.

We investigated the role of mitochondria in the agonist-induced and/or caffeine-induced Ca2+ transients in rat aortic smooth muscle cells. We explored the possibility that proliferation modulates the coupling between mitochondria and endoplasmic reticulum. Ca2+ transients induced by either ATP or caffeine were measured in presence or absence of drugs interfering with mitochondrial activity in freshly dissociated cells (day 1) and in subconfluent primary culture (day 12). We found that the mitochondrial inhibitors, rotenone or carbonyl cyanide m-chlorophenylhydrazone, as well as the permeability transition pore inhibitor, cyclosporin A, had no effect on the ATP-induced Ca2+ transient at either day 1 or day 12, but prevented caffeine-induced cytosolic Ca2+ increase at day 12 but not at day 1. Close connections between ryanodine receptors and mitochondria were observed at both day 1 and 12. Thapsigargin (TG) prevented ATP- and caffeine-induced Ca2+ transients at day 1. At day 12, where only 50% of the cells were sensitive to caffeine, TG did not prevent the caffeine-induced Ca2+ transient, and prevented ATP-induced Ca2+ transient in only half of the cells. Together, these data demonstrate that rat aortic smooth muscle cells at day 1 have an ATP- and caffeine-sensitive pool, which is functionally independent but physically closely linked to mitochondria and totally inhibited by TG. At day 12, we propose the existence of two cell populations: half contains IP3 receptors and TG-sensitive Ca2+ pumps only; the other half contains, in addition to the IP3-sensitive pool independent from mitochondria, a caffeine-sensitive pool. This latter pool is linked to mitochondria through the permeability transition pore and is refilled by both TG-sensitive and insensitive mechanisms.

Functional regions in the essential light chain of smooth muscle myosin as revealed by the mutagenesis approach.

The endogenous essential light chain (LC17) of myosin from intestine smooth muscle was replaced with mutated essential light chains prepared using recombinant techniques. Complete exchange was observed with histidine-tagged derivatives of LC17a, LC17b and E122A-LC17a (LC17a and LC17b are the usual constituants of smooth muscle myosin), with small changes in the ATPase activity of reconstituted myosins. Much less exchange was observed with the light-chain derivative lacking the last 12 amino acid residues, demonstrating the importance of this segment, which may act as one arm of a pair of pincers to bind the myosin heavy chain.

Intracellular Ca(2+) handling in vascular smooth muscle cells is affected by proliferation.

Despite intensive interest in the dedifferentiation process of vascular smooth muscle cells, very little data are available on intracellular Ca(2+) signaling. The present study was designed to investigate the evolution of the intracellular Ca(2+) pools when rat aortic smooth muscle cells (RASMCs) proliferate and to define the mechanisms involved in the functional alterations. RASMCs were cultured in different conditions, and [Ca(2+)](i) was measured by use of fura 2. Expression of the sarco(endo)plasmic reticulum Ca(2+) pumps (SERCA2a and SERCA2b), Ca(2+) channels, the ryanodine receptor (RyR), and the inositol trisphosphate receptor (IP3R) was studied by reverse transcription-polymerase chain reaction and immunofluorescence. Antibodies specific for myosin heavy chain isoforms were used as indicators of the differentiation state of the cell, whereas an anti-proliferating cell nuclear antigen antibody was a marker of proliferation. SERCA2a, SERCA2b, RyR3, and IP3R-1 mainly were present in the aorta in situ and in freshly isolated RASMCs. These cells used the 2 types of Ca(2+) channels to release Ca(2+) from a common thapsigargin-sensitive store. Proliferation of RASMCs, induced by serum or by platelet-derived growth factor-BB, resulted in the disappearance of RyR and SERCA2a mRNAs and proteins and in the loss of the caffeine- and ryanodine-sensitive pool. The differentiated nonproliferative phenotype was maintained in low serum or in cells cultured at high density. In these conditions, RyR and SERCA2a were also present in RASMCs. Thus, expression of RyR and SERCA2a is repressed by cell proliferation, inducing loss of the corresponding Ca(2+) pool. In arterial smooth muscle, Ca(2+) release through RyRs is involved in vasodilation, and suppression of the ryanodine-sensitive pool might thus alter the control of vascular tone.

Role of the C-terminal extremities of the smooth muscle myosin heavy chains: implication for assembly properties.

The two light meromyosin isoforms from rabbit smooth muscle were prepared as recombinant proteins in Escherichia coli. These species which differed only by their C-terminal extremity showed the same circular dichroism spectra and endotherms in measurements of differential scanning calorimetry. Their solubility properties were different at pH 7.0 in the absence of monovalent salts. Their paracrystals formed at low pH differed by their aspect and number. These data suggest a role for the C-terminal extremity of myosin heavy chains in the assembly of myosin molecules in filaments and consequently in the contractility of smooth muscles.

Denervation of rabbit gastrocnemius and soleus muscles: effect on muscle-specific enolase.

We report here, for the first time, the expression of the muscle-specific isoform of the glycolytic enzyme, enolase (EC 4.2.1. 11) (beta enolase), in rabbit skeletal muscles. We have analysed the fast-twitch gastrocnemius and the slow-twitch soleus muscles during normal postnatal development and following denervation. We show that, in rabbit, as already described in rodents, beta enolase gene expression behaves as a good marker of the fast-twitch fibers. In soleus muscle, the beta enolase transcript level is 10-20% of that found in gastrocnemius. Denervation, performed at 8 postnatal days, induces an important drop of beta enolase transcript levels in both developing soleus and gastrocnemius muscles, with a 80% decrease observed 1 week after denervation in the operated muscles, as compared to the corresponding contralateral muscles. Thereafter, the beta enolase transcript level continues to decrease in the fast-twitch muscle, with the beta enolase subunit being detectable only in the atrophic fast-twitch fibers. In contrast, the beta transcript level tends to increase in the denervated slow-twitch muscle, reaching about 50% of that in contralateral soleus, at 7 weeks after surgery. The level of beta enolase transcripts still expressed after denervation seems to stabilize at the same low level in both types of inactive muscles. This suggests that the small fraction of beta enolase expression which is not controlled by the nerve, or by the contractile activity imposed by it, is independent of the muscle phenotype.

Cardiac consequences of prolonged exposure to an isolated increase in aortic stiffness.

In elderly patients, aortic stiffness is a major determinant of increased end-systolic stress leading to left ventricular (LV) hypertrophy with impaired cardiac performance. However, in a rat model of aortic elastocalcinosis (induced by vitamin D(3)-nicotine [VDN] treatment), brief exposure (1 month) to increased aortic stiffness modified neither cardiac function nor cardiac structure. Here we report the impact of longer exposure (3 months) to aortic stiffness. Three months after induction of aortic stiffness, aortic characteristic impedance was measured in awake rats, 8 control and 10 VDN. Stroke volume was measured (electromagnetic probe) at baseline and after acute volume overload. LV weight/body weight ratio, collagen, and myosin heavy chain (MHC) contents were determined. Although aortic characteristic impedance increased (controls, 32+/-2; VDN rats, 50+/-8 10(3) dyne. s/cm(5); P=0.0248), stroke volume was maintained in VDN rats at baseline (controls, 223+/-18; VDN, 211+/-13 microL) and after volume overload (controls, 378+/-14; VDN, 338+/-15 microL). However, LV weight/body weight ratio (controls, 1.54+/-0.07; VDN, 1.73+/-0.05 g/kg; P=0.0397) and LV collagen content (controls, 31+/-4; VDN, 52+/-4 microgram/g dry wt; P=0.0192) increased. A shift from alpha-MHC (controls, 82+/-2%; VDN, 69+/-3%; P=0.0056) to beta-MHC (controls, 18+/-2%; VDN, 31+/-3%; P=0. 0056) was also observed. Three months' exposure to increased aortic stiffness in VDN rats induced LV hypertrophy with moderate interstitial fibrosis and a shift in the MHC-isoform pattern. Such structural adaptation maintains LV performance.

Sarcoplasmic reticulum in vascular cells in hypertension and during proliferation.

1. Multiple sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) and two types of sarcoplasmic reticulum Ca2+ channels, the ryanodine receptor and the inositol 1,4,5 triphosphate (IP3) receptor are expressed. The heterogeneity of the Ca2+ pumps and Ca2+ channels in vascular cells will be discussed. 2. An age-related change in expression of the SERCA isoforms is observed in smooth muscle cells. 3. The sarcoplasmic reticulum Ca(2+)-uptake rate and the level of SERCA 2 mRNA are different in thoracic than in abdominal aortas and in aortas from spontaneously hypertensive rats than from normotensive rats. 4. Proliferation of vascular smooth muscle cells is associated with major changes in intracellular Ca(2+)-handling mechanisms.

Cellular distribution of Ca2+ pumps and Ca2+ release channels in rat cardiac hypertrophy induced by aortic stenosis.

BACKGROUND: The response of ventricular myocytes to pressure overload is heterogeneous and not spatially coordinated. We investigated whether or not the alterations in SERCA and RyR gene expression are homogeneous within the myocardium. METHODS AND RESULTS: The cellular distribution of mRNAs and proteins encoding the 2 sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) isoforms (SERCA 2a and 2b) and 2 Ca2+ release channels (the ryanodine receptor, RyR, and the IP3 receptor, IP3R) were analyzed by in situ hybridization and immunofluorescence, respectively. Analyses were performed during early (1 and 5 days) and late (1 month) stages of cardiac hypertrophy induced in rat by thoracic aortic stenosis (AS). The results indicated that 1 and 5 days after AS, the cellular distribution of SERCA 2a and RyR2 mRNAs in right ventricle and atrium was similar to controls but the mRNA levels appeared to decrease in some areas of the left ventricle (LV). One month after AS, the distribution of SERCA 2a mRNA and protein became heterogeneous throughout the LV, whereas RyR2 mRNA and protein levels were decreased in a homogeneous manner. SERCA 2b, poorly expressed in both cardiomyocytes and vessels of controls, was increased 4-fold 1 month after AS in coronary arteries only. In both sham (Sh) and AS, SERCA 3 and IP3R mRNAs were mainly found in the vessels. CONCLUSIONS: In severe hypertrophy, decreased accumulation of SERCA 2a was heterogeneous and not compensated by an induction of SERCA 2b in the cardiomyocytes. Decrease in RyR2 expression was more homogeneous and not compensated by an increased IP3R expression.

Effects of sustained low-flow ischemia on myocardial function and calcium-regulating proteins in adult and senescent rat hearts.

OBJECTIVE: Both aging and myocardial ischemia are associated with alterations of calcium-regulating proteins. We investigated the effects of graded levels of low-flow ischemia on myocardial function and on SR Ca(2+)-ATPase (SERCA2), Na(+)-Ca2+ exchanger (NCX) and ryanodine receptor (RyR2), at mRNA and protein levels in both adult and senescent myocardium. METHODS: Isolated hearts from 4 and 24 month old (mo) rats were retrogradely perfused during 180 min at 100% (100% CF, n = 11 and n = 11 respectively. 30% (30% CF, n = 10 and n = 12) or 15% (15% CF, n = 13 and n = 8) of their initial coronary flow, and active tension and coronary resistance (in % of their baseline value) were recorded. After 180 min of perfusion. NCX, RyR2 and SERCA2 mRNAs (in % of age-matched 100% CF group value) and protein levels were quantitated in the left ventricles by slot blot and Western blot analysis, respectively. RESULTS: In 24 mo hearts, low-flow ischemia induced a greater fall in active tension (-65 +/- 7% vs. -40 +/- 4% in 4 mo 30% CF, p, 0.01 and -82 +/- 2% vs. -60 +/- 5% in 4 mo 15% CF groups, p < 0.05 after 15 min of ischemia) and a greater increase in coronary resistance (+357 +/- 44% vs. +196 +/- 39% in 4 mo 30% CF, p < 0.05 and +807 +/- 158% vs. +292 +/- 61% in 4 mo 15% CF groups, p < 0.001 after 15 min of ischemia). An increased accumulation of SERCA2 (+36% and NCX (+46%) transcripts, but not RyR2, already occurred in 24 mo 30% CF group while the 3 transcripts accumulated in 24 mo 15% CF group. In 4 mo rats SERCA2 (+26%), NCX (+35%) and RyR2 (+81%) mRNA levels only increased in the 15% CF group. Corresponding calcium-regulating protein levels were unaltered whatever the degree of flow reduction in both 4 mo and 24 mo hearts. CONCLUSION: Low-flow ischemia does not induce calcium-regulating protein loss in both adult and senescent hearts. The increase in mRNAs coding for calcium-handling proteins and the impairment of myocardial function which occur at a lesser degree of coronary flow reduction in senescent hearts, indicate a higher vulnerability to low-flow ischemia during aging.

Characterization of the 3' end of the mouse SERCA 3 gene and tissue distribution of mRNA spliced variants.

The sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA) type 1 and 2 genes are alternatively spliced at their 3' end. We hypothesized that similar mechanism may occur for SERCA 3. Two spliced variants were identified by RNase protection analysis. We then isolated and sequenced the 3' end portion of the mouse SERCA 3 gene, and confirmed the presence of an alternative mRNA transcript by sequencing a cDNA fragment obtained by RT-PCR. Tissue distribution of the alternatively spliced mRNAs was studied by RT-PCR: SERCA 3b was the only isoform expressed in endothelial cells from aorta and heart and also was the major isoform in lung and kidney whereas SERCA 3a and 3b were coexpressed in trachea, intestine, thymus, spleen, and fetal liver.

The Sarco(endo)plasmic Reticulum Ca(2+)-ATPases in the Cardiovascular System During Growth and Proliferation.

The sarco(endo)plasmic reticulum Ca(2+)-ATPase is present in all cell types and is essential in restoring a low cytosolic Ca(2+) concentration after cell activation. In the past years, six different isoforms encoded by three genes have been identified by cDNA cloning. Some of these isoforms are expressed in the cardiovascular system, and their expression is regulated during proliferation that occurs during ontogenic development as well as in pathological cell growth. This article reviews the new features concerning isoform diversity of the sarco(endo)plasmic reticulum Ca(2+)-ATPase and modulation of expression of these isoforms in cardiac and vascular cells.

Sarcoplasmic reticulum function in determining atrioventricular contractile differences in rat heart.

The relationships between the contractile characteristics and the sarcoplasmic reticulum (SR) function of rat atrial and ventricular trabeculae were compared. The isometric developed tension (DT) and the rates of contraction (+ dT/dt) and relaxation (-dT/dt) normalized to cross-sectional area were 3.7, 2.2, and 1.8 times lower, respectively, in intact atrial strips compared with ventricular strips, whereas + dT/dt and -dT/dt (normalized to DT) were 2.3 and 2.8 times higher, respectively, in atria. Atria exhibited a maximal potentiation of DT after shorter rest periods than ventricles and a lower reversal for prolonged rest periods. Caffeine-induced tension transients in saponin-permeabilized fibers suggested that the Ca2+ concentration released in atrial myofibrils reached a lower maximum and decayed more slowly than in ventricular preparations. However, the tension-time integrals indicated an equivalent capacity of sequestrable Ca2+ in SR from both tissues. In atrial, as in ventricular myocardium, the SR Ca2+ uptake was more efficiently supported by ATP produced by the SR-bound MM form of creatine kinase (CK; MM-CK) than by externally added ATP, suggesting a tight functional coupling between the SR Ca2+ adenosinetriphosphatase (ATPase) and MM-CK. The maximal rate of oxalate-supported Ca2+ uptake was two times higher in atrial than in ventricular tissue homogenates. The SR Ca(2+)-ATPase 2a mRNA content normalized to 18S RNA was 38% higher in atria than in ventricles, whereas the amount of mRNA encoding the alpha-myosin heavy chain, calsequestrin, and the ryanodine receptor was similar in both tissues. Thus a lower amount of readily releasable Ca2+ together with a faster uptake rate may partly account for the shorter time course and lower tension development in intact atrial myocardium compared with ventricular myocardium.

Expression of the cardiac ryanodine receptor in the compensated phase of hypertrophy in rat heart.

OBJECTIVES: Abnormal calcium handling is a general feature of cardiac hypertrophy and alteration in the expression of SR proteins has been suggested to be involved in this alteration. To determine the expression of the cardiac ryanodine receptor (Ry2) gene during compensatory hypertrophy, we studied the mRNA and protein accumulation in left ventricles from rats with 30 to 100% hypertrophy. METHODS: Cardiac hypertrophy was obtained after 1 month of aortic constriction. Ry2 mRNA was analyzed by RNase protection assay, Northern and slot blots, and Ry2 protein by high-affinity [3H]ryanodine binding and Western blot. RESULTS: We demonstrate that: (1) the cardiac Ry2 mRNA concentration is decreased by 50% in severe hypertrophy; (2) both the density of the high-affinity sites and the Ry2 protein level are decreased by 25%; (3) the decrease in the mRNA and protein levels and the number of high-affinity sites are highly correlated to the severity of hypertrophy. CONCLUSION: Our results suggest that, as for SR Ca(2+)-ATPase, there is either a downregulation or a lack of upregulation of the gene coding for the Ry2 in compensatory hypertrophy. The decreased density of Ry2 may alter SR Ca2+ transport and contribute to the impaired Ca2+ handling by slowing the Ca2+ movements.

Sarco(endo)plasmic reticulum Ca2+ pump and metabolic enzyme expression in rabbit fast-type and slow-type denervated skeletal muscles. A time course study.

Recent reports by d'Albis et al. have shown that denervation of 8-day-old rabbit fast-twitch muscle (gastrocnemius) leads to the transformation of the muscle towards a slow phenotype but the changes towards slow-type myosin isoforms and contractile properties of the muscle were temporally uncoordinated. We analyzed the time course of the effects of denervation of the gastrocnemius on the expression of the sarcoplasmic reticulum calcium pump isoforms (SERCA) and on the metabolic state of the muscle. Northern-blot analysis showed a rapid loss of the fast Ca2+ pump isoform (SERCA 1) mRNA from the denervated gastrocnemius which became of the oxidative type. The changes observed were complete as early as 35 days post-natal, i.e at the time when changes in contractile properties were previously observed. Denervation of the slow-twitch soleus led to a 50% decrease in the level of the slow Ca2+ pump isoform (SERCA 2) mRNA and was without effect on the metabolic state of the muscle. These findings extend previous results suggesting that in rabbit, continuous innervation is required for differentiation of fast-twitch muscles but is not an absolute requirement for differentiation of the slow-twitch muscle.

Sarcoplasmic reticulum Ca2+ pumps in heart and diaphragm of cardiomyopathic hamster: effects of perindopril.

The polymyopathy of the Syrian hamster is associated with alterations of cellular calcium regulation and contractile performance of cardiac and skeletal muscles and, in particular, the diaphragm. Angiotensin-converting enzyme (ACE) inhibitors have been shown to preserve contractile performance. Therefore we analyzed the expression of the genes coding for the sarco(endo)plasmic reticulum Ca(2+)-adenosinetriphosphatase (SERCA) in heart and diaphragm of the cardiomyopathic Syrian hamster (CSH) from the dilated strain Bio 53-58, and we tested the influence of ACE inhibition on accumulation of the different SERCA mRNAs. In the diaphragm of healthy hamsters, two SERCA mRNA isoforms were present: SERCA 1 and SERCA 2. At 6 mo of age, the myopathic process resulted in decreased levels of SERCA 1, whereas the level of SERCA 2 was unchanged. The ACE inhibitor perindopril (1 mg.kg-1.day-1), administered by force feeding from 1 to 6 mo of age, had no effect on the SERCA 1 mRNA level. In heart, the myopathy was associated with a depressed level of SERCA 2 mRNA in 9- but not in 6-mo-old animals. Perindopril treatment from 6 to 9 mo reversed cardiac hypertrophy and the relative decrease in SERCA 2 mRNA level. Preventive treatment with perindopril from 1 to 9 mo tended to prevent (not significantly) the development of cardiac hypertrophy and reduction in SERCA gene expression. In conclusion, the myopathic process affects SERCA gene expression in the diaphragm and subsequently in the heart. Perindopril treatment can prevent SERCA mRNA loss in heart but not in diaphragm.