Search PubMed⌕ Search

Biomedical subjects

J Schaper

Publications and source records attributed to J Schaper.

At least 55 records · Page 3Linked to original sources
PubMed2000

Antisense oligonucleotide experiments elucidate the essential role of titin in sarcomerogenesis in adult rat cardiomyocytes in long-term culture.

An essential role of titin as a molecular ruler in sarcomerogenesis has been frequently discussed. In this study, we tested the hypothesis that the expression of titin is a prerequisite for thick filament incorporation into sarcomeres by using an antisense oligonucleotide approach to interfere with titin translation in the de-/redifferentiation model of adult rat cardiomyocytes (ARC) in long-term culture. As a first step, the growth pattern ranging from rod shape to round and later to spreading cells and the cell surface area of ARC were quantitatively evaluated and standardized. This represents the basis for experiments interfering with sarcomere formation using three different antisense phosphorothioate oligonucleotides (S-ODN) at a dosage of 10 microM specific for titin mRNA. Presence of fluorescein labeled S-ODN in ARC indicated cellular uptake and both, antisense and random S-ODN, induced a significant increase in cell size as compared with control untreated ARC. At days 12 and 16 in culture, antisense S-ODN treatment resulted in reduced expression of titin and disturbance of myosin incorporation into sarcomeres, evident by diffuse myosin labeling and a significantly decreased area of regular myosin cross-striation (control 75%, day 12 S-ODN 20%, day 16 14%) shown by laser scanning confocal microscopy. Cellular integrity indicated by presence of alpha-actinin was not disturbed. These findings provide evidence for the role of titin as a template for myosin incorporation and therefore as a prerequisite for sarcomerogenesis.

Animals↗
PubMed1999

Spatiotemporal development and distribution of intercellular junctions in adult rat cardiomyocytes in culture.

The mode of development of the intercalated disk (ID) is largely unknown, and the hypothesis was tested that the assembly of cell adhesion junctions may precede the formation of gap junctions (GJ) in developing ID in adult rat cardiomyocyte (ARC) in long-term culture. Immunostaining for connexin 43 (Cx43) and for cell adhesion junction proteins (N-cadherin, catenins, and desmoplakin) in single- and double-label techniques was analyzed and quantified by confocal and electron microscopy. All proteins investigated disappeared 48 hours after ARC isolation and reappeared parallel to redifferentiation of ARC. The newly formed ID, observed after 5 days, showed the presence of N-cadherin, catenins, and desmoplakin, low levels of Cx43, and absence of ultrastructurally discernible gap junctions. A progressive incorporation of Cx43 within ID was observed after 6 days, when cell adhesion junction proteins were already organized as zipper-like structures. Quantitative confocal analysis revealed a progressive augmentation of the fluorescence intensity of Cx43, associated with an increase in both the number and size of GJ, resulting in a substantial increase in the percentage of total GJ length per reassembled ID from 1.67% (day 6) to 15.58% (day 12). In the present study, we show that (1) the formation of the ID can be followed in ARC in culture and (2) the assembly of the adhering type of junction is the prerequisite for subsequent GJ formation within the ID. These findings may have clinical relevance in elaborating strategies for using myocardial grafts and for the potential restoration of GJ communication in cardiac diseases.

Age Factors↗
PubMed1999

Effect of long-term ACE inhibition on myocardial tissue in hypertensive stroke-prone rats.

The aim of the study was to investigate the influence of long-term ACE inhibition with ramipril on myocardial hypertrophy and its molecular background in spontaneously hypertensive stroke-prone rats (SHR-SP). Therefore, 1-month-old pre-hypertensive SHR-SP were randomized into three groups and exposed lifelong via drinking water to 1 mg/kg/day ramipril (anti-hypertensive dose, RHI), 10 micrograms/kg/day ramipril (non-anti-hypertensive dose, RLO) or placebo. After 15 months cardiac tissue was collected from ten rats each for immunohistochemistry and Northern blot analysis of structural proteins, proteins of the extracellular matrix and several growth factors. Results showed that RHI, but not RLO, treatment prevented development of myocyte hypertrophy (ANP). Furthermore, unlike placebo-treated rats, the ramipril-treated animals had no evidence of degeneration and loss of structural proteins (alpha -actinin), inflammatory infiltrates (CD45) and deposition of extracellular matrix proteins (collagen, fibronectin, vimentin). Only in RHI-treated animals, mRNA levels for TGF- beta(1)as well as of collagen alpha(1)(I) and fibronectin were downregulated compared to placebo-treated animals. In contrast, VEGF mRNA levels increased significantly in both groups of ramipril-treated animals v. placebo-treated SHR-SP. Thus, the reported life prolonging effect of high doses of ramipril which is associated with prevention of hypertension and hypertrophy is accompanied by prevention of the development of necrosis and fibrosis. The role of VEGF, however, seems to be independent of this effect.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1999

The role of apoptosis in dilated cardiomyopathy.

Dilated cardiomyopathy is a cardiac disease of unknown origin which is characterized by the gradual development of cardiac failure. Apoptosis, i.e. suicidal programmed cell death, may play a role in the development of heart failure. Only few studies have been carried out until now that describe the rate of apoptosis in human hearts with dilated cardiomyopathy. The numbers reported vary widely. This is also true for studies in different other cardiac diseases such as myocardial infarction or hibernating myocardium. The methods used to identify apoptosis include electron microscopy, labeling of the DNA fragments (TUNEL), staining with the Hoechst dye, annexin V labeling and documentation of DNA fragmentation using gel electrophoresis (laddering). None of these methods are totally reliable in tissue sections in which apoptosis is not a frequent event when they are not combined with another technique, e.g. TUNEL with electron microscopy or laddering. This has, however, only rarely been done. These technical difficulties may be the reason for the wide variation in the rate of apoptosis reported. From our own data we conclude that apoptosis plays a significant role in acute ischemia and in hibernating myocardium but its significance in the progression to heart failure in dilated cardiomyopathy has still to be established.

Apoptosis↗
PubMed1999

Tuberculoma of the pons.

A case of a pontine tuberculoma in a 12-year-old male from Somalia is presented. Cranial magnetic resonance imaging, performed in Abu Dhabi 1 month before admission in Germany revealed a tumorous lesion in the pons with surrounding edema; a presumptive diagnosis of a pontine glioma was made. Chest radiograph disclosed a pulmonary infiltrate with cavitation, as well as hilar and left mediastinal lymphadenopathy, suggestive of active tuberculosis. Tuberculostatic therapy led to an improvement of the patient's clinical status and a significant reduction in the size of the pontine tuberculoma. Intracranial tuberculomas rarely are seen in industrialized countries. They should, however, be considered as an important part of the differential diagnosis of intracranial space-occupying lesions. In most cases, as in this patient, conservative therapy provides good or excellent results.

Antitubercular Agents↗
PubMed1999

Cardiac microvascular endothelial cells express alpha-smooth muscle actin and show low NOS III activity.

We established a culture system of porcine coronary microvascular endothelial cells (MVEC) with high cellular yield and purity >98%. Endothelial origin was confirmed by immunostaining, immunoblotting and fluorescence-activated cell sorter (FACS) analysis using low-density lipoprotein uptake, CD31, von Willebrand factor, and the lectin Dolichos biflorus agglutinin. MVEC were positive for alpha-smooth muscle actin in culture and in myocardium, as confirmed by FACS. Of the primary MVEC, approximately 30% expressed nitric oxide synthase (NOS) III in numbers decreasing from the first passage (6 +/- 1%) to the second passage (4 +/- 1%; P < 0.001 vs. primary isolates), whereas approximately 100% of aortic endothelial cells (AEC) expressed NOS III. In AEC, NOS III activity (pmol citrulline. mg protein-1. min-1) was 80 +/- 10 and was nearly abolished in the absence of calcium (5 +/- 1, P < 0.001). In primary MVEC, however, NOS III activity in the presence and absence of calcium was 20 +/- 4 and 25 +/- 5, respectively. We conclude that cardiac MVEC, in contrast to AEC, contain alpha-smooth muscle actin, show low-grade NOS III activity, and provide a suitable in vitro system for the study of endothelial pathophysiology.

Actinin↗
PubMed1999

Microvascular endothelial cells remodel cultured adult cardiomyocytes and increase their survival.

We investigated the paracrine effect of cardiac microvascular endothelial cells (MVEC) on cultured adult rat cardiomyocytes (ARC). ARC were exposed for 8 days to serum-free medium (CM) conditioned by MVEC. Controls were grown in FCS or FCS-free medium. Protein synthesis of CM-stimulated ARC increased twofold versus 5% FCS-stimulated cells until day 8. Seventy-nine percent of CM-treated myocytes survived, whereas only twenty-four percent of FCS-free ARC retained viability. The phenotype of myocytes exposed to CM was different from control. Analysis by confocal laser microscopy of CM-stimulated myocytes showed actin staining throughout the whole cell body up to the peripheral extensions, with concomitant appearance of myomesin in a cross-striated pattern. The reexpression of fetal alpha-smooth muscle actin determined immunohistochemically and by Western blot increased from day 6 in CM-treated cells, whereas ARC grown in up to 20% serum were negative. These effects could not be mimicked by any of the other cardioactive substances tested here, indicating a novel trophic factor in CM.

Actins↗
PubMed1999

Ultrastructural assessment of myocardial necrosis occurring during ischemia and 3-h reperfusion in the dog.

To determine whether myocardial necrosis may occur during postischemic reperfusion, electron microscopy was used to identify morphological features of irreversible injury in myocardial samples taken from anesthetized dogs with 90-min ischemia and 0-, 5-, 90-, or 180-min reperfusion. In samples without detectable collateral blood flow, necrosis was almost complete, whether or not the myocardium was reperfused. In samples with collateral flow, necrosis was more frequent after 180-min reperfusion than in the absence of reperfusion, despite similar collateral flows in the two groups. Excess of necrosis after 180-min reperfusion was evident in endocardium (ischemia only: 4 of 13, 180-min reflow: 14 of 20; P = 0. 03) and midwall (ischemia only: 9 of 25, 180-min reflow: 29 of 45; P = 0.02). Multiple logistic regression with variables of collateral flow and transmural position was used to determine risk of irreversible injury in 111 samples from ischemic myocardium without reperfusion (model predictive accuracy = 75%, P < 0.00001) and to predict risk of necrosis in myocardium reperfused for 180 min. Of 65 samples from endocardium and midwall with detectable collateral flow, the model predicted necrosis in 23 samples but necrosis was observed in 43 samples (P < 0.01). Reperfusion duration was a determinant of frequency of irreversible injury. Multiple logistic regression for 186 samples from myocardium reperfused for 5, 90, or 180 min showed that reperfusion duration was an independent predictor of irreversible injury (P = 0.0003) when collateral flow and transmural location were accounted for. These findings are consistent with the occurrence of necrosis during reperfusion in myocardium exposed to substantial, prolonged ischemia but with sufficient residual perfusion to avoid necrosis during the period of flow impairment.

Animals↗
PubMed1999

Species differences in arsenic-mediated renal copper accumulation: a comparison between rats, mice and guinea pigs.

1. Administration of arsenite leads to an accumulation of copper in the rat kidney. Owing to the high retention of arsenic in the erythrocytes, however, the rat is considered to possess special toxicokinetics of arsenic and is therefore considered less comparable with other species in this respect. 2. Therefore, we compared the effect of dietary arsenite in mice and guinea pigs with that in rats. Each species was divided into four groups of animals according to the diets fed which contained increasing concentrations of sodium arsenite (NaAsO2; 0, 10, 30 and 60 mg As/kg of diet). Animals were killed after 1, 2 and 3 weeks. Tissues were sampled and analyzed for arsenic and other trace metals (Cu, Fe, Zn and Mn). 3. Compared to controls with copper levels of about 10 microg Cu/g wet wt. in the renal cortex, dietary administration of arsenite up to 60 mg As/kg of diet for 3 weeks to rats increased cortical levels to 65 microg Cu/g wet wt. An increase of renal copper levels similar to that in rats, was only observed in guinea pigs but not in mice. Renal copper accumulation in guinea pigs was time- and concentration-dependent as in rats. Feeding a diet with 60 mg As/kg for 3 weeks increased cortical copper levels from about 6 - 40 microg Cu/g wet wt. Renal copper levels in mice as well as other trace metal levels in guinea pigs and mice were not essentially altered by dietary arsenite. 4. The study shows that the renal copper-arsenic interaction is not restricted to the rat. Since in rats and guinea pigs, but not in mice, arsenic accumulated in the kidney rather similarly, a common mechanism is suggestive. As it was previously shown in rats that only inorganic arsenic is involved in this interaction, a rapid conversion of the inorganic form into methylated metabolites as in mice may diminish the extent of the renal copper accumulation whereas the lack of, or a less efficient, methylation as in guinea pigs or rats increases it.

Animals↗
PubMed1998

Characterization of nebulette and nebulin and emerging concepts of their roles for vertebrate Z-discs.

Nebulin is an 800 kDa large actin-binding protein specific to skeletal muscle and thought to act as a molecular template that regulates the length of thin filaments. Recently, a 100 kDa nebulin-like protein has been described in the avian cardiac muscle and referred to as nebulette. We have determined the full-length (8 kb) cDNA sequence of the human nebulette. Its open reading frame (3044 bp) encodes a 109 kDa protein that shares extensive similarity with the C-terminal region of human nebulin. The C-terminal regions of nebulin and nebulette are identical in domain organization and share a family of highly related C-terminal repeats, a serine-rich domain with potential phosphorylation sites, and an SH3 domain. Immunoelectron-microscopy suggests that the C-terminal 30 kDa of nebulin and nebulette filaments integrate into the Z-disc lattice, whereas their N termini appear to project into the I-band. Gene mapping studies assign the human nebulette gene to chromosome 10p12, whereas the nebulin gene has been previously assigned to 2q21. Evolutionary constraints appear to have maintained identical modular arrangements in these two independent genes. Comparison of nebulin and nebulette cDNAs demonstrates that a subgroup of repeats within the C-terminal regions is regulated tissue-specifically and stage-dependently during development of both molecules. This leads to a substantial diversity of nebulin and nebulette isoforms. Their further study is likely to provide insights into how they contribute to the molecular diversity of Z-discs from different muscle tissues and fiber types.

Actin Cytoskeleton↗
PubMed1998

Progression of myocardial necrosis during reperfusion of ischemic myocardium.

BACKGROUND: The occurrence of myocyte necrosis during reperfusion of ischemic myocardium is controversial. This study measured myocardial 2-deoxyglucose uptake, correlated with histology, to determine whether loss of viability occurred during reperfusion of ischemic myocardium. METHODS AND RESULTS: In 12 anesthetized dogs, the left anterior descending coronary artery was occluded for 90 minutes before 4 hours reperfusion. Myocardial blood flow was measured by microspheres and the tracers 14C-2-deoxyglucose and 18F-2-deoxyglucose were injected intravenously after 5 and 180 minutes of reperfusion, respectively. After 240 minutes, the heart was stained with thioflavin-S (size of no-reflow zone) and triphenyl-tetrazolium chloride (TTC, extent of necrosis). Samples from normal, salvaged, and necrotic myocardium were counted for 14C- and 18F-deoxyglucose and microspheres. With the use of a three-compartment model of 2-deoxyglucose uptake, the rate constant k3 for phosphorylation of 14C- and 18F-2-deoxyglucose was calculated for each sample. Viability was defined as k3> or = 0.125 min(-1) (predictive accuracy 88% versus electron microscopy and 97% versus TTC). Among 58 samples from no-reflow regions, 97% were nonviable after 5 minutes of reperfusion (k3=0.096 +/- 0.027 min[-1]). Among 164 samples from salvaged myocardium, 95% were viable after both 5 and 180 minutes of reperfusion (k3=0.170 +/- 0.056 min[-1] P<.01 versus no-reflow). Among 179 samples from infarcted myocardium, mean k3 after 5 minutes of reperfusion was 0.184 +/- 0.070 min(-1) and 65% of samples were viable, but after 180 minutes of reperfusion mean k3 had decreased to 0.077 +/- 0.032 min(-1) (P<.0001) and 98% of samples were nonviable. CONCLUSIONS: A large proportion of samples from infarcted myocardium are viable at the end of the ischemic period but lose viability during the first hours of reperfusion.

Animals↗
PubMed1998

Monocyte activation in angiogenesis and collateral growth in the rabbit hindlimb.

We have previously shown that monocytes adhere to the vascular wall during collateral vessel growth (arteriogenesis) and capillary sprouting (angiogenesis). In this study we investigated the association of monocyte accumulation with both the production of the cytokines-basic fibroblast growth factor (bFGF) and TNF-alpha-and vessel proliferation in the rabbit after femoral artery occlusion. In particular, we studied the effects of an increase in monocyte recruitment by LPS on capillary density as well as collateral and peripheral conductance after 7 d of occlusion. Monocytes accumulated around day 3 in collateral arteries when maximal proliferation was observed, and stained strongly for bFGF and TNF-alpha. In the lower limb where angiogenesis was shown to be predominant, macrophage accumulation was also closely associated with maximal proliferation (around day 7). LPS treatment significantly increased capillary density (424+/-26.1 n/mm2 vs. 312+/-20.7 n/mm2; P < 0.05) and peripheral conductance (109+/-33.8 ml/min/100 mmHg vs. 45+/-6.8 ml/min/100 mmHg; P < 0.05) as compared with untreated animals after 7 d of occlusion. These results indicate that monocyte activation plays a major role in angiogenesis and collateral artery growth.

Animals↗
PubMed1998

Intramyocardial infusion of FGF-1 mimics ischemic preconditioning in pig myocardium.

Previous studies on the mRNA and protein level suggested a cardioprotective role of FGF-1. These presumed actions of FGF-1 and FGF-2, as well as the underlying mechanisms, were investigated in this study. Human recombinant FGF-1 (0.5 microgram/ml, 20 microliters/min) and FGF-2 (2 micrograms/ml) were applied by means of direct intramyocardial infusion (IM) for 60 min prior to a 60 min LAD-occlusion and 120 min reperfusion. Myocardial infarction compared to the region at risk was significantly decreased by FGF-1 and FGF-2 treatment (FGF-1: 51.8 +/- 7.7%, respectively. FGF-2: 57.3 +/- 6.5% v control 83.4 +/- 2.8%, P < 0.05). The increase in survival time was about 33 min, and equalled that of ischemic preconditioning. This effect was caused by the mitogenic part of the molecule, since infusion of a truncated version of FGF-1 (0.5-1 microgram/ml), lacking mitogenicity but maintaining hemodynamic activity, did not induce cardioprotection (78.3 +/- 0.73% v control 83.4 +/- 2.8%). Suramin (0.5 microgram/ml) prevented the observed cardioprotection (77.0 +/- 1.2% v control 83.4 +/- 2.8%) proving that the cardioprotective effect is receptor-mediated. Genistein (0.5 microgram/ml), an inhibitor of tyrosine kinases, abolished the cardioprotection as well (77.2 +/- 2.4% v control: 83.4 +/- 2.8%). Immunohistochemical staining revealed an uptake and translocation of exogenous FGF-1 to a (peri-)nuclear localization in myocytes and into non-myocytes for FGF-2. We conclude that both FGF-1 and FGF-2 are cardioprotective (FGF-1 being more active on a molar basis), and mimic ischemic preconditioning. Their actions are receptor-mediated and receptor activation is involved. Uptake and transport to a (peri-)nuclear localization, seems to be a pathway of minor relevance, since it could not be blocked by tyrosine kinase receptor inhibition. Tyrosine kinase-coupled receptor occupation in general is not protective as demonstrated by the lack of effect with VEGF-infusion.

Animals↗
PubMed1998

Vascular remodeling and altered protein expression during growth of coronary collateral arteries.

The cellular mechanism of growth of coronary collateral vessels (adaptive arteriogenesis) is still poorly understood. To define a possible role of an altered expression pattern of cellular and matrix proteins in this process we implanted a constricting device around the left circumflex artery in 25 canine hearts and sacrificed the animals at the time of initiation (3 weeks), high activity (6 weeks) and discontinuation (8 weeks) of vessel growth. Methods were electron microscopy, labeling with Ki-67, the TUNEL method and immunofluorescence with confocal laser microscopy. As described earlier, the collateral vessels increased in wall thickness by the formation of a neointima without luminal narrowing. We report here for the first time that extensive vascular remodeling including migration, proliferation and apoptosis in all cell types takes place during the growth phase but not in more mature vessels. The most obvious difference with normal vessels is the reiteration of an embryonal expression pattern in smooth muscle cells of the neointima which includes a significant reduction of desmin and alpha-smooth muscle actin, calponin and vinculin. Fibronectin as a promoter of migration and adhesion was abundant, its antagonist tenascin and chondroitin sulfate showed patchy localization. A completely new finding in arteriogenesis is the involvement of mast cells releasing histamine and serotonin and probably cytokines. Vascular protein expression returned to almost normal at 8 weeks indicating cessation of remodeling. We conclude that in collateral vessel development an altered cellular and matrix protein expression is involved in a drastic case of positive vascular remodeling finally resulting in mature vessels 20-fold increased in size which are capable of maintaining the functional and structural integrity of the myocardium at risk.

Actins↗
PubMed1998

Cardiomyocyte apoptosis in acute and chronic conditions.

Myocytes can die by necrosis or by apoptosis and the characteristics of both kinds of cell death are so typical that a differentiation can be made by histological and molecular-biological methods using electron microscopy, dUTP labeling with fluorescence or peroxidase staining (TUNEL) and the DNA laddering method. However, the problem of quantification of apoptotic cells has not been completely solved because of lack of standardization as well as uncritical use and interpretation of the TUNEL method. Equally, quantification of apoptotic cells is not optimal until now because of three reasons: methodological (overinterpretation of results, no differentiation between myocytes and non-myocytes), experimental (global or regional acute ischemia, chronic conditions such as heart failure or hibernating myocardium), and interpretation (unknown time period for the completion of apoptosis). This problem is reflected in the large differences in incidence of apoptosis reported. Our own data show that in dog myocardium made globally ischemic for 90 min, 8% of the myocytes showed a positive staining for apoptosis (TUNEL method) after 6 h of reperfusion. Despite these results the question of reperfusion injury and the influence of apoptosis still remains open, because it can not be excluded until now that the apoptotic process is initiated during the ischemic period. Studies in hibernating myocardium and chronic heart failure show a similar situation, because of a wide variation of numbers of apoptotic cells and the limited possibility to investigate human tissue. There is no doubt that apoptosis plays an important role in chronic pathophysiological situations such as heart failure and hibernating myocardium but the importance of apoptosis in the acute situation of ischemia/reperfusion still has to be clarified.

Animals↗
PubMed1998

Tumor necrosis factor-alpha is expressed by monocytes/macrophages following cardiac microembolization and is antagonized by cyclosporine.

The time course of expression of TNF-alpha in myocardial wound healing following ischemic injury was investigated in the porcine heart. Microembolization was used to induce focal ischemia and necrosis in hearts of 39 adult pigs. The animals were sacrificed after 3, 6, 12, 24 h, 3 and 7 days, and after 4 weeks, and the myocardial tissue was studied by immunofluorescence using specific antibodies. TNF-alpha containing cells were identified as monocytes/macrophages by double staining with a muramidase antibody. Monocytes/macrophages were the only source of TNF-alpha. Microembolization caused multiple necrotic foci with loss of myocytes in the left ventricular myocardium. These foci contained numerous monocytes/macrophages and showed an inflammatory reaction typical of wound healing followed by replacement with scar tissue. The number of TNF-alpha positive cells increased after 24 h, peaked between 3-7 days and slowly decreased thereafter. Expression of TNF-alpha in monocytes/macrophages was significantly reduced after pretreatment of pigs with cyclosporine or dexamethasone. It is concluded that 1.) in myocardial tissue monocytes/macrophages are the only cell type expressing TNF-alpha, 2.) TNF-alpha is involved in wound healing after ischemia, and 3.) synthesis of TNF-alpha and inflammatory angiogenesis can be inhibited be treatment with either cyclosporine or dexamethasone.

Animals↗
PubMed1998

The internal and external protein scaffold of the T-tubular system in cardiomyocytes.

The transverse tubule system of the cardiomyocyte remains undeformed despite the extreme forces it undergoes during the contraction-relaxation cycle, but the morphological basis for its stability remains unclear. Therefore, we have investigated the architecture and subcellular protein scaffold of the cardiac T-tubules and compared it with that of the costameres and of the free sarcolemma. Tissue samples from normal rat and monkey hearts, and left ventricular tissue from normal and cardiomyopathic human hearts obtained at transplantation surgery were investigated using immunocytochemistry and confocal microscopy and by electron microscopy. In addition, we used a re-differentiation model of isolated, cultured adult rat cardiomyocytes. The cell membrane of the cardiac T-tubules was found to contain the cell-matrix focal adhesion molecules (FAMs) vinculin, talin, the alpha5beta1 integrin and the membrane-associated proteins (MAPs) dystrophin and spectrin. FAMs and MAPs were localized in the T-tubular membrane in a similar pattern: in longitudinally oriented myocytes as transverse punctate lines at the Z-level; in transversally cut myocytes a radial tubular network was found to extend throughout the interior of the cell. Immunolabeling for basement membrane components including collagen IV, fibronectin and laminin showed a colocalization with FAMs and MAPs parallel to the transverse T-tubules. The costameres of the sarcolemma showed a protein composition resembling that of the T-tubules but the intervening segments of free sarcolemma showed absence of FAMs and presence of MAPs. For the first time, we demonstrate the existence and protein composition of the T-tubular scaffold in the human heart. Furthermore, we show that cardiomyocytes from human failing hearts have less abundant but more dilated T-tubules than do experimental animals. These results indicate that the cardiac T-tubular system contains a subcellular scaffold closely resembling that of the costameres. It consists of FAMs, MAPs and basal lamina proteins that confer structural integrity to the cardiac T-tubular membrane during contraction/relaxation cycles.

Animals↗