Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 Apr 25;135(17):1632-1645.
doi: 10.1161/CIRCULATIONAHA.116.024470. Epub 2017 Jan 10.

Sheet-Like Remodeling of the Transverse Tubular System in Human Heart Failure Impairs Excitation-Contraction Coupling and Functional Recovery by Mechanical Unloading

Thomas Seidel  1 , Sutip Navankasattusas  2 , Azmi Ahmad  2 , Nikolaos A Diakos  2 , Weining David Xu  2 , Martin Tristani-Firouzi  2 , Michael J Bonios  2 , Iosif Taleb  2 , Dean Y Li  2 , Craig H Selzman  2 , Stavros G Drakos  1 , Frank B Sachse  1
Affiliations

Sheet-Like Remodeling of the Transverse Tubular System in Human Heart Failure Impairs Excitation-Contraction Coupling and Functional Recovery by Mechanical Unloading

Thomas Seidel et al. Circulation. .

Abstract

Background: Cardiac recovery in response to mechanical unloading by left ventricular assist devices (LVADs) has been demonstrated in subgroups of patients with chronic heart failure (HF). Hallmarks of HF are depletion and disorganization of the transverse tubular system (t-system) in cardiomyocytes. Here, we investigated remodeling of the t-system in human end-stage HF and its role in cardiac recovery.

Methods: Left ventricular biopsies were obtained from 5 donors and 26 patients with chronic HF undergoing implantation of LVADs. Three-dimensional confocal microscopy and computational image analysis were applied to assess t-system structure, density, and distance of ryanodine receptor clusters to the sarcolemma, including the t-system. Recovery of cardiac function in response to mechanical unloading was assessed by echocardiography during turndown of the LVAD.

Results: The majority of HF myocytes showed remarkable t-system remodeling, particularly sheet-like invaginations of the sarcolemma. Circularity of t-system components was decreased in HF versus controls (0.37±0.01 versus 0.46±0.02; P<0.01), and the volume/length ratio was increased in HF (0.36±0.01 versus 0.25±0.02 µm2; P<0.0001). T-system density was reduced in HF, leading to increased ryanodine receptor-sarcolemma distances (0.96±0.05 versus 0.64±0.1 µm; P<0.01). Low ryanodine receptor-sarcolemma distances at the time of LVAD implantation predicted high post-LVAD left ventricular ejection fractions (P<0.01) and ejection fraction increases during unloading (P<0.01). Ejection fraction in patients with pre-LVAD ryanodine receptor-sarcolemma distances >1 µm did not improve after mechanical unloading. In addition, calcium transients were recorded in field-stimulated isolated human cardiomyocytes and analyzed with respect to local t-system density. Calcium release in HF myocytes was restricted to regions proximal to the sarcolemma. Local calcium upstroke was delayed (23.9±4.9 versus 10.3±1.7 milliseconds; P<0.05) and more asynchronous (18.1±1.5 versus 8.9±2.2 milliseconds; P<0.01) in HF cells with low t-system density versus cells with high t-system density.

Conclusions: The t-system in end-stage human HF presents a characteristic novel phenotype consisting of sheet-like invaginations of the sarcolemma. Our results suggest that the remodeled t-system impairs excitation-contraction coupling and functional recovery during chronic LVAD unloading. An intact t-system at the time of LVAD implantation may constitute a precondition and predictor for functional cardiac recovery after mechanical unloading.

Keywords: excitation contraction coupling; heart failure; myocytes, cardiac; recovery of function; ryanodine receptor calcium release channels.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Two-dimensional tile scan of WGA-labeled LV tissue slices obtained from control and HF patient at time of LVAD implantation
Images were acquired with a confocal microscope equipped with a 40× oil immersion lens using a pixel size of 0.2×0.2μm. (A) Tile scan from donor tissue. (B) Magnified view of the boxed region in (A) showing myocytes with dense t-system. (C) Tile scan from HF tissue. (D,E) Magnified views of the boxed region in (C) with myocytes with a sparse and irregular t-system. The remodeled t-system exhibited longitudinal components in the majority of cells. Some examples were marked with arrows. Scale bar in A is 500μm and also applies to C. Scale bar in B is 50μm and also applies to D and E.
Figure 2
Figure 2. Three-dimensional imaging and reconstruction of t-system from control and HF tissue
(A) Confocal microscopic image from control tissue labeled with WGA (red) for extracellular matrix and sarcolemma. A segmented cardiomyocyte is highlighted in green. (B) Magnified view of the boxed regions in (A) showing myocyte with a dense t-system. (C) 3D reconstruction of outer sarcolemma (gray) and t-system (blue) of a section extracted from the cell highlighted in (A). (D) T-tubule highlighted in (C) shown in longitudinal and transverse view. (E) Image from HF tissue labeled as in (A). A segmented cardiomyocyte is highlighted in green. (F) Magnified view of the boxed region in (E) reveals sparse t-system with sheet-like remodeling. Black arrows point to the same sheet-like component of the t-system. (G) 3D reconstruction of outer sarcolemma (gray) and t-system (blue) of a section from the cell highlighted in (E). (H) Remodeled t-system component highlighted in (G) shown in longitudinal and transverse view. Scale bar in A: 40μm. Applies to E. Scale bar in B: 5μm. Applies to F. Scale bar in C: 10μm. Applies to G. Scale bar in D: 2μm. Applies to H.
Figure 3
Figure 3. Three-dimensional analyses of t-tubule geometry in control and HF tissue and imaging of isolated cardiomyocytes
Using an extracellular marker confirm remodeling of t-tubules. (A) Cross-sectional circularity of t-tubules, calculated from the two minor eigenvalues |λ3/λ2|, was lower in HF (n=26) than in control (n=5). (B) Volume-length ratio (V/l) of t-tubules was increased in HF vs control. (C) Mean intracellular sarcolemma distance (ΔSL) was higher in HF cells than in control cells. (D) ΔSL increased with V/l (p<0.01 vs constant model). (E) XY, ZY and XZ cross-sections through three-dimensional image of living isolated cardiomyocyte from HF patient. Dashed blue lines indicate cross-sections. The cell was bathed in 5mg/ml dextran-FITC conjugate (10kDa) as an extracellular marker. Extracellular fluid is shown in white, intracellular space in black. T-sheets were also present in isolated cells (example marked with blue arrow) and exhibited fluorescence, indicating that t-sheet cavities are connected to the extracellular space. Scale bar is 20μm. **p<0.01, ****p<0.0001
Figure 4
Figure 4. Analyses of RyR-sarcolemma distances in tissue from donors and pre-LVAD patients
Confocal microscopic images of (A) control and (C) HF LV tissue labeled with WGA (red) for extracellular matrix and sarcolemma, and for RyRs (green). (B and D) Magnified view of the boxed regions in (A) and (C), respectively. (E) Fraction of RyR fluorescence intensity (IRyR) found at defined sarcolemma distances (ΔSL) in control (black, n=5) and HF (white, n=26). (F) RyR-sarcolemma distance (ΔRyR-SL) in control and HF. Scale bar in A: 40μm. Applies to C. Scale bar in B: 5μm. Applies to D. *p<0.05, **p<0.01
Figure 5
Figure 5. Statistical analyses of clinical and imaging data
Linear regression models of pre-LVAD RyR-sarcolemma distance (ΔRyR-SL) versus (A) pre-LVAD EF (EFpre), (B) post-LVAD EF (EFpost), and (C) EF change during unloading (ΔEF=EFpost−EFpre). Significance of F-statistics against constant model is indicated by p and correlation coefficient by R. Dotted lines indicate 95% confidence intervals. (D) pre-LVAD EF, (E) post-LVAD EF, (F) EF change, (G) HF duration (tHF), and (H) sarcolemma distance (ΔSL) in samples from patients grouped by low (<1μm, black, n=16) and high (>1μm, white, n=10) RyR-sarcolemma distance. **p<0.01 (<1μm vs >1μm), † p<0.001 (EFpost vs EFpre)
Figure 6
Figure 6. Imaging and analyses of calcium transients in HF cardiomyocytes
(A) Confocal microscopic image of Di-8-ANEPPS labeled isolated myoycte with sheet-like remodeled t-system. (B) Fluo-4 images of cell at time 0, 36 and 98ms after stimulation. (C) Map of onset times of Fluo-4 signal (tonset). (D) Map of maximal upstroke velocities in Fluo-4 signal (du/dtmax). (E) Mean±SD of tonset and (F) du/dtmax versus sarcolemma distance (ΔSL). Statistical analysis were performed on 15 HF myocytes. (G) Mean±SD of tonset and (H) du/dtmax versus ΔSL. (I) tonset and (J) its standard deviation (σtonset) in cells grouped by low (<1μm, black) and high (>1μm, white) ΔSL. Scale bar in A: 10μm. Applies to B–D. *p<0.05, **p<0.01
Figure 7
Figure 7. Proposed role of the t-system in functional cardiac recovery in chronic human HF
In some failing hearts, normal t-tubules remodel to t-sheets, t-system components that are significantly widened in the myocyte long axis direction, but still run transversely. Concomitantly, t-system density decreases, which causes a higher fraction of non-junctional ryanodine receptor (RyR) clusters. As a result, triggering of calcium release from the sarcoplasmic reticulum (SR) becomes less efficient. Mechanical unloading by LVAD allows recovery of metabolism, beta-adrenergic response and calcium transporters, but not of t-system structure. Thus, excitation-contraction (EC) coupling can only recover in failing hearts with a preserved t-system, leading to improved contractility and left-ventricular ejection fraction (EF) following unloading, but not in failing hearts with t-sheets and reduced t-system density.
See this image and copyright information in PMC

Comment in

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Mozaffarian D, Benjamin EJ, Go AS, Arnett DK, Blaha MJ, Cushman M, Das SR, de Ferranti S, Despres JP, Fullerton HJ, Howard VJ, Huffman MD, Isasi CR, Jimenez MC, Judd SE, Kissela BM, Lichtman JH, Lisabeth LD, Liu S, Mackey RH, Magid DJ, McGuire DK, Mohler ER, 3rd, Moy CS, Muntner P, Mussolino ME, Nasir K, Neumar RW, Nichol G, Palaniappan L, Pandey DK, Reeves MJ, Rodriguez CJ, Rosamond W, Sorlie PD, Stein J, Towfighi A, Turan TN, Virani SS, Woo D, Yeh RW, Turner MB American Heart Association Statistics C, Stroke Statistics S. Executive Summary: Heart Disease and Stroke Statistics-2016 Update: A Report From the American Heart Association. Circulation. 2016;133:447–454. - PubMed
    1. Drakos SG, Terrovitis JV, Anastasiou-Nana MI, Nanas JN. Reverse remodeling during long-term mechanical unloading of the left ventricle. J Mol Cell Cardiol. 2007;43:231–42. - PubMed
    1. Birks EJ, George RS, Hedger M, Bahrami T, Wilton P, Bowles CT, Webb C, Bougard R, Amrani M, Yacoub MH, Dreyfus G, Khaghani A. Reversal of severe heart failure with a continuous-flow left ventricular assist device and pharmacological therapy: a prospective study. Circulation. 2011;123:381–390. - PubMed
    1. Drakos SG, Kfoury AG, Stehlik J, Selzman CH, Reid BB, Terrovitis JV, Nanas JN, Li DY. Bridge to recovery: understanding the disconnect between clinical and biological outcomes. Circulation. 2012;126:230–241. - PMC - PubMed
    1. Birks EJ, Tansley PD, Hardy J, George RS, Bowles CT, Burke M, Banner NR, Khaghani A, Yacoub MH. Left ventricular assist device and drug therapy for the reversal of heart failure. N Engl J Med. 2006;355:1873–1884. - PubMed

MeSH terms

Substances