Glycation in the cardiomyocyte.

Glycation is a protein post-translational modification that can occur on lysine and arginine residues as a result of a non-enzymatic process known as the Maillard reaction. This modification is irreversible, so the only way it can be removed is by protein degradation and replacement. Small reactive carbonyl species, glyoxal and methylglyoxal, are the primary glycating agents and are elevated in several conditions associated with an increased risk of cardiovascular disease, including diabetes, rheumatoid arthritis, smoking, and aging. Thus, how protein glycation impacts the cardiomyocyte is of particular interest, to both understand how these conditions increase the risk of cardiovascular disease and how glycation might be targeted therapeutically. Glycation can affect the cardiomyocyte through extracellular mechanisms, including RAGE-based signaling, glycation of the extracellular matrix that modifies the mechanical environment, and signaling from the vasculature. Intracellular glycation of the cardiomyocyte can impact calcium handling, protein quality control and cell death pathways, as well as the cytoskeleton, resulting in a blunted contractility. While reducing protein glycation and its impact on the heart has been an active area of drug development, multiple clinical trials have had mixed results and these compounds have not been translated to the clinic-highlighting the challenges of modulating myocyte glycation. Here we will review protein glycation and its effects on the cardiomyocyte, therapeutic attempts to reverse these, and offer insight as to the future of glycation studies and patient treatment.

The structural and functional integrities of porcine myocardium are mostly preserved by cryopreservation.

Structural and functional studies of heart muscle are important to gain insights into the physiological bases of cardiac muscle contraction and the pathological bases of heart disease. While fresh muscle tissue works best for these kinds of studies, this is not always practical to obtain, especially for heart tissue from large animal models and humans. Conversely, tissue banks of frozen human hearts are available and could be a tremendous resource for translational research. It is not well understood, however, how liquid nitrogen freezing and cryostorage may impact the structural integrity of myocardium from large mammals. In this study, we directly compared the structural and functional integrity of never-frozen to previously frozen porcine myocardium to investigate the consequences of freezing and cryostorage. X-ray diffraction measurements from hydrated tissue under near-physiological conditions and electron microscope images from chemically fixed porcine myocardium showed that prior freezing has only minor effects on structural integrity of the muscle. Furthermore, mechanical studies similarly showed no significant differences in contractile capabilities of porcine myocardium with and without freezing and cryostorage. These results demonstrate that liquid nitrogen preservation is a practical approach for structural and functional studies of myocardium.

Pharmacological inhibition of BAG3-HSP70 with the proposed cancer therapeutic JG-98 is toxic for cardiomyocytes.

The co-chaperone Bcl2-associated athanogene-3 (BAG3) maintains cellular protein quality control through the regulation of heat shock protein 70 (HSP70). Cancer cells manipulate BAG3-HSP70-regulated pathways for tumor initiation and proliferation, which has led to the development of promising small molecule therapies, such as JG-98, which inhibit the BAG3-HSP70 interaction and mitigate tumor growth. However, it is not known how these broad therapies impact cardiomyocytes, where the BAG3-HSP70 complex is a key regulator of protein turnover and contractility. Here, we show that JG-98 exposure is toxic in neonatal rat ventricular myocytes (NRVMs). Using immunofluorescence microscopy to assess cell death, we found that apoptosis increased in NRVMs treated with JG-98 doses as low as 10 nM. JG-98 treatment also reduced autophagy flux and altered expression of BAG3 and several binding partners involved in BAG3-dependent autophagy, including SYNPO2 and HSPB8. We next assessed protein half-life with disruption of the BAG3-HSP70 complex by treating with JG-98 in the presence of cycloheximide and found BAG3, HSPB5, and HSPB8 half-lives were reduced, indicating that complex formation with HSP70 is important for their stability. Next, we assessed sarcomere structure using super-resolution microscopy and found that disrupting the interaction with HSP70 leads to sarcomere structural disintegration. To determine whether the effects of JG-98 could be mitigated by pharmacological autophagy induction, we cotreated NRVMs with rapamycin, which partially reduced the extent of apoptosis and sarcomere disarray. Finally, we investigated whether the effects of JG-98 extended to skeletal myocytes using C2C12 myotubes and found again increased apoptosis and reduced autophagic flux. Together, our data suggest that nonspecific targeting of the BAG3-HSP70 complex to treat cancer may be detrimental for cardiac and skeletal myocytes.