Autophagy is altered in skeletal and cardiac muscle of spontaneously hypertensive rats.

AIM: Autophagy is a subcellular degradation mechanism important for muscle maintenance. Hypertension induces well-characterized pathological changes to the heart and is associated with impaired function and increased apoptotic signalling in skeletal muscle. We examined whether essential hypertension affects several autophagy markers in skeletal and cardiac muscle. METHODS: Immunoblotting and qRT-PCR were used to measure autophagy-related proteins/mRNA in multiple skeletal muscles as well as left ventricle (LV) of spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). RESULTS: Skeletal muscles of hypertensive rats had decreased (P < 0.01) cross-sectional area of type I fibres (e.g. soleus WKY: 2952.9 ± 64.4 µm(2) vs. SHR: 2579.9 ± 85.8 µm(2)) and a fibre redistribution towards a 'fast' phenotype. Immunoblot analysis revealed that some SHR skeletal muscles displayed a decreased LC3II/I ratio (P < 0.05), but none showed differences in p62 protein. LC3 and LAMP2 mRNA levels were increased approx. 2-3-fold in all skeletal muscles (P < 0.05), while cathepsin activity, cathepsin L mRNA and Atg7 protein were increased 16-17% (P < 0.01), 2-3-fold (P < 0.05) and 29-49% (P < 0.01), respectively, in fast muscles of hypertensive animals. Finally, protein levels of BAG3, a marker of chaperone-assisted selective autophagy, were 18-25% lower (P < 0.05) in SHR skeletal muscles. In the LV of SHR, LC3I and p62 protein were elevated 34% (P < 0.05) and 47% (P < 0.01), respectively. Furthermore, p62 mRNA was 68% higher (P < 0.05), while LAMP2 mRNA was 45% lower (P < 0.05), in SHR cardiac muscle. There was no difference in Beclin1, Atg7, Bnip3 or BAG3 protein in the LV between strains. CONCLUSION: These results suggest that autophagy is altered in skeletal and cardiac muscle during hypertension.

Evolution of a genetic diagnosis.

Understanding the relationship between genotype and phenotype has become an integral part of the diagnosis and management of patients with inherited arrhythmias and cardiomyopathies. Given the existence of background noise, the majority of genetic testing results should be incorporated into clinical decision making as probabilistic, rather than deterministic, in the diagnosis and management of inherited arrhythmias. This case report captures multiple snapshots of clinical care in the evolution of a diagnosis of a single patient, highlighting the need for repeated phenotypic and genotypic assessment for both the patient and their family.

Effects of atorvastatin on the intracellular stability and secretion of apolipoprotein B in HepG2 cells.

We investigated the effects of atorvastatin, a new 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, on the biogenesis of apolipoprotein B (apoB) in intact and permeabilized HepG2 cells. Intact cells were pretreated either with single or multiple doses of atorvastatin (0.1 to 20 micromol/L) for periods of 6 to 20 hours and pulsed with [35S]methionine. In some cases the cells were permeabilized with digitonin. Experiments were performed to investigate the effects of atorvastatin on (1) the rates of lipid synthesis and secretion, (2) the synthesis and accumulation of apoB, (3) the intracellular stability of apoB, (4) the amount of apoB-containing lipoprotein particles assembled in HepG2 microsomes, and (5) the secretion and accumulation of apoB into the culture medium. ApoB synthesis, degradation, and secretion were measured by pulse-chase experiments with [35S]methionine in both intact and permeabilized HepG2 cells. Lipid synthesis was assessed by pulse-labeling experiments with [3H]acetate or [3H]oleate bound to bovine serum albumin. Comparisons were made under basal conditions and in the presence of oleate (0.36 micromol/L). Atorvastatin acutely inhibited the synthesis of cholesterol and cholesterol ester but did not have a significant effect on triglyceride or phospholipid synthesis. Atorvastatin did not affect the uptake of [35S]methionine by the cells nor did it influence the synthesis of apoB or a control protein, albumin. However, atorvastatin reduced the secretion of apoB into the culture medium, apparently by enhancing the degradation of apoB in the cell under basal and induced conditions with oleate. The stability of apoB associated with the lipoprotein particles was also significantly lowered by atorvastatin. The stimulated degradation of apoB in atorvastatin-treated cells was sensitive to MG132, a proteasome inhibitor. The net effect of atorvastatin was a reduction in the number of apoB-containing lipoprotein particles of different sizes isolated from microsomes and a reduction in apoB secretion into the culture medium. The data suggest that atorvastatin may impair the translocation of apoB into the lumen of the endoplasmic reticulum, thus increasing the amount of apoB degraded intracellularly. It is hypothesized that atorvastatin alters these parameters primarily as a result of inhibiting cholesterol synthesis and limiting the availability of cholesterol and/or cholesterol ester for the normal assembly of apoB-containing lipoprotein particles.

Noninfectious ring-shaped keratitis associated with Pseudomonas aeruginosa.

Ring-shaped keratitis appeared in the left eye of a 37-year-old woman who had worn soft contact lenses for more than five years. The corneal ring began to appear within seven days of a central corneal abrasion. Gram staining of the patient's contact lens cleaning solution showed many gram-negative rods, and microbiologic investigations of the patient's soft contact lens and contact lens case disclosed Pseudomonas aeruginosa. There was no clinical or laboratory evidence of an infectious process. Prompt treatment with polymyxin B-bacitracin ointment and prednisolone acetate 1% eyedrops led to resolution of the opacity and a return to the patient's normal visual acuity. The P. aeruginosa endotoxin may have been transferred through the epithelial break into the superficial corneal stroma, leading to ring formation via endotoxin-initiated, properdin-mediated, antibody-independent complement activation.