Citations

Rewarming patients after profound hypothermia may result in acute heart failure and high mortality (50–80%). However, the underlying pathophysiological mechanisms are largely unknown. We characterized cardiac contractile function in the temperature range of 15–30°C by measuring the intracellular Ca2+concentration ([Ca2+]i) and twitch force in intact left ventricular rat papillary muscles. Muscle preparations were loaded with fura-2 AM and electrically stimulated during cooling at 15°C for 1.5 h before being rewarmed to the baseline temperature of 30°C. After hypothermia/rewarming, peak twitch force decreased by 30–40%, but [Ca2+]iwas not significantly altered. In addition, we assessed the maximal Ca2+-activated force (Fmax) and Ca2+sensitivity of force in skinned papillary muscle fibers. Fmaxwas decreased by ∼30%, whereas the pCa required for 50% of Fmaxwas reduced by ∼0.14. In rewarmed papillary muscle, both total cardiac troponin I (cTnI) phosphorylation and PKA-mediated cTnI phosphorylation at Ser23/24 were significantly increased compared with controls. We conclude that after hypothermia/rewarming, myocardial contractility is significantly reduced, as evidenced by reduced twitch force and Fmax. The reduced myocardial contractility is attributed to decreased Ca2+sensitivity of force rather than [Ca2+]iitself, resulting from increased cTnI phosphorylation.

Abstract Context: A reduction in maximal mitochondrial ATP production rate (MAPR) and mitochondrial DNA (mtDNA) abundance occurs with age in association with muscle weakness and reduced endurance in elderly people. Branched chain amino acids (BCAA) have been extensively used to improve physical performance. Objective: The objective was to determine whether an 8-h infusion of BCAA enhances MAPR equally in healthy young and elderly adults. Methods: Using a crossover study design, we compared the effect BCAA vs. saline infusion in 12 young (23.0 ± 0.8 yr) and 12 elderly (70.7 ± 1.1 yr) participants matched for sex and body mass index. Skeletal muscle MAPR and mtDNA abundance were measured in muscle biopsy samples obtained before and at the end of the 8-h infusion. Results: In young participants, MAPR with the substrates glutamate plus malate (supplying electrons to complex I) and succinate plus rotenone (complex II) increased in response to BCAA infusion, relative to a decline in MAPR in response to the saline infusion. In contrast, MAPR was unaffected by BCAA infusion in the elderly participants. Moreover, mtDNA abundance was lower in the elderly compared with the young participants but was unaffected by the BCAA infusion. Insulin and C-peptide concentrations declined over time during the saline infusion, but these declines were prevented by the BCAA infusion. Conclusions: BCAA increased skeletal muscle MAPR in the young participants in comparison with saline, but this effect was not seen in the elderly participants indicating, that unlike in the young, BCAA does not increase muscle mitochondrial function in the elderly.

AbstractBackground: Parathyroid hormone (PTH) assays able to distinguish between full-length PTH (PTH1–84) and N-terminally truncated PTH (PTH7–84) are of increasing significance in the accurate diagnosis of endocrine and osteological diseases. We describe the discovery of new N-terminal and C-terminal PTH variants and the development of selected reaction monitoring (SRM)-based immunoassays specifically designed for the detection of full-length PTH [amino acid (aa)1–84] and 2 N-terminal variants, aa7–84 and aa34–84.Methods: Preparation of mass spectrometric immunoassay pipettor tips and MALDI-TOF mass spectrometric analysis were carried out as previously described. We used novel software to develop SRM assays on a triple-quadrupole mass spectrometer. Heavy isotope-labeled versions of target peptides were used as internal standards.Results: Top-down analysis of samples from healthy individuals and renal failure patients revealed numerous PTH variants, including previously unidentified aa28–84, aa48–84, aa34–77, aa37–77, and aa38–77. Quantitative SRM assays were developed for PTH1–84, PTH7–84, and variant aa34–84. Peptides exhibited linear responses (R2 = 0.90–0.99) relative to recombinant human PTH concentration limits of detection for intact PTH of 8 ng/L and limits of quantification of 16–31 ng/L depending on the peptide. Standard error of analysis for all triplicate measurements was 3%–12% for all peptides, with <5% chromatographic drift between replicates. The CVs of integrated areas under the curve for 54 separate measurements of heavy peptides were 5%–9%.Conclusions: Mass spectrometric immunoassays identified new clinical variants of PTH and provided a quantitative assay for these and previously identified forms of PTH.

Abstract Heart failure with preserved ejection fraction (HFpEF) is a common but under-recognized cause of pulmonary hypertension (PH), particularly among the elderly. While elevated pulmonary artery pressures are clearly associated with increased mortality in HFpEF, it remains unknown whether or how pulmonary vascular disease in HFpEF should be treated. This case emphasizes the importance of maintaining a high index of suspicion for HFpEF in elderly patients presenting with PH and illustrates potential hazards associated with selective pulmonary vasodilation in this growing population.

Background and Purpose— The role of brain-derived neurotrophic factor (BDNF) and its receptor, tropomyosin receptor kinase B, in control of cerebral circulation is poorly understood. The present study was designed to investigate the cerebral vascular effects of BDNF in vivo. Methods— Replication incompetent adenovirus encoding either rat BDNF (AdBDNF) or green fluorescent protein was injected intracisternally into rabbits. Forty-eight hours later, animals were euthanized. Plasma and cerebrospinal fluid levels of BDNF were measured by enzyme-linked immunosorbent assay, vasomotor function of isolated basilar arteries was studied in organ chambers, protein expression in the basilar arteries was studied by Western blotting, prostanoid levels were measured by enzyme-linked immunosorbent assay, and cyclic adenosine 3′,5′-monophosphate levels were measured by radioimmunoassay. Results— The levels of BDNF in the cerebrospinal fluid were significantly elevated in AdBDNF-treated rabbits as compared with adenovirus encoding green fluorescent protein-treated rabbits (37±5 ng/mL versus 0.006±0.003 ng/mL, respectively; P <0.05; n=14). Western blotting studies revealed that in basilar arteries, AdBDNF increased protein expression of prostacyclin synthase, whereas expression of endothelial nitric oxide synthase and phosphorylated (Ser 1177) endothelial nitric oxide synthase remained unchanged. During incubation with arachidonic acid (1 μmol/L), PGI 2 production and levels of cyclic adenosine 3′,5′-monophosphate were significantly elevated only in AdBDNF-treated rabbit basilar arteries ( P <0.05, n=6). Relaxations to acetylcholine (10 −9 to 10 −5 mol/L) and arachidonic acid (10 −9 to 10 −5 mol/L) were significantly potentiated in basilar arteries from rabbits injected with AdBDNF. Potentiation of relaxations to acetylcholine in AdBDNF-treated basilar arteries was inhibited by the nonselective cyclooxygenase inhibitor, indomethacin (10 −5 mol/L, P <0.05, n=6) and constitutive phospholipase A 2 inhibitor, AACOCF3 (2×10 −5 mol/L, P <0.05, n=5). Conclusion— Our results demonstrate that in cerebral arteries, BDNF-induced activation of tropomyosin receptor kinase B receptor signaling in vivo promotes prostacyclin biosynthesis. These findings provide novel mechanistic insight into the vascular protective effect of BDNF in cerebral circulation.

Abstract Microvesicles (MVs) released by malignant cancer cells constitute an important part of the tumor microenvironment. They can transfer various messages to target cells and may be critical to disease progression. Here, we demonstrate that MVs circulating in plasma of B-cell chronic lymphocytic leukemia (CLL) patients exhibit a phenotypic shift from predominantly platelet derived in early stage to leukemic B-cell derived at advanced stage. Furthermore, the total MV level in CLL was significantly greater compared with healthy subjects. To understand the functional implication, we examined whether MVs can interact and modulate CLL bone marrow stromal cells (BMSCs) known to provide a “homing and nurturing” environment for CLL B cells. We found that CLL-MV can activate the AKT/mammalian target of rapamycin/p70S6K/hypoxia-inducible factor-1α axis in CLL-BMSCs with production of vascular endothelial growth factor, a survival factor for CLL B cells. Moreover, MV-mediated AKT activation led to modulation of the β-catenin pathway and increased expression of cyclin D1 and c-myc in BMSCs. We found MV delivered phospho-receptor tyrosine kinase Axl directly to the BMSCs in association with AKT activation. This study demonstrates the existence of separate MV phenotypes during leukemic disease progression and underscores the important role of MVs in activation of the tumor microenvironment.