Early Stimulation of Phosphatidylcholine Biosynthesis During Wallerian Degeneration of Rat Sciatic Nerve

Phospholipid metabolism was studied in rat sciatic nerve during Wallerian degeneration induced by crush injury. Portions of crushed sciatic nerve, incubated with labeled substrates, showed significantly higher phosphatidylcholine synthesis than normal nerve, prior to any measurable alterations of phospholipid composition. Maximum synthesis occurred 3 days after crush injury, at which time the metabolism of other phospholipids was unchanged. After a rapid decrease in biosynthetic activity, a second phase of enhanced phosphatidylcholine synthesis occurred, beginning 6 days after crush injury. Increased incorporation of [33P]phosphate, [2-3H]glycerol, and [Me-14C]choline indicated stimulation of de novo synthesis of phosphatidylcholine 3 days after injury. Neither base exchange reactions nor sequential methylation of ethanolamine phospholipids contributed significantly to phosphatidylcholine synthesis. Assay of certain key enzymes under optimal conditions in subcellular fractions of sciatic nerve revealed higher activities of cholinephosphate cytidyltransferase, choline phosphotransferase, and acyl-CoA:lysophosphatidylcholine acyltransferase in injured nerve, while choline kinase activity remained unchanged. This indicates that stimulation of phosphatidylcholine synthesis occurs via the cytidine nucleotide pathway, as well as by increased acylation of lysophosphatidylcholine. Although the cause of stimulated phosphatidylcholine synthesis remains unexplained, it is possible that trace amounts of lysophospholipids or other metabolites produced by injury-enhanced phospholipase activity may be responsible.     

Alterations of Inositol Lipid Metabolism of Rat Sciatic Nerve in Streptozotocin-Induced Diabetes

Abstract: The composition and metabolism of rat sciatic nerve phospholipids were studied 20 weeks after induction of chronic diabetes by intraperitoneal injection of streptozotocin (50 mg/kg). On a wet weight basis the nerves from the diabetic animals showed a 7% decrease in total phospholipid from that of controls and a relative decrease in phosphatidylinositol. Incubations of isolated sciatic nerves of diabetic rats in a medium containing [³³P]orthophosphate gave decreased labeling of phosphatidylinositol and substantial changes in the labeling pattern of phosphatidylinositol phosphate and 4,5-bisphosphate from that of controls. The ratio of label in these polyphosphoinositides decreased from 2.5 for normal nerve to about 1.0 for diabetic nerve within a 2-h incubation period. These metabolic alterations were not observed in acutely diabetic animals 5 days after streptozotocin (100 mg/kg) administration. Because polyphosphoinositides may be involved in the control of membrane permeability during axonal conduction, alterations in their relative amounts or turnover rates could be related to the physiological changes of early diabetic neuropathy.     

Serum lipoproteins and albumin in the lecithin: Cholesterol acyltransferase reaction

The unique features of pig ovarian follicular fluids, i.e., presence of high density lipoprotein (HDL) only and lecithin: cholesterol acyltransferase (EC 2.3.1.43; LCAT) activity, provides a good model to study the effect of serum lipoproteins and serum albumin on the LCAT reaction. $\text{In}\text{vitro}$ cholesterol esterification is enhanced when very low density lipoprotein (VLDL) and low density lipoprotein (LDL) fractions are added, but is inhibited when one or the other of these lipoproteins is absent. High concentrations of HDL₂ result in decreased activation which can be compensated for by the addition of the VLDL-LDL mixture. These findings suggest that the rate of cholesterol esterification in ovarian follicular fluid may be enhanced by providing the exogenous VLDL and LDL as the recipients of HDL-cholesteryl ester. The inhibition of LCAT activity caused by free fatty acid and lysophosphatidylcholine can be partially reversed by the addition of serum albumin, suggesting that serum albumin may regulate the LCAT reaction.     

Identification of a small Na/H exchanger-like message in the rabbit myocardium

We examined the Na⁺/H⁺ exchanger message in isolated perfused rabbit hearts using Northern blot analysis with cDNA encoding for the rabbit cardiac Na⁺/H⁺ exchanger. A cDNA probe from the coding region of the rabbit myocardial Na⁺/H⁺ exchanger hybridized to mRNA of 5 kb under high stringency, and to a second 3.8 kb mRNA species under low stringency. When Northern blots were re-probed with a section of the 3′-untranslated region of the cDNA, the 5 kb message was apparent while the smaller 3.8 kb message was not. If isolated working rabbit hearts were subjected to ischemia we observed increases in the 3.8 kb message. Overall, the results show that a 3.8 kb mRNA product, which is homologous to the amiloride sensitive Na⁺/H⁺ exchanger, exists in the myocardium and increases during ischemia in the myocardium.     

Canadian polar bear population structure using genome‐wide markers

Predicting the consequences of environmental changes, including human‐mediated climate change on species, requires that we quantify range‐wide patterns of genetic diversity and identify the ecological, environmental, and historical factors that have contributed to it. Here, we generate baseline data on polar bear population structure across most Canadian subpopulations (n = 358) using 13,488 genome‐wide single nucleotide polymorphisms (SNPs) identified with double‐digest restriction site‐associated DNA sequencing (ddRAD). Our ddRAD dataset showed three genetic clusters in the sampled Canadian range, congruent with previous studies based on microsatellites across the same regions; however, due to a lack of sampling in Norwegian Bay, we were unable to confirm the existence of a unique cluster in that subpopulation. These data on the genetic structure of polar bears using SNPs provide a detailed baseline against which future shifts in population structure can be assessed, and opportunities to develop new noninvasive tools for monitoring polar bears across their range.     

Multiphasic triacylglycerol dynamics in the intact heart during acute in vivo overexpression of CD36

Cardiac triacylglycerol (TAG) stores buffer the intracellular availability of long chain fatty acid (LCFA) that act as nuclear receptor ligands, substrate for lipotoxic derivatives, and high energy-yield fuel. The kinetic characteristics of TAG turnover and homeostatic mechanisms linking uptake and storage dynamics in hearts have until now remained elusive. This work examines TAG pool dynamics in the intact beating heart, under normal conditions and in response to acute gene expression-induced changes in CD36. Dynamic mode ¹³C NMR elucidated multiple kinetic processes in ¹³C-palmitate incorporation into TAG: an initial, saturable exponential component and a slower linear rate. Although previous work indicates the linear component to reflect TAG turnover, we hypothesized the saturable exponential to reflect transport of LCFA across the sarcolemma. Thus, we overexpressed the LCFA transporter CD36 through cardiac-specific adenoviral infection in vivo. Within 72 h, CD36 expression was increased 40% in intact hearts, accelerating the exponential phase relative to PBS-infused hearts. TAG turnover also increased with elevations in adipose triglyceride lipase (ATGL) and a modest increase in diacylglycerol acyltransferase 1 (DGAT1), without a significant expansion of the intracellular lipid pools. The results demonstrate a dynamic system of reciprocal gene regulation that couples saturable LCFA uptake across the sarcolemma to TAG synthesis/lipolysis rates.     

Calreticulin Induces Dilated Cardiomyopathy

Background

Calreticulin, a Ca²⁺-buffering chaperone of the endoplasmic reticulum, is highly expressed in the embryonic heart and is essential for cardiac development. After birth, the calreticulin gene is sharply down regulated in the heart, and thus, adult hearts have negligible levels of calreticulin. In this study we tested the role of calreticulin in the adult heart.

Methodology/Principal Findings

We generated an inducible transgenic mouse in which calreticulin is targeted to the cardiac tissue using a Cre/loxP system and can be up-regulated in adult hearts. Echocardiography analysis of hearts from transgenic mice expressing calreticulin revealed impaired left ventricular systolic and diastolic function and impaired mitral valve function. There was altered expression of Ca²⁺ signaling molecules and the gap junction proteins, Connexin 43 and 45. Sarcoplasmic reticulum associated Ca²⁺-handling proteins (including the cardiac ryanodine receptor, sarco/endoplasmic reticulum Ca²⁺-ATPase, and cardiac calsequestrin) were down-regulated in the transgenic hearts with increased expression of calreticulin.

Conclusions/Significance

We show that in adult heart, up-regulated expression of calreticulin induces cardiomyopathy in vivo leading to heart failure. This is due to an alternation in changes in a subset of Ca²⁺ handling genes, gap junction components and left ventricle remodeling.