Fat-free diet and myocardial excitability, refractoriness and ventricular fibrillation

The lipid composition of the sarcolemma influences its function. The purpose of this study is to investigate the electrophysiological alterations induced in the rat's heart by dietary manipulation of cardiac fatty-acid composition. Strength-duration and strength-interval relationships were used to study excitability and refractoriness respectively. Ventricular fibrillation threshold measured by short bursts of rapid stimulation was used to indicate ventricular vulnerability. Gas liquid chromatography was used to analyse cardiac fatty-acid-composition. We used two-week-old rats fed with fat-free diet and 5% and 10% soya bean oil-supplemented diets for 6-9 weeks. Fat-free diet only was associated with increased eicosatrienoic acid content, a marker of essential fatty-acid deficiency. A decrease in poly-unsaturated to saturated (P:S) fatty-acid ratio was also observed. Five percent oil supplementation was associated with a delay and attenuation of this effect. Fat-free diet was associated with increased excitability, decreased refractoriness and reduced ventricular fibrillation threshold. Ten percent oil supplementation prevented these effects while 5% oil supplementation had only a temporary protective effect. We conclude that manipulation of dietary fat content can affect cardiac fatty-acid composition and electrophysiology.     

Dynamics of thyroid diseases and thyroid‐axis gland masses

Thyroid disorders are common and often require lifelong hormone replacement. Treating thyroid disorders involves a fascinating and troublesome delay, in which it takes many weeks for serum thyroid‐stimulating hormone (TSH) concentration to normalize after thyroid hormones return to normal. This delay challenges attempts to stabilize thyroid hormones in millions of patients. Despite its importance, the physiological mechanism for the delay is unclear. Here, we present data on hormone delays from Israeli medical records spanning 46 million life‐years and develop a mathematical model for dynamic compensation in the thyroid axis, which explains the delays. The delays are due to a feedback mechanism in which peripheral thyroid hormones and TSH control the growth of the thyroid and pituitary glands; enlarged or atrophied glands take many weeks to recover upon treatment due to the slow turnover of the tissues. The model explains why thyroid disorders such as Hashimoto''s thyroiditis and Graves'' disease have both subclinical and clinical states and explains the complex inverse relation between TSH and thyroid hormones. The present model may guide approaches to dynamically adjust the treatment of thyroid disorders.     

Extraction and recombination studies of the interaction of retinol with human plasma retinol-binding protein

Methods have been developed for the removal of retinol from human plasma retinol-binding protein (RBP), so as to form the retinol-free apoprotein, and for the recombination of apo-RBP with retinol to again form the holoprotein. Retinol is removed from RBP by gently shaking a solution of RBP with heptane under controlled conditions. During the shaking, retinol is gradually extracted from the RBP and into the heptane phase. The reassociation of apo-RBP with retinol is achieved by exposing a solution of apo-RBP to Celite coated with a thin film of retinol, followed by isolation of the RBP by gel filtration on Sephadex G-100. This procedure results in the recombination of apo-RBP with an amount of retinol almost identical with that previously removed by extraction. The two-phase extraction procedure was used to explore some of the factors which affect the interaction of retinol with RBP. The retinol-RBP complex was most stable in the lower portion of the pH range 5.6 to 10. The rate of removal of retinol from the RBP-prealbumin complex (the form in which RBP normally circulates in plasma) was markedly less than the rate of its removal from RBP alone. The interaction of retinol with RBP appears to be stabilized by the formation of the RBP-prealbumin complex. The recombination procedure was employed to examine the specificity of the binding of retinol to RBP, by determining whether compounds other than all-trans-retinol would effectively bind to apo-RBP. Apo-RBP did not bind cholesterol, but displayed a slight affinity for phytol. The affinity of RBP for beta-carotene was minimal, whereas both retinyl acetate and retinal were bound about one-third as effectively as all-trans-retinol. In contrast, retinoic acid bound to apo-RBP almost as effectively as did retinol. Each of two isomers of retinol, 13-cis and 11,13-di-cis-retinol, bound to apo-RBP to some extent. The 13-cis isomer appeared to bind somewhat less effectively than did the 11,13-di-cis isomer. The binding of retinol to RBP is highly but not absolutely specific.     

Asymmetric cell division during T cell development controls downstream fate

During mammalian T cell development, the requirement for expansion of many individual T cell clones, rather than merely expansion of the entire T cell population, suggests a possible role for asymmetric cell division (ACD). We show that ACD of developing T cells controls cell fate through differential inheritance of cell fate determinants Numb and α-Adaptin. ACD occurs specifically during the β-selection stage of T cell development, and subsequent divisions are predominantly symmetric. ACD is controlled by interaction with stromal cells and chemokine receptor signaling and uses a conserved network of polarity regulators. The disruption of polarity by deletion of the polarity regulator, Scribble, or the altered inheritance of fate determinants impacts subsequent fate decisions to influence the numbers of DN4 cells arising after the β-selection checkpoint. These findings indicate that ACD enables the thymic microenvironment to orchestrate fate decisions related to differentiation and self-renewal.     

Severe hypoxia induces complete antifolate resistance in carcinoma cells due to cell cycle arrest

Antifolates have a crucial role in the treatment of various cancers by inhibiting key enzymes in purine and thymidylate biosynthesis. However, the frequent emergence of inherent and acquired antifolate resistance in solid tumors calls for the development of novel therapeutic strategies to overcome this chemoresistance. The core of solid tumors is highly hypoxic due to poor blood circulation, and this hypoxia is considered to be a major contributor to drug resistance. However, the cytotoxic activity of antifolates under hypoxia is poorly characterized. Here we show that under severe hypoxia, gene expression of ubiquitously expressed key enzymes and transporters in folate metabolism and nucleoside homeostasis is downregulated. We further demonstrate that carcinoma cells become completely refractory, even at sub-millimolar concentrations, to all hydrophilic and lipophilic antifolates tested. Moreover, tumor cells retained sensitivity to the proteasome inhibitor bortezomib and the topoisomerase II inhibitor doxorubicin, which are independent of cell cycle. We provide evidence that this antifolate resistance, associated with repression of folate metabolism, is a result of the inability of antifolates to induce DNA damage under hypoxia, and is attributable to a hypoxia-induced cell cycle arrest, rather than a general anti-apoptotic mechanism. Our findings suggest that solid tumors harboring a hypoxic core of cell cycle-arrested cells may display antifolate resistance while retaining sensitivity to the chemotherapeutics bortezomib and doxorubicin. This study bears important implications for the molecular basis underlying antifolate resistance under hypoxia and its rational overcoming in solid tumors.     

Automatic scanning of interphase FISH for prenatal diagnosis in uncultured amniocytes

Fluorescence in situ hybridization (FISH) of uncultured amniocytes using chromosome-specific DNA probes offers the opportunity for rapid aneuploidy screening. Between 80 and 95% of all chromosomal disorders expected in the second trimester of pregnancy can be discovered within 24 hr if DNA probes specific for chromosomes 21, 18, 13, X, and Y are used. Rapid results are crucial for clinical decision-making and are helpful in decreasing the anxiety level in most patients. One of the major factors that have been preventing the rapid FISH test from being broadly incorporated into the clinical setting is the limited staff in the cytogenetics laboratories. The present study demonstrates the use of an automated scanning system (Duet, BioView Ltd. Rehovot, Israel) for analyzing FISH in uncultured amniocytes. Fifty-six amniotic fluid samples were evaluated in parallel by karyotyping, manual FISH analysis, and automatic FISH scanning. Automatic scanning provided accurate results compared to both manual FISH scoring and karyotype analysis. The correlation between automatic and manual FISH scanning was found to be very high (r = 0.9, p < 0.0001). The availability of automation for aneuploidy screening in amniotic fluid samples will enable offering this test to a broader patient population while providing fast and reliable results.     

Galectin-3 Stimulates Cell Proliferation

Galectin-3, a β-galactoside-binding protein, has been shown to be involved in multiple biological processes through interaction with its complementary glycoconjugates. Here we provide the first evidence of galectin-3 as a mitogen. Incubation of quiescent cultures of normal human lung fibroblast IMR-90 cells with recombinant galectin-3 (rgalectin-3) stimulated DNA synthesis as well as cell proliferation in a dose-dependent manner. This mitogenic activity was dependent on the lectin property of galectin-3, as it could be significantly inhibited by lactose, a disaccharide competitive for carbohydrate-binding by galectin-3. Chemical cross-linking and affinity-purification experiments identified binding of rgalectin-3 to cell surface glycoproteins, which were not recognized by antibodies directed against lysosome-associated membrane proteins (LAMPs), putative cellular ligands for galectin-3. Moreover, pulse–chase analysis revealed no secretion of galectin-3 by IMR-90 cells. These results indicate that galectin-3 is a mitogen capable of stimulating fibroblast cell proliferation in a paracrine fashion through interaction with cell surface glycoconjugates different from LAMPs and suggest a possible involvement of galectin-3 in tissue remodeling.