Distinct Cell Cycle Proteins Control Schwann Cell Proliferation in Health and Disease ATANASOSKI S University of Basel, Basel, Switzerland
Background: Proliferating Schwann cells, the glial cells of the peripheral nervous system, are a prominent feature during early development and after damage to peripheral nerves. Altered Schwann cell proliferation is also associated with diseases and pathological states including inherited peripheral neuropathies, peripheral nerve tumors, and peripheral neuropathies secondary to diabetes, cancer chemotherapeutic agents, or toxins. To gain more insight into the molecular processes governing Schwann cell proliferation in health and disease, we examined the Schwann cell cycle and its regulation in vivo.
Methods: We have examined the expression, regulation, and localization of cyclins, cyclin-dependent kinases (cdk), and cell cycle inhibitors in Schwann cells of developing and adult peripheral nerves using immunohistochemistry. In addition, we used appropriate mutant mice to examine the functional requirement for the respective cell cycle proteins in Schwann cell proliferation.
Results: Proliferating Schwann cells during development express cyclin D1 in the cytoplasm. After injury, cyclin D1 becomes localized to the nuclei of proliferating Schwann cells. Cyclin D1-deficient animals revealed that developmentally regulated proliferation is not affected by the absence of cyclin D1, whereas injury-induced proliferation is impaired. We further found that the cell cycle inhibitor p21 appears first in the cytoplasm of Schwann cells at postnatal day 7 when most cells have already ceased dividing. After nerve injury, however, p21 is localized mainly in nuclei of dedifferentiated Schwann cells. Consistently, p21-deficient Schwann cells do not undergo proper growth arrest in later phases of nerve development. In contrast, after nerve injury, nuclear p21 is required for correct cell cycle control at the peak of Schwann cell proliferation. We next investigated the requirements for cdk2, 4, and 6 during Schwann cell proliferation. We show that only cdk2 and 4 are expressed in peripheral nerves. Our data from cdk-deficient mice indicate that postnatal Schwann cell proliferation is abolished in the absence of cdk4 but not in the absence of cdk2 or 6.
Conclusions: We find that distinct components of the cell cycle machinery that regulate Schwann cell proliferation during development differ fundamentally from those activated following nerve injury or in peripheral neuropathies.
Magic Bullets for the Treatment of Oxidative Stress-Induced Neurodegenerative Disorders
ATLAS D
Dept. Biological Chemistry The Hebrew University, Jerusalem 91904, Israel
A limited capacity to self-repair and regeneration of cellular damages is the hallmark of many central and peripheral regions of our body. In the last few years, evidence has accumulated that shows major damage to much type of cells that comes from oxidation. The source offree radicals and oxidizing agents are part of the normal cellular machinery, however, failing to remove excess of free radicals or the oxidizing agents creates a situation called oxidative stress (OS), responsible for a devastating complications which end in cell death. In the last decade, clinical trials with various antioxidants failed mainly due to the fact these compounds donot penetrate the cell.
We developed two new series of compounds consisting of small molecular weight thiol compounds that can cross the plasma membrane into the cell as well as the blood brain barrier (BBB).
i) AD4 (N-acetyl cysteine amide) is one of the first series of compounds that was shown to very efficient in vitro and in a large number of animal models, reversing the effects caused by oxidative stress (Ref. 1-10). Among its prominent properties, it is an inhibitor of MAP Kinases such as ERK1/2, JNK, and p38 in vitro it scavenges ROS (Reactive Oxygen Species), regenerate GSH, chelates copper ions, and shows restoration of oxidative stress markers such as cell viability (MTT assay), protein carbonyls, nitrosylation (3-nitrotyrosine), and lipids.
ii) This novel series of low molecular weight peptides is called ”Redox-Cluster” (RC). They offer a prolonged action base on unique properties of cell entry and trapping inside the cell.
These compounds were also shown to be potent inhibitors of the MAP Kinases ERK1/2, JNK, and p38 in vitro. They are scavengers of ROS (reactive oxygen species, and show restoration of all oxidative stress markers such as Cell viability (MTT assay), protein carbonyls, nitrosylation (3-nitrotyrosine) and lipid peroxidation (levels of HNE). In studies in vivo a significant improvement in oxidative markers was observed.
The two families of glutathione precursors are potentially promising “magic” bullets for the treatment of neurodegenerative disorders, as well as other degenerative and multi-system disorders, such as diabetes and asthma.
Small Lipoprotein A-I subclasses (42,000-70,000) are Promising Biomarkers in Cardiovascula Disease ATMEH RF, KASASBEH AZ, ABUODEH MR
Jordan University of Science and Technology, Irbid, Jordan
Background: The predictive value of the high density lipoprotein (HDL) cholesterol as a biomarker for cardiovascular disease is now questionable. HDL contains several subclasses of varying size, composition, and function. Aims: 1) To detect and quantitate the apolipoprotein A-I-containing lipoproteins (LpA-I) directly from fresh plasma instead of HDL subclasses. 2) To study the plasma distribution of these LpAI subclasses in normolipidemics and hyperlipidemics. 3) To follow up the changes in their distribution after a fatty meal and during pregnancy.
Methods: Fresh plasma from 90 normolipidemics and 20 hyperlipidemics was subjected to gradient polyacrylamide gel electrophoresis followed by electrotransfer onto agarose gel-containing anti-apoA-I. Similarly, plasma from 8 normolipidemic sunjects was withdrawn after fasting and 4 and 6 h after a fatty meal. Two pregnant women were followed 4.5, 8.0, and 11.5 months during pregnancy and after delivery.
Results: We detected 12 different LpAI subclasses of molecular mass ranging from 42,000 to > 354,000 with Stokes radius of 2.96 to > 5.8 nm. The percentage of the smallest subclasses SlpAI2, SlpAI3, and SlpAI4 (50,000, 45,000, and 42,000, respectively) was low in normolipidemics (7.8, 3.4, and 2.7%) and significantly (P < 0.01, 0.05, 0.05, respectively) higher in hypelipidemics (18.9, 8.3, 5.2%, respectively). In normolipidemics, the level of SLpAI2 increased significantly (P < 0.05) than the fasting level 4 h after fat ingestion, and decreased after 6 h (14.1, 20.3, 19.5 mg/dL, respectively). The percentage of SLpAI1 increased during pregnancy and decreased after birth (7.8, 9.7, and 6.6%), and SLpAI2 was detected (4.6%) after 8 months and disappeared after birth. A strong positive correlation was observed between the SLpAI subclasses and plasma triacylglycerols (TAGs) in all normolipidemics (r = 0.41, P < 0.01) and in males of < 25 y (r = 0.70, P < 0.01).
Conclusions: 1) Significant differences in the distribution of SLpAI subclasses in plasma were detected in normo- and hypelipidemics. 2) Their plasma level increases with the increase of plasma TAGs. 3) They may be related to the lipolysis of the TAG-rich lipoproteins. 4) The variation of SLpAI2 level was more evident than the others and it may be a good candidate as a biomarker for cardiovascular disease.
Investigation of SGK-1 and Dexras1 expression in Human Embryonic Kidney (HEK 293) cells GHASSEM ATTRAZADEH YAZDI1, MICHAEL SHIPSTON2, FERENC ANTONI
Department of Neuroscience, University of Edinburgh, EH8 9JZ, Scotland, United Kingdom, 1 Hormozgan University of Medical Sciences, Iran, 2Membrane Biology Group, University of Edinburgh Medical School, Hugh Robson Building, Edinburgh, EH8 9XD. Scotland, United Kingdom
Adrenal corticosteroids are involved in multiple aspects of CNS function — for example, the feedback inhibition of the hypothalamic-pituitary adrenocortical axis. Typically, the action of glucocorticoids is mediated by rapid induction of mRNA and protein synthesis. Potential mediators of glucocorticoid action include dexras1, (activator of G protein signaling 1) first reported as a rapidly induced mRNA in pituitary tumour cells. The rapid induction of dexras1 in response to glucocorticoids in pituitary cells raises the possibility that it may be involved in negative feedback regulation of corticotropin secretion. In the present study we examined the induction of dexras1 in human embryonic kidney (HEK293) cells which have been used as a model for glucocorticoid-mediated regulation of corticotropin secretion.
To verify that HEK 293 cells contain functional glucocorticoid receptors (GR) activated by dexamethasone, MMTV-LTR plasmid, which responds to activated-GR by enhancing luciferase expression via the MMTV promoter, was transfected into the HEK 293 cells. 48 h after transfection, the cells were treated with dexamethasone. Analysis of the time-course of dexamethasone action on MMTV regulated luciferase activity revealed that luciferase induction was maximal at 120 min. Exposure to various concentrations of dexamethasone for 120 min produced an increase of MMTV regulated luciferase activity in HEK 293 cells in a concentration -dependent manner. Maximum effect was obtained with 100 nM dexamethasone (up to 5-fold).
The expression of dexras1 was assessed by Northern blot. The results of Northern blot showed a 5 kb dexras1 and a 2.6 kb serum and glucocorticoid-induced protein kinase (SGK-1) mRNA (a positive control for glucocorticoid induction) species in HEK 293 cells. Dexamethasone had no significant effect to amount of dexras1 mRNA induction for varying times (0-120 min) but increased SGK-1 mRNA with maximum effect at 30 min. The analysis of SGK-1 protein in HEK 293 cells also showed significant increase in response to dexamethasone.
The results suggest that; 1) HEK 293 cells respond to dexamethasone via an endogenously expressed GR; 2) Dexras1 is not a glucocorticoid-induced protein in HEK 293 cells; 3) SGK-1 is induced by dexamethasone in HEK 293 cells.
