Cholesterol
Blood. 2006 May
9; [Epub ahead of print] Blood. 2006 May
2; [Epub ahead of print] J
Clin Endocrinol Metab. 2006 May 9; [Epub
ahead of print] : J Mol Neurosci. 2005;25(1):105-17. J Neurosci. 2005 Nov 9;25(45):10502-9. Exp Neurol. 2005 Dec;196(2):452-63. Epub 2005 Oct 20. Parkinsonism
Relat Disord.
2006 Jan;12(1):15-9. Epub 2005 Sep 29.
Constitutive activation of STAT5 and Bcl-xL overexpression can
induce endogenous erythroid colony formation in
human primary cells.
Garcon
L, Rivat C, James
C, Lacout C, Camara-Clayette V, Ugo V, Lecluse Y, Bennaceur-Griscelli A, Vainchenker W.
U 790 INSERM, Institut Gustave
Roussy, 39 rue Camille Desmoulins,
94805
The biological hallmark of PV is the formation of endogenous erythroid colonies (EEC) with an
erythropoietin-independent erythroid
differentiation. Recently it has been shown that an activating mutation of
JAK2 (V617F) was at the origin of PV. In this work we studied whether the
STAT5/Bcl-xL pathway could be responsible for EEC formation. A
constitutively active form of STAT5 was transduced
into human erythroid progenitors and induced an
erythropoietin-independent terminal differentiation and EEC formation.
Furthermore, Bcl-xL overexpression
in erythroid progenitors was also able to induce erythroid colonies despite the absence of
erythropoietin. Conversely, siRNA mediated STAT5
and Bcl- xL knock-down
in human erythroid progenitors inhibited CFU-E
formation in the presence of Epo. Altogether
these results demonstrate that a sustained level of the sole Bcl-xL is capable to give rise to Epo
independent erythroid colony formation and
suggest that in PV patients, JAK2(V617F) may induce EEC via the
STAT5/Bcl-xL pathway.
PMID: 16684963 [PubMed - as supplied by
publisher]
JAK2V617F expression in murine hematopoietic cells
leads to MPD mimicking human PV with secondary myelofibrosis.
Lacout C, Pisani DF, Tulliez M, Moreau
Gachelin F, Vainchenker W, Villeval JL.
Inserm, U790, Universite
Paris XI, Institut Gustave
Roussy (IGR),
A JAK2(V617F) mutation is frequently found in several BCR/ABL-negative myeloproliferative disorders. To address the
contribution of this mutant to the pathogenesis of these different myeloproliferative disorders, we used an adoptive
transfer of marrow cells transduced with a
retrovirus expressing JAK2(V617F) in recipient irradiated mice. Hosts were
analyzed during 6 months after transplantation. For a period of 3 months,
mice developed polycythemia, macrocytosis
and usually peripheral blood granulocytosis.
Transient thrombocytosis was only observed in a
low expresser group. All mice displayed trilineage
hyperplasia in marrow and spleen along with an amplification of myeloid and
erythroid progenitor cells and a formation of
endogenous erythroid colonies. After 3-4 months, polycythemia regressed, abnormally shaped red blood
cells and platelets were seen in circulation and a deposition of reticulin fibers was observed in marrow and spleen.
Development of fibrosis was associated with anemia, thrombocytopenia, high neutrophilia and massive splenomegaly.
These features mimic human Polycythemia Vera and
its evolution towards myelofibrosis. This work
demonstrates that JAK2(V617F) is sufficient for polycythemia
and fibrosis development and offers an in vivo model to assess novel
therapeutic approaches for JAK2(V617F)-positive pathologies. Questions
remain regarding the exact contribution of JAK2(V617F) in other myeloproliferative disorders.
PMID: 16670266 [PubMed - as supplied by publisher]
Leptin Interferes with ACTH/cAMP
Signaling Possibly through a Janus kinase 2-Phosphatidylinositol
3-kinase/Akt-Phosphodiesterase 3-Cyclic AMP Pathway to Down-Regulate
Cholesterol Side-Chain Cleavage Cytochrome P450
Enzyme in Human Adrenocortical NCI-H295 Cell
Line.
Hsu
HT, Chang
YC, Chiu
YN, Liu
CL, Chang
KJ, Guo IC.
Department of Veterinary Medicine, College of Bio-Resources and
Agriculture, National Taiwan University, Taipei, Taiwan; Department of
Surgery, Mackay Memorial Hospital, Taipei, Taiwan; Mackay Medicine, Nursing
and Management College, Taipei, Taiwan; Department of Surgery, National
Taiwan University Hospital, National Taiwan University, Taipei, Taiwan.
Context: Obesity has adverse effects on adrenocortical
functions. Adipocyte-derived leptin,
a biomarker molecule of obesity, may directly control adrenal steroidogenesis via an unclear mechanism. Objective: We
studied the mechanism underlying leptin action on
adrenal steroidogenesis in human adrenocortical NCI-H295 tumor cell line. Methods:
Levels of progesterone, cortisol and cAMP were determined by ELISA. Western blotting was
used to detect protein amounts of P450scc, JAK2, Akt
and their phosphorylated forms. The mRNA
expressions of P450scc and leptin receptors were
measured by RT-PCR and real-time PCR. P450scc promoter activity was
analyzed with a luciferase reporter system.
Results: Cholera toxin mimicked ACTH action by increasing adrenal cAMP levels and steroid secretion. Leptin
did not affect basal release but significantly inhibited ACTH/cholera
toxin-induced steroid secretion. The concomitant inhibitions by leptin on cholera toxin-induced protein and
ACTH/cholera toxin-induced mRNA expression of P450scc were confirmed. Leptin inhibited ACTH/cholera toxin-induced CYP11A1
promoter activity via a known cAMP-responsive
region located between -1.7 kb and -1.5 kb. Leptin
activated phosphorylations of JAK2 and Akt. Inhibitory effects of leptin
on ACTH/cholera toxin-induced cAMP levels,
CYP11A1 promoter activity and steroid secretion were blunted by either
inhibitor of JAK2 (AG490) or PI3K/Akt (Wortmannin),
as well as inhibitors of cAMP-degrading phosphodiesterases (PDEs),
including non-specific IBMX and PDE3-specific SKF94836. Leptin
failed to affect the inductions of CYP11A1 promoter activity and steroid
secretion by PDE-non-hydrolyzable N(6)-MB-cAMP. Conclusions: Leptin
interferes with ACTH/cAMP signaling possibly
through a cAMP-degrading mechanism involving
activation of JAK2, PI3K and PDE3 to down-regulate P450scc expression and
consequent adrenal steroidogenesis.
PMID: 16684834 [PubMed - as supplied by
publisher]
Dystonia-associated forms of torsinA
are deficient in ATPase activity.
Konakova
M, Pulst
SM.
Rose Moss Laboratory for Parkinson's Disease and Neurodegenerative
Disorders, Burns and Allen Research Institute,
Early-onset dystonia is caused by mutations in
the torsinA protein, a putative member of the
AAA+ class of ATPases. In this study we have
evaluated the ATPase activity of bacterially
expressed wild-type torsinA and its
disease-associated mutant forms. Upon overexpression
in Escherichia coli, recombinant torsinA proteins
were accumulated as insoluble inclusion bodies and required refolding to
become soluble and catalytically active. The refolded wild-type and mutant torsinA proteins were capable of hydrolyzing ATP, but
their specific ATPase activities differed
significantly. Deletions of the amino acid residues E302/303 and F323-Y328
resulted in a decrease of ATPase activity to
approximately 35% and approximately 75% of the wild-type level,
respectively. ATPase activity of wild-type and
mutant torsinA proteins was influenced by factors
that varied with cell stress, such as temperature, pH, and ionic strength,
and was inhibited by sodium vanadate. Our results
provide the first direct evidence for a role of torsinA
as an active ATPase and suggest that the
mutations in torsinA might affect normal
functions of the protein by reducing its enzymatic activity.
PMID: 15781971 [PubMed - indexed for MEDLINE]
Silencing primary dystonia:
lentiviral-mediated RNA interference therapy for
DYT1 dystonia.
Gonzalez-Alegre
P, Bode
N, Davidson
BL, Paulson
HL.
Department of Neurology, Carver College of Medicine, University of Iowa,
Iowa City, Iowa 52242, USA. pedro-gonzalez-alegre@uiowa.edu
DYT1 is the most common inherited dystonia.
Currently, there are no preventive or curative therapies for this
dominantly inherited disease. DYT1 dystonia is
caused by a common three-nucleotide deletion in the TOR1A gene that
eliminates a glutamic acid residue from the
protein torsinA. Recent studies suggest that torsinA carrying the disease-linked mutation, torsinA(DeltaE) acts through a
dominant-negative effect by recruiting wild-type torsinA
[torsinA(wt)] into oligomeric
structures in the nuclear envelope. Therefore, suppressing torsinA(DeltaE) expression
through RNA interference (RNAi) could restore the
normal function of torsinA(wt), representing a
potentially effective therapy regardless of the biological role of torsinA. Here, we have generated short hairpin RNAs (shRNAs) that mediate
allele-specific suppression of torsinA(DeltaE) and rescue cells from its dominant-negative
effect, restoring the normal distribution of torsinA(wt).
In addition, delivery of this shRNA by a
recombinant feline immunodeficiency virus effectively silenced torsinA(DeltaE) in a neural
model of the disease. We further establish the feasibility of this viral-mediated
RNAi approach by demonstrating significant
suppression of endogenous torsinA in mammalian
neurons. Finally, this silencing of torsinA is
achieved without triggering an interferon response. These results support
the potential use of viral-mediated RNAi as a
therapy for DYT1 dystonia and establish the basis
for preclinical testing in animal models of the disease.
PMID: 16280588 [PubMed - indexed for MEDLINE]
Generation and characterization of Dyt1 DeltaGAG knock-in mouse as a model for early-onset dystonia.
Dang
MT, Yokoi
F, McNaught KS, Jengelley TA, Jackson
T, Li
J, Li
Y.
Department of Molecular and Integrative Physiology, NeuroTech
Group, Beckman Institute for Advanced Science and Technology,
A trinucleotide deletion of GAG in the DYT1 gene
that encodes torsinA protein is implicated in the
neurological movement disorder of Oppenheim's
early-onset dystonia. The mutation removes a glutamic acid in the carboxy
region of torsinA, a member of the Clp protease/heat shock protein family. The function of
torsinA and the role of the mutation in causing dystonia are largely unknown. To gain insight into
these unknowns, we made a gene-targeted mouse model of Dyt1 DeltaGAG to mimic the mutation found in DYT1 dystonic patients. The mutated heterozygous mice had
deficient performance on the beam-walking test, a measure of fine motor
coordination and balance. In addition, they exhibited hyperactivity in the
open-field test. Mutant mice also showed a gait abnormality of increased
overlap. Mice at 3 months of age did not display deficits in beam-walking
and gait, while 6-month mutant mice did, indicating an age factor in
phenotypic expression as well. While striatal
dopamine and 4-dihydroxyphenylacetic acid (DOPAC) levels in Dyt1 DeltaGAG mice were similar to that of wild-type mice, a
27% decrease in 4-hydroxy, 3-methoxyphenacetic acid (homovanillic
acid) was detected in mutant mice. Dyt1 DeltaGAG
tissues also have ubiquitin- and torsinA-containing aggregates in neurons of the pontine nuclei. A sex difference was noticed in the
mutant mice with female mutant mice exhibiting fewer alterations in behavioral,
neurochemical, and cellular changes. Our results
show that knocking in a Dyt1 DeltaGAG allele in
mouse alters their motor behavior and recapitulates the production of
protein aggregates that are seen in dystonic
patients. Our data further support alterations in the dopaminergic
system as a part of dystonia's neuropathology.
PMID: 16242683 [PubMed - indexed for MEDLINE]
DYT1 mutation in a cohort of Taiwanese
primary dystonias.
Lin
YW, Chang
HC, Chou
YH, Chen
RS, Hsu
WC, Wu
WS, Weng YH, Lu
CS.
Movement Disorders Unit, Department of Neurology,
To investigate the DYT1 gene mutation in Chinese ethnic, we examined a
series of 200 patients with primary dystonias (11
familial and 189 sporadic), 53 of their asymptomatic relatives, 97 patients
with familial or early-onset parkinsonism, and 200 healthy subjects. The
GAG deletion at codon 946 was only found in three
sporadic dystonia patients and seven of their
asymptomatic familial members. The frequency of GAG deletion was 1.5% in dystonia patients, and was 6.7% in early-onset dystonias (< or = 26 years). We conclude that DYT1
mutation is a minor cause of primary dystonias in
a cohort of Taiwanese population.
PMID: 16198613 [PubMed - indexed for MEDLINE