| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
Endocrine Sciences Research Group (J.S.F., A.J.W., P.E.C.), Department of Medicine, University of Manchester, Stopford Building, Oxford Road, Manchester, M13 9PT, United Kingdom; and Division of Endocrinology (C.M.S.), Department of Internal Medicine, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908
Address all correspondence and requests for reprints to: Dr. P. E. Clayton, Endocrine Sciences Research Group, Department of Medicine, University of Manchester, Stopford Building, Oxford Road, Manchester, M13 9PT, United Kingdom. E-mail: peter.clayton{at}man.ac.uk
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
On immunoblots, both normal and LS fibroblasts express JAK2 and STATs 1, 3, and 5. GH induced rapid tyrosine phosphorylation of a protein at approximately 93 kDa in normal fibroblasts, and Western blotting with STAT-specific antibodies revealed STAT5 activation (phosphorylation) by GH. To determine further the identity and the DNA binding characteristics of the STAT proteins that were activated by GH, EMSAs were performed using three DNA elements known to bind STAT proteins; m67, the high affinity c-sis-inducible element (SIE), the interferon response element (IRE), and the lactogenic hormone-responsive region (LHRR). GH failed to induce protein binding to the SIE or IRE in normal skin fibroblasts but did induce the formation of a specific complex with the LHRR. Induction by GH of this LHRR/protein complex, which could be supershifted partially by anti-STAT1 antisera and completely by anti-STAT5 antisera, was transient, maximal between 10 and 30 min and reduced by 60 min. GH also induced distinct LHRR/protein complexes in mouse 3T3-F442A fibroblasts and in human IM-9 lymphocytes, but supershift analysis revealed that these complexes contained STAT5 but not STAT1. Whereas no binding to the LHRR was observed in GH-treated H fibroblasts, GH induced binding to this element in M fibroblasts.
These results demonstrate that 1) the JAK-STAT pathway is activated by GH in normal fibroblasts and that STATs 1 and 5 have a role in GH-dependent signaling in these cells; 2) GH activation of DNA/STAT binding is cell type- and species-specific; and 3) GH failed to activate the STAT pathway in H fibroblasts but induced STAT signaling in M fibroblasts, indicating that the site of GH resistance in the latter is likely to be located within another GH signaling pathway. These fibroblast cultures therefore provide unique models with which to further our understanding of the mechanisms of human GH signaling.
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The rare condition of congenital GH insensitivity (Laron Syndrome, LS) is usually caused by molecular defects within the extracellular domain of the GHR (21). Because the serum GH binding protein (GHBP) shares sequence homology with this portion of the receptor (22), and in humans is probably generated by proteolysis of the GHR (23), LS is often associated with absent or low levels of serum GH binding activity. However, 25% of European LS patients have normal or elevated levels of functional serum GHBP (24). We have previously described four girls with LS from two unrelated families with normal levels of serum GHBP, where a failure to identify GHR gene mutations that account for their GH insensitivity has implicated abnormalities within the intracellular GH signaling pathway (25).
We and others have shown that normal skin fibroblasts express GHRs (26) and specifically bind GH (25, 27) and have demonstrated activation of DNA synthesis and IGFBP-3 gene expression by GH (25, 28, 29). Skin fibroblast cultures established from three of the GHBP-positive LS girls are also capable of normal GH binding but are insensitive to GH at the levels of DNA synthesis and IGFBP-3 gene expression (25). In the present study, we have assessed the ability of GH to activate the STAT signaling pathway in both normal and GHBP-positive LS fibroblasts.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
p91) for use in Western blotting have been described previously
(5). The anti-STAT3 monoclonal antibody was purchased from Transduction
Laboratories (Lexington, KY; S21320), and the anti-JAK2 polyclonal
antibody was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz,
CA) (SC-294). The anti-STAT5B polyclonal antibody used in Western
blotting, which was raised against the C-terminus of STAT5B, was
produced by Dr. C. M. Silva (unpublished data). The following
antibodies were used for supershift analysis: anti-ISGF-3 (STAT91/84)
monoclonal antibody was purchased from Transduction Laboratories
(G16920) and has been described previously (14). Anti-STAT5B polyclonal
antibody (C17), which cross-reacts with human STAT5A and STAT5B, was
obtained from Santa Cruz Biotechnology, Inc. (SC-835X). Anti-IGF-I
receptor monoclonal antibody was from Oncogene Science (Uniondale, NY),
anti-Soy bean polyclonal antibody was from Sigma (S2519), and
antiphosphotyrosine antibody was from Chemicon International Ltd.
(Harrow, UK). Rabbit IgG was purchased from Vector Laboratories
(Peterborough, UK).
2) Cell culture and preparation of cell lysates
Fibroblast cultures were established from skin biopsies taken
from three healthy children of normal stature (NI, NII, and NIII) and
from three children with GHBP-positive Laron Syndrome (HI, MI, and
MII), as described previously (25). Cells were maintained in a 37 C,
5% CO2 incubator in Costar 75 cm2 flasks in
DMEM (Sigma) supplemented with 10% FBS, 1 mM
L-glutamine, 50 IU/ml penicillin, and 50 µg/ml
streptomycin. Upon reaching monolayer confluence, cells were split by
treatment with 1 g/liter trypsin/0.4 g/liter EDTA solution to give a
final concentration of 5 x 104 cells/ml. At 3 days,
cells were washed twice with PBS and incubated for a further 24 h
in DMEM containing 0.1% BSA before treatment with or without hGH (200
ng/ml, unless stated otherwise) at 37 C for the indicated times. The
cells were then rinsed twice in ice-cold PBS to terminate the
incubations.
For Western blotting, cells were scraped on ice into detergent buffer (10 mM Tris, pH 7.4, 150 mM NaCl, 1 mM EGTA, 1 mM EDTA, 1% Triton X-100, 0.5% NP40) containing protease inhibitors (0.2 mM Na3VO4, 0.2 mM phenylmethylsulfonyl fluoride, 10 µg/ml leupeptin, and 10 µg/ml pepstatin A). Cell lysates were quick frozen in liquid nitrogen and stored at -70 C. Before use, lysates were centrifuged at 10,000 x g for 30 min at 4 C. For JAK2 immunoprecipitation, an aliquot of the supernatant was incubated with 1:100 of the JAK2 antibody, before the addition of protein A-agarose (Boehringer Mannheim) for a further 1 h at 4 C. The agarose pellets were washed twice in detergent buffer, and proteins that were specifically bound were removed by boiling the pellet in 1 x Laemmli sample buffer (30).
For EMSA, cells were scraped on ice into an equal volume of 150 mM NaCl/10 mM Tris, pH 8.0, and lysis buffer (0.1% NP40, 1 mM EDTA, 1 mM DTT, 20% glycerol) containing protease inhibitors (1 mM Na3VO4, 0.5 mM phenylmethylsulfonyl fluoride, 1 µg/ml leupeptin, 1 µg/ml aprotinin), and incubated for 1 h at 4 C with occasional mixing. Cell lysates were then microfuged (10,000 x g, 10 min, 4 C), and supernatants were immediately stored at -70 C until use.
3T3-F442A fibroblasts were kindly provided by Dr. N. Anderson (Department of Surgery, University of Manchester, UK), and were grown in DMEM (4.5 mg/ml glucose, GIBCO, BRL, Paisley, UK) supplemented with 2 mM L-glutamine and 10% FBS. Cells were incubated in DMEM/0.1% BSA/0.5% FBS for 24 h before treatment with or without 200 ng/ml hGH for 15 min. Cell lysates were prepared as described above. Human IM-9 cells were purchased from European Collection of Cell Cultures (Salisbury, UK) and were grown in suspension in RPMI 1640 medium, supplemented with 1 mM L-glutamine and 10% FBS. Cells were incubated in RPMI 1640/0.1% BSA for 24 h before treatment with or without hGH (200 ng/ml) for 15 min IM-9 cells were then pelleted by centrifugation at 1000 x g for 3 min, washed in ice-cold PBS, and lysed as described above.
Protein concentration of cell lysates was determined spectrophotometrically using Bio-Rad (Hemel Hempstead, UK) protein assay dye. All experiments were performed on a minimum of two independent lysate preparations from each skin fibroblast line.
3) Western blotting
Western blotting was performed as described previously (5).
Lysates or immunoprecipitates were fractionated through a 7.5%
polyacrylamide gel according to the method of Laemmli (30) and
electrophoretically transferred to nitrocellulose membranes. Membranes
were blocked and then incubated with a polyclonal antiphosphotyrosine
antibody or the specific JAK/STAT antibodies (see Reagents).
Filters were washed, incubated with donkey antirabbit whole antibody
conjugated to horseradish peroxidase (Amersham), and washed again.
Antibody binding was detected using an Amersham ECL kit.
4) EMSAs
The following oligonucleotides were synthesized:
1) The lactogenic hormone responsive region (LHRR) from the bovine ß-casein gene
sense: 5'-gatcAGATTTCTAGGAATTCAAATC
antisense: 5'-gatcGATTTGAATTCCTAGAAATCT
2) m67, the high affinity form of the c-sis inducible element (SIE) from the human c-fos gene
sense: 5'-agctTCATTTCCCGTAAATCCCTA
antisense: 5'-agctTAGGGATTTACGGGAAATGA
3) The interferon responsive element (IRE) from the human interferon regulatory factor 1 gene
sense: 5'-gatcCAGCCTGATTTCCCCGAAATGACG
antisense: 5'-gatcCGTCATTTCGGGGAAATCAGGCTG
Sense and antisense oligonucleotides of each DNA element were annealed,
end-labeled using polynucleotide kinase (Boehringer Mannheim) and
[
-32P]ATP (4500 Ci/mmol, ICN), and purified on a G-25
Quick Spin Column (Boehringer Mannheim, Lewes, UK).
EMSA was performed by adding 10 µl (20 µg) protein lysate and 5
µl H2O (or antibody for supershift experiments [1.25
µg], or unlabeled double-stranded oligonucleotide for competition
experiments) to 8 µl 4 x EMSA buffer (10% glycerol, 1%
Ficoll, 0.06% NP40, 0.2 mg/ml BSA, 8 mM spermidine, 4
mM EDTA, 2 mM DTT, 350 mM KCl, 16
mM HEPES, pH 7.8) containing 3 µg
polydeoxyinosinic-deoxycytidylic acid, and the mixture incubated at
room temperature for 15 min 1 µl (1 ng) of 32P-labeled
oligonucleotide (
1 x 105 cpm) was added to each
reaction to give a final volume of 24 µl, and the mixture incubated
at 30 C for 30 min The samples were immediately electrophoresed on a
4.5% polyacrylamide gel containing 2.5% glycerol and 0.25 x TBE
(22.5 mM Tris borate, 0.5 mM EDTA, pH 8.0) at
200 V. Gels were fixed, dried, and autoradiographed for 1 to 8
days.
5) RT-PCR
Identification of mRNAs for STAT5A and B (18) was achieved by
RT-PCR. Cell pellets were lysed with 0.5 ml RNAzol B (Tel-Test Inc.,
Friendswood, TX), and total cellular RNA was isolated according to
manufacturer’s instructions. Preparation of cDNA from total RNA was
achieved using an RT system (Promega Corporation, Madison, WI). Human
STAT5A and B were amplified from cDNA by 35 cycles of PCR using a
Techne Unit Progene thermocycler (Techne Ltd., Cambridge, UK) (60 sec
denaturation at 95 C, 45 sec annealing at 58 C, and 60 sec elongation
at 72 C).
1) Human STAT5A-specific primers
sense: 5'-CCTTCTTGTTGCGCTTTAGTG-3'
antisense: 5'-TCGAGTACATGGTCAGGGTTC-3'
2) Human STAT5B specific primers
sense: 5'-GTGAGGCGCTCAACATGAAAT-3'
antisense: 5'-AAGCTGAAGATGGAGAGGTCG-3'
PCR products were electrophoresed on a 1.5% agarose gel containing ethidium bromide and visualized by UV light.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
). cDNAs generated from normal, HI, MI,
and MII fibroblasts and IM-9 cells were amplified with primers specific
for hSTAT5A and hSTAT5B. Single PCR products of 400 bp and 865 bp,
respectively, were observed in all cell types (Fig. 1B), confirming the
presence of both isoforms.
|
). Tyrosine phosphorylation
of this protein, characteristic of the STATs, was maximal after 15 min
of GH treatment and still present after 30 min. Direct immunoblotting
of lysates using a STAT5B antibody revealed a GH-induced partial band
shift to a higher mol mass in IM-9 cells (18) and in normal
fibroblasts, representing a phosphorylated form of STAT5B (Fig. 2B). No
GH-induced phosphorylation shift in either STAT1 or STAT3 was detected
in either cell type (Fig. 2B).
|
). Whereas lysates from
GH-treated 3T3-F442A cells formed two distinct complexes with the SIE,
consistent with the studies of Meyer et al. (13), no
GH-induced binding to this element was observed in normal skin
fibroblasts (n = 5 experiments). Furthermore, whereas GH treatment
of 3T3-F442A cells resulted in the formation of a single complex with
the IRE, GH failed to induce binding to this element in normal
fibroblasts (n = 2 experiments). However, GH did induce the
formation of a distinct complex with the LHRR in these cells. This
complex, which was of similar mobility to that formed in GH-treated
3T3-F442A cells (20), was induced within 10 min of GH treatment and
reduced within 60 min (n = 3 experiments). When normal fibroblasts
were treated with a range of GH doses (2–20,000 ng/ml) for 15 min,
optimal DNA-protein binding was induced at 2000 ng/ml GH (Fig. 3B). The
formation of this complex was inhibited by an excess of unlabeled LHRR
in skin and 3T3-F442A fibroblasts and in IM-9 lymphocytes, confirming
the specificity of this complex (Fig. 4A). STAT-specific antibodies were used
to determine the composition of the GH-induced LHRR-protein complex.
Preincubation of GH-treated lysates with anti-ISGF3 (anti-STAT1)
antiserum supershifted the majority of this complex in normal skin
fibroblasts, but not in IM-9 (18) or 3T3-F442A cells (Fig. 4, A and B).
The antibody to STAT5, however, supershifted the GH-induced complex in
all three cell types (Fig. 4, A and B). A complex that migrated faster
through the gel, was competed by excess unlabeled probe, but was not GH
responsive, was seen in all cell types (Fig. 4A, arrowheads). The GH-induced LHRR/STAT1/5 complex in normal
fibroblasts was not supershifted by IgG or by two unrelated control
antibodies but was inhibited by an antiphosphotyrosine antibody (Fig. 4B).
|
|
), binding to the LHRR was induced
within 10 min of GH treatment and was reduced within 60 min in two
independent experiments. The complex was supershifted with both
anti-STAT1 and anti-STAT5 antibodies (Fig. 5B). However, no GH-induced
binding to the LHRR was observed in HI fibroblasts (Fig. 5A) in four
independent experiments.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
We have demonstrated that the components necessary for the activation
of JAK-STAT signaling by GH (i.e. JAK2 and STATs 1, 3, and
5) are expressed by normal fibroblasts. Furthermore, by RT-PCR, we have
shown that these cells (and the LS fibroblasts) express mRNAs for both
STAT5A and STAT5B. When lysates were immunoprecipitated with anti-JAK2
or anti-STAT antibodies, and the immune complexes analyzed by Western
blotting using an antiphosphotyrosine antibody (or vice
versa), no GH-induced JAK2 or STAT tyrosine phosphorylation could
be detected in the normal fibroblasts, in contrast to the 3T3-F442A and
IM-9 cell lines (data not shown). We were also unable to detect
activation of JAK2 in response to GH using a specific JAK2 kinase assay
(data not shown). It is therefore likely that the coimmunoprecipitation
technique and the kinase assay were not sensitive enough to detect the
comparatively small GH responses in the skin fibroblasts. We have,
however, shown that GH induced the rapid tyrosine phosphorylation of a
protein at approximately 93 kDa in normal fibroblasts that comigrated
with the 93-kDa STAT5 band observed in GH-treated IM-9 cells (18). In
addition, by direct Western blotting with an anti-STAT5B antibody, we
have demonstrated that STAT5 is activated (i.e.
phosphorylated) by GH in normal skin fibroblasts. This coincided with
the formation of a specific GH-induced protein complex with the LHRR.
Because this complex was supershifted completely with STAT5 antisera
but only partially with STAT1 antisera, it is possible that STATs 1 and
5 bind to the LHRR as STAT1/5 heterodimers and STAT5/5 homodimers. The
inability of the anti-STAT1 antibody to supershift the GH-induced
LHRR-STAT5-containing complex in IM-9 and 3T3-F442A cells confirmed
that this antibody did not cross-react with STAT5 in normal
fibroblasts. Because STAT1, as well as STAT5, appears to play a role in
GH-dependent signaling in normal human skin fibroblasts, we cannot
explain why GH-induced STAT1 phosphorylation was not detected by
immunoblotting (Fig. 2B
).
The disappearance of the GH-induced LHRR-STAT complex with an antiphosphotyrosine antibody in normal fibroblasts confirms that the bound STAT proteins were indeed tyrosine phosphorylated in response to GH. Similar results have been demonstrated previously in other cell systems (33, 34). Whereas 200 ng/ml GH was sufficient to induce the LHRR-STAT complex in normal fibroblasts, maximal induction of this complex occurred in response to 2000 ng/ml GH. However, at 20,000 ng/ml, induction of the LHRR/STAT complex was reduced, consistent with the observation that high concentrations of GH inhibit GHR dimerization (35, 36). Although GH induced specific binding to the high-affinity SIE (m67) (13) and the IRE in mouse 3T3-F442A fibroblasts, GH failed to induce binding to these elements in normal fibroblasts, indicating species-specific activation of STAT/DNA binding by GH. The lack of GH-induced protein binding to the SIE in normal fibroblasts is consistent with the failure of GH to activate c-fos transcription in these cells (our unpublished data).
The finding that GH induced the formation of a specific protein complex with the LHRR in IM-9 cells, which could be supershifted by STAT5 antisera but not by STAT1 antisera, has been described previously (18). In the same study, Silva et al. demonstrated GH-induced tyrosine phosphorylation of STAT5 in mouse 3T3-F442A cells (18). In accordance with the studies of Han et al. (20), we show that GH induced a specific protein complex with the LHRR in 3T3-F442A cells. Although STAT1 is also known to be tyrosine phosphorylated in response to GH in these cells (13), this complex could only be supershifted with the STAT5 antibody, and not with the STAT1 antibody. These studies in normal human skin fibroblasts, human IM-9 lymphocytes, and mouse 3T3-F442A fibroblasts therefore confirm cell type- and species-specific differences in GH activation of STAT/DNA binding.
We have previously described four girls from two families with GHBP-positive LS. The two affected girls from the first family (HI and HII) are heterozygous for a D152H point mutation in exon 6 of the GHR gene, as are the unaffected father and a brother (25). No GHR gene abnormalities were identified in the two affected girls (MI and MII) from the second family. We have therefore hypothesized that their GH insensitivity is due to a defect(s) within the intracellular GH signaling pathway. In addition, we have previously shown that, in contrast to normal fibroblasts, skin fibroblasts established from HI, MI, and MII are insensitive to GH at the levels of DNA synthesis and IGFBP-3 gene expression (25). The second aim of this study was to examine the effect of GH on STAT signaling in these LS fibroblast cultures in an attempt to identify the site(s) of GH resistance in these children.
Whereas HI fibroblasts express full-length forms of JAK2, STATs 1, 3,
and 5 (data not shown), GH failed to induce STAT binding to the LHRR in
these cells. In a parallel study, we have shown that, whereas GH
induces the phosphorylation and activation of MAP kinase (MAPK) in
normal fibroblasts, GH-dependent activation of MAPK in HI fibroblasts
is reduced (37). Recent studies indicate that two signaling pathways to
MAPK can be activated by GH in 3T3-F442A fibroblasts, one involving a
JAK2
SHCGrb2Sosras rafMEK cascade (9), and the other
by the intermediate activation of phosphoinositide 3-OH kinase by a
mechanism involving JAK2 and IRS-1 (38). Our findings therefore would
suggest that the site of GH resistance in HI lies at the level of JAK2
or between the GHR and JAK2. Further studies are necessary to determine
whether the abnormality is intrinsic to JAK2 itself, or resides within
another, as yet, unidentified protein involved in the GHR/JAK2
complex.
Skin fibroblasts derived from MI and MII expressed the full-length forms of JAK2, STATs 1, 3, and 5. In contrast to HI fibroblasts, GH did induce STAT1 and STAT5 binding to the LHRR in these cells; MI with a time course identical to that seen in normal fibroblasts. In contrast, GH-induced binding and down-regulation occurred more rapidly in MII cells than in MI and normal fibroblasts. As it is likley that the siblings MI and MII have the same signaling defect, we feel that this difference in kinetics is not relevant. Thus, although GH failed to activate DNA synthesis and IGFBP-3 gene expression in MI and MII fibroblasts (25), GH was capable of inducing activation of STAT signaling in these cells. Further studies will determine whether this pathway is functional at the transcriptional level. In the same cells, GH dependent activation of MAPK was significantly reduced compared with normal fibroblasts (37). These data therefore suggest that the intracellular GH signaling defect in MI and MII may lie distal to JAK2 in an ancillary pathway that modulates MAPK activity. Our results indicating that in HI the site of GH resistance lies at the level of JAK2 (central to the initiation of GH signal transduction), whereas in MI and MII it appears to be located along another limb of the GH signaling pathways (affecting MAPK but not STAT activation), could be consistent with the severity of the phenotypes of these girls: HI has a classical LS phenotype (Ht. SDS: -6.8), whereas MI and MII have atypical LS phenotypes, and are less severely growth retarded (Ht. SDS: -4 and -3.4, respectively) (25).
In summary, we have demonstrated that GH rapidly and transiently activates STAT signaling in normal skin fibroblasts, where STATs 1 and 5 appear to bind to DNA. We have shown that GH activates STAT-DNA binding in skin fibroblasts from two sisters with GHBP-positive LS, indicating a molecular defect elsewhere within the GH signaling network. In contrast, GH failed to activate the STAT pathway in fibroblasts derived from another GHBP-positive LS girl. The GH insensitivity syndrome with normal GHBP is therefore associated with diverse abnormalities within the GH intracellular signaling pathways.
Received March 17, 1997.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
in IM-9 cells. J Biol Chem 269:27532–27539 and leukemia
inhibitory factor promoted tyrosyl phosphorylation of insulin receptor
substrate-1. J Biol Chem 270:14685–14692This article has been cited by other articles:
 |
|
 |
|
  J. Xu, S. Ji, D. Y Venable, J. L Franklin, and J. L Messina Prolonged insulin treatment inhibits GH signaling via STAT3 and STAT1 J. Endocrinol., March 1, 2005; 184(3): 481 - 492. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. G. Rosenfeld and V. Hwa Toward a Molecular Basis for Idiopathic Short Stature J. Clin. Endocrinol. Metab., March 1, 2004; 89(3): 1066 - 1067. [Full Text] [PDF]
|
|
|
|
 |
|
 S. R. Edmondson, S. P. Thumiger, G. A. Werther, and C. J. Wraight Epidermal Homeostasis: The Role of the Growth Hormone and Insulin-Like Growth Factor Systems Endocr. Rev., December 1, 2003; 24(6): 737 - 764. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. M. Kofoed, V. Hwa, B. Little, K. A. Woods, C. K. Buckway, J. Tsubaki, K. L. Pratt, L. Bezrodnik, H. Jasper, A. Tepper, et al. Growth Hormone Insensitivity Associated with a STAT5b Mutation N. Engl. J. Med., September 18, 2003; 349(12): 1139 - 1147. [Full Text] [PDF]
|
|
|
|
 |
|
 C. M. Silva, M. T. Kloth, A. J. Whatmore, J. S. Freeth, N. Anderson, K. K. Laughlin, T. Huynh, A. J. Woodall, and P. E. Clayton GH and Epidermal Growth Factor Signaling in Normal and Laron Syndrome Fibroblasts Endocrinology, July 1, 2002; 143(7): 2610 - 2617. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Beauloye, B. Willems, V. de Coninck, S. J. Frank, M. Edery, and J.-P. Thissen Impairment of Liver GH Receptor Signaling by Fasting Endocrinology, March 1, 2002; 143(3): 792 - 800. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Salerno, B. Balestrieri, E. Matrecano, A. Officioso, R. G. Rosenfeld, S. Di Maio, G. Fimiani, M. V. Ursini, and C. Pignata Abnormal GH Receptor Signaling in Children with Idiopathic Short Stature J. Clin. Endocrinol. Metab., August 1, 2001; 86(8): 3882 - 3888. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Baetta, M. Soma, C. De-Fraja, C. Comparato, C. Teruzzi, L. Magrassi, and E. Cattaneo Upregulation and Activation of Stat6 Precede Vascular Smooth Muscle Cell Proliferation in Carotid Artery Injury Model Arterioscler. Thromb. Vasc. Biol., April 1, 2000; 20(4): 931 - 939. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Iida, Y. Takahashi, H. Kaji, N. Onodera, M. O. Takahashi, Y. Okimura, H. Abe, and K. Chihara The C422F Mutation of the Growth Hormone Receptor Gene Is Not Responsible for Short Stature J. Clin. Endocrinol. Metab., November 1, 1999; 84(11): 4214 - 4219. [Abstract] [Full Text]
|
|
|
|
|
|
J. Wojcik, M. A. Berg, N. Esposito, M. E. Geffner, N. Sakati, E. O. Reiter, S. Dower, U. Francke, M.-C. Postel-Vinay, and J. Finidori Four Contiguous Amino Acid Substitutions, Identified in Patients with Laron Syndrome, Differently Affect the Binding Affinity and Intracellular Trafficking of the Growth Hormone Receptor J. Clin. Endocrinol. Metab., December 1, 1998; 83(12): 4481 - 4489. [Abstract] [Full Text]
|
|
|
|
|
|
D. W. Love, A. J. Whatmore, P. E. Clayton, and C. M. Silva Growth Hormone Stimulation of the Mitogen-Activated Protein Kinase Pathway Is Cell Type Specific Endocrinology, April 1, 1998; 139(4): 1965 - 1971. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
