| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
Department of Pathology, Yamanashi Medical University, Tamaho, Yamanashi 409-3898, Japan
Address all correspondence and requests for reprints to: Dr. Xin Li, Department of Pathology, Yamanashi Medical University, 1110 Shimokato, Tamaho, Nakakoma, Yamanashi 409-3898, Japan. E-mail: lixin{at}res.yamanashi-med.ac.jp
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
To further determine a potential role of TSH in regulating the apoptosis of thyroid follicular cells, we attempted to induce apoptosis by the deprivation of serum and hormones in a rat thyroid cell line, FRTL-5 cells. FRTL is a functional thyroid follicular cell line that was established from normal Fisher rat thyroid and requires six hormones, including TSH, for growth (7). Therefore, this cell line is thought to be an ideal model for the investigation of hormone-dependent apoptosis of the thyroid follicular epithelial cells. FRTL-5 cells that were cloned from FRTL cells share many features with FRTL cells, such as the expression of TSH receptor, TSH-dependent growth, synthesis and secretion of thyroglobulin, active trapping of iodine, etc. TSH was particularly important for the growth of FRTL/FRTL-5 cells. Its withdrawal was thought to be an effective means of cell synchronization (8). FRTL cells were found to undergo dramatic morphological changes and ceased growth when cultured in the absence of TSH (9). However, the occurrence of apoptosis in FRTL-5 cells induced by deprivation of TSH has not been reported to date. Recently, there was a report that apoptosis was induced in quiescent FRTL-5 cells by transforming growth factor-ß1. Transforming growth factor-ß1 was found to prevent resting cells from entering the cell cycle when stimulated by serum and a mixture of five hormones, which was added in the usual culture of FRTL-5 cells, but not by TSH and insulin-like growth factor I (IGF-I), accompanied by a reduction in cyclin D and an increase in c-myc (10).
In this study, we demonstrate for the first time that FRTL-5 cells underwent anchorage-dependent apoptosis in the absence of serum and hormones as evaluated by several apoptosis features, and that TSH prevented apoptosis by promoting cell-matrix adhesion and cell cycle progression partly via the cAMP pathway.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
To induce apoptosis, we examined two culture conditions: 1) FRTL-5 cells were cultured in 6H5 for 3 days to reach subconfluence and washed with PBS (-), then further cultured in Ham’s serum-free medium containing 0.2% BSA (Wako, Japan), which was referred to as 0H, for up to 1 week; 2), FRTL-5 cells were cultured in 6H5 medium for 3 days to reach subconfluence and washed with PBS (-), then starved in 0H for 1–2 days. The cells were detached by trypsin treatment and plated at 5 x 105/ml in 0H with or without 10 mU/ml TSH or 1 mM (Bu)2cAMP (Wako) for the indicated times. For blockage of cell-matrix adhesion, FRTL-5 cells were cultured in 2% agarose-coated dishes or flasks.
Cell viability evaluated by
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrasodium bromide (MTT)
assay
The MTT assay was carried out according to the manufacturer’s
instructions (Chemicon International, Inc., Temcula, CA). Briefly,
FRTL-5 cells were passaged into 96-well tissue plates (Falcon) at about
0.5 x 105/100 µl and cultured in 0H with or without
10 mU/ml TSH or 1 mM cAMP for up to 72 h. To determine
the contribution of the cAMP-dependent pathway to the antiapoptotic
effect, FRTL-5 cells were pretreated with a specific protein kinase A
inhibitor H-89
(N-[2-(p-bromocinnamylamino)ethyl-5-isoquinolinesulfonamide;
BIOMOL, Plymouth Meeting, PA; 25 µM) for 30 min (11). On
the day of the experiment, 10 µl 5 mg/ml MTT were added to each pore
and cultured for an additional 4 h. Then 100 µl isopropanol with
0.04 N HCl was added and pipetted thoroughly. Within 1
h, the value of absorbance expressed by the OD at 570 nm by
spectrophotometry was measured on a plate reader (SPECTRAmax 340,
Molecular Devices, Menlo Park, CA). Cell viability at the
indicated times was expressed as a percentage of the absorbance of
cells at time zero. Data were presented as mean ± SD
of triplicate determinations from three independent experiments.
Specific labeling of nuclear DNA fragmentation by the terminal
deoxynucleotidyl transferase-mediated deoxy-UTP nick end labeling
(TUNEL) method (12)
FRTL-5 cells were collected by cell scrapers and washed with PBS
(-). The cells were then fixed with 4% paraformaldehyde for 15 min at
room temperature and spotted onto silane-coated slides. After washing
with PBS, the cells were treated with 2 µg/ml proteinase K for 15 min
at room temperature. After washing with PBS and rinsing with TDT buffer
(30 mM Trizma base, 140 nM sodium cacodylate,
and 1 mM cobalt chloride), pH 7.2, the cells were incubated
with 0.3 U/µl TDT and 0.01 mM biotin-16-deoxy-UTP
(Sigma Chemical Co.) in the above buffer for 1 h at
37 C. The reaction was blocked by TB buffer (300 mM sodium
chloride and 30 mM sodium citrate) for 15 min at room
temperature. After washing with PBS, the cells were incubated with
fluorescein-labeled streptavidin in 1% BSA/PBS containing 2 µg/ml
4',6'-diamido-2 phenylindole hydrochloride (DAPI; Boehringer Mannheim Biochemica) for 60 min at room temperature.
After washing with PBS, the slides were sealed with 50% glycerol in
PBS. Photographs were taken under a fluorescence microscope
(Nikon, Tokyo, Japan).
Transmission electron microscopy
FRTL-5 cells were collected by cell scrapers and centrifuged at
1500 rpm for 10 min. The pellets were fixed with 2.5% glutaraldehyde
in PB buffer, then postfixed with 1% OsO4 in PB
buffer. After having been dehydrated by an ethanol series, the cells
were embedded in Epon 812. Ultrathin sections were cut by an
ultramicrotome (LKB, Rockville, MD), stained with uranyl
acetate and lead citrate, and observed under an electron microscope
(JEM-1005X, JEOL, Tokyo, Japan) at 80 kV.
DNA fragmentation analysis by agarose gel electrophoresis
Low mol wt DNA was isolated as described previously (13). Cells
(2 x 106) were collected by cell scrapers and washed
with PBS (-). The resulting cell pellets were resuspended in lysis
buffer [10 mM Tris-HCl (pH 8.0), 10 mM EDTA,
and 0.5% Triton X-100] and centrifuged at about 13,000 x
g for 20 min at 4 C to separate the fragmented DNA (soluble)
from intact chromatin. The supernatant was treated with 0.1 mg/ml
deoxyribonuclease-free ribonuclease for 1 h at 37 C, followed by
0.2 mg/ml proteinase K and 1% SDS for 2 h at 50 C. The DNA was
then extracted with phenol and phenol/chloroform and precipitated
overnight with isopropanol at -20 C. The DNA samples were analyzed
with 1.8% agarose gel.
Flow cytometric analysis for DNA fragmentation and cell cycle
Flow cytometric analysis was performed as described previously
(14). Briefly, FRTL-5 cells were cultured in tissue flasks under the
conditions described above. The cells were collected by cell scrapers
and centrifuged. Then the cell pellets were gently resuspended in 1.2
ml hypotonic fluorochrome solution containing propidium iodide (PI; 50
µg/ml) in 0.1% sodium citrate and 0.1% Triton X-100 and stained
overnight at 4 C. The cells were analyzed on a fluorescence-activated
cell sorter (Becton Dickinson and Co., Rutherford, NJ).
The data were analyzed using ModFitLT V1.00 (Macintosh)
software.
Indirect immunofluorescence
Antibodies were obtained from the following sources: rabbit
antimouse laminin antibody (Sigma Chemical Co.), rabbit
antimouse type IV collagen (LSL, Japan), rabbit antirat
fibronectin (Life Technologies, Inc.), rabbit antihuman
5 integrin (Chemicon), mouse antihuman p27 monoclonal
antibody (Transduction Laboratories, Inc., Lexington, KY),
anti-cyclin D1 and 2/bcl-1 monoclonal antibody (MBL, Japan),
antifocal adhesion kinase [anti-FAK; monoclonal antibody from
Upstate Biotechnology, Inc. (Lake Placid, NY), and
polyclonal antibody from Santa Cruz Biotechnology, Inc.
(Santa Cruz, CA)], and antiphosphorylated tyrosine (4G10,
Upstate Biotechnology, Inc.; PY20, Transduction Laboratories, Inc.).
The cells for immunofluorescence tests were seeded onto glass
coverslips of 1.2 cm in diameter, fixed with 4% paraformaldehyde in
PBS for 15 min at room temperature, and permeabilized with 0.1% Triton
X-100 in PBS for 30 min. For 5 integrin, the cells were
fixed with cold 100% acetone for 3 min. The cells were incubated with
primary antibody in 1% BSA/PBS overnight at 4 C. The coverslips were
then washed with PBS and allowed to react with fluorescein-labeled
secondary antibody (DAKO Corp.) for 45 min at room
temperature. After washing with PBS, the coverslips were mounted on
slides using 50% glycerol in PBS and observed using a confocal
scanning microscope (Leica Corp., Heidelberg, Germany).
For actin filament staining, paraformaldehyde-fixed and permeabilized
cells were stained with 2 U/ml rhodamine-phalloidin (Molecular Probes, Inc., Eugene, OR) for 1 h at room temperature.
The cells stained by anti-5 integrin antibody were also
analyzed using a fluorescence-activated cell sorter (Becton Dickinson and Co.).
Immunoprecipitation and Western blot analysis
FRTL-5 cells were seeded in 75-ml flasks in 6H5 medium for 3
days to subconfluence and starved in 0H for 1–2 days. The cells were
then detached by trypsin treatment and seeded at about 5 x
106/10 ml in 10-cm dishes in 0H with or without 1 mU/ml TSH
or 1 mM cAMP for the indicated times. The cells were
collected by cell scrapers and washed once with cold PBS. After
centrifuging, the cell pellet was lysed at 4 C in 0.3 ml lysis buffer
containing 50 mM Tris-HCl (pH 7.4), 1 mM sodium
vanadate, 1 mM EDTA, 1% Triton X-100, 25 µg/ml
leupeptin, and 0.5 mM phenylmethylsulfonylfluoride;
sonicated briefly; and centrifuged. After adding 0.1 ml of 3 x
Laemmli’s sample buffer to 0.2 ml of the above suspension, the mixture
was incubated for 3 min at 100 C. Protein assay was carried out by the
method of Lowry. For immunoprecipitation, the lysate (
500 µg
protein) was precleared with 50 µl protein A-Sepharose beads, and the
supernatant was incubated with 3 µg anti-FAK monoclonal antibody for
2 h at 4 C. The immunocomplex was recovered by the addition of 50
µl protein A-Sepharose beads, incubated for 1 h at 4 C, and
centrifuged. The beads were washed with lysis buffer three times and
eluted by boiling in sample buffer.
For Western blot analysis, approximately 50 µg immunoprecipitated protein or whole cell lysate were run on 8% or 12% SDS-PAGE and transferred to a polyvinylidene difluorene membrane (ATTO, Japan). Blots were blocked with 1% BSA in PBS overnight at 4 C. After washing with PBS containing 0.4% Tween-80 three times, the blots were incubated with 1 µg/ml antiphosphotyrosine antibody (4G10 and PY20) or anti-FAK polyclonal antibody for 2 h, rinsed, and incubated with a 1:10,000 dilution of horseradish peroxidase-labeled goat secondary antibody for 2 h. The blots were again rinsed and visualized using enhanced chemiluminescence (ECL kit, Amersham Pharmacia Biotech, Arlington Heights, IL) according to the manufacturer’s instructions. For cell cycle-relevant proteins, the membrane was first immunoblotted with a monoclonal antibody against cyclin D (MBL) at a dilution of 1:100. After having been developed by ECL, the membrane was deprobed using stripping buffer (62.5 mM Tris-HCl, 2% SDS, and 0.4 M 2-mercaptoethanol) at 60 C for 30 min and reprobed by a monoclonal antibody against p27 (Transduction Laboratories, Inc.) at a dilution of 1:2500.
Statistics
Statistical analysis was performed using ANOVA, followed by
Fisher’s protected least significant difference test (PLSD; StatView
4.0). P < 0.01 was selected as the level of
significance.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
) when cultured by
method 1, in which FRTL-5 cells had reached subconfluence in the
presence of serum and hormones, then starved in 0H for up to 1 week,
the percentage of apoptotic cells was quite low as evaluated by flow
cytometry (not more than 20%). It was difficult to induce apoptosis by
deprivation of serum and hormones if FRTL-5 cells became attached to
the substrate, suggesting that cell-matrix adhesion might play an
important role in maintaining the viability of FRTL-5 cells. Therefore,
we tried to induce apoptosis by method 2, i.e. deprivation
of serum and hormones as soon as subpassage was performed (see
Materials and Methods). Sustained cell attachment was not
obtained in the absence of TSH. Most cells floated in the serum-free
medium and seemed to undergo apoptosis after 24 h (Fig. 2a). In contrast, in the presence of TSH,
the cells attached to the polystyrene substrate and survived, forming
follicle-like colonies, although the cells did not spread well without
serum (Fig. 2b). cAMP showed an effect similar to TSH on maintenance of
cell adhesions and cell viability (data not shown). Because apoptosis
seemed to be satisfactorily induced by the latter method, we used this
culture condition throughout the following experiments.
|
|
The results showed that the cell viability was significantly reduced
time dependently when FRTL-5 cells were cultured in 0H without TSH. In
contrast, TSH and cAMP apparently maintained the viability of FRTL-5
cells during the first 48 h of culture (p <
0.0001; Fig. 3a). The apparent increase
in cell viability at 72 h might result from the increase in cell
number due to the mitogenic effect of TSH or cAMP. To further determine
the contribution of the cAMP/PKA pathway to the maintenance of cell
viability, we pretreated FRTL-5 cells with a specific protein kinase A
inhibitor, H-89, 30 min before TSH or cAMP challenge. H-89 (25
µM) almost completely inhibited the effect of TSH or cAMP
on the maintenance of cell viability at 48 h (p <
0.01; Fig. 3b). The inhibition was not due to the cytotoxic effect of
this inhibitor, as it did not affect the viability of cells cultured in
0H. The above results suggested that TSH might maintain cell viability
and prevent apoptosis in FRTL-5 cells mainly via the cAMP/PKA
pathway.
|
). The fragmented nuclei were positive
for TUNEL (arrow in Fig. 4b), indicating the presence of
nicks in DNA strands of these cell nuclei. Gel electrophoretic analysis
of DNA revealed ladders of 180–200 bp in the cells cultured in 0H
without TSH. This behavior was not observed in cells cultured in the
presence of TSH or cAMP (Fig. 5).
|
|
) or disrupted (arrowhead in
Fig. 6) in clear contrast to nonapoptotic nuclei
(asterisk in Fig. 6). Apoptotic bodies were frequently seen.
In addition, these cells seemed to have more lysosomes and autophagic
activity.
|
). The percentages of apoptotic cells
and the cells in different phases of the cell cycle were derived from
the cell cycle analysis software ModFit LTV 1,00. As shown in Fig. 7b, the percentage of apoptotic cells increased time dependently in 0H
without TSH. Further, we found that the percentage of cells in the S
phase decreased, whereas the cells in the G2/M phase increased
simultaneously (Fig. 7a), suggesting that there was cell cycle arrest
of the G1 transition. TSH inhibited the appearance of
subdiploidy cells and promoted the progression of the G1/S
phase transition. cAMP showed effects similar to TSH (Fig. 7, a and
b).
|
),
implying that anchorage-dependent FRTL-5 cells cultured in the absence
of serum and hormones might activate the pathway of growth arrest and
induce apoptosis, and that TSH could prompt the cell cycle progression
partly via the cAMP pathway.
|
). As for the integrin receptors, we
observed the expression of 5 integrin. There was also
not a large difference between the groups with and without TSH by
immunofluorescence and flow cytometry (data not shown).
|
), suggesting that cell-matrix
interaction is important for the activation of survival signals. As the
cell-matrix interaction is known to stimulate a series of kinases and
induce tyrosine phosphorylation of proteins, we then examined the
tyrosine phosphorylation of protein of FRTL-5 cell lysates using
Western blotting analysis. As shown in Fig. 11, TSH apparently increased the
tyrosine phosphorylation of 120- to 125-kDa substrates after 24 h
(arrowhead). We further examined the expression of FAK and
its phosphorylation. TSH was found to increase the expression and
phosphorylation of FAK in our culture system (Fig. 12, a and b).
|
|
|
). We colocalized actin filament with
FAK at the contact plaques (arrowheads in Fig. 13, a and c)
by double staining the FAK-immunolabeled cells with rhodamine-phaloidin
(red sites in Fig. 13). Similar to another report (18), we
also observed the enhanced immunoreactivity of FAK in the cell nuclei
(arrow in Fig. 13b) and membrane blebs (arrowhead
in Fig. 13b) in apoptotic cells, but we did not detect fragments from
the cleavage of FAK by immunoblot analysis (data not shown).
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
As early as 1978, Folkman and Moscona proposed that appropriate cell shape was critical for DNA synthesis by normal cells, which was explained as the theory of anchorage dependence (19). This means that normal cells will not grow unless attached to a substratum. When the interaction between normal cells and the extracellular matrix is blocked, apoptosis occurs. This phenomenon was named anoikis, a Greek word meaning homelessness, by Frisch and Francis (20). Anoikis was first reported by Meredith et al. (21), who induced apoptosis by incubating human umbilical vein endothelial cells by plating the cells on agarose-coated dishes or in the absence of serum and growth factors. In addition to endothelial cells, anchorage-dependent apoptosis was also observed in kidney cells (18), mammary cells (22), ovarian cells (23, 24), melanocytes (25), and oncogene-transformed fibroblasts (26). To our knowledge, this is the first report of anchorage-dependent apoptosis induced in thyroid cells.
Extracellular matrix (ECM) proteins such as fibronectin, laminin, and
type IV collagen have been considered to be survival factors for many
cell types. Cell-ECM adhesion is mainly involved in the interaction of
integrins and these extracellular proteins. The downstream signaling
events may associate or activate a number of nonreceptor protein
tyrosine kinases, including FAK, which might provide the cell survival
signal or facilitate gene expression and cell cycle progression events
(27). Consistent with another report (17), we demonstrated that FRTL-5
cells synthesized and secreted the main ECM proteins. On the other
hand, thyroid cells were reported to express integrins such as
1ß1, 3ß1,
5ß1, and 6ß1
(28). 5 integrin was localized in FRTL-5 cells by
immunofluorescence in our study (data not shown). As apoptosis still
occurred despite the binding of ECM to integrin receptors when TSH and
serum were absent, we hypothesized that the simple ligation of integrin
was not sufficient for the maintenance of viability of FRTL-5 cells
under these conditions. Other events that are activated by TSH may be
needed. FAK, which is thought to act as an integrin-mediated signal
transducer, was reported to play a role in the prevention of apoptosis
in diverse cell types (29, 30, 31). Inhibition of FAK resulted in apoptosis
in fibroblasts (29), and its overexpression prevented anoikis (30, 31).
Immunoblot analysis in our study demonstrated that TSH increased the
expression of phosphorylated FAK, suggesting that FAK may also be
involved in the maintenance of TSH-dependent survival in FRTL-5
cells.
Protein tyrosine phosphorylation was found to regulate the TSH-induced increase in proliferation and iodide uptake through the cAMP pathway in FRTL-5 cells (32). While exploring the effect of TSH on the regulation of growth in FRTL-5 cells, Takahashi et al. (33) also demonstrated that TSH potentiated IGF-I-dependent tyrosine phosphorylation of 175-kDa substrate and stimulated IGF-I-independent accumulation of tyrosine phosphorylation. The latter TSH-dependent phosphorylation was largely confined to the band of 120–125 kDa. The 175-kDa substrate was believed to be related to cytoskeletal proteins, whereas the properties of the 120- to 125-kDa substrate were not determined. Our findings obtained from Western blot analysis of whole cell lysates by antityrosine phosphorylation antibody also demonstrated that TSH strengthened or probably stimulated the tyrosine phosphorylation of 120–125 kDa substrate. The result of immunoprecipitation with anti-FAK antibody showed increased expression of FAK and its tyrosine phosphorylation after TSH stimulation, suggesting that at least FAK may be included in this band.
Recent studies have established the role of cell cycle relevant proteins in the regulation of apoptosis (34, 35). A cyclin-dependent kinase inhibitor, p27, was reported to result in cell cycle arrest of the G1 to S phase transition by associating with cyclin D-CDK4 complex and potentially inhibiting Rb phosphorylation (36). Overexpression of p27 resulted in the G1-S arrest and triggered apoptosis in cancer cells (34). Recently, Carneiro et al. reported the induction of apoptosis in FRTL-5 cells by transforming growth factor-ß1 in the absence of TSH (10), accompanied by increased expression of p15 and c-myc messenger RNA and decreased expression of cyclin D1 and P27. Apoptosis induced in our study was accompanied by decreased expression of cyclin D without a decreased level of p27, but even with a slightly increased expression of p27. These changes were coincident with the onset of apoptosis (i.e. after 24 h; data not shown). Similarly, recent papers using FRTL-5 cells also demonstrated the relation of p27 accumulation to the arrest of G1/S transition (37, 38), in which degradation of p27 was induced during G1/S transition by growth stimulation, including calf serum, insulin, and TSH. In the present study, we showed that TSH alone had apparent effects on the cell cycle-relevant proteins in FRTL-5 cells.
Several pieces of evidence indicated that cross-talk may exist between the pathway of cell adhesion and cell cycle regulation (27, 39, 40, 41). The cyclin E-CDK2 complex was activated in late G1 phase in attached human fibroblasts, but not in suspension-cultured cells. This decreased cyclin E-CDK2 activity in suspension cells resulted from an increase in the amount of CDK2 inhibitors, such as p27 and p21 (39). Also, forced expression of exogenous cyclin A made NRK fibroblasts partially independent of anchorage (40). These data imply a dependence of cyclin/cyclin-dependent kinase activity on cell anchorage. More recently, overexpression of wild-type FAK was reported to accelerate the G1 to S phase transition by increasing cyclin D1 expression and decreasing p21 expression (41). Our results also suggested that increased expression of phosphorylated FAK could be involved in the promotion of cell cycle progression in FRTL-5 cells stimulated by TSH. The factors residing between the proximal cell adhesion events and distal cell cycle machinery were not clear. Lately, the Rho family of small guanosine triphosphatases, which includes Rho, Rac, and Cdc42, was reported to be a molecular switch controlling a signal transduction pathway that links membrane receptors to gene transcription (42). Rho-related kinases exert their biological functions in the rearrangement of the cellular skeleton and enhancement of focal adhesion (43, 44). In addition, the Rho family plays a pivotal role in G1 progression and DNA synthesis in response to mitogenic stimulator (45). It was shown that the geranylation of Rho A protein and its translocation to membranes were essential for the degradation of p27 and G1/S transition in growth-stimulated FRTL-5 cells (37, 38). The effect of the Rho family on regulating apoptosis remains to be studied.
In conclusion, TSH prevents apoptosis by increasing cell-matrix adhesion and facilitating cell cycle progression partly via the cAMP pathway in FRTL-5 cells, thus suggesting that in addition to regulating growth and differentiation, TSH may function as a survival factor in thyroid cells.
Received April 30, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
5ß1 integrin supports survival of cells on
fibronectin and up-regulates bcl-2 expression. Proc Natl
Acad Sci USA 92:6161–6165This article has been cited by other articles:
 |
|
 |
|
  S. Ayturk, A. Gursoy, A. Kut, C. Anil, A. Nar, and N. B. Tutuncu Metabolic syndrome and its components are associated with increased thyroid volume and nodule prevalence in a mild-to-moderate iodine-deficient area Eur. J. Endocrinol., October 1, 2009; 161(4): 599 - 605. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Satoh, X. Li, H. Yokota, M. Osada, Y. Ozaki, R. Katoh, and Y. Yatomi Regulation by Sphingolipids of the Fate of FRTL-5 Cells J. Biochem., January 1, 2009; 145(1): 31 - 36. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Lania, M. Filopanti, S. Corbetta, M. Losa, E. Ballare, P. Beck-Peccoz, and A. Spada Effects of Hypothalamic Neuropeptides on Extracellular Signal-Regulated Kinase (ERK1 and ERK2) Cascade in Human Tumoral Pituitary Cells J. Clin. Endocrinol. Metab., April 1, 2003; 88(4): 1692 - 1696. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Bell, A. Gagnon, P. Dods, D. Papineau, M. Tiberi, and A. Sorisky TSH signaling and cell survival in 3T3-L1 preadipocytes Am J Physiol Cell Physiol, October 1, 2002; 283(4): C1056 - C1064. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Kimura, A. Van Keymeulen, J. Golstein, A. Fusco, J. E. Dumont, and P. P. Roger Regulation of Thyroid Cell Proliferation by TSH and Other Factors: A Critical Evaluation of in Vitro Models Endocr. Rev., October 1, 2001; 22(5): 631 - 656. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. S. Park, H. Kim, J. M. Suh, S. J. Park, S. H. You, H. K. Chung, K. W. Lee, O-Y. Kwon, B. Y. Cho, Y. K. Kim, et al. Involvement of JAK/STAT (Janus Kinase/Signal Transducer and Activator of Transcription) in the Thyrotropin Signaling Pathway Mol. Endocrinol., May 1, 2000; 14(5): 662 - 670. [Abstract] [Full Text]
|
|
|
|
 |
|
 H. Kim, T.-H. Lee, E. S. Park, J. M. Suh, S. J. Park, H. K. Chung, O-Y. Kwon, Y. K. Kim, H. K. Ro, and M. Shong Role of Peroxiredoxins in Regulating Intracellular Hydrogen Peroxide and Hydrogen Peroxide-induced Apoptosis in Thyroid Cells J. Biol. Chem., June 9, 2000; 275(24): 18266 - 18270. [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 |
