| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Department of Pharmacology, Case Western Reserve University, Cleveland, Ohio 44106
Address all correspondence and requests for reprints to: John H. Nilson, Ph.D., Director, School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4234. E-mail: jhn{at}wsu.edu.
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
LH overexpressing mice (LHCTP) develop ovary-dependent pituitary adenomas preceded by hyperproliferation of cells expressing the transcription factor, Pit-1 (11). The pituitary glands of LHCTP mice also have an altered responsiveness to estradiol and develop prolactinomas when exposed to elevated levels of this hormone (11). Conversely, ovariectomy prevented the development of pituitary adenomas. These data suggest that chronic ovarian hyperstimulation provides the critical trophic support for the development of pituitary tumors in LHCTP mice. It is likely that this trophic support must ultimately be accompanied by changes in gene expression that result in tumor formation.
To identify genes that may be involved in pituitary tumorigenesis, we carried out global gene expression profiling on pituitary adenomas derived from LHCTP mice. We have identified a number of genes with altered expression in all pituitary tumors analyzed. One of the most highly expressed genes in LHCTP pituitary tumors was represented by an expressed sequence tag (EST) that encodes p8. Herein, we have correlated expression of p8 with a subset of prolactinomas and characterized the role of p8 or candidate of metastasis-1, a HMGA-like nuclear protein (12, 13) in tumorigenicity of pituitary cells.
p8 Was originally identified in a cDNA library of genes expressed in the acute phase of pancreatitis (14). Based on structural and biochemical analysis, p8 appears to belong to the HMGA1 (HMG-I/Y-like) subfamily of HMG proteins (12). Notably, p8 lacks the AT-hook DNA binding domain, characteristic of HMGA proteins, yet it still possesses the ability to bind DNA sequence nonspecifically (12), another property associated with HMGA proteins. HMGA proteins have been characterized as architectural transcription factors that are involved in chromatin remodeling and enhance some formation (reviewed in Refs. 15 and 16). Genes encoding these proteins are highly expressed during embryonic development but are low or undetectable in adult tissues (reviewed in Ref. 17). Accordingly, the gene for p8 is expressed in the developing pituitary gland but is silent in the adult (18).
Although normally expressed only in embryonic tissues, HMGA gene expression has been correlated with a number of different types of cancers. In various cancer subtypes, studies have found both overexpression of full-length HMGA proteins as well as chromosomal rearrangements involving the formation of chimeric proteins (reviewed in Ref. 16). In addition to the correlation between HMGA proteins and cancer, functional studies have shown that overexpression of HMGA in nontumorigenic mammary cells results in tumors in nude mice (19). Further supporting the role of HMGA proteins in oncogenesis, transgenic mice overexpressing HMGA2 under the control of the cytomegalovirus promoter develop spontaneous tumors in various tissues. Most importantly, the majority of these tumors occurred in the pituitary gland (20).
Although many studies have implicated HMGA proteins in tumorigenesis, the role of p8, specifically, has yet to be fully elucidated. In fact, reports have implicated p8 in both promotion of growth as well as inhibition of cellular growth (14, 21) underscoring a paradoxical feature of the protein. Herein, we report the first association between expression of p8 and pituitary tumorigenesis both in vivo and in a cell culture model.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
) and a group of 53 genes that were decreased in tumors by up to 50% of baseline (Table 2).
|
|
HMGA proteins are typically expressed maximally in the developing embryo but are extinguished in adult tissues (reviewed in Ref. 16). The reexpression of this family of nuclear factors is associated with cells that have transitioned from quiescent to actively growing and proliferating (24). Previously, we have shown, by in situ hybridization, that p8 mRNA is transiently expressed in developing pituitary glands but becomes undetectable in adult pituitaries (18). Therefore, we sought to address whether reexpression of p8 in LHCTP pituitaries correlates with development of pituitary adenomas.
p8 Expression Is Increased in Pituitary Tumors
Microarray analysis revealed an average 12.3-fold increase in expression of an EST corresponding to p8 in tumors compared with wild-type pituitaries (Table 1
). The range of increase was between 11.8- and 12.6-fold (Fig. 1A). We confirmed the increase in p8 expression by semiquantitative RT-PCRs. Total RNA from pituitary glands of LHCTP tumors and age-matched wild-type littermates was used in duplex RT-PCR using p8 specific primers and primers for 18S rRNA (Fig. 1B). We found a 12-fold increase in p8 expression in pituitary adenomas in LHCTP mice when compared with pituitaries from wild-type littermates (Fig. 1C), which is remarkably consistent with the change observed on the microarray. Notably, these tumors and wild-type samples were distinct from those used in the initial microarray studies. Together, these data suggest that increases in p8 expression consistently correlate with the presence of pituitary adenomas in the LHCTP mouse model of LH hypersecretion.
|
To determine which tumor subtypes express p8, we performed in situ hybridization studies with pituitary sections from mice bearing adenomas and from wild-type littermates. As expected from previous studies (18), p8 expression was undetectable in pituitary sections from adult wild-type mice whereas PRL was robustly expressed (Fig. 2). Conversely, distinct foci of p8 are present in pituitary tumor sections from LHCTP transgenics (Fig. 2). In addition to p8, we used probes against PRL, GH, and TSH (data not shown) on adjacent sections to determine which cell type was present within p8-containing tumor foci. Thus far, all tumor foci that we have found contain p8 also produce PRL, but not all PRL-containing foci express p8 (Table 3). These data suggest that reexpression of p8 may be one of many potential insults that may lead to transformation of lactotropes.
|
|
, A and B). Using this approach, we generated lines with decreases in p8 expression ranging from 45% of control (GHM206) to approximately 61% of control (GHM222). Thus, we have attenuated expression of p8 in GH3 cells, providing us with a tool to determine the functional consequences of changing p8 expression in a cell line representing the most affected cell type in LHCTP pituitary glands.
|
). Mice injected with empty expression vector (GHM103)-transfected cells developed palpable tumors beginning at 27 d. In contrast, mice injected with clonal lines with reduced expression of p8 did not form tumors as quickly as mice injected with cells containing the empty expression vector. In addition, these had a smaller final volume (Fig. 4B). Notably, we also observed a positive correlation between the relative reduction of p8 expression and extent of tumorigenicity. For example, nude mice injected with the line with the greatest reduction in p8 expression (GHM206) did not develop tumors at all (Fig. 4, A and B). This contrasts with GHM275, which had slightly higher p8 expression, and exhibited attenuated tumorigenicity. Further, GHM222, a line with the least reduction in p8 expression, developed tumors at a rate similar to mice injected with GHM103 controls. Thus, the rank order of p8 expression in these cells (GHM103>222>275>206) correlates positively with extent of tumorigenicity. Together, these data suggest that maintenance of the GH3 transformed phenotype displays a concentration dependence on p8 mRNA.
|
). Similar to the results observed for GHM206 with a 42% reduction in p8 expression, the LßT2 cells with no detectable p8 expression were no longer tumorigenic. These studies suggest that p8 expression in both somatolactotrope and gonadotrope-derived cell lines is critical for maintaining the transformed phenotype of these cells and that the loss of p8 expression can reverse the tumorigenicity of both cell types.
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
p8 expression is increased in every pituitary adenoma analyzed in LHCTP females (Fig. 1). Additionally, p8 selectively colocalizes with PRL-containing foci in our model (Fig. 2). We have also shown that the presence of p8 is critical for maintaining the transformed phenotype of cell lines derived from both somatolactotropes (GH3) and gonadotropes (LßT2) (Figs. 4 and 5).
Although our studies suggest that p8 is critical for maintaining the tumorigenicity of somatolactotrope and gonadotrope cell lines, we suspect that alterations in p8 expression may be one of several distal events involved in cellular transformation. For example, LHCTP mice develop pituitary hyperplasia before the presence of adenomas (11), yet we have not been able to document a concomitant increase in p8 expression before the appearance of tumors (data not shown). Additionally, in situ hybridization studies demonstrate that p8 is expressed in only 21% of PRL-positive tumor foci in vivo (Table 3). It is tempting to use this data to suggest that whereas p8 reexpression may be implicated in the pathogenesis of some prolactinomas, other mediators of adenoma development are likely involved as well. Alternatively, it is also possible that our assessment of p8 expression by in situ hybridization captures a limited time point in the transformation of lactotropes where each PRL-positive focus may have already expressed p8 or would have gone on to express p8 based on its particular stage of transformation. Thus, all PRL-positive foci in LHCTP tumors may, at discrete times, express p8 mRNA. Clearly, additional studies will be required to distinguish between these possibilities.
p8 has been shown to be involved in transformation and proliferation of cell types other than pituitary cells. In a study by Iovanna and colleagues (30), p8 was necessary for transformation of mouse embryonic fibroblasts (MEFs) by E1A and ras. p8 has also been implicated in pancreatic carcinoma (31, 32) and thyroid neoplasms (33). Although p8 also affects proliferation, this may be cell specific. For example, p8 induces proliferation in COS-7 cells (14), whereas MEFs isolated from mice with a targeted disruption in p8 grow at a more rapid rate and are resistant to apoptosis (21).
Our data further underscore the oncogenic connection with p8 by demonstrating, for the first time, a link between p8 and maintenance of pituitary cell tumorigenicity. Because p8 is not increased in hyperproliferative pituitaries as well as tumors, we suggest that p8 expression is important in pituitary tumorigenesis through processes that may be distinct from those that simply provide an increased mitotic drive. p8 and other HMGA proteins have been implicated both in proliferation and differentiation (15). Although it remains unclear how p8 maintains a transformed phenotype in pituitary cells and whether it functions to simply alter proliferation or through other mechanisms such as altering the efficiency of DNA repair, both the LHCTP transgenic model and the pituitary-derived stable cell lines developed here will allow us to explore these questions further.
In addition to its proposed roles in proliferation and tumorigenesis, p8 also plays discrete roles during embyrogenesis. Recently, we reported (18) that p8 appears to be a stage-specific component of the cellular proteome, playing a functional role in gonadotrope differentiation. Specifically, these studies suggest that p8 may play a critical role in establishing the temporal pattern of expression of the LHß gene during embryogenesis (18). This study also underscores the notion that p8 has multiple functions (differentiation, proliferation, apoptosis, or transformation) that are probably dependent on the specific cellular environment. Most importantly, because p8 is normally only expressed during embryogenesis in the pituitary gland, our data provide further evidence that its reexpression in other pituitary cell types during adulthood is linked to a pathogenic outcome.
Although multiple functions are associated with p8, its specific targets have not yet been elucidated. It has been shown previously that the presence of p8 is necessary for the growth-promoting effects of TGFß in MEFs and that p8 expression enhances SMAD (similar to mothers against decapentaplegic) activity (34). On the other hand, MEFs from p8-knockout mice have lower levels of p27, suggesting p8 may regulate cell growth in part through this cyclin-dependent kinase inhibitor (21). p8 also has the ability to bind DNA (12). Thus, although p8 may bind promoters of genes that promote tumorigenesis, it is equally possible that p8 acts through a cohort of protein binding partners such as p300 (35). Another HMGA family member has been shown to act by enhancing the binding of serum response factor to DNA and serum response factor-dependent transcription (36). Therefore, it is also likely that p8 may serve as a scaffold for other proteins and alter expression of target genes in a DNA-binding independent manner. Identifying the molecular targets of p8 will be an important avenue for determining its mechanism of action in the discrete cell types of the pituitary.
In summary, we have identified a new HMGA protein (p8) that may be functionally linked to formation of pituitary prolactinomas by using gene expression profiling of pituitary adenomas from LHCTP mice. Our in situ hybridization studies clearly indicate that p8 is localized to prolactinomas, and we have defined a critical role for p8 in maintaining the tumorigenicity of both somatolactotrope and gonadotrope cell lines. Together, these data suggest that p8 is necessary for maintaining the transformed status of multiple pituitary cell types. These data also further solidify a link between HMGA proteins and the genesis of pituitary tumors.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
Nude/athymic (NCR nu/nu) mice were maintained in the CWRU Athymic Animal Facility. Four-week old females were used in all studies. Athymic animals were housed in microisolator units with a 12-h light, 12-h dark schedule, constant temperature, and given food ad libitum. Mice were injected with 106 cells of each line. Cells were injected sc in PBS in both side flanks of each mouse; five mice in each group were injected, with the exception of lines 103 and 206. These two lines were injected in both side flanks of five mice in two separate experiments, for a total of 10 mice in each of these groups. Tumors appeared at early as 27 d, and measurements were taken through 42 d post injection. Tumor volumes were determined using the formula described previously (37): V = A X B2 x 0.4 where A and B represent the larger and smaller axes of the tumor, respectively.
RNA Isolation
RNA was isolated using TRizol according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA).
Affymetrix GeneChip Analysis
The MIAME checklist containing all information is included as supplemental Fig. 1 published as supplemental data on The Endocrine Society’s Journals Online web site at http://mend.endojournals.org.
Semiquantitative RT-PCR
Total RNA isolated from pituitary glands of five transgenic females at 9–12 months of age and six age-matched wild-type littermates was used in individual reverse transcription reactions to generate single-stranded cDNA. Briefly, 1 µg total RNA, 20 pmol of random hexamer primers (Invitrogen), final concentrations of 1x first-strand buffer (Invitrogen), 0.01M dithiothreitol, 0.5 mM deoxynucleotide triphosphates (Invitrogen), 1 µl RNase Out (Invitrogen), and 1 µl of Superscript II (Invitrogen) in a final volume of 20 µl were incubated for 1 h at 50 C. The reaction was stopped by incubating at 94 C for 5 min and then diluted to 100 µl with diethylpyrocarbonate-treated water for use in PCR.
Single-strand cDNA was used in a duplex PCR with p8 primers (forward: 5'-ATGGCCACCTTGCCACCA-3' and reverse: 5'-TCAGCGCCAGGCTTTTTTCCT-3') and Quantum RNA 18S Classic Primers (Ambion, Inc., Austin, TX) as an internal control. Initially, the optimal dilution of 18S primer to competimer (3:7) was determined according to the manufacturer’s instructions (Ambion), using annealing temperature of 55 C, and [
32P]dCTP detection. PCRs were performed in a volume of 50 µl containing 5 µl of cDNA template from the reverse transcription reaction, a final concentration of 1x PCR buffer (Invitrogen), 0.2 mM deoxynucleotide triphosphates (Invitrogen), 2.5 mM MgCl2, 4 µl of 18S primers-competimers (3:7,Ambion), 20 pmol of forward p8 primer, 20 pmol of reverse p8 primer, 0.5 µl Taq Polymerase (Invitrogen), and 0.5 µl [32P]dCTP (Perkin-Elmer Corp., Norwalk, CT). Using the manufacturer’s guidelines, cycle 25 was determined to be within the linear range for each primer pair. This experiment was repeated and all subsequent PCRs used 25 cycles. PAGE and autoradiography were used to visualize the PCR products, which were then quantified using the Ambis System of radioanalysis as previously described (38). Each sample was assayed in at least two separate experiments.
Plasmids
p8 cDNA was isolated by PCR from mouse pituitary-derived cDNA (described above). Forward: 5'-CACAGGCAAGACTTTGGAGAG-3' and reverse: 5'-AGGTGGGAAAGAATCAAGCA-3' primers for p8 were used to generate a 446-bp product containing the entire open reading frame. The PCR product was subsequently cloned in the reverse (p8AS) orientation into the EcoRI site of pcDNA3. The clone was sequenced to verify accuracy of the PCR.
In Situ Hybridization
In situ hybridization was performed as described previously with minor modifications (39). Briefly, 15-µm sections were warmed to 42 C for 20 min, fixed in 4% paraformaldehyde/PBS at 37 C, and washed in PBS. Sections were acetylated using a 0.1 M triethanolamine/0.25% acetic anhydride mixture and incubated with hybridization solution minus the probe [50% formamide, 5x standard sodium citrate (SSC), 2% Boehringer blocking powder, 0.1% Triton X-100, 0.5% 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonate, 1 mg/ml yeast RNA, 5 mM EDTA, 50 mg/ml heparin in diethylpyrocarbonate-treated water] at 55 C. Riboprobe was diluted in hybridization solution and hybridized overnight at 55 C in a chamber humidified with 5x SSC. The next morning, sections were washed in a 50% formamide/0.5x SSC mixture at 55 C followed by a wash in 0.5x SSC at room temperature and blocked with the following solution in a chamber humidified with water: 10% heat-inactivated sheep serum; 2% BSA; 0.02% sodium azide in 50 mM Tris-Cl (pH 7.5), 100 mM NaCl, 0.1% Triton X-100. Antidigoxigenin Fab fragments (Roche Molecular Biochemicals, Indianapolis, IN) were diluted 1:1000 in blocking buffer and incubated on sections for 1 h at room temperature in a humidified chamber and washed in PBS. After equilibration in several washes of chromagen buffer [100 mM Tris-Cl (pH 9.5), 100 mM NaCl, 50 mM MgCl2], sections were developed overnight in the chromagen buffer with 4.5 µl/ml nitro blue tetrazolium chloride and 3.5 µl/ml 5-bromo-4-chloro-3-indolyl-phosphate added as substrate for alkaline phosphatase. Sections were rinsed with PBS, fixed in 4% paraformaldehyde/PBS, washed, and mounted. In situ hybridization was repeated at least two times per animal with n = 6 for wild type and n = 8 for transgenic mice.
Probes for in Situ Hybridization
Plasmids containing cDNA for murine TSHß and murine LHß were described previously (39). Rat PRL probe was a gift from Sally Camper’s laboratory, and p8 plasmid was described above. Plasmids were linearized to generate sense (S) and antisense (AS) riboprobes: probe sizes were: 233 bp, LHß; 496 bp, TSHß; 735 bp, PRL; and 446 bp, p8. GH cDNA was obtained from mouse pituitary tissue using RT-PCR with the following primers: forward, 5'-TGGCCTCAGGAGGCTAGTG-3'; and reverse, 5'-ATCCTGCTGAGGAACTGCAC-3'. The PCR product was cloned into pCRII-TOPO vector using the TOPO-TA system (Invitrogen). Digoxigenin-labeled riboprobe was generated by in vitro transcription using 10x DIG RNA labeling mix (Roche Molecular Biochemicals) and purified using the RNeasy Mini Kit (QIAGEN, Valencia, CA).
Generation of Stable Cell Lines
LßT2 stably transfected lines were maintained as previously described (18). GH3 cells were maintained in high-glucose DMEM containing 2 mM L-glutamine, supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. To generate stable cell lines, 2 x 105 GH3 cells in 35-mm culture dishes were transfected overnight with 2 µg of pcDNA3 or p8AS-pcDNA3, using 10 µl of LipofectAMINE (Invitrogen) in serum and DMEM without antibiotic. Transfection medium was removed, and cells were returned to antibiotic-free growth medium the next day. Thus, 48 h after transfection, cells were passaged to 100-mm culture dishes and maintained in selection media containing 1 mg/ml G418 for 4 wk. Clonal lines were generated by using sterile cloning disks soaked in trypsin to transfer individual clones to 15-mm dishes, which were further propagated. Cell lines were always maintained in selection medium.
Northern Blot Analysis
For all Northern blots, 15 µg of total RNA were separated by electrophoresis in a 1% 3[N-morpholino]propanesulfonic acid/formaldehyde gel. RNA was transferred to a nylon membrane (Hybond-N+, Amersham Biosciences, Arlington Heights, IL). Prehybridization, hybridization, and washes were carried out in Quickhyb (Stratagene, La Jolla, CA) according to the manufacturer’s instructions. Random labeling of probes using 32P-dCTP (PerkinElmer) was carried out using DECAprime II (Ambion) according to the manufacturer’s protocol. The murine p8 probe consisted of the 446-bp fragment described above, and the ß-actin probe represented 600 bp. Northern blots were repeated at least three times with at least two separate RNA samples for each cell line. Phosphor imager (Packard Cyclone, Boston, MA) analysis was used to visualize and quantify the bands (Optiquant).
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
This work was supported by National Institutes of Health Grants DK28859 and CA86387 (to J.H.N.).
Abbreviations: EST, Expressed sequence tag; HMG, high-mobility group; LHCTP, LH overexpressing mice; MEF, mouse embryonic fibroblast; PRL, prolactin; SSC, standard sodium citrate.
Received for publication April 19, 2004. Accepted for publication July 1, 2004.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
|---|
20/+mice are predisposed to tumors of the pituitary gland with a nearly complete penetrance. Oncogene 9:1021–1027[Medline]
subunit gene. Emergence of a new element. J Biol Chem 269:19360–19368This article has been cited by other articles:
 |
|
 |
|
  C. M. Million Passe, C. R. White, M. W. King, P. L. Quirk, J. L. Iovanna, and C. C. Quirk Loss of the Protein NUPR1 (p8) Leads to Delayed LHB Expression, Delayed Ovarian Maturation, and Testicular Development of a Sertoli-Cell-Only Syndrome-Like Phenotype in Mice Biol Reprod, October 1, 2008; 79(4): 598 - 607. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Feng, M. A. Lawson, and P. Melamed A Proteomic Comparison of Immature and Mature Mouse Gonadotrophs Reveals Novel Differentially Expressed Nuclear Proteins that Regulate Gonadotropin Gene Transcription and RNA Splicing Biol Reprod, September 1, 2008; 79(3): 546 - 561. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Fang, M. Menon, W. Kapelle, O. Bogacheva, O. Bogachev, E. Houde, S. Browne, P. Sathyanarayana, and D. M. Wojchowski EPO modulation of cell-cycle regulatory genes, and cell division, in primary bone marrow erythroblasts Blood, October 1, 2007; 110(7): 2361 - 2370. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Path, A. Opel, M. Gehlen, V. Rothhammer, X. Niu, C. Limbert, L. Romfeld, S. Hugl, A. Knoll, M. D. Brendel, et al. Glucose-dependent expansion of pancreatic beta-cells by the protein p8 in vitro and in vivo Am J Physiol Endocrinol Metab, December 1, 2006; 291(6): E1168 - E1176. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W E Farrell Pituitary tumours: findings from whole genome analyses. Endocr. Relat. Cancer, September 1, 2006; 13(3): 707 - 716. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Hernandez-Gonzalez, I. Gonzalez-Robayna, M. Shimada, C. M. Wayne, S. A. Ochsner, L. White, and J. S. Richards Gene Expression Profiles of Cumulus Cell Oocyte Complexes during Ovulation Reveal Cumulus Cells Express Neuronal and Immune-Related Genes: Does this Expand Their Role in the Ovulation Process? Mol. Endocrinol., June 1, 2006; 20(6): 1300 - 1321. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. G Morris, M. Musat, S. Czirjak, Z. Hanzely, D. M Lillington, M. Korbonits, and A. B Grossman Differential gene expression in pituitary adenomas by oligonucleotide array analysis Eur. J. Endocrinol., July 1, 2005; 153(1): 143 - 151. [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 |
