Insulin-like growth factor I induces migration and invasion of human multiple myeloma cells

Multiple myeloma (MM) is an incurable form of cancer characterized by accumulation of malignant plasma cells in the bone marrow. During the course of this disease, tumor cells cross endothelial barriers and home to the bone marrow. In latter stages, myeloma cells extravasate through blood vessels and may seed a variety of organs. Insulin-like growth factor I (IGF-I) is one of several growth factors shown to promote the growth of MM cells. In the current study, we have assessed the ability of IGF-I to serve additionally as a chemotactic factor affecting the mobility and invasive properties of these cells. Results indicate that IGF-I promotes transmigration through vascular endothelial cells and bone marrow stromal cell lines. Analysis of endogenous signaling pathways revealed that protein kinase D/protein kinase Cmicro (PKD/PKCmicro) and RhoA were both activated in a phosphatidylinositol 3-kinase (PI-3K)-dependent manner. Inhibition of PI-3K, PKCs, or Rho-associated kinase by pharmacologic inhibitors abrogated migration, whereas mitogen-activated protein kinase (MAPK), Akt, and p70S6 kinase inhibitors had no effect. These results suggest that IGF-I promotes myeloma cell migration by activation of PI-3K/PKCmicro and PI-3K/RhoA pathways independent of Akt. The identification of IGF-I as both a proliferative and migratory factor provides a rational basis for the development of targeted therapeutic strategies directed at IGF-I in the treatment of MM.     

Growth factor-induced phosphoinositide 3-OH kinase/Akt phosphorylation in smooth muscle cells: induction of cell proliferation and inhibition of cell death

OBJECTIVE: The signaling pathways mediating proliferation and apoptosis in vascular smooth muscle cells (VSMC) are not well established. It has previously been shown that activation of the phosphoinositide 3-OH kinase (PI3K)/Akt pathway or the ERK 1/2 pathway can mediate anti-apoptotic function in different cell types. This study determined the specific contribution of the PI3K/Akt and ERK pathway in the regulation of apoptosis and proliferation of VSMC. METHODS AND RESULTS: Incubation of rat VSMC with FCS, insulin or IGF-1 time-dependently stimulated the phosphorylation of Akt, however FCS but not insulin or IGF-1 activated the MAP-kinase ERK 1/2. Moreover, insulin inhibited H(2)O(2)-induced apoptosis via the Akt pathway as demonstrated by pharmacological inhibition of the PI3K or overexpression of a dominant negative Akt mutant. In contrast, FCS inhibited H(2)O(2)-induced apoptosis via the Akt and also the ERK pathway. FCS, but not insulin or IGF-1 induced VSMC proliferation, suggesting that Akt activation is necessary but not sufficient for VSMC proliferation. FCS-induced proliferation of VSMC was only mediated via the Akt pathway and not the ERK pathway. CONCLUSIONS: These results define a link between cell proliferation and programmed cell death in VSMC via the same signal transduction pathway, namely activation of the serine/threonine kinase Akt, which may have significant implication for the development of vascular diseases or remodeling.     

Saw palmetto extract suppresses insulin-like growth factor-I signaling and induces stress-activated protein kinase/c-Jun N-terminal kinase phosphorylation in human prostate epithelial cells

A common alternative therapy for benign prostatic hyperplasia (BPH) is the extract from the fruit of saw palmetto (SPE). BPH is caused by nonmalignant growth of epithelial and stromal elements of the prostate. IGF action is important for prostate growth and development, and changes in the IGF system have been documented in BPH tissues. The main signaling pathways activated by the binding of IGF-I to the IGF-I receptor (IGF-IR) are the ERK arm of the MAPK cascade and the phosphoinositol-3-kinase (PI3K)/protein kinase B (PKB/Akt) cascade. We tested the hypothesis that SPE suppresses growth and induces apoptosis in the P69 prostate epithelial cell line by inhibiting IGF-I signaling. Treatment with 150 microg/ml SPE for 24 h decreased IGF-I-induced proliferation of P69 cells and induced cleavage of the enzyme poly(ADP-ribose)polymerase (PARP), an index of apoptosis. Treatment of serum-starved P69 cells with 150 microg/ml SPE for 6 h reduced IGF-I-induced phosphorylation of Akt (assessed by Western blot) and Akt activity (assessed by an Akt kinase assay). Western blot analysis showed that SPE reduced IGF-I-induced phosphorylation of the adapter protein insulin receptor substrate-1 and decreased downstream effects of Akt activation, including increased cyclin D1 levels and phosphorylation of glycogen synthase kinase-3 and p70(s6k). There was no effect on IGF-I-induced phosphorylation of MAPK, IGF-IR, or Shc. Treatment of starved cells with SPE alone induced phosphorylation the proapoptotic protein JNK. SPE treatment may relieve symptoms of BPH, in part, by inhibiting specific components of the IGF-I signaling pathway and inducing JNK activation, thus mediating antiproliferative and proapoptotic effects on prostate epithelia.     

Stimulation of insulin-like growth factor (IGF) binding protein-3 synthesis by IGF-I and transforming growth factor-alpha is mediated by both phosphatidylinositol-3 kinase and mitogen-activated protein kinase pathways in mammary epithelial cells

IGF binding protein (IGFBP)-3 is an important regulator of mammary epithelial cell (MEC) growth and can enhance the ability of both IGF-I and epidermal growth factor ligands such as TGFalpha to stimulate MEC proliferation. Here we investigate the role of the phosphatidylinositol-3 kinase (PI3K) and MAPK pathways in the regulation of IGFBP-3 expression by IGF-I and TGFalpha in bovine MECs. Both growth factors stimulated DNA synthesis, although IGF-I was the stronger mitogen. IGF-I and TGFalpha also stimulated IGFBP-3 mRNA and protein levels. TGFalpha stimulated rapid, transient activation of Akt that was maximal at 5 min and diminished by 15 min. In contrast, IGF-I-induced Akt activation was maximal between 15 and 90 min and was sustained for 6 h. Although ERK 1/2 was maximally stimulated by TGFalpha between 5 and 15 min, IGF-I did not stimulate discernible activation of ERK 1/2. In addition, TGFalpha but not IGF-I induced rapid phosphorylation of Shc, whereas only IGF-I activated insulin receptor substrate-1. Pretreatment with the PI3K inhibitor LY294002 or knockdown of p85 with small interfering RNA inhibited IGF-I or TGFalpha-stimulated IGFBP-3 expression. Similarly, MAPK kinase-1 inhibitors PD98059 and U0126 each abolished TGFalpha-stimulated increases in IGFBP-3 mRNA levels. In contrast to TGFalpha, IGF-I retained the ability to partially increase IGFBP-3 mRNA levels in the presence of MAPK kinase-1 inhibitors, indicating that IGF-I may activate alternative substrates of the PI3K pathway that are involved in IGFBP-3 regulation. In conclusion, stimulation of IGFBP-3 mRNA levels by mitogens is regulated through both the PI3K and MAPK pathways in bovine MECs.     

Molecular aspects of gefitinib antiproliferative and pro-apoptotic effects in PTEN-positive and PTEN-negative prostate cancer cell lines

To date, no effective therapeutic treatment allows abrogation of the progression of prostate cancer (PCa) to more invasive forms. One of the major targets for the therapy in PCa can be epidermal growth factor receptor (EGFR), which signals via the phosphoinositide 3'-kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK) pathways, among others. Despite multiple reports of overexpression in PCa, the reliance on activated EGFR and its downstream signalling to the PI3K and/or MAPK/extracellular signal-regulated kinase (ERK) pathways has not been fully elucidated. We reported that the EGFR-selective tyrosine kinase inhibitor gefitinib (ZD1839; Iressa) is able to induce growth inhibition, G(1) arrest and apoptosis in PCa cells and that its effectiveness is associated primarily with phosphatase and tensin homologue deleted from chromosome 10 (PTEN) expression (and thus Akt activity). In fact PTEN-negative PCa cells are slowly sensitive to gefitinib treatment, because this molecule is unable to downregulate PI3K/Akt activity. PI3K inhibition, by LY294002 or after PTEN transfection, restores EGFR-stimulated Akt signalling and sensitizes the cells to pro-apoptotic action of gefitinib. The MAPK pathway seems to be involved primarily on cell-growth modulation because dual blockade of EGFR and ERK1/2 phosphorylation potentiates growth inhibition (both not cell apoptosis) in PTEN-positive PCa cells and reduced EGF-mediated growth in PTEN-negative cells. Thus the effectiveness of gefitinib requires growth factor receptor-stimulated PI3K/Akt and MAPK signalling to be intact and functional. The loss of the PTEN activity leads to uncoupling of this signalling pathway, determining a partial gefitinib resistance. Moreover, gefitinib sensitivity may be maintained in these cells through its inhibitory potential in MAPK/ERK pathway activity, modulating proliferative EGFR-triggered events. Therefore, our data suggest that the inhibition of EGFR signalling can result in a significant growth reduction and in increased apoptosis in EGFR-overexpressing PCa cells with different modalities, which are regulated by PTEN status, and this may have relevance in the clinical setting of PCa.     

Involvement of protein kinase C and phosphatidylinositol 3-kinase pathways in the survival of B-cell chronic lymphocytic leukemia cells

B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of long-lived CD5(+) B lymphocytes. TPA (12-O-tetradecanoylphorbol 13- acetate) and interleukin-4 (IL-4) inhibit apoptosis of B-CLL lymphocytes ex vivo. We used specific inhibitors of protein kinase C (PKC), extracellular-regulated kinase (ERK), and phosphatidylinositol 3-kinase (PI3-kinase) to study their involvement in TPA- and IL-4-induced survival of B-CLL lymphocytes. BisI, a specific inhibitor of PKC, induced apoptosis and inhibited the antiapoptotic activity of TPA and IL-4. B-CLL cells have a basal PKC activity that was increased by TPA but not by IL-4. TPA, but not IL-4, induced ERK activation. However, the inhibition of ERK activation did not affect the viability of B-CLL lymphocytes, demonstrating that this pathway is not involved in their survival. Inhibition of PI3-kinase by LY294002 induced apoptosis of B-CLL cells and inhibited the survival effect of IL-4 and TPA. In addition, Akt, a downstream effector of PI3-kinase activity, was phosphorylated by TPA and IL-4 in B-CLL cells, though PI3-kinase had no effect on PKC-dependent phosphorylation of Akt. Furthermore, the inhibition of PKC or PI3-kinase increased dexamethasone- and fludarabine-induced apoptosis ex vivo in the presence of survival factors. These results demonstrate that PKC and PI3-kinase are involved in the survival of B-CLL cells and suggest that inhibitors of these pathways could be combined with the drugs used in the treatment of B-CLL.     

IGF-I promotes Schwann cell motility and survival via activation of Akt

We previously reported insulin-like growth factor-I (IGF-I) promotes Schwann cell (SC) motility and rescues SC from apoptosis induced by serum deprivation. This effect is mediated by phosphatidylinositol-3 (PI-3) kinase. In the current study, we examined the role of Akt, a downstream kinase of PI-3K, in SC motility and IGF-I mediated protection from apoptosis. IGF-I induces Akt phosphorylation at Ser473, an event which may be blocked by pretreatment with a PI-3K inhibitor, LY294002. In dominant negative K179M Akt (K179M) transfected SC, however, Akt is not activated in response to IGF-I. In addition, IGF-I is unable to promote SC motility and survival in K179M SC. These results suggest a critical role for Akt in IGF-I mediated motility and survival in SC.     

Hyperglycemia enhances adipogenic induction of lipid accumulation: involvement of extracellular signal-regulated protein kinase 1/2, phosphoinositide 3-kinase/Akt, and peroxisome proliferator-activated receptor gamma signaling

The molecular events of hyperglycemia-triggered increase in adipogenic induction of lipid accumulation remain unclear. We examined the effects of hyperglycemia on adipogenic induction of lipid accumulation and its involved signaling molecules, such as phosphoinositide 3-kinase (PI3K), ERKs, and peroxisome proliferator-activated receptor gamma (PPAR gamma). Bone marrow-derived mesenchymal stem cells (MSCs) isolated from FVB/N mice were capable of differentiating into adipocytes in adipogenic medium. The effects of high glucose (HG) (25.5 mm) were assessed in vitro by RT-PCR, ELISA, flow cytometry, immunostaining, and immunoblotting. The in vivo effect of hyperglycemia was further studied in streptozotocin (STZ)-induced diabetic FVB/N mice. Exposure of MSCs to HG enhanced adipogenic induction of lipid accumulation as compared with 5.5 mm glucose. HG increased PPAR gamma expression and PI3K activity and its downstream effector Akt phosphorylation during adipogenesis. Inhibition of PI3K/Akt activity with PI3K inhibitor LY294002 or by expressing the dominant negative p85 or Akt prevented the HG-enhanced PPAR gamma-dependent adipogenic induction of lipid accumulation. Moreover, HG increased the phosphorylation of ERK1/2 during adipogenesis. MAPK/ERK inhibitor PD98059 inhibited the PI3K activity, Akt phosphorylation, and lipid accumulation triggered by HG. PI3K inhibitor LY294002 did not affect the HG-increased ERK1/2 phosphorylation during adipogenesis. We next observed that adipogenic induction of lipid accumulation of MSCs isolated from STZ-induced diabetic mice is enhanced. Moreover, triglyceride, PPAR gamma expression, phosphorylated Akt and ERK1/2, and marrow fat in bones of STZ-diabetic mice were also increased. These results suggest that hyperglycemia enhances the adipogenic induction of lipid accumulation through an ERK1/2-activated PI3K/Akt-regulated PPAR gamma pathway.     

Lipid rafts are required for Kit survival and proliferation signals

In addition to its physiologic role as central regulator of the hematopoietic and reproductive systems, the Kit receptor tyrosine kinase (RTK) is pathologically overexpressed in some forms of leukemia and constitutively activated by oncogenic mutations in mast-cell proliferations and gastrointestinal stromal tumors. To gain insight into the general activation and signaling mechanisms of RTKs, we investigated the activation-dependent dynamic membrane distributions of wild-type and oncogenic forms of Kit in hematopoietic cells. Ligand-induced recruitment of wild-type Kit to lipid rafts after stimulation by Kit ligand (KL) and the constitutive localization of oncogenic Kit in lipid rafts are necessary for Kit-mediated proliferation and survival signals. KL-dependent and oncogenic Kit kinase activity resulted in recruitment of the regulatory phosphatidylinositol 3-kinase (PI3-K) subunit p85 to rafts where the catalytical PI3-K subunit p110 constitutively resides. Cholesterol depletion by methyl-beta-cyclodextrin prevented Kit-mediated activation of the PI3-K downstream target Akt and inhibited cellular proliferation by KL-activated or oncogenic Kit, including mutants resistant to the Kit inhibitor imatinib-mesylate. Our data are consistent with the notion that Kit recruitment to lipid rafts is required for efficient activation of the PI3-K/Akt pathway and Kit-mediated proliferation.     

IL-2- and IL-15-induced activation of the rapamycin-sensitive mTORC1 pathway in malignant CD4+ T lymphocytes

We examined functional status, activation mechanisms, and biologic role of the mTORC1 signaling pathway in malignant CD4(+) T cells derived from the cutaneous T-cell lymphoma (CTCL). Whereas the spontaneously growing CTCL-derived cell lines displayed persistent activation of the TORC1 as well as the PI3K/Akt and MEK/ERK pathways, the IL-2-dependent cell lines activated the pathways in response to IL-2 and IL-15 but not IL-21. Activation of mTORC1 and MEK/ERK was nutrient dependent. The mTORC1, PI3K/Akt, and MEK/ERK pathways could also be activated by IL-2 in the primary leukemic, mitogen-preactivated CTCL cells. mTORC1 activation was also detected in the CTCL tissues in the lymphoma stage-dependent manner with the highest percentage of positive cells present in the cases with a large cell transformation. Rapamycin inhibited mTORC1 signaling and suppressed CTCL cell proliferation but showed little effect on their apoptotic rate when used as a single agent. Activation of the mTORC1, PI3K/Akt, and MEK/ERK pathways was strictly dependent on the Jak3 and Jak1 kinases. Finally, mTORC1 activation was transduced preferentially through the PI3K/Akt pathway. These findings document the selective gammac-signaling cytokine-mediated activation of the mTORC1 pathway in the CTCL cells and suggest that the pathway represents a therapeutic target in CTCL and, possibly, other T-cell lymphomas.     

Involvement of Akt kinase in the action of STI571 on chronic myelogenous leukemia cells

To elucidate the role of mitogen-activated protein kinases (MAPKs) and Akt kinase in leukemogenesis caused by the breakpoint cluster region (BCR)-Abelson (ABL) tyrosine kinase oncoprotein, we examined the activities of MAPKs and Akt kinase and their roles in the action of STI571, a specific inhibitor of BCR-ABL tyrosine kinase, in chronic myelogenous leukemia (CML) cells. We found that extracellular signal-regulated kinase (ERK) 1/2 and Akt kinase are constitutively active in the chronic phase of CML, blast crisis of CML, and the CML-derived K562 cell line. Both interferon-alpha and STI571 suppressed ERK1/2 activity in K562 cells. In contrast, Akt kinase activity was inhibited only by STI571. K562 cell proliferation was markedly suppressed by LY294002, a specific inhibitor of PI3K/Akt kinase, and STI571 but not by PD98059, a specific inhibitor of MEK1/2. In addition, caspase-3 was activated by treatment of cells with STI571 and LY294002 but not with PD98059. These data indicate that Akt kinase may play a role in the proliferation of CML leukemia cells and the action of STI571. Primary leukemia cells from patients with CML blast crisis did not show inhibition of ERK1/2 or Akt kinase activity and were resistant to caspase-3-associated apoptosis after treatment with STI571. These findings suggest that STI571 does not effectively block signaling molecules downstream of the BCR-ABL tyrosine kinase in some cases of CML blast crisis.     

Combined functional and molecular analysis of tumor cell signaling defines 2 distinct myeloma subgroups: Akt-dependent and Akt-independent multiple myeloma

Although the phosphatidylinositide 3-kinase (PI3K)/Akt pathway has been reported to contribute to the malignant growth of multiple myeloma (MM), the true relevance of Akt kinases for this disease is still unclear. In particular, functional analyses in primary tumor cells and genetic target validation experiments are missing. Here, we used combined functional and molecular analyses to determine the importance of Akt activity in a large panel of primary MM samples and in MM cell lines. Akt down-regulation with isoform-specific siRNA constructs or with an Akt1/2-specific pharmacologic inhibitor strongly induced apoptosis in approximately half of the primary MM samples analyzed. Sensitivity to Akt inhibition strongly correlated with the activation status of Akt as determined by immunohistochemistry, phospho-Akt-specific flow cytometry, and Western analysis. Additional blockade of the MAPK and the IL-6R/STAT3 pathways was often not sufficient to decrease the viability of MM cells resilient to Akt inhibition. Taken together, these experiments led to the identification of 2 myeloma subgroups: Akt-dependent and Akt-independent MM.     

Overexpression of insulin-like growth factor binding protein-5 helps accelerate progression to androgen-independence in the human prostate LNCaP tumor model through activation of phosphatidylinositol 3'-kinase pathway

Although insulin-like growth factor (IGF) binding protein-5 (IGFBP-5) is highly up-regulated in normal and malignant prostate tissues after androgen withdrawal, its functional role in castration-induced apoptosis and androgen-independent progression remains undefined. To analyze the functional significance of IGFBP-5 overexpression in IGF-I-mediated mitogenesis and progression to androgen-independence, IGFBP-5-overexpressing human androgen-dependent LNCaP prostate cancer cells were generated by stable transfection. The growth rates of IGFBP-5-transfected LNCaP cells were significantly faster, compared with either the parental or vector-only transfected LNCaP cells in both the presence and absence ofdihydrotestosterone. IGFBP-5-induced increases in LNCaP cell proliferation occurs through both IGF-I-dependent and -independent pathways, with corresponding increases in the cyclin D1 messenger RNA expression and the fraction of cells in S + G2/M phases of the cell cycle. Changes in Akt/protein kinase B, a downstream component of phosphatidylinositol 3'-kinase (PI3K) pathway, in the LNCaP sublines also paralleled changes in their growth rates. Although treatment with a PI3K inhibitor induced apoptosis in both control and IGFBP-5-overexpressing LNCaP cells, this PI3K inhibitor-induced apoptosis was prevented by exogenous IGF-I treatment only in IGFBP-5 transfectants, suggesting that IGFBP-5 overexpression can potentiate the antiapoptotic effects of IGF-I. Furthermore, tumor growth and serum prostate-specific antigen levels increased several fold faster in mice bearing IGFBP-5-transfected LNCaP tumors after castration, despite having similar tumor incidence and tumor growth rates with controls when grown in intact mice before castration. Collectively, these data suggest that IGFBP-5 overexpression in prostate cancer cells after castration is an adaptive cell survival mechanism that helps potentiate the antiapoptotic and mitogenic effects of IGF-I, thereby accelerating progression to androgen independence through activation of the PI3K-Akt/ protein kinase B signaling pathway.     

Signal transduction pathways in androgen-dependent and -independent prostate cancer cell proliferation

In a previous report, we showed that increased activation of Akt, a downstream effector of phosphoinositide 3-kinase (PI3K) together with decreased activation of extracellular-signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase (MAPK) family, predicted poor clinical outcome in prostate cancer (Kreisberg et al. 2004 Cancer Research 64 5232-5236). We now show that Akt activation, but not ERK activation, is correlated with proliferation in human prostate tumors as estimated by the expression of the cell proliferation antigen Ki67. We verified these results in vitro, using the androgen-dependent prostate cancer cell line LNCaP and its androgen-independent clone C4-2 as models of prostate cancer of good and poor clinical outcome, respectively. C4-2 cells expressed higher Akt activation, lower ERK activation and increased proliferation compared with LNCaP cells, similar to cases of poor clinical outcome. The PI3K inhibitor LY294002, but not the MAPK/ERK kinase inhibitor PD98059, induced growth arrest in both cell lines. Transient transfection with constitutively active Akt increased proliferation while dominant negative Akt decreased it, thus showing that Akt plays an important role in prostate cancer proliferation. Akt regulates the expression and activation of the androgen receptor. Androgen receptor inhibition with Casodex induced growth arrest in LNCaP cells, but not in C4-2 cells. Another PI3K downstream effector, p70 S6 kinase, requires prior phosphorylation by mammalian target of rapamycin (mTOR) for complete activation. Activation of p70 S6 kinase was higher in C4-2 compared with LNCaP cells. Rapamycin, an mTOR inhibitor, had a growth-inhibitory effect in C4-2 cells, but not in LNCaP cells. Our data suggest a shift from a Casodex-sensitive proliferation pathway in LNCaP cells to a rapamycin-sensitive pathway in C4-2 cells.     

Dexamethasone induces apoptosis in proliferative chondrocytes through activation of caspases and suppression of the Akt-phosphatidylinositol 3'-kinase signaling pathway

Although glucocorticoids are known to induce apoptosis in chondrocytes, the mechanisms for this effect and the potential antiapoptotic role of IGF-I are unknown. To address this, we studied the effects of dexamethasone (Dexa) on apoptosis in the HCS-2/8 chondrocytic cell line. Dexa (25 microm) increased apoptosis (cell death ELISA) by 39% and 45% after 48 and 72 h, respectively (P < 0.01 and P < 0.05, respectively). IGF-I (100 ng/ml) decreased Dexa-induced apoptosis to levels similar to control cells. Apoptosis was associated with cleavage of poly-ADP-ribose polymerase (PARP) and alpha-fodrin and activation of caspases-8, -9, and -3 (Western), an effect that was counteracted when chondrocytes were cocultured with Dexa + IGF-I. Inhibitors for caspases-8, -9, and -3 (50 microm each) equally suppressed Dexa-induced apoptosis (P < 0.01). Time-response experiments showed that caspase-8 was activated earlier (at 12 h) than caspase-9 (at 36 h). We studied the phosphatidylinositol 3'-kinase (PI3K) pathway to further investigate the mechanisms of Dexa-induced apoptosis. Dexa decreased Akt phosphorylation by 93% (P < 0.001) without affecting total Akt and increased the p85alpha subunit 4-fold. The Akt inhibitor SH-6 (10 microm) increased apoptosis by 54% (P < 0.001). When combining Dexa with SH-6, apoptosis was not further increased, showing that Dexa-induced apoptosis is mediated through inhibition of the PI3K pathway. Addition of IGF-I to SH-6- or Dexa + SH-6-treated cells decreased apoptosis by 21.2% (P < 0.001) and 20.6% (P < 0.001), respectively. We conclude that Dexa-induced apoptosis is caspase dependent with an early activation of caspase-8. IGF-I can rescue chondrocytes from Dexa-induced apoptosis partially through the activation of other pathways than the PI3K signaling pathway. Based on our in vitro data, we speculate that in vivo treatment with glucocorticoids may diminish longitudinal growth by increasing apoptosis of proliferative growth plate chondrocytes.