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Endocrinology Vol. 139, No. 3 1300-1305
Copyright © 1998 by The Endocrine Society

ARTICLES

Triiodothyronine Modulates Interleukin-6 Synthesis in Osteoblasts: Inhibitions in Protein Kinase A and C Pathways1

Haruhiko Tokuda, Osamu Kozawa, Atsushi Harada, Ken-ichi Isobe and Toshihiko Uematsu

Department of Internal Medicine (H.T.) and Department of Orthopedics (A.H.), Chubu National Hospital, National Institute for Longevity Sciences, Obu, Aichi 474; Department of Pharmacology, Gifu University School of Medicine (O.K., T.U.), Gifu 500; and Department of Basic Gerontology, National Institute for Longevity Sciences (K.I.), Obu, Aichi 474, Japan

Address all correspondence and requests for reprints to: Haruhiko Tokuda, Department of Internal Medicine, Chubu National Hospital, National Institute for Longevity Sciences, Obu, Aichi 474, Japan.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
In osteoblast-like MC3T3-E1 cells, we recently reported that PGE1 and PGF2{alpha} induce interleukin (IL)-6 synthesis via activation of protein kinase A and protein kinase C, respectively. Moreover, in the case of IL-1-induced IL-6 synthesis in these cells, we showed that protein kinase C activation by IL-1 limits the IL-6 synthesis. In the present study, we investigated the effect of T3 on IL-6 synthesis induced by these agonists in MC3T3-E1 cells. T3, which by itself had little effect on IL-6 synthesis, significantly reduced the IL-6 synthesis induced by PGE1 in a dose-dependent manner in the range between 10 pM and 10 nM. T3 also reduced PGE1-induced activation of protein kinase A. T3 inhibited the IL-6 synthesis induced by cholera toxin, an activator of Gs, or forskolin, which directly activates adenylate cyclase. However, T3 did not affect (Bu)2cAMP-induced IL-6 synthesis. In addition, T3 reduced PGF2{alpha}-induced IL-6 synthesis dose dependently in the range between 10 pM and 10 nM. T3 also inhibited IL-6 synthesis induced by 12-O-tetradecanoylphorbol-13-acetate, an activator of protein kinase C. On the other hand, T3 markedly enhanced IL-1-induced IL-6 synthesis. This enhancement by T3 was potentiated in protein kinase C down-regulated cells. T3 hardly affected the protein kinase C activation induced by PGF2{alpha} or IL-1. These results strongly suggest that T3 modulates IL-6 synthesis at two points in osteoblasts as follows; one is exerted at the point between adenylate cyclase and protein kinase A, and the other is at a point downstream from protein kinase C activation.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
IT IS well known that hyperthyroidism is a major cause of secondary osteoporosis (1). The receptor of thyroid hormone belongs to steroid hormone receptor superfamily (2). It is generally accepted that the effects of thyroid hormone, as well as other steroid hormones, are exerted through binding to its specific intracellular receptors and subsequently activating the expression of the gene network (2). Bone metabolism is maintained by two types of functional cells, osteoblasts and osteoclasts, which are responsible for bone formation and bone resorption, respectively (3). Accumulating evidence indicates that osteoblasts, rather than osteoclasts, possess receptors for many bone resorptive agents (3), suggesting that osteoblasts also play an important role in the regulation of bone resorption. In hyperthyroidism, the serum levels of osteocalcin and alkaline phosphatase, markers of mature osteoblast phenotype (4, 5), and the excretion of pyridinoline and hydroxypyridinoline cross-link, which reflects bone resorption, are elevated (5). It is clinically recognized that both bone formation and resorption are increased in hyperthyroidism, and the imbalance results in the bone loss. By histomorphometric analysis, it has been shown that osteoclast numbers and resorbing surfaces with loss of trabecular bone are increased (5). T3 receptor is recognized in osteoblasts including osteoblast-like MC3T3-E1 cells (6, 7). It has been reported that thyroid hormone increases alkaline phosphatase activity and secretion of osteocalcin, insulin-like growth factors, and insulin-like growth factor-binding proteins in osteoblasts, and that it modulates proliferation of osteoblasts (5, 6, 7). Therefore, it is probable that the effects of thyroid hormone on bone metabolism are mainly attributable to the modulation of osteoblast functions.

It is well known that interleukin (IL)-6 is a pleiotropic multifunctional cytokine that regulates diverse cell functions such as promotion of B cell differentiation and T cell activation and induction of acute-phase proteins (8, 9). It is recognized that IL-6 is produced and secreted in a variety of cells. As for bone metabolism, it has been reported that IL-6 stimulates bone resorption and induces osteoclast formation (10, 11). Bone resorptive agents such as IL-1, PTH, tumor necrosis factor-{alpha}, and platelet-derived growth factor have been reported to stimulate IL-6 production and its secretion in cultured osteoblasts (10, 12, 13, 14). Therefore, accumulating evidence suggests that IL-6 secreted from osteoblasts is an important downstream effector of bone resorptive agents. PGs are well recognized as important modulators of bone metabolism (3). We reported that PGE1 stimulates IL-6 synthesis via activation of protein kinase A in osteoblast-like MC3T3-E1 cells (15). In addition, we recently showed that PGF2{alpha} induces IL-6 synthesis through protein kinase C, which is activated through both phosphoinositide hydrolysis by phospholipase C and phosphatidylcholine hydrolysis by phospholipase D in these cells (16). As for T3 effect on IL-6 synthesis in bone cells, it has only been reported that T3 potentiates the stimulatory effect of IL-1 on IL-6 production in fetal rat limb bone cultures (17). However, the detailed mechanism of the effect of T3 on IL-6 synthesis has not yet been clarified.

In the present study, we investigated the effect of T3 on IL-6 synthesis induced by several agonists in osteoblast-like MC3T3-E1 cells. Herein, we show that T3 modulates IL-6 synthesis at two points in osteoblasts as follows: one is exerted at the point between adenylate cyclase and protein kinase A, and the other is at a point downstream of protein kinase C activation.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Materials
T3, PGE1, PGF2{alpha}, cholera toxin, forskolin, (Bu)2cAMP, and 12-O-tetradecanoylphorbol-13-acetate (TPA) were purchased from Sigma Chemical Co. (St. Louis, MO). IL-1 was purchased from Funakoshi Pharmaceutical Co. (Tokyo, Japan). Mouse IL-6 enzyme immunoassay (EIA) kit, an assay system for protein kinase C and [{gamma}-32P]ATP (3000 Ci/mmol) was purchased from Amersham Japan (Tokyo, Japan). An assay system for protein kinase A was purchased from Promega Corp. (Madison, WI). Other materials and chemicals were obtained from commercial sources. T3 was dissolved in 0.1 M NaOH. PGE1, PGF2{alpha}, and forskolin were dissolved in ethanol. TPA was dissolved in dimethyl sulfoxide. The maximum concentration of ethanol or dimethyl sulfoxide was 0.1%, which did not affect the assay for IL-6.

Cell culture
Cloned osteoblast-like MC3T3-E1 cells were maintained as previously described (15). In brief, the cells (5 x 104) were seeded into 35-mm diameter dishes in 2 ml {alpha}-MEM containing 10% FCS. After 5 days, the medium was exchanged for 2 ml {alpha}-MEM containing 0.3% FCS. The cells were used for experiments after 24 h. When indicated, the cells were pretreated with 0.1 µM TPA for 24 h, as previously reported (18).

Assay for IL-6
The cultured cells were pretreated with T3 for 8 h, and then stimulated by PGE1, PGF2{alpha}, cholera toxin, forskolin, (Bu)2cAMP, TPA, or IL-1 in 1 ml {alpha}-MEM containing 0.3% FCS for 48 h. T3 was maintained in the culture medium during the treatment with agonists. The conditioned medium was collected, and IL-6 in the medium was measured by an IL-6 EIA kit.

Assay for protein kinase A activity
The cultured cells were pretreated with T3 for 8 h, and then stimulated by PGE1 at 37 C for 10 min in 1 ml of an assay buffer (5 mM HEPES, pH 7.4, 150 mM NaCl, 5 mM KCl, 5.5 mM glucose, 0.8 mM MgSO4 and 1 mM CaCl2) containing 0.01% BSA. The reaction was terminated by aspirating the medium, and the protein kinase A activity was determined by a protein kinase A assay system.

Assay for protein kinase C activity
The cultured cells were pretreated with T3 for 8 h, and then stimulated by PGF2{alpha} or IL-1 at 37 C for 10 min in 1 ml of an assay buffer (5 mM HEPES, pH 7.4, 150 mM NaCl, 5 mM KCl, 5.5 mM glucose, 0.8 mM MgSO4 and 1 mM CaCl2) containing 0.01% BSA. The reaction was terminated by aspirating the medium, and the protein kinase C activity was determined by a protein kinase C assay system.

Determination
The absorbance of EIA samples was measured at 450 nm with EL 340 Bio Kinetic Reader (Bio-Tek Instruments, Inc., Winooski, VT). The radioactivity of 32P samples for protein kinase A and protein kinase C activity was determined with a Beckman LS-6500IC liquid scintillation spectrometer (Palo Alto, CA).

Statistical analysis
The data were analyzed by one-way ANOVA followed by Bonferroni method for multiple comparison between pairs, and P < 0.05 was considered significant. All data are presented as the mean ± SEM of triplicate determinations.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Effect of T3 on PGE1-induced IL-6 synthesis in MC3T3-E1 cells
We recently reported that PGE1-stimulated IL-6 synthesis is mediated by activation of protein kinase A in osteoblast-like MC3T3-E1 cells (15). We first examined the effect of T3 on the IL-6 synthesis induced by PGE1. T3 (10 nM), which by itself had little effect on IL-6 synthesis, significantly inhibited the PGE1-induced IL-6 synthesis (Fig. 1Go). The inhibitory effect of T3 was dose dependent in the range between 10 pM and 10 nM (Fig. 1Go). The maximum effect of T3 was observed at 10 nM, and the IL-6 synthesis was suppressed to about 10% of control (Fig. 1Go). The viability of the cells treated with 10 nM T3 for 48 h was more than 90%, as assessed by trypan blue staining.



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  		Figure 1. Effect of T3 on PGE1-induced IL-6 synthesis in MC3T3-E1 cells. Cultured cells were pretreated with various doses of T3 for 8 h and then stimulated by 10 µM PGE1 (•) or vehicle ({circ}) for 48 h. Each value represents mean ± SEM of triplicate independent cell preparations. Similar results were obtained with two additional and different cell preparations. *, P < 0.05 compared with value of PGE1 alone.

 
Effect of T3 on PGE1-induced protein kinase A activation in MC3T3-E1 cells
We confirmed that H-89, a specific protein kinase A inhibitor (19), markedly reduced PGE1-induced IL-6 synthesis (data not shown), as previously reported (15). So, we next examined the effect of T3 on PGE1-induced protein kinase A activation in these cells. PGE1 (10 µM) significantly activated protein kinase A (Table 1Go). T3 (10 nM), which by itself hardly affected protein kinase A activity, markedly reduced PGE1-induced activation of protein kinase A (Table 1Go).


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  		Table 1. Effect of T3 on PGE1-induced protein kinase A activation in MC3T3-E1 cells

 
Effects of T3 on cholera toxin-, forskolin-, or (Bu)2cAMP-induced IL-6 synthesis in MC3T3-E1 cells
To clarify whether or not the inhibitory effect of T3 on the PGE1-induced IL-6 synthesis is exerted at a point upstream from Gs or adenylate cyclase, we next examined the effect of T3 on cholera toxin- or forskolin-induced IL-6 synthesis. Cholera toxin is known to be an activator for Gs, a GTP-binding protein that mediates stimulative signals to adenylate cyclase (20). Forskolin is a direct activator of adenylate cyclase (21). T3 markedly reduced the IL-6 synthesis induced by cholera toxin (1 µg/ml) dose dependently in the range between 10 pM and 10 nM (Fig. 2AGo). The maximum effect was observed at 10 nM, and the cholera toxin-induced IL-6 synthesis was reduced to about 50% of control. Forskolin-induced IL-6 synthesis was also inhibited by T3 in a dose-dependent manner in the range between 10 pM and 10 nM, and the maximum effect of T3 was observed at 10 nM, a dose that caused a 60% reduction (Fig. 2BGo). However, T3 hardly affected (Bu)2cAMP-induced IL-6 synthesis up to 10 nM (Fig. 2CGo).



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  		Figure 2. Effects of T3 on cholera toxin-, forskolin-, and (Bu)2cAMP-induced IL-6 synthesis in MC3T3-E1 cells. Cultured cells were pretreated with various doses of T3 for 8 h and then stimulated by 1 µg/ml cholera toxin (•) or vehicle ({circ}) (A), 50 µM forskolin (•) or vehicle ({circ}) (B), or 3 mM (Bu)2cAMP (•) or vehicle ({circ}) (C) for 48 h. Each value represents mean ± SEM of triplicate independent cell preparations. Similar results were obtained with two additional and different cell preparations. *, P < 0.05, compared with value of cholera toxin alone (A). *, P < 0.05 compared with value of forskolin alone (B).

 
Effect of T3 on PGF2{alpha}-induced IL-6 synthesis in MC3T3-E1 cells
We recently demonstrated that PGF2{alpha} stimulates the synthesis of IL-6 through activation of protein kinase C in osteoblast-like MC3T3-E1 cells (16). Thus, we examined the effect of T3 on the PGF2{alpha}-induced IL-6 synthesis. T3 (10 nM) markedly suppressed the IL-6 synthesis induced by PGF2{alpha} (Fig. 3Go). The inhibitory effect of T3 was dose dependent in the range between 10 pM and 10 nM, and the maximum effect was observed at 10 nM (Fig. 3Go). The PGF2{alpha}-induced IL-6 synthesis was reduced to about 30% of control by 10 nM T3. We confirmed that calphostin C, a highly potent and specific inhibitor of protein kinase C (22), significantly inhibited PGF2{alpha}-induced IL-6 synthesis (data not shown), as previously described (16).



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  		Figure 3. Effect of T3 on PGF2{alpha}-induced IL-6 synthesis in MC3T3-E1 cells. Cultured cells were pretreated with various doses of T3 for 8 h and then stimulated by 10 µM PGF2{alpha} (•) or vehicle ({circ}) for 48 h. Each value represents mean ± SEM of triplicate independent cell preparations. Similar results were obtained with two additional and different cell preparations. *, P < 0.05 compared with value of PGF2{alpha} alone.

 
Effect of T3 on TPA-induced IL-6 synthesis in MC3T3-E1 cells
To clarify the point at which the inhibitory effect of T3 on PGF2{alpha}-induced IL-6 synthesis is exerted, we next examined the effect of T3 on the IL-6 synthesis induced by TPA, which is known to be a direct activator of protein kinase C (23). As previously reported (16), TPA (0.1 µM) alone significantly stimulated IL-6 synthesis via activation of protein kinase C in MC3T3-E1 cells. T3 significantly reduced the TPA-induced IL-6 synthesis in a dose-dependent manner in the range between 10 pM and 10 nM (Fig. 4Go). The maximum effect of T3 on the TPA-induced IL-6 synthesis was observed at 10 nM, and the synthesis was reduced to about 30% of control.



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  		Figure 4. Effect of T3 on TPA-induced IL-6 synthesis in MC3T3-E1 cells. Cultured cells were pretreated with various doses of T3 for 8 h and then stimulated by 0.1 µM TPA (•) or vehicle ({circ}) for 48 h. Each value represents mean ± SEM of triplicate independent cell preparations. Similar results were obtained with two additional and different cell preparations. *, P < 0.05 compared with value of TPA alone.

 
Effect of T3 on the IL-6 synthesis induced by IL-1 in MC3T3-E1 cells
It has been reported that T3 potentiates the stimulatory effect of IL-1 on IL-6 production in fetal rat bone cultures (17). Recently, we reported that IL-1 activates protein kinase C through phosphatidylcholine-specific phospholipase C in these cells, and the activation of protein kinase C limits IL-6 synthesis induced by IL-1 itself (24). Therefore, we further examined the effect of T3 on IL-1-induced IL-6 synthesis in osteoblast-like MC3T3-E1 cells. T3 significantly enhanced the IL-1-induced IL-6 synthesis (Fig. 5Go). The effect of T3 on the IL-1-induced IL-6 synthesis was dose dependent in the range between 10 pM and 10 nM (Fig. 5Go). We confirmed that calphostin C significantly enhanced IL-1-induced IL-6 synthesis (data not shown), as previously reported (24).



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  		Figure 5. Effect of T3 on IL-1-induced IL-6 synthesis in MC3T3-E1 cells. Cultured cells were pretreated with various doses of T3 for 8 h and then stimulated by 30 ng/ml IL-1 (•) or vehicle ({circ}) for 48 h. Each value represents mean ± SEM of triplicate independent cell preparations. Similar results were obtained with two additional and different cell preparations. *, P < 0.05 compared with value of IL-1 alone.

 
Effect of T3 on IL-1-induced IL-6 synthesis in protein kinase C down-regulated MC3T3-E1 cells
It has been shown that 24-h pretreatment of TPA (0.1 µM) down-regulates protein kinase C in MC3T3-E1 cells (18). We previously confirmed that the binding capacity of [3H]phorbol-12, 13-dibutylate, a protein kinase C-activating phorbol ester (23), in TPA-pretreated cells was reduced to approximately 30% of the capacity in intact cells (16). Thus, we examined the effect of down-regulation of protein kinase C on the enhancement by T3 of IL-1-induced IL-6 synthesis in these cells. As previously described (21), IL-1-induced IL-6 synthesis was significantly amplified in protein kinase C down-regulated cells (Table 2Go). The enhancement of IL-1-induced IL-6 synthesis by 10 nM T3 was markedly potentiated in protein kinase C down-regulated cells (Table 2Go).


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  		Table 2. Effect of protein kinase C down-regulation on T3-induced enhancement of IL-1-induced IL-6 synthesis in MC3T3-E1 cells

 
Effects of T3 on PGF2{alpha}- or IL-1-induced activation of protein kinase C in MC3T3-E1 cells
We further examined the effects of T3 on the activation of protein kinase C induced by PGF2{alpha} or IL-1 in these cells. PGF2{alpha} (10 µM) and IL-1 (30 ng/ml) truly activated protein kinase C (Table 3Go). T3 (10 nM), which by itself had no effect on protein kinase C activity, affected neither PGF2{alpha} nor IL-1-induced protein kinase C activation in these cells (Table 3Go).


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  		Table 3. Effect of T3 on PGF2{alpha}- or IL-1-induced protein kinase C activation in MC3T3-E1 cells

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
In the present study, we showed that PGE1-induced IL-6 synthesis was significantly inhibited by T3 in osteoblast-like MC3T3-E1 cells. We previously reported that PGE1 stimulates alkaline phosphatase activity via cAMP production without affecting phosphoinositide-hydrolyzing phospholipase C in these cells (25). Moreover, we recently demonstrated that PGE1 induces IL-6 synthesis via activation of protein kinase A in these cells (15). We demonstrated in this study the reduction by T3 of PGE1-induced protein kinase A activation, suggesting that the inhibitory effect of T3 on PGE1-induced IL-6 synthesis results from the decrease of protein kinase A activity. To clarify the point at which T3 inhibits the IL-6 synthesis induced by the activation of cAMP-mediating pathway, we first examined the effect of T3 on cholera toxin-induced IL-6 synthesis in these cells. It is well known that Gs functions as an intermediary in transmembrane signaling from the receptor to adenylate cyclase, and mediates stimulation of this enzyme (20). Cholera toxin ADP-ribosylates the {alpha}-subunit of Gs and elicits continuous activation of Gs, resulting in cAMP production (20). We showed that T3 inhibited the IL-6 synthesis stimulated by cholera toxin. Additionally, we demonstrated that the synthesis of IL-6 induced by forskolin, was also inhibited by T3. Thus, these findings suggest that the inhibitory effect of T3 is exerted at a point downstream from adenylate cyclase in osteoblast-like MC3T3-E1 cells. Furthermore, we examined the effect of T3 on the (Bu)2cAMP-induced IL-6 synthesis, and showed that T3 did no longer affect the IL-6 synthesis by (Bu)2cAMP. On the basis of our findings, it is most likely that the inhibitory effect of T3 on the PGE1-induced IL-6 synthesis is exerted at the point between adenylate cyclase and protein kinase A in osteoblast-like MC3T3-E1 cells.

We next examined the effect of T3 on PGF2{alpha}-induced IL-6 synthesis in MC3T3-E1 cells. T3 significantly inhibited the IL-6 synthesis induced by PGF2{alpha}. As for the signaling mechanism of PGF2{alpha}, we previously reported that PGF2{alpha} stimulates protein kinase C through both activation of phosphoinositide-hydrolyzing phospholipase C and phosphatidyl-choline-hydrolyzing phospholipase D in MC3T3-E1 cells (26, 27). In addition, we recently reported that PGF2{alpha} stimulates IL-6 synthesis via activation of protein kinase C in these cells (16). To clarify whether the inhibition by T3 of PGF2{alpha}-induced IL-6 synthesis is exerted at a point downstream from protein kinase C activation or not, we next examined the effect of T3 on the IL-6 synthesis induced by TPA. We demonstrated in this study that T3 also inhibited TPA-induced IL-6 synthesis without affecting PGF2{alpha}-induced activation of protein kinase C, suggesting that the inhibitory effect of T3 is exerted at a point downstream from protein kinase C in osteoblast-like MC3T3-E1 cells. This notion is also supported by the ineffectiveness of T3 on PGF2{alpha}-induced activation of protein kinase C. Taken together, it is probable that T3 inhibits IL-6 synthesis at two points as follows: one is exerted at the point between adenylate cyclase and protein kinase A, and the other is at a point downstream from protein kinase C. In addition, we showed that T3 did not suppress but potentiates the IL-1-induced IL-6 synthesis. This finding is consistent with an earlier observation that T3 enhances IL-1-induced IL-6 production in the cultures of fetal rat limb bone cultures (17). The precise mechanism of the induction of IL-6 production by IL-1 has not yet been clarified; however, we have recently reported that IL-1-induced activation of protein kinase C negatively regulates IL-6 synthesis stimulated by IL-1 itself in MC3T3-E1 cells (24). We also demonstrated in this study that the enhancement of IL-1-induced IL-6 synthesis by T3 was potentiated in protein kinase C down-regulated cells. Thus, it is probable that T3 still increases IL-1-stimulated IL-6 synthesis under the condition by which protein kinase C is down-regulated. As for the effect of protein kinase C in IL-6 synthesis in osteoblasts, directly activated protein kinase C has a stimulatory effect on IL-6 synthesis as previously reported (16). It has been reported that protein kinase C mediates IL-6 synthesis induced by PTH, platelet-derived growth factor, or PGF2{alpha} in osteoblasts (13, 14, 16). As a whole, it is probable that protein kinase C could regulate IL-6 synthesis both positively and negatively in osteoblasts, and that the regulatory fashion of protein kinase C might depend on the circumstances of intracellular signaling mechanism of each agonist. Taking into account of the ineffectiveness of T3 on IL-1-induced protein kinase C activation, our findings suggest that T3 affects IL-1-induced IL-6 synthesis at a point downstream from protein kinase C in osteoblasts, and relieves the inhibitory effect of this protein kinase. Currently, it is recognized that numerous signal transduction pathways other than protein kinase C, such as mitogen-activated protein kinase, nuclear factor for IL-6, nuclear factor-{kappa}B, etc. are involved in IL-1-induced IL-6 gene expression (28). So, the enhancement by T3 of IL-1-induced IL-6 synthesis shown in this study might also result from the modulation of one (or some) of them. Further investigation would be required to clarify the details. The potential effects of T3 on IL-6 synthesis induced by PGE1, PGF2{alpha} and IL-1 shown in this study are summarized in Fig. 6Go.



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  		Figure 6. Diagram of effects of T3 on IL-6 synthesis induced by PGE1, PGF2{alpha} and IL-1 in MC3T3-E1 cells. -, Negative regulation; AC, adenylate cyclase; PC-PLD, phosphatidylcholine-specific phospholipase D; PI-PLC, phosphatidylinositol-specific phospholipase C; PC-PLC, phosphatidylcholine-specific phospholipase C; PKA, protein kinase A; PKC, protein kinase C.

 
It is well known that T3 stimulates the bone resorption in bone organ cultures (5). However, it has been reported that T3 at higher doses can have an inhibitory effect on bone resorption (29), suggesting that the effect of T3 on bone resorption is biphasic. It is well recognized that IL-6 secreted from osteoblasts acts as an important downstream effector of bone resorbing agents (10, 12, 13, 14). As shown in this study, it is probable that, through the inhibitory effects on both protein kinase A and protein kinase C pathways, T3 could modulate IL-6 synthesis induced by various agonists in osteoblasts. Therefore, it is highly possible that T3 modulates IL-6 synthesis induced by ubiquitous bioactive substances such as autocoids, cytokines, and hormones through the mechanisms shown in this study, and affects bone metabolism. These effects of T3 on osteoblasts might contribute to the pathogenesis of osteopathy observed in patients with hyperthyroidism.

In conclusion, our results strongly suggest that T3 modulates IL-6 synthesis at two points in osteoblasts as follows: one is exerted at the point between adenylate cyclase and protein kinase A, and the other is at a point downstream of protein kinase C activation.


   Footnotes
 
1 This work was supported by a Grant-in-Aid for Scientific Research (09671041) for the Ministry of Education, Science and Culture of Japan, the Research Grant for Longevity Sciences (8A-2), and the Research Grant of Cooperative Studies for Longevity Sciences for National Sanatoria from the Ministry of Health and Welfare of Japan. Back

Received September 22, 1997.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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A. Shih, S. Zhang, H. J. Cao, H.-Y. Tang, F. B. Davis, P. J. Davis, and H.-Y. Lin
Disparate Effects of Thyroid Hormone on Actions of Epidermal Growth Factor and Transforming Growth Factor-{alpha} Are Mediated by 3',5'-Cyclic Adenosine 5'-Monophosphate-Dependent Protein Kinase II
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