C-jun Gene Expression Research Articles

Signal transduction in arteriosclerosis: mechanical stress-activated MAP kinases in vascular smooth muscle cells (review).

Vascular smooth muscle cell (SMC) proliferation is a key event in the development of (spontaneous) atherosclerosis, hypertension-related arteriosclerosis, angioplasty-induced restenosis and venous bypass graft arteriosclerosis. Many factors or environmental stimuli are believed to be responsible for SMC growth or hypertrophy in the vessel wall. How these environmental stimuli or signals applied onto the surface of SMCs are transduced into the cell nucleus resulting in quantitative and qualitative changes in gene expression in SMCs of arterial walls is largely unknown. Mitogen-activated protein (MAP) kinases are rapidly activated in cells stimulated with various extracellular signals by dual phosphorylation of tyrosine and threonine residues. They are thought to play a pivotal role in transmitting transmembrane signals required for cell growth and differentiation. Recent studies have focused on the signalling events in vascular tissues in vivo and in cultured SMCs in vitro. It has been demonstrated that acute hypertension and angioplasty rapidly induced MAP kinase activation in the arterial wall. Kinase activation is followed by an increase in c-fos and c-jun gene expression and enhanced transcription factor AP-1 DNA-binding activity. A similar MAP kinase activation can be mimicked in in vitro cultured SMCs stimulated by either shear stress or cyclic strain stretch, suggesting direct effects of mechanical force. Interestingly, physical forces rapidly resulted in phosphorylation of platelet-derived growth factor (PDGF) receptor, an activated state, in cultured SMCs. Thus, mechanical stresses may directly perturb the cell surface or alter receptor conformation, thereby initiating signalling pathways usually used by growth factors. These findings have significantly enhanced our knowledge concerning the pathogenesis of arteriosclerosis and provide a basis for therapeutic intervention on vascular diseases.

Chronic alcohol intake inhibits retinoic acid synthesis and up-regulates AP-1 (c-Jun and c-Fos) gene expression in rats

Chronic alcohol intake inhibits retinoic acid synthesis and up-regulates AP-1 (c-Jun and c-Fos) gene expression in rats

Retinoic acid inhibits induction of c-Jun protein by ultraviolet radiation that occurs subsequent to activation of mitogen-activated protein kinase pathways in human skin in vivo.

Human skin is exposed daily to solar ultraviolet (UV) radiation. UV induces the matrix metalloproteinases collagenase, 92-kD gelatinase, and stromelysin, which degrade skin connective tissue and may contribute to premature skin aging (photoaging). Pretreatment of skin with all-trans retinoic acid (tRA) inhibits UV induction of matrix metalloproteinases. We investigated upstream signal transduction pathways and the mechanism of tRA inhibition of UV induction of matrix metalloproteinases in human skin in vivo. Exposure of human skin in vivo to low doses of UV activated EGF receptors, the GTP-binding regulatory protein p21Ras, and stimulated mitogen-activated protein (MAP) kinases, extracellular signal-regulated kinase (ERK), c-Jun amino-terminal kinase (JNK), and p38. Both JNK and p38 phosphorylated, and thereby activated transcription factors c-Jun and activating transcription factor 2 (ATF-2), which bound to the c-Jun promoter and upregulated c-Jun gene expression. Elevated c-Jun, in association with constitutively expressed c-Fos, formed increased levels of transcription factor activator protein (AP) 1, which is required for transcription of matrix metalloproteinases. Pretreatment of human skin with tRA inhibited UV induction of c-Jun protein and, consequently, AP-1. c-Jun protein inhibition occurred via a posttranscriptional mechanism, since tRA did not inhibit UV induction of c-Jun mRNA. These data demonstrate, for the first time, activation of MAP kinase pathways in humans in vivo, and reveal a novel posttranscriptional mechanism by which tRA antagonizes UV activation of AP-1 by inhibiting c-Jun protein induction. Inhibition of c-Jun induction likely contributes to the previously reported prevention by tRA of UV induction of AP-1-regulated matrix-degrading metalloproteinases in human skin.

O-92 - Transforming growth factor-β stimulates c-jun gene expression through AP-1 binding site

O-92 - Transforming growth factor-β stimulates c-jun gene expression through AP-1 binding site

Posttranscriptional regulation of collagenase-1 gene expression in synoviocytes by adenosine receptor stimulation.

To characterize the transcriptional and posttranscriptional regulation of collagenase-1 by adenosine receptor stimulation in interleukin-1 (IL-1)-stimulated fibroblast-like synoviocytes (FLS). FLS were stimulated with IL-1 and either the nonselective adenosine agonist 5'-N-ethylcarboxamidoadenosine (NECA) or the adenylate cyclase activator forskolin. Electrophoretic mobility shift assays were performed to determine AP-1 and cAMP-responsive element binding protein (CREB) activation. Transcriptional activation was determined by transfecting HS68 dermal fibroblasts with a collagenase-chloramphenicol acetyltransferase construct. Finally, collagenase messenger RNA (mRNA) half-life was determined by activating cells in the presence of IL-1, IL-1 + NECA, or IL-1 + forskolin and culturing cells in the presence of actinomycin D. NECA and forskolin had no effect on AP-1 activation, c-jun or c-fos gene expression, or CREB phosphorylation. IL-1 markedly increased collagenase promoter activity, and neither NECA nor forskolin blocked this action. Studies of mRNA half-life showed that both NECA and forskolin decreased the half-life of collagenase mRNA in IL-1-stimulated FLS and HS68 cells. The findings of this study demonstrate that NECA and forskolin decrease collagenase gene expression in FLS and dermal fibroblasts due to enhanced mRNA degradation.

Induction in human osteoblastic cells (SaOS2) of the early response genes fos, jun, and myc by the amino terminal fragment (ATF) of urokinase.

Previous studies have demonstrated that overexpression of urinary plasminogen activator (uPA) in rat prostate cancer cells results in increased skeletal metastases, which are primarily of the osteoblastic variety. The osseous activation induced by the metastases appears to be mediated through the amino terminal fragment (ATF) of uPA, which lacks the catalytic domain and can act as a growth factor for osteoblasts. To explore further the mechanism of action of uPA in bone cells, we evaluated the effects of ATF on modulating the expression of various proto-oncogenes. Human-osteoblast-derived osteosarcoma cells, SaOS2, were treated with graded doses of ATF for 10-120 min, and effects on early response proto-oncogenes were monitored. ATF increased c-myc, c-jun, and c-fos gene expression in a time-dependent manner for up to 60 min, after which mRNA levels fell. The maximum induction was seen in c-fos gene expression, which was found to be dose dependent. This effect of ATF was localized to its growth-factorlike domain. Examination of the half life of these transcripts in the presence of the transcriptional inhibitor actinomycin D demonstrated that ATF does not alter the stability of c-fos mRNA in these bone cells. Nuclear run-off assays indicated that ATF effects were due to stimulation of c-fos gene transcription. An increase in c-fos protein levels was correlated with the augmentation of its mRNA in ATF-treated SaOS2 cells. Pretreatment of SaOS2 cells with the protein tyrosine kinase inhibitor herbimycin and recombinant soluble uPA receptor (uPAR) caused a significant reduction in the ability of ATF to induce c-fos expression. These results demonstrate a novel role for uPA in activating early response proto-oncogenes, in particular c-fos, which plays an important role in bone cell growth and differentiation and may be a key factor in the signal transduction pathway of ATF.

The protein kinase A pathway inhibits c-jun and c-fos protooncogene expression induced by the protein kinase C and tyrosine kinase pathways in cultured human thyroid follicles.

We have previously demonstrated antagonistic interactions between the major signal transduction pathways in human thyroid follicles: TSH acting via protein kinase A (PKA) attenuated phorbol ester [acting via protein kinase C (PKC)] as well as epidermal growth factor (EGF)-protein tyrosine kinase (PTK)-mediated cell proliferation, whereas the PKC and PTK pathways inhibited PKA-mediated cell differentiation. In view of the key role played by the protooncogenes c-jun and c-fos in the cascade of events leading to cell proliferation and differentiation, we examined whether the antagonism we observed between the pathways could be related to changes in the expression of these genes. The experimental model used was the same in vitro system as that used in the above study on cell growth and differentiation: thyroid follicles of human origin cultured in suspension under serum-free conditions. Both EGF (1-50 ng/mL) and the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA; 10(-11)-10(-7) mol/L) dose and time dependently stimulated c-jun and c-fos messenger ribonucleic acid (mRNA) expression. The c-jun and c-fos mRNA stimulation elicited by TPA was reduced by the PKC inhibitors, chelerythrine and staurosporine, and could not be mimicked by 4alpha-phorbol 12,13-didecanoate (a phorbol ester that fails to activate PKC), whereas the stimulation induced by EGF was diminished by the PTK inhibitor, genistein. This indicates a PKC- and PTK-mediated pathway triggered by TPA and EGF, respectively. TSH induced an increase in c-jun and c-fos mRNA, which, though significant, was small compared to that elicited by TPA or EGF. Addition of TSH (0.1-0.5 mU/mL), however, to either TPA or EGF dose dependently inhibited the c-jun and c-fos mRNA elicited by these agents. The repressive action of TSH on the effects of TPA and EGF mRNA were mimicked by forskolin and 8-bromo-cAMP, suggesting that the TSH inhibitory action is PKA mediated. The TSH inhibitory action seems to require de novo protein synthesis, as it was abrogated in the presence of cycloheximide. In conclusion, the present study provides novel data on c-jun and c-fos gene expression and their modulation by the major signal transduction pathways operating in human thyrocytes. Moreover, using the same serum-free system of human thyroid follicles cultured with the same agents and at the same doses as in our previous study on cell growth and differentiation, we found the TSH/PKA pathway to inhibit PKC- and EGF/tyrosine kinase-induced c-jun and c-fos mRNA, i.e. antagonistic effects parallel to those previously observed measuring cell proliferation. The findings suggest an association between human thyroid cell proliferation and c-jun and c-fos gene expression.

Regulation of c-jun gene expression in endothelial cells by the protein kinase inhibitor staurosporine.

The proto-oncogene c-jun, a member of the family of immediate-early genes, is transcriptionally induced in different cell types by a variety of stimuli, including mitogens, tumor promoters, growth factors. We show here that the protein kinase inhibitor staurosporine, which inhibits both the serine-threonine and tyrosine specific protein kinases, also causes differential regulation of the c-jun gene in endothelial cells. Increasing concentrations of staurosporine modulated the steady-state levels of c-jun mRNA in bovine aortic endothelial (BAE) cells in a multiphasic manner. The half-life of c-jun mRNA did not change significantly under these conditions, suggesting that the modulations in the mRNA levels were caused primarily by differential transcriptional activity of the gene. The expression of c-jun gene is believed to be regulated by its own product, the JUN protein, which constitutes a major component of the inducible transcription factor AP-1. In order to test whether the differential regulation of c-jun gene was caused by the differential activation (or inactivation) of the AP-1 transcription factor, the DNA-binding activity of this transcription factor in staurosporine-treated cells was measured. Gelshift analysis with a synthetic oligonucleotide probe showed modest effects of staurosporine on the DNA-binding activity of the transcription factor AP-1. The changes observed in the DNA-binding activity of AP-1 did not parallel the changes observed in the steady-state levels of c-jun mRNA. Similarly, the expression of an AP-1 dependent reporter gene construct was regulated in a fashion entirely different from the c-jun gene during the same protein kinase inhibitory conditions. These results suggest the existence of an alternative pathway that regulates the c-jun gene expression in endothelial cells independent of both the protein kinase and AP-1 transcription factor activation steps.

Inhibition of protein kinase C and proto-oncogene expression by crocetin in NIH/3T3 cells.

Crocetin, a carotenoid isolated from the seeds of Gardenia jasminoides, was found to be a potent inhibitor of tumor promotion induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in mouse skin. When mouse fibroblast NIH/3T3 cells were treated with TPA alone, protein kinase C (PKC) translocated from the cytosolic fraction to the particulate fraction. Pretreatment with 60 and 120 microM crocetin for 15 min inhibited the TPA-induced PKC activity in the particulate fraction by 50% and 66%, respectively, but did not affect the level of PKC protein. Crocetin also reduced the level of TPA-stimulated phosphorylation of cellular proteins. Cells pretreated with crocetin (120 microM) had 55% less PKC [3H]phorbol dibutyrate-binding capacity. Suppression of TPA (100 ng/mL)-induced c-jun and c-fos gene expression was also observed in the mouse fibroblast cells pretreated with crocetin (30, 60, and 120 microM). Our results provided a basis for understanding the inhibitory effect of crocetin on TPA-mediated tumor promotion.

Inhibition of protein kinase C and proto‐oncogene expression by crocetin in NIH/3T3 cells

Crocetin, a carotenoid isolated from the seeds of Gardenia jasminoides, was found to be a potent inhibitor of tumor promotion induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in mouse skin. When mouse fibroblast NIH/3T3 cells were treated with TPA alone, protein kinase C (PKC) translocated from the cytosolic fraction to the particulate fraction. Pretreatment with 60 and 120 μM crocetin for 15 min inhibited the TPA-induced PKC activity in the particulate fraction by 50% and 66%, respectively, but did not affect the level of PKC protein. Crocetin also reduced the level of TPA-stimulated phosphorylation of cellular proteins. Cells pre-treated with crocetin (120 μM) had 55% less PKC [3H]phorbol dibutyrate-binding capacity. Suppression of TPA (100 ng/mL)-induced c-jun and c-fos gene expression was also observed in the mouse fibroblast cells pre-treated with crocetin (30, 60, and 120 μM). Our results provided a basis for understanding the inhibitory effect of crocetin on TPA-mediated tumor promotion. © 1996 Wiley-Liss, Inc.

The tumor promoter arsenite stimulates AP-1 activity by inhibiting a JNK phosphatase.

Trivalent arsenic (As3+) is highly carcinogenic, but devoid of known mutagenic activity. Therefore, it is likely to act as a tumor promoter. To understand the molecular basis for the tumor-promoting activity of As3+, we examined its effect on transcription factor AP-1, whose activity is stimulated by several other tumor promoters. We found that As3+, but not As5+, which is toxic but not carcinogenic, is a potent stimulator of AP-1 transcriptional activity and an efficient inducer of c-fos and c-jun gene expression. Induction of c-jun and c-fos transcription by As3+ correlates with activation of Jun kinases (JNKs) and p38/Mpk2, which phosphorylate transcription factors that activate these immediate early genes. No effect on ERK activity was observed. As5+, on the other hand, had a negligible effect on JNK or p38/Mpk2 activity. Biochemical analysis and co-transfection experiments strongly suggest that the primary mechanism by which As3+ stimulates JNK activity involves the inhibition of a constitutive dual-specificity JNK phosphatase. This phosphatase activity appears to be responsible for maintaining low basal JNK activity in non-stimulated cells and its inhibition may lead to tumor promotion through induction of proto-oncogenes such as c-jun and c-fos, and stimulation of AP-1 activity. The same phosphatase may also regulate p38/Mpk2 activity.

Coincidental expression of E5a and c-jun in human papillomavirus type 6/11-infected condylomata

Previously, we have shown that E5a can induce expression of the c-jun gene in human papillomavirus (HPV)-11 E5a transformed NIH 3T3 cells and human epidermal keratinocytes. In this study, we investigated the relationship between expression of the E5a gene and c-jun in pathologically confirmed condylomata specimens using mRNA hybridization in situ. The c-jun RNA concentration was significantly higher in condylomata specimens with E5a mRNA expression than in specimens without E5a mRNA expression, or in normal cervical specimens. The cells with c-jun expression were located predominantly in the basal and parabasal cell layers. These layers were also the primary location of E5a-expressing cells. This is the first demonstration of a strong correlation (74%) between expression of the E5a and c-jun genes in condylomata specimens. This correlation might reflect regulation by HPV-11 E5a of c-jun gene expression in condyloma.

The regulation mechanism of c-jun and junB by human papillomavirus type 16 E5 oncoprotein.

In this study, we show that HPV-16 E5 induced anchorage-independent growth in immortalized human epidermal keratinocytes and that HPV-16 E5 in human keratinocytes had higher expression of c-jun and junB; also, we investigated the role of transcriptional initiation pathways in the expression elevation. In addition, Ras-dependent pathway, as well as PKC-dependent pathway, leads to HPV-16 E5-induced c-jun gene expression.

Heme Oxygenase-1 Gene Expression by a Glutathione Depletor, Phorone, Mediated through AP-1 Activation in Rats

This study shows that induction of heme oxygenase-1 (HO-1) gene expression by a glutathione (GSH) depletor, phorone, is inhibited by cycloheximide pretreatment and involves changes in c-jun, not c-fos, mRNA. Buthionine sulfoximine (BSO) enhanced markedly both c-jun and HO-1 gene expression evoked by phorone. Phorone dramatically increased AP-1 binding activity, which was blocked by unlabeled AP-1 oligonucleotide and abolished by anti-Jun antibodies, but not anti-Fos antibodies. In addition, pretreatment with dexamethasone, an inhibitor of AP-1 DNA binding, inhibited phorone-mediated HO-1 mRNA induction. These findings suggest that HO-1 induction by phorone is likely to involve in the activation of AP-1(Jun/Jun) binding, which could be associating with GSH depletion.

1,25(OH)2 vitamin D3, and retinoic acid antagonize endothelin-stimulated hypertrophy of neonatal rat cardiac myocytes.

1,25(OH)2 Vitamin D3 (VD3) and retinoic acid (RA) function as ligands for nuclear receptors which regulate transcription. Though the cardiovascular system is not thought to represent a classical target for these ligands, it is clear that both cardiac myocytes and vascular smooth muscle cells respond to these agents with changes in growth characteristics and gene expression. In this study we demonstrate that each of these ligands suppresses many of the phenotypic correlates of endothelin-induced hypertrophy in a cultured neonatal rat cardiac ventriculocyte model. Each of these agents reduced endothelin-stimulated ANP secretion in a dose-dependent fashion and the two in combination proved to be more effective than either agent used alone (VD3: 49%; RA:52%; VD3 + RA:80% inhibition). RA, at concentrations known to activate the retinoid X receptor, and, to a lesser extent, VD3 effected a reduction in atrial natriuretic peptide, brain natriuretic peptide, and alpha-skeletal actin mRNA levels. Similar inhibition (VD3:30%; RA:33%; VD3 + RA:59% inhibition) was demonstrated when cells transfected with reporter constructs harboring the relevant promoter sequences were treated with VD3 and/or RA for 48 h. These effects were not accompanied by alterations in endothelin-induced c-fos, c-jun, or c-myc gene expression, suggesting either that the inhibitory locus responsible for the reduction in the mRNA levels lies distal to the activation of the immediate early gene response or that the two are not mechanistically coupled. Both VD3 and RA also reduced [3H]leucine incorporation (VD3:30%; RA:33%; VD3 + RA:45% inhibition) in endothelin-stimulated ventriculocytes and, once again, the combination of the two was more effective than either agent used in isolation. Finally, 1,25(OH)2 vitamin D3 abrogated the increase in cell size seen after endothelin treatment. These findings suggest that the liganded vitamin D and retinoid receptors are capable of modulating the hypertrophic process in vitro and that agents acting through these or similar signaling pathways may be of value in probing the molecular mechanisms underlying hypertrophy.

Overexpression of the proto-oncogene c-jun in association with low-risk type specific human papillomavirus infection in condyloma acuminata.

Infection with different types of human papillomavirus (HPV) is associated with neoplasia at different anatomic sites. The "low-risk" HPVs (LR-HPV) are responsible for benign genital lesions such as condyloma acuminata. In order to clarify the tumorigenic mechanism of LR-HPV, the HPV infection status was investigated and the expression of the c-jun proto-oncogene in different HPV-related skin and genital lesions analyzed. Of the 17 condyloma specimens analyzed by Western blotting, 13 cases (76.5%) exhibited overexpression of the c-jun gene. All 13 cases harbored high copy numbers of the LR-HPV genome with an average of 926 copies per cell, whereas the other four cases had an average of 12 copies of LR-HPV per cell (P < 0.001). Further typing of HPV by Southern blotting revealed that HPV-6 and HPV-11 infections predominated in c-jun positive cases. The c-jun protein was detected much less frequently in cervical cancers (three of 29, or 10.3%) and skin warts (one of 10), and was not detected in five genital polyps or in five normal cervical tissues. These findings suggest a type 6/11-specific induction of c-jun gene expression in HPV-related neoplastic lesions.

Temporal changes in insulin-like growth factor I, c-fos, and c-jun gene expression during hyperplastic kidney growth in weanling rats.

We have previously determined that compensatory renal growth (CRG) during the initial 24-48 h after uninephrectomy (UNX) is GH independent in weanling animals, but associated with significant increases in insulin-like growth factor I (IGF-I) and IGF-I receptor gene expression. The purpose of the present study was to determine the temporal sequence of molecular and cellular events that occur at various time points (1, 6, 12, 18, 24, 48, and 72 h post-UNX) during this early period of accelerated renal growth in the weanling (21- to 25-day-old) rat. Rapid and sustained increases in steady state renal IGF-I receptor and IGF-I messenger RNA (mRNA) were observed at 1 and 6 h, respectively, and remained elevated in the remnant kidneys until 72 h post-UNX. The mRNAs for the early response genes, c-fos and c-jun, were not induced in the remnant kidneys from weanling rats until between 12-18 h, but were also sustained through 48 h post-UNX. Increases in remnant kidney DNA content and [3H]thymidine incorporation also occurred from 18-48 h post-UNX and returned to baseline levels by 72 h post-UNX, indicating that the hyperplastic response in the weanling remnant kidney occurs over a discrete period early after UNX. Neither IGF-I nor early response genes were elevated in kidneys from adult animals, which exhibited only hypertrophic renal growth at those early time points after UNX. These findings suggest that early CRG in the weanling rat is associated with rapid increases in IGF-I mRNA followed by a rise in c-fos and c-jun gene expression and a mitogenic response. Furthermore, when the mRNA levels of IGF-I and early response genes returned to baseline levels, mitogenic growth stopped, and slower prolonged hypertrophic renal growth ensued.

Temporal changes in insulin-like growth factor I, c-fos, and c-jun gene expression during hyperplastic kidney growth in weanling rats

Temporal changes in insulin-like growth factor I, c-fos, and c-jun gene expression during hyperplastic kidney growth in weanling rats

Age-related Decline in Mitogen-activated Protein Kinase Activity in Epidermal Growth Factor-stimulated Rat Hepatocytes

A number of studies have demonstrated that the proliferative capacity of cells declines with aging. In particular, epidermal growth factor (EGF)-stimulated DNA synthesis is reduced in hepatocytes from aged rats relative to young rats. Growth factor stimulation activates a genetic program in large part regulated by a family of mitogen-activated protein kinases (MAPK) that phosphorylate and thereby activate transcription factors involved in controlling the expression of proliferation-associated genes. In the present study, we compared the activation of the extracellular signal-regulated kinase 2 (ERK2) and c-Jun N-terminal kinase 1 (JNK1) MAPK in EGF-stimulated hepatocytes derived from young (6-month) and aged (24-month) rats. JNK activity was not appreciably altered by EGF treatment of cells from either age group. In contrast, ERK2 was highly activated by EGF treatment, but the magnitude of activation was significantly lower in hepatocytes of aged animals compared to those of young animals (7-fold versus 20-fold, respectively). The reduced ERK2 activity in response to EGF was associated with decreased c-fos and c-jun mRNA expression and lower levels of AP-1 transcription factor DNA binding activity in the aged hepatocytes. Finally, the basal expression of MAPK phosphatase 1, a MAPK-regulated gene involved in regulating MAPK activity, was higher in aged hepatocytes. Taken together, these findings suggest that an alteration in the balance between MAP kinase-phosphatase activities could contribute to the age-related decline in proliferative capacity.

Hypoglycemia-induced AP-1 transcription factor and basic fibroblast growth factor gene expression in multidrug resistant human breast carcinoma MCF-7/ADR cells.

We investigated the effect of hypoglycemic treatment on the activation of the AP-1 transcription factors and the regulation of basic fibroblast growth factor (bFGF) gene expression in multidrug resistant human breast carcinoma MCF-7/ADR cells. Northern blot and gel mobility shift assays showed that hypoglycemic treatment induced c-jun and c-fos gene expression, AP-1 binding activity, as well as bFGF gene expression. Moreover, transfected cells expressing high levels of abnormal c-Jun protein exhibited a reduction in the bFGF protein levels compared to parental cells. A potent protein kinase C (PKC) inhibitor, H-7 (60 micrograms/ml) suppressed the stress-induced bFGF gene expression. Our study also demonstrated that H-7 did not facilitate the decay of bFGF mRNA. Thus, the suppression of bFGF gene expression by treatment with H-7 was due to the effect of the drug on the synthesis of bFGF mRNA rather than the stability of bFGF mRNA. Our data suggest that hypoglycemia-induced bFGF gene expression is mediated through the activation of PKC and the AP-1 transcription factors.