Interleukin-6 Synthesis In Cells Research Articles

Synergistic effect of vasoactive intestinal peptides on TNF-α-induced IL-6 synthesis in osteoblasts: amplification of p44/p42 MAP kinase activation

We previously showed that tumor necrosis factor-alpha (TNF-alpha) stimulates synthesis of interleukin-6 (IL-6), a potent bone resorptive agent, via p44/p42 mitogen-activated protein (MAP) kinase and phosphatidylinositol 3-kinase/Akt in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the effect of vasoactive intestinal peptide (VIP) on TNF-alpha-induced IL-6 synthesis in these cells. VIP, which by itself slightly stimulated IL-6 synthesis, synergistically enhanced the TNF-alpha-induced IL-6 synthesis in MC3T3-E1 cells. The synergistic effect of VIP on the TNF-alpha-induced IL-6 synthesis was concentration-dependent in the range between 1 and 70 nM. We previously reported that VIP stimulated cAMP production in MC3T3-E1 cells. Forskolin, a direct activator of adenylyl cyclase, or 8-bromoadenosine-3',5'-cyclic monophosphate (8bromo-cAMP), a plasma membrane-permeable cAMP analogue, markedly enhanced the TNF-alpha-induced IL-6 synthesis as well as VIP. VIP markedly up-regulated the TNF-alpha-induced p44/p42 MAP kinase phosphorylation. The Akt phosphorylation stimulated by TNF-alpha was only slightly affected by VIP. PD98059, a specific inhibitor of MEK1/2, significantly suppressed the enhancement of TNF-alpha-induced IL-6 synthesis by VIP. The synergistic effect of a combination of VIP and TNF-alpha on the phosphorylation of p44/p42 MAP kinase was diminished by H-89, an inhibitor of cAMP-dependent protein kinase. These results strongly suggest that VIP synergistically enhances TNF-alpha-stimulated IL-6 synthesis via up-regulating p44/p42 MAP kinase through the adenylyl cyclase-cAMP system in osteoblasts.

Involvement of p42/p44 MAP kinase in endothelin-1-induced interleukin-6 synthesis in osteoblast-like cells

We previously reported that endothelin-1 induces synthesis of interleukin-6 (IL-6) via activation of protein kinase C in osteoblast-like MC3T3-E1 cells. In the present study, we further investigated whether p42/p44 mitogen-activated protein (MAP) kinase is involved in endothelin-1-induced IL-6 synthesis in these cells. Endothelin-1 stimulated p42/p44 MAP kinase activation in a dose-dependent manner in the range between 0.1 nmol/L and 0.1 μmol/L. PD98059, a specific inhibitor of the upstream kinase that activates p42/p44 MAP kinase, suppressed endothelin-1-induced IL-6 synthesis as well as endothelin-1-activated p42/p44 MAP kinase. Both p42/p44 MAP kinase activation and IL-6 synthesis induced by 12- O-tetradecanoylphorbol-13-acetate (TPA), a protein kinase C-activating phorbol ester, were reduced by PD98059. Calphostin C, a highly specific inhibitor of protein kinase C, suppressed endothelin-1-stimulated p42/p44 MAP kinase activation as well as endothelin-1-induced IL-6 synthesis. These results strongly suggest that protein kinase C-dependent p42/p44 MAP kinase activation is involved in endothelin-1-induced IL-6 synthesis in osteoblast-like cells.

Prostaglandin F2alpha stimulates interleukin-6 synthesis via activation of PKC in osteoblast-like cells.

In previous studies, we reported that prostaglandin F2alpha (PGF2alpha) stimulates phosphoinositide hydrolysis by phospholipase C and phosphatidylcholine hydrolysis by phospholipase D in osteoblast-like MC3T3-E1 cells. In the present study, we examined the effect of PGF2alpha on synthesis of interleukin-6 (IL-6) and the involvement of protein kinase C (PKC) activation in the IL-6 synthesis in these cells. PGF2alpha significantly stimulated IL-6 synthesis in a dose-dependent manner in the range between 10 nM and 10 microM. A PKC-activating phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), induced IL-6 synthesis. On the contrary, 4alpha-phorbol 12,13-didecanoate, a PKC-nonactivating phorbol ester, had no effect. The synthesis of IL-6 stimulated by a combination of PGF2alpha and TPA was not additive. Staurosporine, an inhibitor for protein kinases that suppressed the TPA-induced IL-6 synthesis, significantly inhibited the PGF2alpha-induced IL-6 synthesis. Calphostin C, a highly specific PKC inhibitor, also suppressed the PGF2alpha-stimulated synthesis of IL-6. The effect of PGF2alpha on IL-6 synthesis in PKC-downregulated cells was much weaker than that in intact cells. These results strongly suggest that PGF2alpha induces IL-6 synthesis via PKC activation in osteoblast-like cells.

Prostaglandin E2 Induces Interleukin‐6 Synthesis in Human Astrocytoma Cells

Prostaglandins (PGs) and cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6), have been implicated in the etiopathology of various inflammatory and degenerative disorders, including Alzheimer's disease (AD) and prion diseases. Nonsteroidal antiinflammatory drugs (NSAIDs), potent inhibitors of PG synthesis, appear to be beneficial in the treatment of AD. To assess whether PGs are able to induce IL-6 synthesis in cells of the CNS, IL-6 mRNA and protein syntheses were measured in a human astrocytoma cell line after stimulation with different PGs. PGE1 and PGE2, but not PGD2 and PGF2 alpha, led to a rapid and transient induction of IL-6 mRNA, followed by IL-6 protein synthesis. Furthermore, PGE2 potentiated IL-1 beta-induced IL-6 mRNA synthesis. These results are discussed with respect to the participation of PGs in neurodegenerative diseases (and its inhibition by NSAIDs) by affecting cytokine expression.

Platelet-derived growth factor stimulates the synthesis of interleukin-6 in cells of the osteoblast lineage.

Platelet-derived growth factor (PDGF) increases bone resorption and the number of osteoclasts in calvarial sections, and it may regulate local cytokines involved in bone remodeling. Interleukin-6 (IL-6), a cytokine secreted by osteoblasts, osteoclasts, and stromal cells, is known to increase osteoclast recruitment. We tested the effects of PDGF on IL-6 expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). Treatment of Ob cells with PDGF BB caused a time- and dose-dependent induction of IL-6 messenger RNA (mRNA), as determined by Northern blot analysis. The effect was maximal after 1 h of treatment and was observed with PDGF BB at 0.3-3.3 nM. Treatment with PDGF BB for 24 h also increased IL-6 polypeptide levels in the culture medium, as determined by a specific bioassay. Although PDGF AA increased IL-6 mRNA levels, its effect was less pronounced than that of PDGF BB. Phorbol 12-myristate 13-acetate (PMA) induced IL-6 transcripts, and the effect of PDGF BB was inhibited in the presence of the protein kinase C (PKC) inhibitor, sangivamycin, or after down-regulation of PKC by PMA preincubation. Although forskolin increased IL-6 mRNA levels, PDGF BB did not induce cAMP production in Ob cells. The calcium ionophore, ionomycin, enhanced IL-6 transcripts in Ob cells and the intracellular calcium chelator, 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra-(acetoxymethyl)-ester, inhibited the induction of IL-6 transcripts by PDGF BB, PMA, and PTH. In conclusion, PDGF BB stimulates IL-6 expression in Ob cells, a response that is PKC and calcium dependent. The increase in IL-6 expression may be relevant to the actions of PDGF BB on bone resorption.