Atorvastatin inhibits osteoclast differentiation by suppressing NF-κB and MAPK signaling during IL-1ß-induced osteoclastogenesis.

BACKGROUND/AIMS: To define the effect of statins on interleukin 1ß (IL-1ß)-induced osteoclastogenesis and elucidate the underlying mechanisms. METHODS: Bone marrow cells were obtained from 5-week-old male ICR (Institute for Cancer Research) mice, and they were cultured to differentiate them into osteoclasts with macrophage colony-stimulating factor and the receptor activator of nuclear factor (NF)-κB ligand in the presence or absence of IL-1ß or atorvastatin. The formation of osteoclasts was evaluated by tartrate-resistant acid phosphatase (TRAP) staining and resorption pit assay with dentine slice. The molecular mechanisms of the effects of atorvastatin on osteoclastogenesis were investigated using reverse transcription polymerase chain reaction and immunoblotting for osteoclast specific molecules. RESULTS: Atorvastatin significantly reduced the number of TRAP-positive multinucleated cells as well as the bone resorption area. Atorvastatin also downregulated the expression of the NF of activated T-cell c1 messenger RNA and inhibited the expression of osteoclast-specific genes. A possible underlying mechanism may be that atorvastatin suppresses the degradation of the inhibitors of NF-κB and blocks the activation of the c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38; thus, implicating the NF-κB and mitogen-activated protein kinases pathway in this process. CONCLUSIONS: Atorvastatin is a strong inhibitor of inflammation-induced osteoclastogenesis in inflammatory joint diseases.

A pathogenic role for ER stress-induced autophagy and ER chaperone GRP78/BiP in T lymphocyte systemic lupus erythematosus.

Abnormal regulation of ER stress and apoptosis has been implicated in autoimmune disorders. Particularly, ER stress-induced autophagy and the role of GRP78, or BiP in T lymphocyte survival and death in SLE are poorly understood. This study investigated the pathogenic roles of ER stress-induced autophagy and GRP78/BiP in apoptosis of T lymphocytes. We compared spontaneous and induced autophagy and apoptosis of T lymphocytes in healthy donors and patients with SLE. The molecular mechanism of altered autophagy and apoptosis was investigated in T lymphocytes transfected with siRNA for beclin 1 and CHOP and T lymphocytes overexpressing GRP78. Decreased autophagy and increased apoptosis in response to TG-induced ER stress were observed in lupus T lymphocytes. GRP78 and ER stress-signaling molecules, such as PERK, p-eIF2α, IRE1, and ATF6 decreased, whereas CHOP levels increased in lupus T cells in response to TG. The levels antiapoptotic molecules, Bcl-2 and Bcl-XL decreased, whereas the proapoptotic molecules, Bax and caspase 6, increased in lupus T cells. The TG-induced ER stress altered autophagy and apoptosis, which in turn, led to abnormal T cell homeostasis with increased apoptotic T cell death. We hypothesize that aberrant autophagy of T lymphocytes as a result of ER stress and decreased GRP78 expression is involved in the pathogenesis of SLE and might serve as important therapeutic targets.

Ethyl acetate fraction from Angelica sinensis inhibits IL-1ß-induced rheumatoid synovial fibroblast proliferation and COX-2, PGE2, and MMPs production.

BACKGROUND: The root of Angelica sinensis (AS), also known as "Dang-gui," was a popular herbal medicine widely used in the treatment of gynecological diseases in China, Korea, and Japan for a long time. This study aimed to determine the effects of ethyl acetate fraction from Angelica sinensis (EAAS) on the interleukin-1ß (IL-1ß)-induced proliferation of rheumatoid arthritis synovial fibroblasts (RASFs), and production of matrix metalloproteinases (MMPs), cyclooxygenase (COX) 2, and prostaglandin E2 (PGE2), involved in articular bone and cartilage destruction, by RASFs. RESULTS: RASF proliferation was evaluated with cholecystokinin octapeptide (CCK-8) reagent in the presence of IL-1ß with/without EAAS. Expression of MMPs, tissue inhibitor of metalloproteinases-1 (TIMP-1), COXs, PGE2, and intracellular mitogen-activated protein kinase (MAPK) signaling molecules, including p-ERK, p-p38, p-JNK, and NF-κB, were examined using immunoblotting or semi-quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. EAAS inhibited IL-1ß-induced RASF proliferation; MMP-1, MMP-3, and COX-2 mRNA and protein expressions; and PGE2 production. EAAS also inhibits the phosphorylation of ERK-1/2, p38, and JNK, and activation of NF-κB by IL-1ß. CONCLUSION: EAAS might be a new therapeutic modality for rheumatoid arthritis management.

Increased RANKL-mediated osteoclastogenesis by interleukin-1ß and endoplasmic reticulum stress.

OBJECTIVE: The mechanism by which IL-1ß and thapsigargin (TG)-induced endoplasmic reticulum (ER) stress modulate the receptor activator of nuclear factor kappa-B ligand (RANKL)-mediated osteoclastogenesis remains elusive. Thus, we investigated the osteoclast-specific and ER signals in osteoclastogenesis of bone marrow-derived cells. METHODS: Bone marrow cells (BMCs) were obtained from 5-week-old male ICR mice and cultured to be differentiated into osteoclasts with M-CSF and RANKL in the presence or absence of IL-1ß, TG, or 4-phenylbutyric acid (PBA), an ER stress-reducing drug. The formation of osteoclasts was evaluated by tartrate-resistant acid phosphatase (TRAP) staining and resorption pit assay with a dentine slice. The molecular mechanism of IL-1ß and ER stress in osteoclastogenesis was investigated in BMCs transfected with siRNA for GRP78, PERK and IRE1 using reverse transcription-polymerase chain reaction and immunoblotting for osteoclast-specific and ER stress signaling molecules. RESULTS: IL-1ß and ER stress induced by TG-augmented the formation of osteoclasts, which was significantly inhibited by PBA and was mediated with osteoclast-specific signals, including c-Fos, NFATc1, and ER stress- associated signaling pathways, such as PERK, IRE1, GRP78, and eIF2α. siRNA-mediated knockdown of ER stress signals inhibited the expression of NFATc1 and c-Fos, thus reducing IL-1ß and/or TG-induced formation of osteoclasts. CONCLUSIONS: Osteoclastogenesis by IL-1ß and/or ER stress is mainly associated with upregulation of eIF2α, GRP78, PERK and IRE1. These results suggest that the signaling pathway of ER stress-induced osteoclast formation might be a new therapeutic target to prevent inflammatory and destructive arthritic disease such as RA and diverse osteoporosis.

Kaempferol inhibits IL-1ß-stimulated, RANKL-mediated osteoclastogenesis via downregulation of MAPKs, c-Fos, and NFATc1.

Kaempferol is one of the most common flavonoid that is present in a variety of vegetables and fruits and has effects on bone metabolism. The present study was performed to define the effects of kaempferol on interleukin (IL)-1ß-stimulated receptor activator of NF-κB ligand (RANKL)-mediated osteoclast differentiation. Bone marrow cells were harvested from 6-week-old male imprinting control region mice, and the differentiation of osteoclasts from these cells was evaluated by tartrate-resistant acid phosphatase staining and resorption pit formation assay. Phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated p38, phosphorylated c-Jun amino-terminal kinase, NF-κB (p65), IκBα, c-Fos, and nuclear factor of activated T cells c1 (NFATc1) expressions were examined by Western blotting and quantitative RT-PCR. Kaempferol inhibits IL-1ß-stimulated, RANKL-mediated osteoclast differentiation and also inhibits IL-1ß-stimulated, RANKL-mediated phosphorylation of ERK 1/2, p38 and JNK MAP kinases, and expressions of c-Fos and NFATc1. These results indicate that kaempferol has an inhibitory role in the bone loss by preventing osteoclast formation and suggest that it might be a novel therapeutic agent for the treatment of inflammatory arthritis by managing bone destruction.

Sulforaphane inhibits IL-1ß-induced proliferation of rheumatoid arthritis synovial fibroblasts and the production of MMPs, COX-2, and PGE2.

This study was performed to define the effects of sulforaphane on interleukin-1ß (IL-1ß)-induced proliferation of rheumatoid arthritis synovial fibroblasts (RASFs), the expression of matrix metalloproteinases (MMPs) and cyclooxygenase (COX), and the production of prostaglandin E2 (PGE2) by RASFs. The proliferation of RASFs was evaluated with CCK-8 reagent in the presence of IL-1ß with/without sulforaphane. The expression of MMPs, tissue inhibitor of metalloproteinase-1, COXs, intracellular mitogen-activated protein kinase signalings, including p-ERK, p-p38, p-JNK, and nuclear factor-kappaB (NF-kB), and the production of PGE2 were examined by Western blotting or semi-quantitative RT-PCR and ELISA. Sulforaphane inhibits unstimulated and IL-1ß-induced proliferation of RASFs; the expression of MMP-1, MMP-3, and COX-2 mRNA and protein; and the PGE2 production induced by IL-1ß. Sulforaphane also inhibits the phosphorylation of ERK-1/2, p-38, and JNK and activation of NF-kB by IL-1ß. These results indicate that sulforaphane inhibits the proliferation of synovial fibroblasts, the expression of MMPs and COX-2, and the production of PGE2, which are involved in synovitis and destruction of RA, and suggest that sulforaphane might be a new therapeutic agent for RA.

Ethyl acetate fraction from Angelica sinensis inhibits IL-1ß-induced rheumatoid synovial fibroblast proliferation and COX-2, PGE2, and MMPs production

BACKGROUND: The root of Angelica sinensis (AS), also known as "Dang-gui," was a popular herbal medicine widely used in the treatment of gynecological diseases in China, Korea, and Japan for a long time. This study aimed to determine the effects of ethyl acetate fraction from Angelica sinensis (EAAS) on the interleukin-1ß (IL-1ß)-induced proliferation of rheumatoid arthritis synovial fibroblasts (RASFs), and production of matrix metalloproteinases (MMPs), cyclooxygenase (COX) 2, and prostaglandin E2 (PGE2), involved in articular bone and cartilage destruction, by RASFs. RESULTS: RASF proliferation was evaluated with cholecystokinin octapeptide (CCK-8) reagent in the presence of IL-1ß with/without EAAS. Expression of MMPs, tissue inhibitor of metalloproteinases-1 (TIMP-1), COXs, PGE2, and intracellular mitogen-activated protein kinase (MAPK) signaling molecules, including p-ERK, p-p38, p-JNK, and NF-κB, were examined using immunoblotting or semi-quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. EAAS inhibited IL-1ß-induced RASF proliferation; MMP-1, MMP-3, and COX-2 mRNA and protein expressions; and PGE2 production. EAAS also inhibits the phosphorylation of ERK-1/2, p38, and JNK, and activation of NF-κB by IL-1ß. CONCLUSION: EAAS might be a new therapeutic modality for rheumatoid arthritis management.

Kaempferol inhibits IL-1ß-induced proliferation of rheumatoid arthritis synovial fibroblasts and the production of COX-2, PGE2 and MMPs.

Inflammatory cytokines, matrix metalloproteinases (MMPs) and cyclooxygenase (COX)-2 released from rheumatoid arthritis synovial fibroblasts (RASFs) are involved in the destruction of both articular bone and cartilage. Kaempferol has been reported to act as an antioxidant and anti-inflammatory agent by inhibiting nitric oxide synthase and COX enzymes. The aim of the present study was to determine the effects of kaempferol on the interleukin-1ß (IL-1ß)-induced proliferation of RASFs and the production of MMPs, COX and prostaglandin E2 (PGE2) by RASFs. The proliferation of the RASFs stimulated with IL-1ß and treated with/without kaempferol was evaluated by CCK-8 assay. The expression of MMPs, TIMP metallopeptidase inhibitor-1 (TIMP-1), COXs, PGE2 and that of intracellular MAPK signaling molecules, including p-ERK, p-p38, p-JNK and nuclear factor-κB (NF-κB) was examined by immunoblotting or semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and ELISA under the conditions described above. Kaempferol inhibited the proliferation of both unstimulated and IL-1ß­stimulated RASFs, as well as the mRNA and protein expression of MMP-1, MMP-3, COX-2 and PGE2 induced by IL-1ß. Kaempferol also inhibited the phosphorylation of ERK-1/2, p38 and JNK, as well as the activation of NF-κB induced by IL-1ß. These results indicate that kaempferol inhibits synovial fibroblast proliferation, as well as the production of and MMPs, COX­2 and PGE2, which is involved in articular inflammation and destruction in rheumatoid arthritis (RA). Our data suggest that kaempferol may be a novel therapeutic agent for the treatment of RA.

Quercetin inhibits IL-1ß-induced proliferation and production of MMPs, COX-2, and PGE2 by rheumatoid synovial fibroblast.

This study was aimed to determine the effects of quercetin on the interleukin-1ß (IL-1ß)-induced proliferation of rheumatoid synovial fibroblasts (RASFs) and production of matrix metalloproteinases (MMPs), cyclooxygenase (COX), and prostaglandin E2 (PGE2) by RASFs. The proliferation and apoptosis of RASFs was evaluated with CCK-8 reagent and flow cytometry in the presence of IL-1with CCK-8 reagquercetin. The expression of MMPs, IL-1ß enhanced the expression of MMP-1, MMP-3, tissue inhibitor of metalloproteinase (TIMP)-1, COXs, PGE2, and intracellular mitogen-activated protein kinase (MAPK) signalings including phosphorylated extracellular signal-regulated kinase (p-ERK), p-p38, phosphorylated c-Jun N-terminal kinase (p-JNK), and nuclear factor kB (NF-kB) were examined by immunoblotting or semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) in conditions as described above. Quercetin inhibits unstimulated and IL-1ß-induced proliferation of RASFs and MMP-1, 3, COX-2 messenger ribonucleic acid and protein expression, PGE2 production induced with IL-1ß. Quercetin also inhibits the phosphorylation of ERK-1/2, p38, JNK and activation of NF-kB by IL-1ed. These results indicate that quercetin inhibits synovial fibroblasts proliferation and MMPs, COX-2, and PGE2 production, which involved joint destruction in rheumatoid arthritis (RA), and suggest that it might be a new therapeutic agent for management of RA.

Effects of Ficus carica paste on loperamide-induced constipation in rats.

OBJECTIVE: Constipation is one of the most common gastrointestinal complaints worldwide. This study examined the effects of fig (Ficus carica L.) paste for the treatment of loperamide-induced constipation in a rat model. METHODS: Animals were divided into one normal control group and four experimental groups (0, 1, 6, and 30 g/kg). Loperamide (2 mg/kg, twice per day) was injected intraperitoneally to induce constipation in the four experimental groups. Fig paste was administered for 4 weeks to assess its anti-constipation effects. RESULTS: Fecal pellet number, weight and water content were increased in the fig-treated groups as compared to the control group. Reductions in body weight and increased intestinal transit length were observed in the fig-treated groups. Fecal pellet number was reduced in the distal colons of the fig-treated rats. Exercise and ileum tension increased in the experimental groups as compared to the control group. According to histological analyses, the thickness of the distal colon and areas of crypt epithelial cells that produce mucin were increased in the fig-treated groups in a dose-dependent manner. CONCLUSION: Constipation was decreased when fig fruit was fed to rats. Specifically, fecal number, weight, and water content, as well as histological parameters such as thickness and mucin areas in the distal colon were improved. Fig treatment may be a useful therapeutic and preventive strategy for chronic constipation.