Seon-Yle Ko

Effect of Rehmannia glutinosa Libosch extracts on bone metabolism

BACKGROUND Rehmannia glutinosa Libosch extracts (RGX) were investigated to determine if they play roles in bone metabolism. METHODS The effects on osteoblasts were determined by measuring (1) cell proliferation, (2) alkaline phosphatase (ALP) activity, (3) mRNA expression of bone-related proteins, (4) transcriptional activity of Runx2, and (5) osteoprotegerin (OPG) secretion. The effects on the osteoclasts were investigated by measuring (1) tartrate-resistant acid phosphatase-positive [TRAP(+)] multinucleated cell (MNC) formation and (2) resorption areas after culturing osteoclast precursors. Bone mineral density (BMD) measurements and histological observations on rats were also carried out. RESULTS RGX treatment showed a significant increase in both the proliferation and ALP activity of osteoblasts. RGX increased the expression of the bone-related genes. OPG secretion was markedly increased after RGX treatment. In addition, RGX treatment decreased the number of TRAP(+) MNCs and the resorption areas. In vivo studies using ovariectomy-induced osteoporotic rats revealed that RGX alleviated the decrease in the trabecular BMD, and increased the cortical bone thickness and trabeculation of the bone marrow spaces. CONCLUSIONS RGX stimulates the proliferation and activities of osteoblasts, while inhibiting the generation and resorptive activities of osteoclasts. It also shows preventive effects on osteoporotic bone loss induced by an ovariectomy. Although the active substances have not yet been identified, it is believed that the RGX seems to contain active components that have a potential to enhance the bone metabolism in osteoporosis.

Regulation of Polo-like Kinase 1 by DNA Damage in Mitosis INHIBITION OF MITOTIC PLK-1 BY PROTEIN PHOSPHATASE 2A

DNA damage triggers multiple checkpoint pathways to arrest cell cycle progression. Polo-like kinase 1 (Plk1) is an important regulator of several events during mitosis. In addition to Plk1 functions in cell cycle, Plk1 is involved in DNA damage check-point in G2 phase. Normally, ataxia telangiectasia-mutated kinase (ATM) is a key enzyme involved in G2 phase cell cycle arrest following DNA damage, and inhibition of Plk1 by DNA damage during G2 occurs in a ATM/ATR-dependent manner. However, it is still unclear how Plk1 is regulated in response to DNA damage in mitosis in which Plk1 is already activated. Here, we show that treatment of mitotic cells with doxorubicin and γ-irradiation inhibits Plk1 activity through dephosphorylation of Plk1, and cells were arrested in G2 phase. Treatments of the phosphatase inhibitors and siRNA experiments suggested that PP2A pathway might be involved in regulating mitotic Plk1 activity in mitotic DNA damage. Finally, we propose a novel pathway, which is connected between ATM/ATR/Chk and protein phosphatase-Plk1 in DNA damage response in mitosis.

Myricetin suppresses LPS-induced MMP expression in human gingival fibroblasts and inhibits osteoclastogenesis by downregulating NFATc1 in RANKL-induced RAW 264.7 cells

OBJECTIVE Periodontitis is an inflammatory disease that affects connective tissue attachments and the supporting bone that surrounds the teeth. Gingival fibroblasts induce the overexpression of matrix metalloproteinase (MMP), which is involved in inflammatory progression in periodontitis. Osteoclasts are responsible for skeletal modeling and remodeling but may also destroy bone in several bone diseases, including osteoporosis and periodontitis. This study examined the anti-destructive effects of myricetin on human gingival fibroblasts (HGF) under lipopolysaccharide- (LPS-) induced inflammatory conditions, and the anti-osteoclastogenetic effect of myricetin on the receptor activator of NF-κB ligand (RANKL) induced RAW264.7 cells was also investigated. DESIGN The effects of myricetin on HGF were determined by measuring the cell viability and mRNA expression and enzyme activity of tissue-destructive proteins, including MMP-1, MMP-2 and MMP-8. The effects of myricetin on osteoclasts were examined by measuring the following: (1) the cell viability, (2) the formation of tartrate-resistant acid phosphatase (TRAP)(+) multinucleated cells, (3) MAPK signalling pathways (4) mRNA expression of osteoclast-associated genes and (5) tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) secretion. RESULTS The myricetin had no effects on the cell viability of the HGF and decreased the mRNA expression and enzyme activity of MMP-1, MMP-2 and MMP-8 in the HGF. Myricetin inhibited the formation of RANKL-stimulated TRAP(+) multinucleated cells. Myricetin also inhibited the RANKL-stimulated activation of p-38, ERK and cSrc signaling, and inhibited the RANKL-stimulated degradation of I(k)B in the RAW264.7 cells. In addition, the RANKL-stimulated induction of NFATc1 transcription factors was abrogated by myricetin. Myricetin decreased the mRNA expression of osteoclast-associated genes, including cFOS, TRAP and cathepsin K in the RAW264.7 cells. Myricetin inhibited the secretion of LPS-induced TNF-α and IL-1β in the RAW264.7 cells. CONCLUSIONS These findings suggest that myricetin has therapeutic effects on bone-destructive processes, such as those that occur in periodontal diseases.

n-Butanol extracts of Panax notoginseng suppress LPS-induced MMP-2 expression in periodontal ligament fibroblasts and inhibit osteoclastogenesis by suppressing MAPK in LPS-activated RAW264.7 cells

OBJECTIVE Periodontitis is a group of inflammatory diseases that affect connective tissue attachments and the supporting bone that surround the teeth. Osteoclasts are responsible for skeletal modeling and remodeling but may also destroy bone in several bone diseases, including osteoporosis and periodontitis. This study examined the anti-inflammatory effects of Panax notoginseng (PN) on periodontal ligament fibroblasts (PDLFs) and RAW264.7 cells under lipopolysaccharide (LPS) induced inflammatory conditions. DESIGN The effects of PN on PDLFs were determined by measuring the cell viability and mRNA expression of tissue-destructive proteins. The effects of PN on osteoclasts were examined by measuring the following: (1) the cell viability, (2) the formation of Tartrate-resistant acid phosphatase (TRAP)(+) multinucleated cells, (3) MAPK signaling pathways, (4) mRNA expression of inflammatory-related proteins and (5) nitric oxide (NO) production. RESULTS The n-butanol extracts of PN (bPN) increased the cell proliferation of the PDLFs and decreased the mRNA expression of matrix metalloproteinase (MMP)-2 in the PDLFs. bPN inhibited the formation of LPS-stimulated TRAP(+) multinucleated cells. bPN also inhibited the LPS-stimulated activation of JNK and ERK signaling, and inhibited the LPS-stimulated degradation of I(K)B in the RAW264.7 cells. In addition, bPN decreased the mRNA expression of MMP-9 and iNOS, which are involved in the range of pathophysiological processes, such as inflammation in the RAW264.7 cells. NO production was also decreased via the inhibition of iNOS. CONCLUSIONS These findings suggest that bPN has therapeutic effects on bone-destructive processes, such as those that occur in periodontal diseases.

The polyphenol resveratrol promotes skeletal growth in mice through a sirtuin 1‐bone morphogenic protein 2 longevity axis

The polyphenol resveratrol (RSV) exists in high quantities in certain foods (e.g. grapes and nuts). However, the capacity of RSV to confer physiological health benefits and a biological mechanism through which this might occur remains unclear.

A monoclonal antibody recognizes undifferentiation-specific carbohydrate moieties expressed on cell surface of the human dental pulp cells

Human dental pulp cells are obtained from dental pulp tissue, and have the ability to form dentin and a pulp-like complex. Although adult stem cells have been identified from the primary culture by using specific cell surface markers, the identity of surface markers for the purification of stem cells within the dental pulp population are still unclear. Previously, we had constructed monoclonal antibodies against the undifferentiated cell-specific surface markers of human dental pulp cells (hDPCs) by performing decoy immunization. Among them, a monoclonal antibody against the cell surface antigen of the undifferentiated hDPCs (named UPSA-1) was purified and its heavy and light chain consensus regions were analyzed. The cell surface binding affinity of UPSA-1 mAb on the undifferentiated hDPCs was stronger than that on the differentiated cells. When tunicamycin was applied to hDPSCs during culture, the cell surface binding affinity of the antibody was dramatically decreased, and dentinogenic differentiation was reduced. The purified UPSA-1 antigen band resulting from immunoprecipitation disappeared or shifted down on the SDS-PAGE by deglycosylation. These data suggested that glycosylation on the cell surface might be a marker of an undifferentiated state, and that UPSA-1 mAb might be useful for identifying the carbohydrate moiety on the cell surface of undifferentiated pulp cells.

Inhibitory Effect of Myricetin on Matrix Metalloproteinase Expression and Activity in Periodontal Inflammation

Flavonoid myricetin, usually found in tea and medicinal plants, has antioxidant and anti-inflammatory effects. Our objectives in this study were to verify the effects of myricetin on periodontal ligament fibroblasts (PDLFs) under inflammatory conditions and to observe its effects on osteoclast generation and on cytokine expression in RAW264.7 cells. To determine the effects of myricetin on PDLFs, we examined the expression and activity of proteolytic enzymes, including MMP-1, MMP-2, and MMP-8, which all play an important role in chronic periodontitis. We observed the effects of myricetin on intracellular signal transduction to verify the molecular mechanism involved. By measuring the formation of TRAP–positive multinucleated cells and the expression and activity of MMP-8, we were able to assess the effects of myricetin on osteoclast generation. In addition, by measuring the secretion of IL-6 and NO, we could evaluate the effects of myricetin on inflammatory mediators. We found that Myricetin had no effect on the viability of the PDLFs in the presence of inflammation, but it did decrease both the expression of MMP-1 and MMP-8 and the enzyme activity of MMP-2 and MMP-8 in these fibroblasts. Myricetin also decreased the lipopolysaccharide-stimulated phosphorylation of JNK, p38 signaling, IKKB, AKT, and p65RelA in the PDLFs. In the RAW264.7 cells, myricetin inhibited both the expression and the activity of MMP-8. Furthermore, Myricetin not only suppressed the generation of LPS-stimulated osteoclasts, but it also slightly inhibited LPS-stimulated degradation of IkB and decreased the release of LPS-induced IL-6 and NO. These findings suggest that myricetin alleviates the tissue-destructive processes that occur during periodontal inflammation.