Characteristics of blood tests in patients with acute cerebral infarction who developed symptomatic intracranial hemorrhage after intravenous administration of recombinant tissue plasminogen activator.

OBJECTIVE: Patients suspected as having acute ischemic stroke usually undergo blood tests, including coagulation-related indexes, because thrombocytopenia and coagulopathy are contraindications for recombinant tissue plasminogen activator (rtPA) administration. We aimed to identify blood test indexes associated with symptomatic intracranial hemorrhage (sICH) in patients with acute ischemic stroke who received intravenous rtPA. METHODS: This retrospective observational study included patients diagnosed with acute ischemic stroke who were treated with intravenous rtPA at the emergency department of a tertiary hospital in Seoul between February 2008 and January 2018. Blood test indexes were compared between the sICH and non-sICH groups. Logistic regression and receiver-operating characteristic curve analyses were performed. RESULTS: In this study, 375 patients were finally included. Of 375 patients, 42 (11.2%) showed new intracranial hemorrhage on follow-up brain computed tomography, of whom 14 (3.73%) had sICH. Platelet count, aspartate aminotransferase and lactate dehydrogenase levels were significantly different between the sICH and non-sICH groups, and platelet count showed statistical significance in the regression analysis. Significantly lower platelet counts were observed in the sICH group than in the non-sICH group (174,500 vs. 228,000/mm3, P=0.020). The best cutoff platelet count was 195,000/mm3, and patients with platelet counts of <195,000/mm3 had a 5.4- times higher risk of developing sICH than those with platelet counts of ≥195,000/mm3. CONCLUSION: Platelet count was the only independent parameter associated with sICH among the blood test indexes. Mild thrombocytopenia may increase the risk of sICH after intravenous administration of rtPA.

Enhancement of active compound, genipin, from Gardeniae Fructus using immobilized glycosyl hydrolase family 3 ß-glucosidase from Lactobacillus antri.

Geniposide is an iridoid glycoside, which is abundant in Gardeniae Fructus. Despite the various pharmaceutical effects of geniposide on a human body, its hydrolysis into a smaller molecule, genipin, by ß-glucosidase produced by bacteria in the intestines is particularly important to improve geniposide uptake into the body. Since geniposide is much more abundant in Gardeniae Fructus than its aglycone genipin, we herein transformed geniposide into genipin using purified recombinant ß-glucosidase from Lactobacillus antri (rBGLa), which was expressed in Escherichia coli to enhance the genipin content. Purified rBGLa was characterized using p-nitrophenyl ß-D-glucopyranoside, and the optimal temperature and pH for its ß-glucosidase activity were found to be 45 °C and 6.0. When the enzyme was immobilized, rBGLa was active at higher temperatures than the free enzyme, and we confirmed that its stability upon changes in pH and temperature was highly improved. Using 0.5 µg/mL free rBGLa, single compound of 0.4 mM geniposide was efficiently converted into genipin within 2 h, and the immobilized rBGLa also successfully transformed geniposide in a hot-water extract of Gardeniae Fructus into the aglycone, which makes it applicable to the food and pharmaceutical industries.

Artemisia capillaris Alleviates Bone Loss by Stimulating Osteoblast Mineralization and Suppressing Osteoclast Differentiation and Bone Resorption.

Artemisia capillaris has been used to treat jaundice and relieve high liver-heat in traditional medicine. In this study, we found that the administration of a water extract from A. capillaris (WEAC) to the receptor activator of nuclear factor kappa-B ligand (RANKL)-induced bone loss model significantly prevents osteoporotic bone loss, increasing bone volume/trabecular volume by 22% and trabecular number by 24%, and decreasing trabecular separation by 29%. WEAC stimulated in vitro osteoblast mineralization from primary osteoblasts in association with increasing expression of osterix, nuclear factor of activated T cells cytoplasmic 1, and activator protein-1, as well as phosphorylation of extracellular signal-regulated kinase. In contrast to the anabolic effect of WEAC, WEAC significantly suppressed in vitro osteoclast formation from bone marrow macrophages by inhibiting the RANKL signaling pathways and bone resorption by downregulating the expression of resorption markers. Therefore, this study demonstrated that WEAC has a beneficial effect on bone loss through the regulation of osteoblast mineralization, as well as osteoclast formation and bone resorption. These results suggest that A. capillaris may be a promising herbal candidate for therapeutic agents to treat or prevent osteoporotic bone diseases.

Alpinia officinarum Stimulates Osteoblast Mineralization and Inhibits Osteoclast Differentiation.

Alpinia officinarum rhizome has been used as a traditional herbal remedy to treat inflammatory and internal diseases. Based on the previously observed inhibitory effect of A. officinarum rhizome in an arthritis model, we evaluated whether a water extract of A. officinarum rhizome (WEAO) would enhance in vitro osteoblast mineralization using calvarial osteoblast precursor cells or would inhibit in vitro osteoclast differentiation and bone resorption using bone marrow derived macrophages. In osteoblasts, WEAO enhanced the mRNA levels of transcription factor (runt-related transcription factor 2, smad1, smad5, and junB) and marker (bone morphogenetic protein-2, collagen type 1alpha1, and osteocalcin) genes related to osteoblast mineralization, consistent with increased alizarin red S staining intensity. WEAO markedly inhibited osteoclast differentiation by suppressing the receptor activator for nuclear factor-[Formula: see text]B ligand-induced downregulation of inhibitor of DNA binding 2 and V-maf musculoaponeurotic fibrosarcoma oncogene homolog B and the phosphorylation of c-Jun N-terminal kinase, p38, nuclear factor-[Formula: see text]B, c-Src, and Bruton's tyrosine kinase to induce nuclear factor of activated T cells cytoplasmic 1 expression. WEAO also suppressed the resorbing activity of mature osteoclasts by altering actin ring formation. Therefore, the results of this study demonstrate that WEAO stimulates osteoblast mineralization and inhibits osteoclast differentiation. Thus, WEAO may be a promising herbal candidate to treat or prevent pathological bone diseases by regulating the balance between osteoclast and osteoblast activity.

A water extract of Malva verticillata seeds suppresses osteoclastogenesis and bone resorption stimulated by RANK ligand.

BACKGROUND: Malva verticillata seeds are used as a therapeutic medicine to treat kidney dysfunction in traditional Chinese medicine (TCM). TCM has suggested that herbal medicine tonifying kidney function may have beneficial effect on bone metabolism. METHODS: Osteoclastogenesis was examined in bone marrow macrophages by measuring tartrate-resistant acid phosphatase (TRAP) activity and counting the number of TRAP-stained multinuclear cells. The activation of receptor activator of nuclear factor-kB (RANK) ligand signaling, and the expression of c-Fos and nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1) were investigated by western blot analysis. Transcription factor and bone resorption marker mRNA levels were evaluated using real-time quantitative polymerase chain reaction. The bone resorption activity of mature osteoclast was examined in osteoclasts cultured on a hydroxyapatite-coated culture plate. RESULTS: A water extract of M. verticillata seeds (WEMV) inhibited osteoclastogenesis stimulated by RANKL. WEMV also strongly inhibited expression of c-Fos and NFATc1 as well as phosphorylation of c-Jun N-terminal kinase, p38, I-kBα, and phospholipase γ2. Furthermore, WEMV significantly attenuated osteoclast resorption activity and downregulated mRNA expression of resorption markers. CONCLUSION: These results demonstrate that WEMV inhibits osteoclastogenesis and bone resorption by suppressing the RANKL signaling pathway and suggest that M. verticillata seeds may be used as a therapeutic candidate in complementary alternative medicine to treat pathological bone diseases.

Orostachys japonicus Suppresses Osteoclast Differentiation by Inhibiting NFATc1 Expression.

The herb Orostachys japonicus has been traditionally used to treat chronic diseases, such as hepatitis, hemorrhoids, and cancer, in Asia. In this study, we investigated the effect of Orostachys japonicus water extract (OJWE) on the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and bone loss. We found that OJWE inhibited RANKL-induced osteoclast differentiation in a dose-dependent manner without affecting bone resorption in bone marrow-derived macrophage cells. Interestingly, OJWE significantly reduced serum levels of C-terminal telopeptide of type 1 collagen and tartrate-resistant acid phosphatase (TRAP) 5b, markers of bone resorption and osteoclast number, respectively, in an animal model of bone loss. Furthermore, OJWE suppressed the RANKL-induced up-regulation of nuclear factor of activated T cells cytoplasmic 1 (NFATc1) expression, and activation of the p38 signaling pathway, but prevented the RANKL-mediated down-regulation of interferon regulatory factor-8 (IRF-8), which is known to be an anti-osteoclastogenic factor that represses NFATc1 expression. We also identified gallic acid and quercetin-3-O-ß-D-glucoside as the OJWE components that inhibit RANKL-induced osteoclast differentiation. These results suggest that OJWE inhibits osteoclast differentiation by inhibiting RANKL-induced NFATc1 expression, which prevents osteoclast differentiation and bone loss. The present study elucidated a mechanism of action underlying the inhibitory effect of OJWE on osteoclast differentiation. Our findings suggest that O. japonicus has therapeutic potential for use in the treatment of bone diseases.

Water extract of Magnolia officinalis cortex inhibits osteoclastogenesis and bone resorption by downregulation of nuclear factor of activated T cells cytoplasmic 1.

BACKGROUND: Magnolia officinalis cortex has been traditionally used to treat stomach and intestine diseases in traditional Korean medicine. In this study, we investigated the effect of water extract of M. officinalis cortex (WEMC) on osteoclast differentiation and function. METHODS: Phytochemical characterization of WEMC was performed by high-performance liquid chromatography analysis. Osteoclast differentiation of bone marrow-derived macrophages was determined by tartrate-resistant acid phosphatase activity assay. Receptor activator of nuclear factor-κB ligand (RANKL) signaling factors and transcription factors regulating osteoclast differentiation were analyzed by Western blot and real-time polymerase chain reaction. Bone resorption function of mature osteoclasts was examined by using culture plate coated with inorganic crystalline calcium phosphate. Furthermore, the in vivo effect of WEMC on osteoporosis was examined using RANKL-induced bone loss model, characterized by micro-computed tomography and bone metabolism marker analysis. RESULTS: WEMC inhibited RANKL-induced osteoclast differentiation and the bone resorbing activity of mature osteoclasts. WEMC contains gallic acid and honokiol as active constituents contributing to the inhibitory effect of WEMC on osteoclast differentiation. Further, WEMC suppressed RANKL-induced activation of p38 and nuclear factor-κB pathways and expression of osteoclastogenic transcription factors such as c-Fos for AP-1 and nuclear factor of activated T cells cytoplasmic 1. Ectopic overexpression of a constitutive active form of nuclear factor of activated T cells cytoplasmic 1 rescued the antiosteoclastogenic effect of WEMC. Consistent with the in vitro results, WEMC suppressed RANKL-induced trabecular bone loss in mice. CONCLUSION: WEMC might have a therapeutic potential to treat pathological bone diseases due to increased osteoclast differentiation and function.

Water extract of the fruits of Alpinia oxyphylla inhibits osteoclast differentiation and bone loss.

BACKGROUND: Excessive bone resorption by osteoclasts causes pathological bone destruction, seen in various bone diseases. There is accumulating evidence that certain herbal extracts have beneficial effects on bone metabolism. The fruits of Alpinia oxyphylla has been traditionally used for the treatment of diarrhea and enuresis. In this study, we investigated the effects of water extract of the fruits of Alpinia oxyphylla (WEAO) on osteoclast differentiation and osteoclast-mediated bone destruction. METHODS: For osteoclast differentiation assay, mouse bone marrow-derived macrophages (BMMs) were cultured in the presence of RANKL and M-CSF. RANKL signaling pathways and gene expression of transcription factors regulating osteoclast differentiation were investigated by real-time PCR and Western blotting. A constitutively active form of NFATc1 was retrovirally transduced into BMMs. Bone resorbing activity of mature osteoclast was examined on a plate coated with an inorganic crystalline calcium phosphate. The in vivo effect against bone destruction was assessed in a murine model of RANKL-induced osteoporosis by micro-computed tomography and bone metabolism marker analyses. RESULTS: WEAO dose-dependently inhibited RANKL-induced osteoclast differentiation from BMMs by targeting the early stages of osteoclast differentiation. WEAO inhibited RANKL-induced expression of NFATc1, the master regulator of osteoclast differentiation. Overexpression of a constitutively active form of NFATc1 blunted the inhibitory effect of WEAO on osteoclast differentiation, suggesting that NFATc1 is a critical target of the inhibitory action of WEAO. WEAO inhibited RANKL-induced expression of c-Fos, an upstream activator of NFATc1, by suppressing the classical NF-κB signaling pathway. WEAO also inhibited RANKL-induced down-regulation of Id2 and MafB, negative regulators of NFATc1. WEAO does not directly affect bone resorbing activity of mature osteoclasts. In accordance with the in vitro results, WEAO attenuated RANKL-induced bone destruction in mice by inhibiting osteoclast differentiation. CONCLUSIONS: This study demonstrates that WEAO exhibits a protective effect against bone loss by inhibiting RANKL-induced osteoclast differentiation. These findings suggest that WEAO might be useful for the prevention and treatment of bone diseases associated with excessive bone resorption.

Water extract of Acer tegmentosum reduces bone destruction by inhibiting osteoclast differentiation and function.

The stem of Acer tegmentosum has been widely used in Korea for the treatment of hepatic disorders. In this study, we investigated the bone protective effect of water extract of the stem of Acer tegmentosum (WEAT). We found that WEAT inhibits osteoclast differentiation induced by receptor activator of nuclear factor-κB ligand (RANKL), an essential cytokine for osteoclast differentiation. In osteoclast precursor cells, WEAT inhibited RANKL-induced activation of JNK, NF-κB, and cAMP response element-binding protein, leading to suppression of the induction of c-Fos and nuclear factor of activated T cells cytoplasmic 1, key transcription factors for osteoclast differentiation. In addition, WEAT inhibited bone resorbing activity of mature osteoclasts. Furthermore, the oral administration of WEAT reduced RANKL-induced bone resorption and trabecular bone loss in mice. Taken together, our study demonstrates that WEAT possesses a protective effect on bone destruction by inhibiting osteoclast differentiation and function.