Formononetin, an isoflavone, activates AMP-activated protein kinase/ß-catenin signalling to inhibit adipogenesis and rescues C57BL/6 mice from high-fat diet-induced obesity and bone loss.

Balance between adipocyte and osteoblast differentiation is the key link of disease progression in obesity and osteoporosis. We have previously reported that formononetin (FNT), an isoflavone extracted from Butea monosperma, stimulates osteoblast formation and protects against postmenopausal bone loss. The inverse relationship between osteoblasts and adipocytes prompted us to analyse the effect of FNT on adipogenesis and in vivo bone loss, triggered by high-fat diet (HFD)-induced obesity. The anti-obesity effect and mechanism of action of FNT was determined in 3T3-L1 cells and HFD-induced obese male mice. Our findings show that FNT suppresses the adipogenic differentiation of 3T3-L1 fibroblasts, through down-regulation of key adipogenic markers such as PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα) and sterol regulatory element-binding protein (SREBP) and inhibits intracellular TAG accumulation. Increased intracellular reactive oxygen species levels and AMP-activated protein kinase (AMPK) activation accompanied by stabilisation of ß-catenin were attributed to the anti-adipogenic action of FNT. In vivo, 12 weeks of FNT treatment inhibited the development of obesity in mice by attenuating HFD-induced body weight gain and visceral fat accumulation. The anti-obesity effect of FNT results from increased energy expenditure. FNT also protects against HFD-induced dyslipidaemia and rescues deterioration of trabecular bone volume by increasing bone formation and decreasing bone resorbtion caused by HFD. FNT's rescuing action against obesity-induced osteoporosis commenced at the level of progenitors, as bone marrow progenitor cells, obtained from the HFD mice group supplemented with FNT, showed increased osteogenic and decreased adipogenic potentials. Our findings suggest that FNT inhibits adipogenesis through AMPK/ß-catenin signal transduction pathways and protects against HFD-induced obesity and bone loss.

Constituents of Dalbergia sissoo Roxb. leaves with osteogenic activity.

One new isoflavone glucoside, caviunin 7-O-[ß-d-apiofuranosyl-(1→6)-ß-d-glucopyranoside] (10) and a new itaconic derivative, (E)-4-methoxy-2-(3,4-dihydroxybenzylidene)-4-oxobutanoic acid (15) along with series of isoflavones and flavonols with their glucosides (1-9 and 11-14) and a lignan glucoside (16) were isolated from the ethanolic extract of Dalbergia sissoo leaves. The structures of these compounds were established on the basis of IR, UV, (1)H and (13)C NMR, DEPT, COSY, HSQC, HMBC and MS data. All compounds (1-16) were assessed for osteogenic activity in primary calvarial osteoblast cultures. Compounds 1-4 and 10 increased alkaline phosphatase activity and mineralization thus resulting in significant osteogenic activity.

Osteogenic constituents from Pterospermum acerifolium Willd. flowers.

Phytochemical investigation of ethanol extracts of the Pterospermum acerifolium flowers led to the isolation and identification of two new flavones, 4'-(2-methoxy-4-(1,2,3-trihydroxypropyl) phenoxy luteolin (1) and 5,7,3'-trihydroxy-6-O-ß-D-glucopyranosyl flavone (2), and one new lactone, 3,5-dihydroxyfuran-2(5H)-one (3) along with 14 known compounds (4-17). The structure of compounds 1-17 was established based on MS, 1D and 2D NMR, spectroscopic analysis. Eight of these compounds (1-6, 8 and 9) were assessed for osteogenic activity by using primary cultures of rat osteoblast. The compounds 1, 3 and 4 significantly stimulated osteoblast differentiation and mineralization as evident from a marked increase in expression of alkaline phosphatase and alizarin red-S staining of osteoblasts.

(6)-Gingerolinduced myeloid leukemia cell death is initiated by reactive oxygen species and activation of miR-27b expression.

The natural polyphenolic alkanone (6)-gingerol (6G) has established anti-inflammatory and antitumoral properties. However, its precise mechanism of action in myeloid leukemia cells is unclear. In this study, we investigated the effects of 6G on myeloid leukemia cells in vitro and in vivo. The results of this study showed that 6G inhibited proliferation of myeloid leukemia cell lines and primary myeloid leukemia cells while sparing the normal peripheral blood mononuclear cells, in a concentration- and time-dependent manner. Mechanistic studies using U937 and K562 cell lines revealed that 6G treatment induced reactive oxygen species (ROS) generation by inhibiting mitochondrial respiratory complex I (MRC I), which in turn increased the expression of the oxidative stress response-associated microRNA miR-27b and DNA damage. Elevated miR-27b expression inhibited PPARγ, with subsequent inhibition of the inflammatory cytokine gene expression associated with the oncogenic NF-κB pathway, whereas the increased DNA damage led to G2/M cell cycle arrest. The 6G induced effects were abolished in the presence of anti-miR-27b or the ROS scavenger N-acetylcysteine. In addition, the results of the in vivo xenograft experiments in mice indicated that 6G treatment inhibited tumor cell proliferation and induced apoptosis, in agreement with the in vitro studies. Our data provide new evidence that 6G-induced myeloid leukemia cell death is initiated by reactive oxygen species and mediated through an increase in miR-27b expression and DNA damage. The dual induction of increased miR-27b expression and DNA damage-associated cell cycle arrest by 6G may have implications for myeloid leukemia treatment.

N-Methyl-6, 7-dimethoxyisoquinolone in Annona squamosa twigs is the major immune modifier to elicit polarized Th1 immune response in BALB/c mice.

Annona squamosa (AS) has traditionally been used as ethnomedicine. We have earlier extracted and fractionated the twigs of AS based upon its bioactivity and observed its immune potentiating activity that was localized in its three fractions. Present communication deals with the phytochemical analysis and pharmacological investigation of the most active chloroform fraction that led to isolation and identification of a number of compounds whose structures were elucidated using 1D and 2D NMR spectroscopic analysis. Amongst the twelve pure compounds isolated, five compounds Lanuginosine (1), (+)-O-methylarmepavine (2), (+)-anomuricine (3), Isocorydine (4), and N-methyl-6, 7-dimethoxyisoquinolone (5) were evaluated in vivo for their immune modifier activities in BALB/c mice after oral administration at three log doses of 0.3, 1.0 and 3.0mg/kg for 14 consecutive days. Of these, three compounds (1, 2 and 5) showed dose dependent immune stimulating activity. However, the uppermost activity was noted in the compound N-methyl-6, 7-dimethoxyisoquinolone at the 3.0mg/kg oral dose. The activity was assessed in the form of increased splenic T and B cellular proliferation, up-regulated CD4+, CD8+ and CD19+ cell population and accentuation in the peritoneal macrophage function. The compound possibly acted modifying the expression of Th1- and Th2- cytokines via stimulation of pro-inflammatory Th1 cytokines IL-2 and IFN-γ. These results warrant the use of the above compounds as an efficient immune-stimulant or immune-adjuvant against diseases with immune suppression. The analogs of the compound may further be chemically synthesized to achieve desired immune modifying activity.

Chemical constituents of Kigelia pinnata twigs and their GLUT4 translocation modulatory effect in skeletal muscle cells.

Phytochemical investigation of the ethanolic extract of twigs of Kigelia pinnata DC. afforded one new iridoid 7-hydroxy eucommiol (1), and nine known compounds (2-10). The structure of compounds was elucidated by extensive spectroscopic methods, including 1D, 2D NMR experiments and MS analysis. All these compounds were evaluated for GLUT4 translocation modulatory effect in skeletal muscle cells. Four of the tested compounds 1, 5, 6 and 7 showed significant stimulation of GLUT4 translocation to cell surface in skeletal muscle cells without any adverse effect on cell viability. Effect of these four compounds was concentration-dependent and comparable to standard drug rosiglitazone. These findings indicate that constituents of K. pinnata may provide leads for the therapeutics for insulin resistance and diabetes.

[6]-Gingerol induces bone loss in ovary intact adult mice and augments osteoclast function via the transient receptor potential vanilloid 1 channel.

SCOPE: [6]-Gingerol, a major constituent of ginger, is considered to have several health beneficial effects. The effect of 6-gingerol on bone cells and skeleton of mice was investigated. METHODS AND RESULTS: The effects of 6-gingerol on mouse bone marrow macrophages and osteoblasts were studied. 6-Gingerol-stimulated osteoclast differentiation of bone marrow macrophages but had no effect on osteoblasts. Capsazepine, an inhibitor of TRPV1 (transient receptor potential vanilloid 1) channel, attenuated the pro-osteoclastogenic effect of 6-gingerol or capsaicin (an agonist of TRPV1). Similar to capsaicin, 6-gingerol stimulated Ca(2) + influx in osteoclasts. The effect of daily feeding of 6-gingerol for 5 wk on the skeleton of adult female Balb/cByJ mice was investigated. Mice treated with capsaicin and ovariectomized (OVx) mice served as controls for osteopenia. 6-Gingerol caused increase in trabecular osteoclast number, microarchitectural erosion at all trabecular sites and loss of vertebral stiffness, and these effects were comparable to capsaicin or OVx group. Osteoclast-specific serum and gene markers of 6-gingerol-treated mice were higher than the OVx group. Bone formation was unaffected by 6-gingerol. CONCLUSION: Daily feeding of 6-gingerol to skeletally mature female mice caused trabecular osteopenia, and the mechanism appeared to be activation of osteoclast formation via the TRPV1 channel.

Phytoceramides and acylated phytosterol glucosides from Pterospermum acerifolium Willd. seed coat and their osteogenic activity.

Phytochemical investigation of seed coats of Pterospermum acerifolium afforded two phytoceramides (1, 2) and two acylated phytosterol glucosides (3, 4) together with five known compounds (5-9). Their structures were elucidated on the basis of extensive spectroscopic analysis using 1D, 2D NMR and Mass spectrometry. Compounds 1, 2, 3, and 4 were assessed for their osteogenic activity using primary cultures of osteoblasts harvested from neonatal rat calvaria. Among these compounds, 1 and 2 markedly stimulated osteoblast differentiation assessed by alkaline phosphatase production and osteoblast mineralization by alizarin red-S staining.

A standardized phytopreparation from an Indian medicinal plant (Dalbergia sissoo) has antiresorptive and bone-forming effects on a postmenopausal osteoporosis model of rat.

OBJECTIVE: The aim of this study was to evaluate the skeletal effects of an extract made from the leaves and pods of Dalbergia sissoo (butanol-soluble standardized fraction [BSSF]) on ovariectomized rats, a model for postmenopausal osteopenia. METHODS: Adult Sprague-Dawley rats were ovariectomized and administered BSSF (50 and 100 mg/kg per day) or 17ß-estradiol orally for 12 weeks. The sham-operated group and the ovariectomy + vehicle group served as controls. Bone microarchitecture, bone turnover markers (serum osteocalcin and C-telopeptide fragment of collagen type I), biomechanical strength, new bone formation (based on mineral apposition rate and bone formation rate), and skeletal expressions of osteogenic and resorptive gene markers were studied. Uterine histomorphometry was used to assess estrogenicity. Bioactive marker compounds in BSSF were analyzed by high-performance liquid chromatography. One-way analysis of variance was used to test the significance of effects. RESULTS: In comparison with ovariectomized rats treated with vehicle, BSSF treatment in ovariectomized rats resulted in an improved trabecular microarchitecture of the long bones, increased biomechanical strength parameters of the vertebra and femur, decreased bone turnover markers (osteocalcin and type I collagen) and expression of skeletal osteoclastogenic genes, and increased new bone formation and expression of osteogenic genes in the femur. Overall, the osteoprotective effects of BSSF were comparable to those of 17ß-estradiol. BSSF did not exhibit uterine estrogenicity. Analysis of marker compounds revealed the presence of osteogenic methoxyisoflavones, including caviunin 7-O-[ß-D-apiofuranosyl-(1→6)-ß-D-glucopyranoside] (a novel compound), biochanin A, and pratensin. CONCLUSIONS: Oral doses of BSSF in the preclinical setting are effective in preventing estrogen deficiency-induced bone loss by dual action: inhibition of bone resorption and stimulation of new bone formation.

Constituents of Tinospora sinensis and their antileishmanial activity against Leishmania donovani.

Two new compounds 4-methyl-heptadec-6-enoic acid ethyl ester (2) and 3-hydroxy-2,9,11-trimethoxy-5,6-dihydro isoquino[3,2-a]isoquinolinylium (7) were isolated from an ethanolic extract of the stems of Tinospora sinensis, along with six known compounds (1, 3-6 and 8). The structures of new compounds were established on the basis of detailed spectroscopic studies. Compound 7 exhibited the highest in vitro antileishmanial activity against Leishmania donovani promastigotes and intracellular amastigotes, whereas compounds 2, 4, 5 and 6 demonstrated moderate activity. Other compounds were found to be inactive.