We recently reported that extracts made from the stem bark of Ulmus wallichiana promoted peak bone mass achievement in growing rats and preserved trabecular bone mass and cortical bone strength in ovariectomized (OVX) rats. Further, 6-C-b-D-glucopyranosyl-(2S,3S)-(þ)-3',4',5,7-tetrahydroxyflavanol (GTDF), a novel flavonol-C-glucoside isolated from the extracts, had a nonestrogenic bonesparing effect on OVX rats. Here we studied the effects of GTDF on osteoblast function and its mode of action and in vivo osteogenic effect. GTDF stimulated osteoblast proliferation, survival, and differentiation but had no effect on osteoclastic or adipocytic differentiation. In cultured osteoblasts, GTDF transactivated the aryl hydrocarbon receptor (AhR). Activation of AhR mediated the stimulatory effect of GTDF on osteoblast proliferation and differentiation. Furthermore, GTDF stimulated cAMP production, which mediated osteogenic gene expression. GTDF treatments given to 1-to 2-day-old rats or adult rats increased the mRNA levels of AhR target genes in calvaria or bone marrow stromal cells. In growing female rats, GTDF promoted parameters of peak bone accrual in the appendicular skeleton, including increased longitudinal growth, bone mineral density, bone-formation rate (BFR), cortical deposition, and bone strength. GTDF promoted the process of providing newly generated bone to fill drill holes in the femurs of both estrogen-sufficient and -deficient rats. In osteopenic OVX rats, GTDF increased BFR and significantly restored trabecular bone compared with the ovaries-intact group. Together our data suggest that GTDF stimulates osteoblast growth and differentiation via the AhR and promotes modeling-directed bone accrual, accelerates bone healing after injury, and exerts anabolic effects on osteopenic rats likely by a direct stimulatory effect on osteoprogenitors. Based on these preclinical data, clinical evaluation of GTDF as a potential bone anabolic agent is warranted. ß

Diabetes is a major risk factor of stroke and is associated with increased frequency of stroke and a poorer prognosis for recovery. In earlier studies we have utilized Type 2 diabetic mouse models of stroke and demonstrated that diabetic db/db and ob/ob mice experience larger infarct volumes and impaired recovery associated with greater infiltration of macrophage following hypoxic-ischemic (H/I) insult than their heterozygous non-diabetic db/+ & ob/+ littermates. To obtain a better understanding of the pathogenesis of the impaired recovery, we have investigated the role of matrix metalloproteases and their endogenous inhibitors in the breakdown of the blood-brain barrier (BBB) following H/I. Diabetic db/db mice showed a significant and more rapid increase in MMP-9 mRNA, protein and gelatinolytic activity compared to db/+, which resulted in an increased degradation of occludin & collagen IV and subsequently an increased BBB permeability and greater infiltration of neutrophils into the infarct area. The expression of the MMPs, especially in the db/+ mice, is preceded by an elevated expression of their endogenous tissue inhibitors of metalloproteases (TIMPs) 1, 2, & 3, while in the db/db mice, a lower expression of the TIMPs is associated with greater MMP 3 & 9 expression. These results suggest that an imbalance in the MMPs/TIMPs cascade in the diabetic mouse, particularly MMP-9, results in a greater neutrophil invasion, a compromised BBB and consequently a greater insult.

Impaired peripheral wound healing is a hallmark of diabetis pathology and has been attributed to compromised macrophage activation. Stroke is another component of diabetic pathology, with increased tissue infarction and worsened recovery although the mechanisms remain unresolved. In this study, we investigated whether a compromised glial/macrophage response might contribute to cerebral hypoxic-ischemic (H/I) brain damage in diabetic (db/db), relative to their normoglycemic db/ + mice. Hypoxia-ischemia was induced in 8-week-old male db/db and db/ + mice by the ligation of right common carotid artery followed by systemic hypoxia (8% O 2 : 92% N 2 ) for 17 mins. Mice were killed at specific intervals of reperfusion/recovery and the brains analyzed by in situ hybridization or total RNA isolation. In situ hybridization using bfl-1 (microglia) and glial fibrillary acidic protein (GFAP) (astrocytes) revealed expression of both bfl-1 and GFAP in the ipsilateral hemisphere at 4 h in the db/ + mice, which was delayed and minimal in the db/db mice. RNase protection assays showed a robust increase in expression of the proinflamatory cytokines tumor necrosis factor-a (TNFa), interleukin-1 IL-1a, and IL-1b mRNA in the db/ + mice at 6 to 8 h of reperfusion peaking at 8 to 12 h; in db/db mice expression was markedly delayed and diminished. Real-time-polymerase chain reaction (RT-PCR) confirmed the reduced and delayed expression TNFa, IL-1a, IL-1b, and the growth factors insulin-like growth factor-1 and ciliary neurotrophic factor in the db/db mice; enzyme-linked immunosorbent assays confirmed the reduced and delayed translation of IL-1b protein. These findings suggest that a compromised inflammatory response may underlie the greater infarct associated with diabetic db/db mice compared with their nondiabetic littermates following a hypoxic/ischemic insult.

Two experiments were conducted to study the effect of supplementing organic forms of zinc (Zn), selenium (Se) and chromium (Cr) on performance, anti-oxidant activities and immune responses in broiler chickens from 1 to 21 days of age, which were reared in cyclic heat-stressed condition under tropical summer in open-sided poultry house. A total of 200 (experiment I) and 450-day-old (experiment II) broiler male chicks (Cobb 400) were randomly distributed in stainless steel battery brooders (610 mm × 762 mm × 475 mm) at the rate of five birds per pen. A maize-soybean meal-based control diet (CD) containing recommended (Vencobb 400, Broiler Management Guide) concentrations of inorganic trace minerals and other nutrients was prepared. The CD was supplemented individually with organic form of selenium (Se, 0.30 mg/kg), chromium (Cr, 2 mg/kg) and zinc (Zn, 40 mg/kg) in experiment I. In experiment II, two concentrations of each Zn (20 and 40 mg/kg), Se (0.15 and 0.30 mg/kg) and Cr (1 and 2 mg/kg) were supplemented to the basal diet in 2 × 2 × 2 factorial design. A group without supplementing inorganic trace minerals was maintained as control group in both experiments. Each diet was allotted randomly to ten replicates in both experiments and fed ad libitum from 1 to 21 days of age. At 19th day of age, blood samples were collected for estimation of anti-oxidant and immune responses. Supplementation of Se, Cr and Zn increased (P < 0.05) body mass gain (BMG) and feed intake compared to those fed the CD in experiment I. The feed efficiency (FE) in Cr-fed group was higher (P < 0.05) compared to the CD-fed group. Se or Cr supplementation reduced lipid peroxidation (LP) compared to broilers fed the CD. In experiment II, BMG was not affected (P > 0.05) by the interaction between levels of Zn, Se and Cr in broiler diet. The FE improved (P < 0.05) with supplementation of the trace minerals tested at both concentrations except in group fed 40 mg Zn, 0.5 mg Se and 1 mg Cr/kg. Reduction in lipid peroxidation (LP, P < 0.05) and increased (P < 0.05) activity of superoxide dismutase were observed in broiler fed organic Zn, Se and Cr compared to the CD-fed group. The dietary concentrations of Zn, Se and Cr did not influence (P > 0.05) the immune responses (Newcastle disease titre and cell-mediated immune response to phytohaemagglutinin-P) in both the experiments. Based on the results, it is concluded that supplementation of organic form of Se, Cr and Zn (0.30, 2 and 40 mg/kg, respectively) either alone or in combination significantly improved performance and anti-oxidant responses (reduced LP and increased superoxide dismutase) in commercial broiler chicks (21 days of age) reared in cyclic heat stress conditions in open-sided poultry house during summer.