The proteosome inhibitor MG132 attenuates Retinoic Acid Receptor trans-activation and enhances trans-repression of Nuclear Factor κB. Potential relevance to chemo-preventive interventions with retinoids

BACKGROUND: Nuclear factor kappa B (NFκB) is a pro-malignant transcription factor with reciprocal effects on pro-metastatic and anti-metastatic gene expression. Interestingly, NFκB blockade results in the reciprocal induction of retinoic acid receptors (RARs). Given the established property of RARs as negative regulators of malignant progression, we postulated that reciprocal interactions between NFκB and RARs constitute a signaling module in metastatic gene expression and malignant progression. Using Line 1 tumor cells as a model for signal regulation of metastatic gene expression, we investigated the reciprocal interactions between NFκB and RARs in response to the pan-RAR agonist, all-trans retinoic acid (at-RA) and the pan-RAR antagonist, AGN193109. RESULTS: At-RA [0.1–1 μM] dose-dependently activated RAR and coordinately trans-repressed NFκB, while AGN193109 [1–10 μM] dose-dependently antagonized the effects of at-RA. At-RA and AGN193109 reciprocally regulate pro-metastatic matrix metalloprotease 9 (MMP 9) and its endogenous inhibitor, the tissue inhibitor of metalloprotease 1 (TIMP 1), in a manner consistent with the putative roles of NFκB and RAR in malignant progression. Activation of RAR concurs with its ubiquitination and proteosomal degradation. Accordingly, the proteosome inhibitor, MG132 [5 μM], blocked RAR degradation, quelled RAR trans-activation and enhanced RAR trans-repression of NFκB. CONCLUSION: We conclude that reciprocal interactions between NFκB and RARs constitute a signaling module in metastatic gene expression and malignant progression and propose that the dissociative effect of proteosome inhibitors could be harnessed towards enhancing the anticancer activity of retinoids

The Association between Environmental Lead Exposure and Bone Density in Children

Osteoporosis is a decrease in bone mineral density (BMD) that predisposes individuals to fractures. Although an elderly affliction, a predisposition may develop during adolescence if a sufficient peak BMD is not achieved. Rat studies have found that lead exposure is associated with decreased BMD. However, human studies are limited. We hypothesized that the BMD of children with high lead exposure would be lower than the BMD of children with low lead exposure. We collected data on 35 subjects; 16 had low cumulative lead exposure (mean, 6.5 μg/dL), and 19 had high exposure (mean, 23.6 μg/dL). All were African American; there was no difference between the groups by sex, age, body mass index, socioeconomic status, physical activity, or calcium intake. Significant differences in BMD between low and high cumulative lead exposure were noted in the head (1.589 vs. 1.721 g/cm(2)), third lumbar vertebra (0.761 vs. 0.819 g/cm(2)), and fourth lumbar vertebra (0.712 vs. 0.789 g/cm(2)). Contrary to our hypothesis, subjects with high lead exposure had a significantly higher BMD than did subjects with low lead exposure. This may reflect a true phenomenon because lead exposure has been reported to accelerate bony maturation by inhibiting the effects of parathyroid hormone–related peptide. Accelerated maturation of bone may ultimately result in a lower peak BMD being achieved in young adulthood, thus predisposing to osteoporosis in later life. Future studies need to investigate this proposed model

Lead Exposure Inhibits Fracture Healing and Is Associated with Increased Chondrogenesis, Delay in Cartilage Mineralization, and a Decrease in Osteoprogenitor Frequency

Lead exposure continues to be a significant public health problem. In addition to acute toxicity, Pb has an extremely long half-life in bone. Individuals with past exposure develop increased blood Pb levels during periods of high bone turnover or resorption. Pb is known to affect osteoblasts, osteoclasts, and chondrocytes and has been associated with osteoporosis. However, its effects on skeletal repair have not been studied. We exposed C57/B6 mice to various concentrations of Pb acetate in their drinking water to achieve environmentally relevant blood Pb levels, measured by atomic absorption. After exposure for 6 weeks, each mouse underwent closed tibia fracture. Radiographs were followed and histologic analysis was performed at 7, 14, and 21 days. In mice exposed to low Pb concentrations, fracture healing was characterized by a delay in bridging cartilage formation, decreased collagen type II and type X expression at 7 days, a 5-fold increase in cartilage formation at day 14 associated with delayed maturation and calcification, and a persistence of cartilage at day 21. Fibrous nonunions at 21 days were prevalent in mice receiving very high Pb exposures. Pb significantly inhibited ex vivo bone nodule formation but had no effect on osteoclasts isolated from Pb-exposed animals. No significant effects on osteoclast number or activity were observed. We conclude that Pb delays fracture healing at environmentally relevant doses and induces fibrous nonunions at higher doses by inhibiting the progression of endochondral ossification

Cyclooxygenase-2 regulates mesenchymal cell differentiation into the osteoblast lineage and is critically involved in bone repair

Preclinical and clinical studies suggest a possible role for cyclooxygenases in bone repair and create concerns about the use of nonsteroidal antiinflammatory drugs in patients with skeletal injury. We utilized wild-type, COX-1(–/–), and COX-2(–/–) mice to demonstrate that COX-2 plays an essential role in both endochondral and intramembranous bone formation during skeletal repair. The healing of stabilized tibia fractures was significantly delayed in COX-2(–/–) mice compared with COX-1(–/–) and wild-type controls. The histology was characterized by a persistence of undifferentiated mesenchyme and a marked reduction in osteoblastogenesis that resulted in a high incidence of fibrous nonunion in the COX-2(–/–) mice. Similarly, intramembranous bone formation on the calvaria was reduced 60% in COX-2(–/–) mice following in vivo injection of FGF-1 compared with either COX-1(–/–) or wild-type mice. To elucidate the mechanism involved in reduced bone formation, osteoblastogenesis was studied in bone marrow stromal cell cultures obtained from COX-2(–/–) and wild-type mice. Bone nodule formation was reduced 50% in COX-2(–/–) mice. The defect in osteogenesis was completely rescued by addition of prostaglandin E2 (PGE(2)) to the cultures. In the presence of bone morphogenetic protein (BMP-2), bone nodule formation was enhanced to a similar level above that observed with PGE(2) alone in both control and COX-2(–/–) cultures, indicating that BMPs complement COX-2 deficiency and are downstream of prostaglandins. Furthermore, we found that the defect in COX-2(–/–) cultures correlated with significantly reduced levels of cbfa1 and osterix, two genes necessary for bone formation. Addition of PGE(2) rescued this defect, while BMP-2 enhanced cbfa1 and osterix in both COX-2(–/–) and wild-type cultures. Finally, the effects of these agents were additive, indicating that COX-2 is involved in maximal induction of osteogenesis. These results provide a model whereby COX-2 regulates the induction of cbfa1 and osterix to mediate normal skeletal repair

Synergism of aminobisphosphonates and farnesyl transferase inhibitors on tumor metastasis

Aminobisphosphonates have shown significant antitumor activity in vitro and in vivo with selective pharmacodistribution to bone, and an established role in the treatment of malignant bone disease. Given that the mode of action of aminobisphosphonates involves decreasing the prenylation of the Rho family of proteins, through decreasing the availability of prenyl groups (farnesyl and geranylgeranyl isoprenoids), the authors sought the inhibition of Rho protein prenylation at two points, by using an aminobiphosphonate (alendronate) in conjunction with a prenyl transferase inhibitor (R115777, a specific farnesyl transferase inhibitor with limited effects in geranylgeranyl transferase). The authors show synergistic inhibition of the prenylation dependent membrane association and migratory function of Rho proteins, translating into a suppressive effect on in vitro tumor cell invasiveness and in vivo metastasis. The findings support the use of aminobisphosphonates in conjunction with farnesyl transferase inhibitors in the prevention of metastatic progression and suggest that metastatic progression is a valid end point in assessing the antitumor activity of farnesyl transferase inhibitors