Vectorization of morpholino oligomers by the (R-Ahx-R)4 peptide allows efficient splicing correction in the absence of endosomolytic agents.

The efficient and non-toxic nuclear delivery of steric-block oligonucleotides (ON) is a prerequisite for therapeutic strategies involving splice correction or exon skipping. Cationic cell penetrating peptides (CPPs) have given rise to much interest for the intracellular delivery of biomolecules, but their efficiency in promoting cytoplasmic or nuclear delivery of oligonucleotides has been hampered by endocytic sequestration and subsequent degradation of most internalized material in endocytic compartments. In the present study, we compared the splice correction activity of three different CPPs conjugated to PMO(705), a steric-block ON targeted against the mutated splicing site of human beta-globin pre-mRNA in the HeLa pLuc705 splice correction model. In contrast to Tat48-60 (Tat) and oligoarginine (R(9)F(2)) PMO(705) conjugates, the 6-aminohexanoic-spaced oligoarginine (R-Ahx-R)(4)-PMO(705) conjugate was able to promote an efficient splice correction in the absence of endosomolytic agents. Our mechanistic investigations about its uptake mechanisms lead to the conclusion that these three vectors are internalized using the same endocytic route involving proteoglycans, but that the (R-Ahx-R)(4)-PMO(705) conjugate has the unique ability to escape from lysosomial fate and to access to the nuclear compartment. This vector, which has displays an extremely low cytotoxicity, the ability to function without chloroquine adjunction and in the presence of serum proteins. It thus offers a promising lead for the development of vectors able to enhance the delivery of therapeutic steric-block ON in clinically relevant models.     

Thyroid-stimulating hormone restores bone volume, microarchitecture, and strength in aged ovariectomized rats.

We show the systemic administration of low levels of TSH increases bone volume and improves bone microarchitecture and strength in aged OVX rats. TSH's actions are mediated by its inhibitory effects on RANKL-induced osteoclast formation and bone resorption coupled with stimulatory effects on osteoblast differentiation and bone formation, suggesting TSH directly affects bone remodeling in vivo. INTRODUCTION: Thyroid-stimulating hormone (TSH) receptor haploinsufficient mice with normal circulating thyroid hormone levels have reduced bone mass, suggesting that TSH directly affects bone remodeling. We examined whether systemic TSH administration restored bone volume in aged ovariectomized (OVX) rats and influenced osteoclast formation and osteoblast differentiation in vitro. MATERIALS AND METHODS: Sprague-Dawley rats were OVX at 6 months, and TSH therapy was started immediately after surgery (prevention mode; n = 80) or 7 mo later (restoration mode; n = 152). Hind limbs and lumbar spine BMD was measured at 2- or 4-wk intervals in vivo and ex vivo on termination at 8-16 wk. Long bones were subjected to microCT, histomorphometric, and biomechanical analyses. The direct effect of TSH was examined in osteoclast and osteoblast progenitor cultures and established rat osteosarcoma-derived osteoblastic cells. Data were analyzed by ANOVA Dunnett test. RESULTS: In the prevention mode, low doses (0.1 and 0.3 microg) of native rat TSH prevented the progressive bone loss, and importantly, did not increase serum triiodothyroxine (T3) and thyroxine (T4) levels in aged OVX rats. In restoration mode, animals receiving 0.1 and 0.3 microg TSH had increased BMD (10-11%), trabecular bone volume (100-130%), trabecular number (25-40%), trabecular thickness (45-60%), cortical thickness (5-16%), mineral apposition and bone formation rate (200-300%), and enhanced mechanical strength of the femur (51-60%) compared with control OVX rats. In vitro studies suggest that TSH's action is mediated by its inhibitory effects on RANKL-induced osteoclast formation, as shown in hematopoietic stem cells cultivated from TSH-treated OVX rats. TSH also stimulates osteoblast differentiation, as shown by effects on alkaline phosphatase activity, osteocalcin expression, and mineralization rate. CONCLUSIONS: These results show for the first time that systemically administered TSH prevents bone loss and restores bone mass in aged OVX rats through both antiresorptive and anabolic effects on bone remodeling.     

Clinical risk factors for fractures in multi-ethnic women: the Women's Health Initiative.

To identify risk factors for fractures in multi-ethnic women, we studied 159,579 women enrolled in the Women's Health Initiative. In general, risk factors for fractures were similar across ethnic groups. However, irrespective of their ethnicity, women with multiple risk factors have a high risk of fracture. Targeting these high-risk women for screening and intervention could reduce fractures. INTRODUCTION: Fracture rates tend to be lower in minority women, but consequences may be greater. In addition, the number of fractures is expected to increase in minority women because of current demographic trends. There are limited prospective data on risk factors for fractures in minority women. MATERIALS AND METHODS: We studied 159,579 women 50-79 yr of age enrolled in the Women's Health Initiative. Information on risk factors was obtained by questionnaire or examination. Nonspine fractures that occurred after study entry were identified over an average follow-up of 8 +/- 2.6 (SD) yr. RESULTS: Annualized rates (%) of fracture in whites, blacks, Hispanics, Asians, and American Indians were 2.0, 0.9, 1.3, 1.2, and 2.0, respectively. Significant predictors [HR (95% CI)] of fractures by ethnic group were as follows: blacks: at least a high school education, 1.22 (1.0, 1.5); (+) fracture history, 1.7 (1.4, 2.2); and more than two falls, 1.7 (1.9, 2.0); Hispanics: height (>162 cm), 1.6 (1.1, 2.2); (+) fracture history, 1.9 (1.4, 2.5); more than two falls, 1.8 (1.4, 2.3); arthritis, 1.3 (1.1, 1.6); corticosteroid use, 3.9 (1.9, 8.0); and parental history of fracture, 1.3 (1.0, 1.6); Asians: age (per 5 yr), 1.2 (1.0, 1.3); (+) fracture history, 1.5 (1.1, 2.0); current hormone therapy (HT), 0.7 (0.5, 0.8); parity (at least five), 1.8 (1.1, 3.0); more than two falls, 1.4 (1.1, 1.9); American Indian: (+) fracture history, 2. 9 (1.5, 5.7); current HT, 0.5 (0.3, 0.9). Women with eight or more risk factors had more than a 2-fold higher rate of fracture compared with women with four or fewer risk factors. Two ethnicity x risk factor interactions were identified: age and fall history. CONCLUSIONS: Irrespective of their ethnicity, women with multiple risk factors have a high risk of fracture. Targeting these high-risk women for screening and intervention could reduce fractures.     

Predictors of Partner Notification for C. trachomatis and N. gonorrhoeae: An Examination of Social Cognitive and Psychological Factors

Efforts to control chlamydial and gonococcal infections include notifying eligible sexual partners of possible infection, primarily by asking the diagnosed patient to notify their partners. This approach, known as patient referral, is widely used but poorly understood. The current study examined psychosocial and cognitive factors associated with patient referral among an urban, minority sample of 168 participants recently diagnosed with Chlamydia trachomatis or Neisseria gonorrhoeae. At a follow-up interview 1-month from diagnosis, participants were more likely to have notified all eligible partners if they had greater intention to notify at baseline (OR = 3.72; 95% CI = 1.34, 10.30) and if they had only one partner at baseline (OR = 4.08; 95% CI = 1.61, 10.31). There were also gender differences as well as differences based on type of partner (i.e., regular, casual, one-time). The implications of these findings for the design of programs to promote patient referral for sexually transmitted infections are discussed. Schwartz, Malka, Augenbraun, McCormack, and Wilson are with the State University of New York, Downstate Medical Center, Brooklyn, NY, USA; Rubin is with the New York City Department of Health, Bureau of STD Control, New York, NY, USA; Rubin, Hogben, and Liddon are with the Centers for Disease Control and Prevention, Atlanta, GA, USA; Schwartz is with the Department of Preventive Medicine and Community Health, SUNY Downstate Medical Center, Box 1240, 450 Clarkson Avenue, Brooklyn, NY 11203, USA.