Differential susceptibility of primary and established human glioma cells to adenovirus infection: targeting via the epidermal growth factor receptor achieves fiber receptor-independent gene transfer

Adenovirus (Ad) vectors are promising for gene therapy of glioma due to their ability to achieve efficient gene transfer upon intratumoral administration. Yet in this context, Ad mediates widespread gene transfer to both tumor and surrounding parenchyma. Ad entry is dependent upon the expression of fiber receptors, such as coxsackie/adenovirus receptor, and alpha(v) integrins on the target cells for binding and internalization, respectively. We hypothesized that the susceptibility of human gliomas to Ad would likely be heterogeneous due to variable expression of these receptors. It was found that established human glioma cell lines exhibited differential susceptibility to Ad-mediated gene transfer, which correlated directly with the level of radiolabeled Ad binding and with the expression of coxsackie/adenovirus receptor but not with the expression of alpha(v) integrins. To circumvent the lack of fiber receptors and to target Ad gene transfer specifically to tumor cells, we used a bispecific antibody conjugate to ablate Ad binding to fiber receptors and retarget binding to the epidermal growth factor receptor (EGFR), a tumor-associated marker negligibly expressed in normal, mitotically quiescent neural tissues. The results demonstrate that EGFR-targeted Ad gene transfer was EGFR specific and independent of fiber-fiber receptor interactions. Furthermore, EGFR targeting significantly enhanced Ad gene delivery to 7 of 12 established glioma cell lines and to 6 of 8 cultured primary gliomas. Interestingly, EGFR-targeted Ad gene transfer did not correlate with EGFR expression across cell lines, suggesting the importance of other factors. This study establishes that fiber receptor expression limits the utility of Ad vectors for gene transfer to glioma cells and suggests that targeting Ad via EGFR may prove valuable for tumor-specific gene transfer to high-grade gliomas. These findings have key relevance in the context of Ad vector-based approaches for glioma gene therapy.     

Declining sperm counts in the United States? A critical review

PURPOSE: Recent reports suggest declining sperm counts in the United States. These reports did not include all available data and did not account for geographic variations noted in prior studies. We examined all available data on U.S. sperm counts and evaluated whether geographic variations account for the decline suggested. MATERIALS AND METHODS: We reviewed all 29 U.S. studies from 1938 to 1996 reporting manually counted semen analyses of 9,612 fertile or presumably fertile men. We determined mean sperm concentrations by geographic location with weighted analysis of variance, and assessed any changes with time by linear regression analysis. RESULTS: Mean sperm concentrations from New York were significantly higher than from all other U.S. cities (98.6 versus 71.6 x 10(6) sperm per cc, respectively, p = 0.006). There has been no statistically significant change with time for mean sperm concentrations reported from New York (p = 0.49) or from U.S. cities other than New York (p = 0.62). Analysis without separating by location revealed a decline (p = 0.047). CONCLUSIONS: Sperm concentrations are highest in New York compared to other U.S. cities. When accounting for this geographic difference and examining all available data, there appears to be no significant change in sperm counts in the U.S. during the last 60 years. Further studies addressing the causes of geographic variations are needed.     

Association of human leucocyte antigen phenotype with vaccine efficacy in patients receiving vaginal mucosal immunization for recurrent urinary tract infection

CONTEXT: One or both commercial tuberculin skin test reagents (Aplisol and Tubersol) may have a high rate of false-positive reactions. OBJECTIVE: To compare the reaction size and specificity of skin testing with Aplisol, Tubersol, and the standard purified protein derivative (PPD-S1). DESIGN: Double-blind trial, conducted between May 14, 1997, and October28, 1997, in which each individual received 4 tuberculin skin reagents at sites assigned at random. SETTING: Health departments and universities in 6 US cities. PARTICIPANTS: A total of 1555 persons at low risk of latent tuberculosis infection. INTERVENTION: Simultaneous skin tests with Aplisol, Tubersol, PPD-S1, and either a second PPD-S1 or PPD-S2 (a proposed new standard). MAIN OUTCOME MEASURE: Reaction size at each injection site measured by 2 investigators blinded to type of reagent. RESULTS: Aplisol produced slightly larger reactions than Tubersol, but this difference did not significantly change skin test interpretation. The mean +/- SD reaction sizes were 3.4+/-4.2 mm with Aplisol, 2.1+/-3.2 mm with Tubersol, and 2.5+/-3.6 mm with PPD-S1. Assuming that all participants were uninfected and using a 10-mm cutoff, the specificities of the tests were high: Aplisol, 98.2%; Tubersol, 99.2%; and PPD-S1, 98.9%. Significant variability was not detected in interobserver, host, and lot-to-lot reagent comparisons. CONCLUSION: Using a cutoff of at least 10 mm, testing with 3 different PPD reagents resulted in similar numbers of uninfected persons being correctly classified.     

Maternal regulation of embryonic growth: the role of vasoactive intestinal peptide

Vasoactive intestinal peptide (VIP) is an important growth regulator of the embryonic day (E)9-E11 mouse. In comparably aged rat embryos, VIP messenger RNA (mRNA) is not detectable; however, peak concentrations of VIP in maternal rat serum indicate a nonembryonic source. In the current study, mouse maternal and embryonic tissues were examined from E6-E12. Although RT-PCR revealed VIP mRNA in E6-E7 conceptuses, by E8 (when extraembryonic tissues could be separated from the embryo), VIP mRNA was detected only in the decidua/trophoblast. Decidual/trophoblastic VIP mRNA decreased until E10, after which it was not detectable. VIP mRNA was not apparent in the embryo until E11-E12. At E9, VIP immunoreactivity was localized to abundant, diffuse cells in the decidua basalis, which were also immunoreactive for T cell markers. VIP binding sites were dense in the decidua/trophoblast at E6, but gradually decreased until E10, after which they were not apparent. VIP binding sites were detected in embryonic neuroepithelium by E9. The transient presence of VIP binding sites and mRNA in the decidua/trophoblast correlate with the critical period of VIP growth regulation, when VIP mRNA is absent in the embryo. These findings suggest that maternal lymphocytes are the source of VIP's regulating early postimplantation embryonic growth.     

Double-strand break repair by Ku70 requires heterodimerization with Ku80 and DNA binding functions

Heterodimers of the 70 and 80 kDa Ku autoantigens (Ku70 and Ku80) activate the DNA-dependent protein kinase (DNA-PK). Mutations in any of the three subunits of this protein kinase (Ku70, Ku80 and DNA-PKcs) lead to sensitivity to ionizing radiation (IR) and to DNA double-strand breaks, and V(D)J recombination product formation defects. Here we show that the IR repair, DNA end binding and DNA-PK defects in Ku70-/- embryonic stem cells can be counteracted by introducing epitope-tagged wild-type Ku70 cDNA. Truncations and chimeras of Ku70 were used to identify the regions necessary for DNA end binding and IR repair. Site-specific mutational analysis revealed a core region of Ku70 responsible for DNA end binding and heterodimerization. The propensity for Ku70 to associate with Ku80 and to bind DNA correlates with the ability to activate DNA-PK, although two mutants showed that the roles of Ku70 in DNA-PK activation and IR repair are separate. Mutation of DNA-PK autophosphorylation sites and other structural motifs in Ku70 showed that these sites are not necessary for IR repair in vivo. These studies reveal Ku70 features required for double-strand break repair.     

A prospective evaluation of a diffractive versus a refractive designed multifocal intraocular lens

OBJECTIVE: To evaluate prospectively a diffractive (811E, Pharmacia; power add +4.0 D) versus a refractive (PA154N, Allergan; power add +3.5 D) designed multifocal lens. PARTICIPANTS: Eighty patients planned for cataract surgery without additional ocular pathologies were randomized into the diffractive or refractive group, respectively. INTERVENTION: A standardized no-stitch phacoemulsification with implantation of one of the two multifocal lenses was performed in each patient. MAIN OUTCOME MEASURES: Distance and near-visual acuity, contrast sensitivity, low contrast visual acuity, glare visual acuity, and depth of focus were measured after surgery. RESULTS: All treated patients had best-corrected visual acuities of 20/30 or better. Near-uncorrected vision was significantly better (P < 0.0001) with the diffractive lens (mean, J1) than with the refractive lens (mean, J4). Low contrast visual acuity (61 +/- 12% versus 59 +/- 9%), glare visual acuity (39 +/- 19% versus 38 +/- 14%), and contrast sensitivity (1.48 +/- 0.08 versus 1.50 +/- 0.12) were not significantly different between the groups. CONCLUSIONS: Both lens designs showed satisfactory functional results with advantages for the diffractive lens design.