An in vitro assessment of the accuracy of new and in-use torque-limiting devices.

STATEMENT OF PROBLEM: Recent studies have shown that torque-limiting devices (TLDs) do not meet their torque targets and are affected by factors such as the use of the TLDs and the sterilization processes used. PURPOSE: The purpose of this in vitro study was to investigate the accuracy of TLDs currently in use in the Advance Education Program in Prosthodontics at the New York University College of Dentistry. MATERIAL AND METHODS: Five new and 46 in-use TLDs (Nobel BioCare) from the implant kits of the graduate students were evaluated. One investigator was trained and calibrated before testing and after every 10 evaluations. A 3-jaw chuck was mounted on the center of a cap testing device by using the provided mounting screws. A LOCATOR torque driver was clamped into the chuck, and a torque wrench driver was attached to it. The device was placed on a flat table with direct overhead lighting that allowed the investigator to have a consistent view of the notches on the TLDs. A piece of cardboard was attached to the device to ensure that the investigator could not see the readouts. The blinded researcher inserted the wrench and applied the appropriate torque force at the designated notches while another researcher recorded the results. Two readings were made for each TLD at 15, 35, and 45 Ncm. A 2-way ANOVA and an intraclass correlation coefficient to test for intraclinician reliability were performed by using a statistical software program (α=.05). RESULTS: Two TLDs were damaged and not tested. The 2-way ANOVA demonstrated no significant difference (P>.05) between graduate students in year 1, 2, and 3 or between the autoclaved and new TLDs. The intraclass correlation coefficient was 0.861 for 15 Ncm, 0.589 for 35 Ncm, and 0.764 for 45 Ncm. CONCLUSIONS: In this in vitro study, new and used TLDs all met the recommended torque values. No significant differences were found among groups, suggesting that autoclaving and use did not affect the accuracy of the TLDs tested.

ESCRT-II's involvement in HIV-1 genomic RNA trafficking and assembly.

BACKGROUND INFORMATION: Several host proteins play crucial roles in the HIV-1 replication cycle. The endosomal sorting complex required for transport (ESCRT) exemplifies a large, multi-component host machinery that is required by HIV-1 for viral budding. ESCRT promotes the inward budding of vesicles from the membranes of late endosomes to generate multi-vesicular bodies. However, HIV-1 co-opts the ESCRT to enable outwards budding of virus particles from the plasma membrane, a phenomenon that is topologically similar to multi-vesicular body biogenesis. A role for ESCRTII in mRNA trafficking has been established in Drosophila in which the ESCRT-II components, Vps22 and Vps36, promote the localisation of the bicoid mRNA in the fertilised egg. This is achieved via specific interactions with the Staufen protein. In this work, we investigated a possible implication of ESCRT-II in the HIV-1 replication cycle. RESULTS: Co-immunoprecipitation analyses and live cell tri-molecular fluorescence complementation assays revealed that interactions between EAP30 and Gag and another between EAP30 and Staufen1 occur in mammalian cells. We then depleted EAP30 (the orthologue for Vps22) by siRNA to target ESCRT-II in HIV-1 expressing cells. This treatment disrupted ESCRT-II function and leads to the degradation of the two other ESCRT-II complex proteins, EAP45 and EAP20, as well as the associated Rab7-interacting lysosomal protein. The depletion of EAP30 led to dramatically reduced viral structural protein Gag and virus production levels, without any effect on viral RNA levels. On the contrary, the overexpression of EAP30 led to a several-fold increase in virus production. Unexpec-tedly, siRNA-mediated depletion of EAP30 led to a block to HIV-1 genomic RNA trafficking and resulted in the accumulation of genomic RNA in the nucleus and juxtanuclear domains. CONCLUSIONS: Our data provide the first evidence that the Staufen1-ESCRT-II interaction is evolutionarily conserved from lower to higher eukaryotes and reveal a novel role for EAP30 in the control of HIV-1 RNA trafficking and gene expression.