Vitronectin and its fragments purified as serum inhibitors of Staphylococcus aureus gamma-hemolysin and leukocidin, and their specific binding to the hlg2 and the LukS components of the toxins.

Staphylococcal gamma-hemolysin and leukocidin are bi-component cytolysins, consisting of LukF (or Hlg1)/Hlg2 and LukF/LukS, respectively. Here, we purified serum inhibitors of gamma-hemolysin and leukocidin from human plasma. Protein sequencing showed that the purified inhibitors of 62, 57, 50 and 38 kDa were the vitronectin fragments with truncation(s) of the C-terminal or both N- and C-terminal regions. The purified vitronectin fragments specifically bound to the Hlg2 component of gamma-hemolysin and the LukS component of leukocidin to form high-molecular-weight complexes with them, leading to inhibition of the toxin-induced lysis of human erythrocytes and human polymorphonuclear leukocytes, respectively. Intact vitronectin also showed inhibitory activity to the toxins. The ability of gamma-hemolysin and leukocidin to bind vitronectin and its fragments is a novel function of the pore-forming cytolysins.     

Markers for trans-Golgi Membranes and the Intermediate Compartment Localize to Induced Membranes with Distinct Replication Functions in Flavivirus-Infected Cells

Replication of the flavivirus Kunjin virus is associated with virus-induced membrane structures within the cytoplasm of infected cells; these membranes appear as packets of vesicles associated with the sites of viral RNA synthesis and as convoluted membranes (CM) and paracrystalline arrays (PC) containing the components of the virus-specified protease (E. G. Westaway, J. M. Mackenzie, M. T. Kenney, M. K. Jones, and A. A. Khromykh, J. Virol. 71:6650-6661, 1997). To determine the cellular origins of these membrane structures, we compared the immunolabelling patterns of several cell markers in relation to these sites by immunofluorescence and immunoelectron microscopy. A marker for the trans-Golgi membranes and the trans-Golgi network, 1,4-galactosyltransferase (GalT), was redistributed to large foci in the cytoplasm of Kunjin virus-infected cells, partially coincident with immunofluorescent foci associated with the putative sites of viral RNA synthesis. As determined by immunoelectron microscopy, the induced vesicle packets contained GalT, whereas the CM and PC contained a specific protein marker for the intermediate compartment (ERGIC53). A further indicator of the role of cellular organelles in their biogenesis was the observation that the Golgi apparatus-disrupting agent brefeldin A prevented further development of immunofluorescent foci of induced membranes if added before the end of the latent period but that once formed, these membrane foci were resistant to brefeldin A dispersion. Reticulum membranes emanating from the induced CM and PC were also labelled with the rough endoplasmic reticulum marker anti-protein disulfide isomerase and were obviously redistributed during infection. This is the first report identifying trans-Golgi membranes and the intermediate compartment as the apparent sources of the flavivirus-induced membranes involved in events of replication.     

Phorbol ester facilitates apoptosis in murine fibroblasts pretreated by mild ultraviolet radiation.

Although phorbol 12-myristate 13-acetate (PMA) inhibits apoptosis and promotes the growth of some types of cells, it induces apoptosis in other cells. We evaluated the apoptotic effects of PMA on murine fibroblasts (L-929) that had been exposed to ultraviolet-B (UV-B) radiation at 312 nm, which promotes tumor cell growth. Exposure to PMA alone did not induce Fas, Fas-L, or apoptosis. Cells exposed to mild UV-B irradiation (80 J/m(2)) alone exhibited a slight expression of Fas and Fas-L 36 to 48 h after the exposure, and exhibited apoptosis as evidenced by DNA fragmentation 72 h after exposure. The addition of PMA (0.8 x 10(-5) to 3.2 x 10(-5) M) to the medium 24 h after the UV-B exposure markedly and dose-dependently enhanced these cell responses. Confluent untreated cells, cells cocultured with PMA, and cells cocultured with PMA for 24 h after the UV-B exposure consistently expressed mRNAs for wild-type p53, bcl-2, and ICE. Expression of c-myc mRNA was initially observed, but became undetectable in the cells cocultured for 24 h with a high concentration of PMA (3.2 x 10(-5) M) following UV-B exposure. Such cells subsequently exhibited the maximal apoptotic response. We conclude that mild exposure to UV-B altered murine fibroblast cells in such a way as to facilitate their death by apoptosis upon addition of PMA.     

A finite element based method to determine the properties of planar soft tissue.

A finite element based method to determine the incremental elastic material properties of planar membranes was developed and evaluated. The method is applicable to tissues that exhibit inhomogeneity, geometric and material nonlinearity, and anisotropy. Markers are placed on the tissue to form a four-node quadrilateral element. The specimen is loaded to an initial reference state, then three incremental loading sets are applied and the nodal displacements recorded. One of these loadings must include shear. These data are used to solve an over-determined system of equations for the tangent stiffness matrix. The method was first verified using analytical data. Next, data obtained from a latex rubber sheet were used to evaluate experimental procedures. Finally, experiments conducted on preconditioned rat skin revealed nonlinear orthotropic behavior. The vector norm comparing the applied and calculated nodal force vectors was used to evaluate the accuracy of the solutions.     

An implantable transducer for measuring tension in an anterior cruciate ligament graft.

The goal of this study was to develop a new implantable transducer for measuring anterior cruciate ligament (ACL) graft tension postoperatively in patients who have undergone ACL reconstructive surgery. A unique approach was taken of integrating the transducer into a femoral fixation device. To devise a practical in vivo calibration protocol for the fixation device transducer (FDT), several hypotheses were investigated: (1) The use of a cable versus the actual graft as the means for applying load to the FDT during calibration has no significant effect on the accuracy of the FDT tension measurements; (2) the number of flexion angles at which the device is calibrated has no significant effect on the accuracy of the FDT measurements; (3) the friction between the graft and femoral tunnel has no significant effect on measurement accuracy. To provide data for testing these hypotheses, the FDT was first calibrated with both a cable and a graft over the full range of flexion. Then graft tension was measured simultaneously with both the FDT on the femoral side and load cells, which were connected to the graft on the tibial side, as five cadaver knees were loaded externally. Measurements were made with both standard and overdrilled tunnels. The error in the FDT tension measurements was the difference between the graft tension measured by the FDT and the load cells. Results of the statistical analyses showed that neither the means of applying the calibration load, the number of flexion angles used for calibration, nor the tunnel size had a significant effect on the accuracy of the FDT. Thus a cable may be used instead of the graft to transmit loads to the FDT during calibration, thus simplifying the procedure. Accurate calibration requires data from just three flexion angles of 0, 45, and 90 deg and a curve fit to obtain a calibration curve over a continuous range of flexion within the limits of this angle group. Since friction did not adversely affect the measurement accuracy of the FDT, the femoral tunnel can be drilled to match the diameter of the graft and does not need to be overdrilled. Following these procedures, the error in measuring graft tension with the FDT averages less than 10 percent relative to a full-scale load of 257 N.     

Trigger finger secondary to anomalous lumbrical insertion: a case report and review of the literature

Trigger finger is a relatively common clinical entity, most frequently caused by stenosing tenosynovitis. Several other conditions not related to tenosynovitis also have been described as a cause of triggering, and these have been reviewed. We present a rare anomaly of the fourth lumbrical muscle insertion as a cause of triggering of the right little finger. This was completely relieved following excision of the anomalous muscle. This rare anatomic variant should be added to the list of potential causes of trigger finger.