Linkage guide for rotational alignment during total knee arthroplasty

A linkage guide was devised for use in conjunction with knee arthroplasty instruments to achieve proper component rotation. The femoral component was rotationally aligned to the surgical epicondylar axis using one guide. The other guides were used after all bone surfaces were cut and soft tissue balancing was completed. A Kirschner wire was guided into the proximal tibial aspect parallel to the sagittal plane of the femoral component with the patella in its normal position and the knee in full extension. The wire was used as a reference to determine tibial component rotation. The relative rotational alignment of 21 knees in 19 patients who had undergone cruciate-retaining total knee arthroplasty utilizing this guide was assessed using the modified Eckhoff method. The results of radiographic analysis were compared with those in a control group of 25 knees where the guide was not employed. The external rotation of the femoral component relative to the tibial component was 1.3° ± 2.0° (mean ± SD) in the guided group and 1.1° ± 4.4° in the control group. The relative rotational angle was significantly more consistent using the guide. Implant malrotation can be reduced using our technique.     

Distribution characteristics of immunosuppressants FK506 and cyclosporin A in the blood compartment

Using two representative immunosuppressants, FK506 (FK) and cyclosporin A (CyA), of which the mechanism of pharmacological action is the same although there is a great difference in the pharmacological intensity, the distribution characteristics were studied in both in vivo and in vitro experiments using rat, dog, and human blood. Blood samples were fractionated by means of sedimentation in Ficoll-Paque®, and the drug contents in the diluted plasma fraction, erythrocyte fraction, and lymphocyte fraction were measured by an HPLC method. FK distributes to the lymphocyte fraction to a level about three times greater than that of CyA, while CyA distributes to the erythrocyte fraction to a level ten times that of FK. The distribution pattern of these fractions was independent of the drug concentration and species after correcting the drug concentration in each fraction with the blood drug concentration. The uptakes of FK and CyA in the isolated lymphocytes obtained from the rat spleen and human peripheral blood were also studied. The amount of FK taken up by the spleen lymphocytes is five times greater than that of CyA. In the case of the uptake study using human peripheral blood lymphocytes, the concentration of FK in the lymphocyte is 100-fold higher than that of CyA. This difference in the lymphocyte level between the two immunosuppressants is thought to be one of the reasons why FK is more potent than CyA, a difference of about 100-fold in the in vitro pharmacological study and about tenfold in the in vivo organ transplantation experiments.     

The cleavage and inactivation of plasminogen activator inhibitor type 1 by neutrophil elastase: the evaluation of its physiologic relevance in fibrinolysis

The effect of the proteolytic cleavage of plasminogen activator inhibitor type 1 (PAI-1) by human neutrophil elastase (HNE) on fibrinolysis was investigated. HNE cleaved active PAI-1 and produced low molecular weight forms of inactive PAI-1, as previously reported. Latent PAI-1 was resistant to HNE treatment. Vitronectin (VN) partially protected the cleavage. NH2-terminal sequence analysis indicated that the cleavage site was Val355-Ser356 (P4-P3). The effects of PAI-1 cleavage by HNE on clot lysis was studied in a purified system. Clot lysis time without PAI-1 was 20.0 +/- 5.0 minutes and was prolonged to 86.7 +/- 2.9 minutes by 68 nmol/L of PAI-1. It was shortened when HNE (from 0.6 nmol/L to 80 nmol/L) was added and returned to the value obtained without PAI-1 by 80 nmol/L of HNE (20.0 +/- 5.8 minutes). However, in the absence of PAI-1, elastase did not enhance clot lysis at all. Euglobulin clot lysis time was also shortened after HNE treatment. The cleavage and inactivation of PAI-1 by HNE was shown to be a novel pathway to enhance fibrinolysis.     

The sites of origin of dopaminergic afferent fibers to the lateral habenular nucleus in the rat

The lateral habenular nucleus of the rat contains a dense plexus of dopaminergic fibers, which are more marked in the medial part of the lateral habenular nucleus than in its lateral counterpart. Employing a combination of fluorescent retrograde axonal tracing with fluorogold and tyrosine hydroxylase immunofluorescence histochemistry, we investigated the distribution of cells of origin of the dopaminergic afferent fibers to the lateral habenular nucleus in the rat. The cells double-labeled with both fluorogold injected into the lateral habenular nucleus and tyrosine hydroxylase antisera were seen in a variety of fore- and midbrain regions, including the bed nucleus of the stria terminalis, medial preoptic area, periventricular, ventromedial, and dorsomedial hypothalamic nuclei, ventral tegmental area, interfascicular nucleus, substantia nigra pars compacta, ventrolateral division of the midbrain periaqueductal gray, and dorsal raphe nucleus. The double-labeled cells were located bilaterally with an ipsilateral predominance, and constituted approximately 10% of the total fluorogold-positive cell population. We have further observed by anterograde axonal tracing with Phaseolus vulgaris–leucoagglutinin that projection fibers arising from the sites of origin of the dopaminergic afferent fibers to the lateral habenular nucleus terminate mainly in the medial part of the lateral habenular nucleus, and to a lesser extent in its lateral conterpart. Thus, we have found in the present study that the dopaminergic neurons sending their axons to the lateral habenular nucleus are widely distributed in the A9, A10, A14, and A15 dopaminergic cell groups. Such dopaminergic neurons may exert regulatory influences upon many limbic-associated brain regions via the lateral habenular nucleus. © 1993 Wiley-Liss, Inc.     

Multiplex PCR assay for rapid identification of oculopathogenic adenoviruses by amplification of the fiber and hexon genes

Eye infections caused by adenovirus (Ad) often result in nosocomial infections and community epidemics with significant rates of morbidity. No antiviral agent effective against Ad is yet available for clinical use. Therefore, early diagnosis is still the mainstay for patient management and the prevention of epidemics. A multiplex PCR assay based on amplification of a combination of the fiber and hexon genes which can identify the six important oculopathogenic serotypes of Ads (Ad serotype 3[Ad3], Ad4, Ad7, Ad8, Ad19, and Ad37) in a single-tube amplification reaction was developed. Ad serotypes could be distinguished by the different amplicon sizes. The assay correctly identified prototype strains as well as isolates in clinical specimens. In comparison with a previously described PCR-restriction fragment polymorphism method, our assay gave unequivocal results for clinical specimens. Our multiplex PCR has the potential to serve as a rapid and cost-effective tool for the typing of important ocular Ads.     

Development of enrofloxacin ELISA using a monoclonal antibody tolerating an organic solvent with broad cross-reactivity to other newquinolones

Antibacterial reagents, especially quinolones, are widely used in animals and humans, and have caused serious problems to human health because of their residual contaminants in food. In order to screen for different kinds of newquinolones at the same time, a sensitive and specific enzyme-linked immunoassay (ELISA) has been developed. The anti-enrofloxacin monoclonal antibody was selected because of its ability to react with structurally related newquinolones in organic solvent. The antibody has 100% cross-reactivity with norfloxacin, ciprofloxacin and other newquinolones at 50% inhibition of control values IC₅₀, but not with nitroflazone, sulphadimethoxine. The lowest detection limit of this ELISA was 0.7 ng/ml (ppb) when enrofloxacin was used as the calibrator. Eel extracts were spiked with enrofloxacin and the average recoveries at 10, 50, 100 ng/ml were 98, 102 and 91%, respectively. The proposed ELISA is a useful method for the practical microquantitation of various newquinolones in biological and environmental specimens.     

Establishment of an HIV cell-cell fusion assay by using two genetically modified HeLa cell lines and reporter gene

Infection of human cells with the human immunodeficiency virus type I (HIV-1) can be mimicked by a fusion process between cells expressing the HIV envelope protein (Env) and cells expressing both human CD4 together with the appropriate human chemokine receptors. In this study, a T-tropic HIV cell-cell fusion assay was established that utilized CD4, human CXCR4 and HIV NL4-3 gp160 as fusion components and a T7 polymerase-activated luciferase as a reporter system. The HeLa T4 cells used, expressed CD4 and CXCR4, and the applied HeLa KS386 cells expressed HIV NL4-3 gp160. By combining HeLa T4 cells with HeLa KS386 cells, an approximately about 100- to 300-fold increase in luciferase activity could be elicited relative to the control. The addition of anti-CD4 monoclonal antibody (Mab) (RPA-T4) or anti-CXCR4 Mab (12G5) in the assay significantly inhibited the fusion event; in contrast, an anti-CCR5 Mab (2D7) had no effect, indicating that the fusion assay was CD4 and CXCR4 dependent. In this report, fusion events could be monitored by both the luciferase reporter system and syncytia formation. Fusion events were monitored and compared using these two approaches. The luciferase reporter system was found to be more sensitive than syncytia formation. Moreover, compared with previous HIV fusion models, such as using recombinant vaccinia viruses, this system has several advantages, including simplicity and sensitivity. Finally, the system provides a powerful tool to study fusion mechanisms mediated by T-tropic HIV gp160, as well as to screen for fusion-blocking antibodies and antiviral agents.     

Liberation of serotonin from rabbit blood platelets by bacterial cell walls and related compounds.

A study was made on the activity of various bacterial cell walls and peptidoglycans to liberate serotonin from rabbit blood platelets. All of the test cell walls or peptidoglycans prepared from 27 strains of 21 bacterial species were shown to cause a marked release of serotonin, regardless of differences in types of peptidoglycan and non-peptidoglycan moieties and in some biological properties. The assay made with the water-soluble "digests" of Staphylococcus epidermidis cell wall peptidoglycans, which were prepared by use of appropriate enzymes, revealed that a polymer of peptidoglycan subunits (a disaccharide-stempeptide) was definitely active in the release of serotonin, but a structural unit monomer was inactive. Among a variety of synthetic muramylpeptides and their 6-O-acyl derivatives, only 6-O-(3-hydroxy-2-docosylhexacosanoyl)-N-acetylmuramyl-L-alanyl-D-isoglutaminyl- L-lysyl-D-alanine was found to hold a strong serotonin-liberating activity.