Expansions of T-cell subsets expressing particular T-cell receptor variable regions in chronic beryllium disease

Bisphosphonates are currently the most important class of antiresorptive drugs used for the treatment of metabolic bone diseases. Although the molecular targets of bisphosphonates have not been identified, these compounds inhibit bone resorption by mechanisms that can lead to osteoclast apoptosis. Bisphosphonates also induce apoptosis in mouse J774 macrophages in vitro, probably by the same mechanisms that lead to osteoclast apoptosis. We have found that, in J774 macrophages, nitrogen-containing bisphosphonates (such as alendronate, ibandronate, and risedronate) inhibit post-translational modification (prenylation) of proteins, including the GTP-binding protein Ras, with farnesyl or geranylgeranyl isoprenoid groups. Clodronate did not inhibit protein prenylation. Mevastatin, an inhibitor of 3-hydroxy-3-methylglutatyl (HMG)-CoA reductase and hence the biosynthetic pathway required for the production of farnesyl pyrophosphate and geranylgeranyl pyrophosphate, also caused apoptosis in J774 macrophages and murine osteoclasts in vitro. Furthermore, alendronate-induced apoptosis, like mevastatin-induced apoptosis, could be suppressed in J774 cells by the addition of farnesyl pyrophosphate or geranylgeranyl pyrophosphate, while the effect of alendronate on osteoclast number and bone resorption in murine calvariae in vitro could be overcome by the addition of mevalonic acid. These observations suggest that nitrogen-containing bisphosphonate drugs cause apoptosis following inhibition of post-translational prenylation of proteins such as Ras. It is likely that these potent antiresorptive bisphosphonates also inhibit bone resorption by preventing protein prenylation in osteoclasts and that enzymes of the mevalonate pathway or prenyl protein transferases are the molecular targets of the nitrogen-containing bisphosphonates. Furthermore, the data support the view that clodronate acts by a different mechanism.     

Identification and characterization of novel antimicrobial decapeptides generated by combinatorial chemistry

Novel combinatorial libraries consisting of simplified amino acid sequences were designed to screen for peptides active against the Candida albicans membrane. A novel decapeptide, KKVVFKVKFK, that had a unique primary amino acid sequence was identified in this work. This peptide irreversibly inhibited the growth of C. albicans and showed a broad range of antibacterial activity but no hemolytic activity. Circular dichroism spectra revealed that the predominant secondary structure of this peptide strongly depended on the membrane-mimetic environments; the peptide preferred to form an amphipathic alpha-helical structure in the presence of 50% trifluoroethanol, while it preferred to adopt a distorted alpha-helical structure in the presence of sodium dodecyl sulfate micelles. Experiments in which dye was released from vesicles indicated that this novel antimicrobial peptide killed microorganisms through the action on the membrane as its primary target. Replacement of amino acids in this active decapeptide on the basis of information from the libraries could provide unique information about factors affecting its antimicrobial activity such as its secondary structure, net positive charge, and hydrophobicity.     

A functional role for specific spliced variants of the alpha7beta1 integrin in acetylcholine receptor clustering

The clustering of acetylcholine receptors (AChR) on skeletal muscle fibers is an early event in the formation of neuromuscular junctions. Recent studies show that laminin as well as agrin can induce AChR clustering. Since the alpha7beta1 integrin is a major laminin receptor in skeletal muscle, we determined if this integrin participates in laminin and/or agrin-induced AChR clustering. The alternative cytoplasmic domain variants, alpha7A and alpha7B, and the extracellular spliced forms, alpha7X1 and alpha7X2, were studied for their ability to engage in AChR clustering. Immunofluorescence microscopy of C2C12 myofibers shows that the alpha7beta1 integrin colocalizes with laminin-induced AChR clusters and to a much lesser extent with agrin-induced AChR clusters. However, together laminin and agrin promote a synergistic response and all AChR colocalize with the integrin. Laminin also induces the physical association of the integrin and AChR. High concentrations of anti-alpha7 antibodies inhibit colocalization of the integrin with AChR clusters as well as the enhanced response promoted by both laminin and agrin. Engaging the integrin with low concentrations of anti-alpha7 antibody initiates cluster formation in the absence of agrin or laminin. Whereas both the alpha7A and alpha7B cytoplasmic domain variants cluster with AChR, only those isoforms containing the alpha7X2 extracellular domain were active. These results demonstrate that the alpha7beta1 integrin has a physiologic role in laminin-induced AChR clustering, that alternative splicing is integral to this function of the alpha7 chain, and that laminin, agrin, and the alpha7beta1 integrin interact in a common or convergent pathway in the formation of neuromuscular junctions.     

Osteoprotegerin production by human osteoblast lineage cells is stimulated by vitamin D, bone morphogenetic protein-2, and cytokines

Rheumatoid arthritis(RA) is a chronic, deforming and destructive arthritis of unknown etiology. For the medical treatment of RA, NSAID has been the first choice of drug. Recently it has been known that early use of DMARD may result in clinical remission. Understanding of the pivotal role of cytokines and adhesion molecules for the rheumatoid joint destruction enabled us to target these cytokines and molecules as therapeutic measures. Monoclonal antibodies were produced against the cytokines and adhesion molecules such as IL-1, IL-6, IL-6R, TNF-alpha, as well as CD4 molecules. Clinical use of these monoclonal antibodies was found to be effective for rheumatoid arthritis. However these therapeutic measures have several disadvantages such as transient efficacy and side effect.     

Thrombin receptor (PAR-1) antagonists. Heterocycle-based peptidomimetics of the SFLLR agonist motif

Phorbol ester treatment of MCF-7 cells led to the tyrosine phosphorylation and activation of PKC delta. However, through Western blot analysis and in vitro immunecomplex kinase assays, we detected a differential localization of tyrosine-phosphorylated PKC delta and catalytically active PKC delta. Catalytically active PKC delta was concentrated in Triton X-100 solubilized-membrane fractions while tyrosine-phosphorylated PKC delta was localized to the cytosol fraction. Phorbol ester treatment of MCF-7 cells stimulated both the time-dependent in vivo association of Src with PKC delta, evidenced in Src immunoprecipitates by the co-immunoprecipitation of PKC delta, and activation of Src, evidenced in Src immunoprecipitates as an increase in reactivity with a Src antibody (clone 28) reactive only with active Src (dephosphorylated on residue 530) and in Src and PKC delta immunoprecipitates by an increase in Src kinase activity. While our data are consistent with reports in the literature showing the activator/stimulus-dependent tyrosine phosphorylation of PKC delta, our data show that the tyrosine phosphorylation of PKC delta is not essential for kinase activity. These results are the first to demonstrate an in vivo association between PKC delta and active Src in the absence of over-expression of either PKC delta or Src, and support the association of Src and PKC delta towards a physiological function.     

Penicillin vs. erythromycin in the treatment of diphtheria

PURPOSE: To review the University of Florida experience in treating ependymomas, analyze prognostic factors, and provide treatment recommendations. METHODS AND MATERIALS: Forty-one patients with ependymoma and no metastases outside the central nervous system received postoperative radiotherapy with curative intent between 1966 and 1989. Ten patients had supratentorial lesions, 22 had infratentorial lesions, and 9 had spinal cord lesions. All patients had surgery (stereotactic biopsy, subtotal resection, or gross total resection). Most patients with high-grade lesions received radiotherapy to the craniospinal axis. Low-grade intracranial lesions received more limited treatment. Spinal cord lesions were treated using either partial spine or whole spine fields. RESULTS: Of 32 intracranial tumors, 21 recurred, all at the primary site; no spinal cord tumors recurred. Overall 10-year survival rates were 51% (absolute) and 46% (relapse-free); by tumor site: spinal cord, 100%; infratentorial, 45%; supratentorial, 20% (p = 0.002). On multivariate analysis, tumor site was the only factor that influenced absolute survival (p = 0.0004); other factors evaluated included grade, gender, age, duration of symptoms, resection extent, primary tumor dose, treatment field extent, surgery-to-radiotherapy interval, and days under radiotherapy treatment. CONCLUSIONS: Patients with supratentorial or infratentorial tumors receive irradiation, regardless of grade. Craniospinal-axis fields are used when spinal seeding is radiographically or pathologically evident. Spinal cord tumors are treated using localized fields to the primary site if not completely resected. Failure to control disease at the primary site remains the main impediment to cure.     

Coronary bypass in vascular patients: a relatively high-risk procedure

A premise of cardiac risk stratification is that the added risk of coronary artery bypass grafting (CABG) is offset by the improved safety of subsequent vascular reconstruction (VR). We questioned if elective CABG is patients with severe peripheral vascular disease (PVD) is a relatively high-risk procedure. A cohort study of 680 elective CABG patients from January 1993 to December 1994 was performed using three mutually exclusive outcomes of complication-free survival, morbidity, and mortality. Patient characteristic, operative, and outcome data were prospectively collected. Retrospective review determined that 58 patients had either a standard indication for or a history of VR. Overall CABG mortality was 2.5%, with statistically similar but relatively higher rates for PVD as compared to non-PVD patients. In contrast, major morbidity occurred at rates 3.6-fold higher in PVD patients (39.7%) than in disease-free patients (16.7%) after adjustment for the effects of patient and operative variables (odds ratio [OR] 3.67, 95% confidence interval [CI] 1.93-6.99). CABG morbidity in the PVD patient was most likely in those patients with aortoiliac (OR 9.51, CI 3.20-28.27) and aortic aneurysmal (OR 5.24, CI 1.28-21.41) disease types. CABG in PVD patients is associated with significant major morbidity. Such morbidity may preclude or alter the timing of subsequent VR.