A 76-amino acid disulfide loop in the Yersinia pseudotuberculosis invasin protein is required for integrin receptor recognition

The Yersinia pseudotuberculosis invasin protein is a 986-amino acid protein that promotes bacterial penetration into mammalian cells by avidly binding multiple beta 1-chain integrins. A 192-amino acid carboxyl-terminal domain of invasin was previously shown to be sufficient for binding. Evidence is presented here that a 76-amino acid disulfide loop in the integrin binding domain of invasin is required for invasin-mediated cell binding and entry. Bacterial mutants that were altered at either of 2 cysteine residues in the binding domain of invasin were completely defective for entry. Purified invasin protein derivatives altered at either of these cysteines, in contrast to the wild-type invasin, did not promote either cell binding or penetration. Analysis of proteolytic products of invasin in the presence or absence of reducing agent provided evidence of an intra-chain disulfide bond near the carboxyl terminus of the protein. Alkylation of invasin derivatives with [3H]iodoacetate indicated that these 2 cysteines were normally disulfide-bonded. A treatment that resulted in the maximal reduction of the disulfide bond also resulted in maximal loss of cell attachment activity. These results indicate that the 76-amino acid disulfide loop at the carboxyl terminus of invasin is required for recognition by integrins.     

Activation and detoxification of dinitropyrenes by cytosol and microsomes from Aroclor-pretreated rats in the Ames and umu assays

PURPOSE: To evaluate the efficacy of topical mitomycin C in treating conjunctival and corneal epithelial dysplasia and neoplasia. METHODS: Seven eyes of seven patients with conjunctival and corneal epithelial dysplasia and neoplasia were treated with one drop of topical mitomycin C 0.04% four times a day for 7 days in alternate weeks. The patients' charts were reviewed retrospectively. Patients with either multiple recurrences or extensive ocular surface involvement were treated. In all eyes, the diagnosis of epithelial dysplasia or neoplasia was confirmed by histopathology before the onset of therapy. Patients were examined at least every 14 days during treatment and examined at intervals after completion of treatment. RESULTS: With topical mitomycin C, six eyes of seven patients had complete clinical regression of their conjunctival and corneal epithelial dysplasia and neoplasia. One eye of one patient had partial clinical regression of conjunctival and corneal epithelial dysplasia. Follow-up after completion of topical mitomycin C therapy and excision of residual disease ranged from 2 to 16 months (mean, 9 months; SD, 4.3 months) and was without clinical sign of recurrence. Topical mitomycin C therapy was associated with transitory ocular discomfort, conjunctival injection, tearing, photophobia, and punctate epithelial keratopathy. CONCLUSION: In this small series of eyes, topical mitomycin C was effective as a treatment for conjunctival and corneal epithelial dysplasia and neoplasia.     

Studies on anti-inflammatory agents. V. Synthesis and pharmacological properties of 3-(difluoromethyl)-1-(4-methoxyphenyl)-5- [4-(methylsulfinyl)phenyl]pyrazole and related compounds

A series of novel 1,5-diphenylpyrazole derivatives bearing hydrophilic substituents was prepared. The anti-inflammatory and analgesic activities of these compounds were evaluated by using the adjuvant arthritis and Randall-Selitto assays in rats, and the structure-activity relationships were studied. The optimal compound was 3-(difluoromethyl)-1-(4-methoxyphenyl)-5-[4-(methylsulfinyl)phenyl]pyraz ole (10) with oral ED50 values of 0.31 and 2.6 mg/kg on adjuvant-induced arthritis and carrageenin-induced foot edema, respectively. Compound 10 showed analgesic activities not only toward inflamed paw but also toward normal paw (ED30 = 0.55 and 1.8 mg/kg, respectively) in the Randall-Selitto assay, and moreover, 10 was effective in the tail-pinch assay (ED50 = 21 mg/kg) similarly to morphine. The asymmetric synthesis and pharmacological properties of the enantiomers of 10 are also reported.     

Nonclinical model for assessing gastric effects of bisphosphonates

Mechanisms by which ketones potentiate manganese-bilirubin (Mn-BR)-induced cholestasis are unknown. The purpose of the present study was to investigate the effect of methyl isobutyl ketone (MiBK), a widely used ketonic solvent, at the level of the bile canalicular membrane (BCM) and to verify if altered membrane lipid dynamics could be involved in MiBK-potentiated Mn-BR cholestasis. Male Sprague-Dawley rats were exposed 4 hr/day for 3 days to MiBK vapors (200 or 600 ppm). Eighteen hours after the last exposure, manganese (Mn, 4.5 mg/kg) was given i.v. followed 15 min later by bilirubin (BR, 25 mg/kg). Rats were killed 30 min after BR; liver cell plasma membranes (bile canalicular and sinusoidal), microsomes, mitochondria, and cytosol were isolated by differential centrifugation. Lipids were extracted and cholesterol was measured in each fraction. After Mn-BR and MiBK exposure (600 ppm), results indicated a marked increase in BCM cholesterol content compared to rats exposed to air only. This increase was greater than that due to Mn-BR or MiBK given alone. Also, results indicated that cholesterol increased in a dose-related fashion in BCM after MiBK exposure, whereas PM cholesterol remained unaltered. To identify the source of the increased BCM cholesterol and to permit distinction between de novo cholesterol synthesis and subcellular shifts, the hepatic lipid pool was labeled in vivo with [3H]-cholesterol and [2-14C]-mevalonic acid, a cholesterol synthesis precursor. Results showed that after 600 ppm MiBK exposure, 14C-labeled cholesterol was greater than 3H-labeled cholesterol, indicating that the contribution of de novo cholesterol synthesis to the total cholesterol content of the various isolated hepatocellular fractions was more important than the contribution of intracellular pools. Therefore, increased BCM cholesterol content and enhanced accumulation of newly synthesized cholesterol appear to be involved in MiBK potentiation of Mn-BR-induced cholestasis.     

Compartment-ablation studies of GLUT4 distribution in adipocytes: evidence for multiple intracellular pools

The available data suggest that GLUT4 does populate the recycling endosomal system to some extent, but that a large proportion of the intracellular GLUT4 resides in a compartment that is devoid of transferrin receptors and may have properties more akin to specialized secretory vesicles. The study of the nature and biogenesis of this compartment will provide important insight into the mechanism by which insulin stimulates glucose transport. Further study of the role of the synaptobrevins in these distinct subcellular compartments will probably shed further light on the mechanism by which insulin stimulates GLUT4 translocation.     

Perioperative risk factors affecting hospital stay and hospital costs in open heart surgery for patients > or = 65 years old

OBJECTIVE: Demographic changes, associated with increased demands for open heart surgery in the elderly, place increased burden on financial resources. To evaluate perioperative risk factors affecting incidence of hospital events and estimation of hospital charges, 2577 patients > or = 65 years (range 65-91), operated on from January 1991 to December 1994, were compared with a concurrent cohort of 2642 younger patients. METHODS: Statistical analysis, by surgical procedure, focused on hospital mortality, key postoperative complications affecting length of hospital stay and hospital charges. RESULTS: Overall hospital mortality was 4.7%, 3.5% in younger patients versus 6.1% in the older group (P < 0.01). Mortality was significantly lower in patients less than 65 years undergoing coronary artery bypass grafting (3% versus 5%, P < 0.01) and valve replacement (4% versus 9%, P = 0.01). Significant risk factors for hospital death in the elderly: diabetes (P < 0.01), hypertension (P < 0.01), myocardial infarction (P < 0.01) and congestive heart failure (P < 0.01). Significant postoperative events, more common in older patients, included prolonged ventilation (P < 0.01), congestive heart failure (P < 0.01), infection (P < 0.01), cerebrovascular accident (P < 0.01), and intra aortic balloon pump (P < 0.01). Incremental risk factors for morbidity in the elderly were: higher New York Heart Association class, congestive heart failure, emergent operation, and female gender. Mean length of hospital stay for the < 65 group was 15.3 versus > 19.5 days for the > 65 group (P < 0.01). Length of stay over 18 days positively correlated with increased morbidity in both age groups. For patients > or = 65 years of age, the average hospital charge for open heart surgery was 172% higher for patients with a length of stay greater than 18 days compared with 165% for patients less than 65 years of age. CONCLUSIONS: Higher operative mortality and longer length of stay in elderly patients, resulting in increased health care costs, was associated with more co-morbidities. These results suggest interventions designed to reduce congestive heart failure and other co-morbidities may improve patient's recovery and reduce costs.