Studies of the denaturation patterns of bovine alpha-crystallin using an ionic denaturant, guanidine hydrochloride and a non-ionic denaturant, urea

The effects of non-ionic and ionic denaturation and denaturation/renaturation on the native structure of alpha-crystallin at room temperature were examined. Native alpha-crystallin, at concentrations above and below the previously reported critical micelle concentration (CMC) range, was denatured by varying concentrations of urea and guanidine hydrochloride. The resulting denatured samples were examined by gel filtration fast performance liquid chromatography (FPLC), circular dichroism spectropolarimetry (CD), and transmission electron microscopy. Elution peak samples from gel filtration chromatography with sufficiently high concentrations were examined for subunit composition by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The studies presented herein demonstrate that the denaturation and renaturation of alpha-crystallin via non-ionic urea denaturation results in different renaturation species, depending upon the initial concentration of alpha-crystallin which is denatured and the concentration of urea, including certain species which, by gel filtration FPLC, have an apparent molecular weight greater than the native 800 kD aggregate. Transmission electron microscopy has also demonstrated the existence of a high molecular weight aggregate form for denatured samples. Ionic dissociation, in contrast, proceeds much in the same manner above and below the CMC range, the major difference occurring at 2 M guanidine hydrochloride. alpha B-crystallin is preferentially removed from the native alpha-crystallin aggregate upon treatment with 2 M guanidine hydrochloride indicating, once again, differences between the two subunits. Above and below the CMC range, dissociation with guanidine hydrochloride appears to plateau after 4 M guanidine hydrochloride as indicated by the presence of two apparent homotetrameric species and no further dissociation of these species with increasing guanidine hydrochloride concentrations. CD demonstrates that some secondary structure, which is lost with lower concentrations of alpha-crystallin, is still present when concentrations of alpha-crystallin, well above the critical micelle concentration range, are treated with high concentrations of urea at room temperature. In contrast, concentrations both above and below the CMC range demonstrate a significant loss of secondary structure upon treatment with 2 M guanidine hydrochloride. Finally, ionic denaturation and subsequent renaturation results in the formation of a species which is functionally incapable of protecting gamma-crystallin from heat-induced aggregation.     

Saccharide-assisted delivery of cytotoxic liposomes to human malignant cells

The overexpression of lectins by malignant cells was applied for in vitro targeting of liposomes equipped with a saccharide vector and loaded in the lipid phase with a lipid derivative of anticancer agent sarcolysine. The lectin specificity of human leukemia HL-60 and human lung adenocarcinoma ACL cells was revealed by tests with fluorescein-labeled sugar probes. With the help of fluorescent lipid dye it was shown that active saccharide ligands increased the level of the vectored liposome binding to malignant cells by 50-80% as compared to liposomes without vector or with inactive one. The degree of liposome/cell membrane fusion was monitored fluorometrically and was shown to be complete and independent of the vectors. The targeted drug-loaded liposomes had the cytotoxic activity 2-4 times higher as compared to the vector-free ones.     

Changes in dental caries prevalence of school children in Araraquara, SP, Brazil

Previous reports indicate that venous thrombosis is an infrequent problem in patients with HIV infection. Despite this, various HIV-related factors have been proposed as potentially thrombogenic and an HIV-related hypercoagulability has been suggested. At the present time, there exists no consensus of opinion regarding prophylaxis against venous thrombosis for hospitalized patients with HIV. This article aims to provide an overview of venous thrombosis in HIV infection with particular reference to published and personal evidence for possible risk factors and their implications for prophylaxis.     

Conservative management of penetrating and blunt Type III renal injuries

This study examines whether, while controlling for local economic conditions, job design and other organizational factors affect facility aide turnover rates in a sample of 250 nursing homes from 10 states. Facility characteristics were largely based on administrator and director of nursing interviews conducted in 1993 as part of the Health Care Financing Administration's evaluation of the Resident Assessment Instrument (RAI) implementation. Aide turnover was significantly reduced by involvement in interdisciplinary care plan meetings. Turnover rates were higher in for-profit homes. Aide involvement in assessments, aide training and workload, case mix severity, payor source mix, and facility size were not significantly related to aide turnover.     

The P300 component of the auditory evoked potential in the posttraumatic vegetative state

The dynamic studies of P300 component of the acoustic evoked potential were conducted in 12 patients in posttraumatic state of the autonomic system. The amplitude and time parameters of P300 were analyzed, its spatial distribution over the cortex, and features of generation. The obtained results were compared with the normative data. Significant changes in P300 in comparison with the normal characteristics were revealed in patients with the absence of conscious mental activity. These changes were maximally expressed when the state of patients was irreversible. In case of patients' outcome from the studied state the amplitude-temporal and topographic response characteristics tended to normalization but did not reach it completely. The obtained results allow us to consider the P300 component as one of the most informative indices in consciousness recovery after severe brain injury.     

Vascular protection with newer antihypertensive agents

ROLE OF CALCIUM: Alterations in the structure and function of small arteries may contribute to elevated blood pressure in patients with essential hypertension and the cardiovascular complications of this disease. Angiotensin II, endothelin 1, and other peptides, as well as catecholamines, may contribute to the altered structure and function of small arteries, in part through their effects on intracellular calcium. Intracellular calcium is implicated in signal transduction pathways that result in smooth muscle cell growth and contraction and in the release of endothelium-dependent agents involved in the regulation of vascular function. VASCULAR EFFECTS OF NEWER ANTIHYPERTENSIVE AGENTS: The newer antihypertensive agents that affect intracellular calcium pathways either directly or indirectly, such as calcium antagonists or angiotensin converting enzyme (ACE) inhibitors, may exert vascular protective effects, partly by regressing vascular remodeling and by correcting endothelial dysfunction. A double-blind trial comparing the effects of the ACE inhibitor cilazapril and the beta-blocker atenolol on media: lumen ratio of small gluteal subcutaneous arteries, which was significantly higher in hypertensive patients before treatment than in normotensive subjects, was corrected after treatment with cilazapril. In contrast, no improvement was seen in the media: lumen ratio of small arteries in the patients receiving atenolol, even after 2 years of treatment. In another study, treatment with the intrinsically long-acting calcium antagonist amlodipine resulted in a reduction in forearm minimal vascular resistance. Patients treated with amlodipine also showed normalized responses of forearm blood flow to nitric oxide synthase inhibition. Furthermore, patients treated with a slow-release formulation of nifedipine exhibited normal media: lumen ratio of small arteries and normal endothelium-dependent relaxation. CONCLUSIONS: These findings appear very promising; however, it remains to be demonstrated whether the apparently beneficial effects of treatment with long-acting calcium antagonists and ACE inhibitors will lead to reductions in the cardiovascular morbidity and mortality associated with hypertension.     

Root stimulation studies in the evaluation of patients with motor neuron disease

Nerve root stimulation may be employed in patients with motor neuron disease (MND) to rule out motor neuropathy with conduction block. The diagnostic utility of these studies is unknown, in part because the range of amplitude changes across nerve root segments in patients with active neuronal degeneration has not been well studied. We reviewed root stimulation studies in 32 patients (59 nerves) with MND and found segmental amplitude reduction from 0 to 45%, a range similar to values reported for normal subjects; there was no suggestion of conduction block based on our usual criteria.