Distribution and type of morphological damage in human nuclear age-related cataracts

The distribution and type of fiber cell damage was evaluated in human age-related nuclear cataracts and in aged normal (non-cataractous) lenses. Ten age-related nuclear cataracts (53 to 89 years old) and four normal lenses (59 to 67 years old) were examined by electron microscopy of fixed Vibratome sections. Images from the adult, juvenile, fetal and embryonic nuclear regions were compared. Each cataractous lens contained a central region of increased light scattering which involved the embryonic and fetal regions with progressively less involvement in the juvenile and adult nuclear regions. Some damaged fiber cells were observed in all specimens, although damage was minor and infrequent in the normal lenses. Degeneration of single or groups of fiber cells was noted in all the adult nuclei of the cataractous lenses, becoming less frequent in the juvenile nuclei. The types of damage included localized voids, multilamellar membrane aggregates, globular bodies, enlarged cells and regions of highly convoluted membranes. The fetal and embryonic nuclei of the cataractous lenses exhibited rare and minor morphological defects, and were virtually identical to the equivalent regions of the normal aged lenses. Examination of cell interfaces in opaque regions of cataractous lenses revealed that the oldest fiber cells sustained apparent membrane loss. Extracellular spaces in the embryonic, fetal and juvenile regions of the cataractous lenses often contained dense deposits, presumably cytoplasmic material lost from adjacent fibers. The results indicate that the region of greatest nuclear opacity, located in the lens center, does not contain any significant cellular damage. This suggests that older fiber cells respond differently to pathological and senescent changes than younger cells made after fetal development. The observed loss of membranes and cytoplasmic material from the oldest fiber cells may be a contributory mechanism in the formation of age-related human nuclear cataracts.     

Two partially unfolded states of Torpedo californica acetylcholinesterase

Chemical modification with sulfhydryl reagents of the single, nonconserved cysteine residue Cys231 in each subunit of a disulfide-linked dimer of Torpedo californica acetylcholinesterase produces a partially unfolded inactive state. Another partially unfolded state can be obtained by exposure of the enzyme to 1-2 M guanidine hydrochloride. Both these states display several important features of a molten globule, but differ in their spectroscopic (CD, intrinsic fluorescence) and hydrodynamic (Stokes radii) characteristics. With reversal of chemical modification of the former state or removal of denaturant from the latter, both states retain their physiochemical characteristics. Thus, acetylcholinesterase can exist in two molten globule states, both of which are long-lived under physiologic conditions without aggregating, and without either intraconverting or reverting to the native state. Both states undergo spontaneous intramolecular thioldisulfide exchange, implying that they are flexible. As revealed by differential scanning calorimetry, the state produced by chemical modification lacks any heat capacity peak, presumably due to aggregation during scanning, whereas the state produced by guanidine hydrochloride unfolds as a single cooperative unit, thermal transition being completely reversible. Sucrose gradient centrifugation reveals that reduction of the interchain disulfide of the native acetylcholinesterase dimer converts it to monomers, whereas, after such reduction, the two subunits remain completely associated in the partially unfolded state generated by guanidine hydrochloride, and partially associated in that produced by chemical modification. It is suggested that a novel hydrophobic core, generated across the subunit interfaces, is responsible for this noncovalent association. Transition from the unfolded state generated by chemical modification to that produced by guanidine hydrochloride is observed only in the presence of the denaturant, yielding, on extrapolation to zero guanidine hydrochloride, a high free energy barrier (ca. 23.8 kcal/mol) separating these two flexible, partially unfolded states.     

The use of nurse practitioners in the acute care setting

A collaborative practice model was initiated in a university hospital to assist resident physicians to coordinate patient care on specialty services. Nurse practitioner (NP) data were collected on daily work activities and categorized as direct care, indirect care, administration, education, and research. Satisfaction surveys were collected from patients, physicians and nursing staff. Data on clinic evaluation and management service provided by the NPs were reported. The study supported the appropriateness of NPs in the acute care setting.     

Respiratory changes due to long-term exposure to urban levels of air pollution: a histopathologic study in humans

STUDY OBJECTIVES: To evaluate the potential associations between long-term exposure to air pollution and histopathologic evidence of damage to the lungs in humans. DESIGN: Lung tissue samples were collected during necropsies of individuals who died due to violent causes, selected on the basis of their exposure background. PATIENTS: The exposed group was composed of individuals who lived in Guarulhos, an area with high mean levels of inhalable particles. The control group was composed of individuals who lived in two cities with economies based on agricultural activities: Ribeir?o Preto and Ourinhos. Interventions: Information about cigarette smoking and occupational exposure was obtained from family members. MEASUREMENTS AND RESULTS: Morphometric evaluation of the main bronchus was conducted to determine the volume ratio of submucosal glands. Histopathologic alterations of the bronchioli were evaluated by scoring the presence of inflammatory reaction, wall thickening, and secretory hyperplasia. The number of spots of carbon deposition was counted along the regions of lymphatic drainage (visceral pleura and axial connective tissue around bronchi and blood vessels). Statistical analysis was done by means of regression models controlled for age, smoking, and occupational exposure. Lungs collected from the high pollution area presented evidence of more histopathologic damage in comparison to those from the clean environments. These effects were observed even after controlling for individual differences in age, sex, and cigarette smoking levels. CONCLUSIONS: These results suggest that long-term exposure to air pollution may contribute to the pathogenesis of airway disease, and that urban levels of air pollution have adverse effects on the respiratory tract.     

Detection of resistance to amphotericin B among Cryptococcus neoformans clinical isolates: performances of three different media assessed by using E-test and National Committee for Clinical Laboratory Standards M27-A methodologies

Although reliable detection of resistance in vitro is critical to the overall performance of any susceptibility testing method, the recently released National Committee for Clinical Laboratory Standards M27-A methodology for susceptibility testing of yeasts discriminates poorly between resistant and susceptible isolates of Candida spp. We have previously shown that both substitution of antibiotic medium 3 for RPMI 1640 medium in the microdilution variant of the M27-A method and use of the E-test agar diffusion methodology permit detection of amphotericin B-resistant Candida isolates. To determine the relevance of these observations to Cryptococcus neoformans, we have evaluated the performances of both the M27-A and the E-test methodologies with this yeast using three different media (RPMI 1640 medium, antibiotic medium 3, and yeast nitrogen base). As with Candida, we found that only antibiotic medium 3 permitted consistent detection of resistant isolates when testing was performed in broth by the M27-A method. When testing was performed by the E-test agar diffusion method, both RPMI 1640 medium and antibiotic medium 3 agar permitted ready detection of the resistant isolates. Reading of the results after 48 h of incubation was required for testing in broth by the M27-A method, while the MIC could be determined after either 48 or 72 h when the agar diffusion method was used.     

Arachidonic acid is an autocoid mediator of the differential action of 1,25-(OH)2D3 and 24,25-(OH)2D3 on growth plate chondrocytes

Prior studies have shown that 24,25-(OH)2D3 and 1,25-(OH)2D3 regulate protein kinase C (PKC) in costochondral chondrocytes in a cell maturation-dependent manner, with 1,25-(OH)2D3 affecting primarily growth zone (GC) cells and 24,25-(OH)2D3 affecting primarily resting zone (RC) cells. In addition, 1,25-(OH)2D3 has been shown to increase phospholipase A2 activity in GC, while 24,25-(OH)2D3 has been shown to decrease phospholipase A2 activity in RC. Stimulation of phospholipase A2 in GC caused an increase in PKC, whereas inhibition of phospholipase A2 activity in RC cultures increased both basal and 24,25-(OH)2D3-induced PKC activity, suggesting that phospholipase A2 may play a central role in mediating the effects of the vitamin D metabolites on PKC. To test this hypothesis, RC and GC cells were cultured in the presence and absence of phospholipase A2 inhibitors (quinacrine and oleyloxyethylphosphorylcholine [OEPC]), phospholipase A2 activators (melittin and mastoparan), or arachidonic acid alone or in the presence of the target cell-specific vitamin D metabolite. PKC specific activity in the cell layer was determined as a function of time. Phospholipase A2 inhibitors decreased both basal and 1,25-(OH)2D3-induced PKC activity in GC. When phospholipase A2 activity was activated by inclusion of melittin or mastoparan in the cultures, basal PKC activity in RC was reduced, while that in GC was increased. Similarly, melittin and mastoparan decreased 24,25-(OH)2D3-induced PKC activity in RC and increased 1,25-(OH)2D3-induced PKC activity in GC. For both cell types, the addition of arachidonic acid to the culture media produced an effect on PKC activity that was similar to that observed when phospholipase A2 activators were added to the cells. These results demonstrate that vitamin D metabolite-induced changes in phospholipase A2 activity are directly related to changes in PKC activity. Similarly, exogenous arachidonic acid affects PKC in a manner consistent with activation of phospholipase A2. These effects are cell maturation- and time-dependent and metabolite-specific.