Retinal pigment epithelium is protected against apoptosis by alphaB-crystallin

PURPOSE: The degeneration of retinal pigment epithelial (RPE) cells is considered to be a crucial event in the pathophysiology of age-related macular degeneration (AMD). Cumulative oxidative damage has been implicated in the development of the changes seen in AMD. The present study was undertaken to evaluate the expression of the small heat shock protein alphaB-crystallin in the RPE in response to oxidative stress and to explore whether alphaB-crystallin expression confers an antiapoptotic cytoprotective effect on RPE cells. METHODS: Native human RPE cells from the macula and retinal periphery were analyzed by RT-PCR and Western blot analysis for expression of alphaB-crystallin. Monolayer cultures of human RPE cells were stressed by heat shock (42 degrees C for 20 minutes) or oxidant-mediated injury (50-300 micro M H(2)O(2) for 1 hour). Induction of alphaB-crystallin and the corresponding mRNA was assessed by Western and Northern blot analyses. To study the cytoprotective effect of alphaB-crystallin, human RPE cells were transfected with either a neomycin-selectable expression vector containing alphaB-crystallin cDNA or a control vector without alphaB-crystallin cDNA. Caspase-3 activity was determined by observing the cleavage of a colorimetric peptide substrate. Cell viability was quantified by combined propidium iodide and Hoechst 33342 staining. RESULTS: alphaB-crystallin is constitutively expressed in RPE under in vivo and in vitro conditions. Western blot analysis of freshly isolated RPE showed greater baseline expression levels in RPE derived from the macular area than in that from the more peripheral regions. Heat shock treatment and oxidative stress caused a significant increase in alphaB-crystallin mRNA and protein. Oxidant-mediated injury in RPE cells with baseline expression levels of alphaB-crystallin resulted in apoptotic cell death, as measured by caspase-3 activity, whereas RPE cells that had been stably transfected with alphaB-crystallin were more resistant to H(2)O(2)-induced cellular injury. CONCLUSIONS: alphaB-crystallin may function as a stress-inducible antiapoptotic protein in human RPE and is inducible by oxidative stress, a condition implicated in the pathogenesis of AMD. Overexpression of alphaB-crystallin may be an important mechanism for the RPE to prevent apoptotic cell death in response to cellular stress.     

The flavonol quercetin activates basolateral K(+) channels in rat distal colon epithelium

1. The flavonol quercetin has been shown to activate a Cl(-) secretion in rat colon. Unlike the secretory activity of the related isoflavone genistein, quercetin's secretory activity does not depend on cyclic AMP; instead, it depends on Ca(2+). We investigated the possible involvement of Ca(2+) dependent basolateral K(+) channels using apically permeabilized rat distal colon epithelium mounted in Ussing chambers. 2. In intact epithelium, quercetin induced an increase in short-circuit current (I(sc)), which was diminished by the Cl(-) channel blockers NPPB and DPC, but not by glibenclamide, DIDS or anthracene-9-carboxylic acid. The effect of the flavonol was also inhibited by several serosally applied K(+) channel blockers (Ba(2+), quinine, clotrimazole, tetrapentylammonium, 293B), whereas other K(+) channel blockers failed to influence the quercetin-induced increase in I(sc) (tetraethylammonium, charybdotoxin). 3. The apical membrane was permeabilized by mucosal addition of nystatin and a serosally directed K(+) gradient was applied. The successful permeabilization was confirmed by experiments demonstrating the failure of bumetanide to inhibit the carbachol-induced current. 4. In apically permeabilized epithelium, quercetin induced a K(+) current (I(K)), which was neither influenced by ouabain nor by bumetanide. Whereas DPC, NPPB, charybdotoxin and 293B failed to inhibit this I(K), quinine, Ba(2+), clotrimazole and tetrapentylammonium were effective blockers of this current. 5. We conclude from these results that at least part of the quercetin-induced Cl(-) secretion can be explained by an activation of basolateral K(+) channels.     

Gender differences in coping with tension-type headaches

OBJECTIVE: To study gender differences of coping with illness strategies in tension-type headaches. METHOD: We enrolled 89 subjects (50 women, 39 men) suffering from episodic (n = 37) and chronic (n = 52) tension-type headaches (TTH). Patients were required to answer a Freiburg Questionnaire of Coping with Illness (FQCI), a Von Zerssen Depression Scale (D-S), quality-of-life questionnaires, and a headache home diary (over 4 weeks). In addition, pressure pain thresholds (temporal muscles) and Total Tenderness Scores were obtained. RESULTS: While pain intensity, frequency and quality-of-life parameters were basically the same for female and male EPISODIC TTH sufferers, women scored significantly higher on the F3 subscale (distracting and encouraging) of the FQCI and tended to score higher on the F1 subscale (depressive). Among CHRONIC TTH patients, women reported the pain to be more intense (VAS), were more depressed (D-S), and scored lower on several quality-of-life scores. Female chronic TTH sufferers scored significantly lower on the F2 subscale (active coping) and tended to score higher on F5 (denying). CONCLUSIONS: We conclude that pessimistic coping with illness strategies are more frequent in female episodic and chronic TTH sufferers. We would like to recommend special psychologic intervention in particular to female chronic TTH sufferers which would offer counseling in developing active coping skills.     

Reduced levels of specific autobiographical memories in schizophrenia

Autobiographical memory is intrinsically related to the self and personal identity. This study investigated whether both personal episodic memory and semantic memory are impaired in schizophrenia, a disease characterized by an abnormal personal identity. Personal episodic memory and personal semantic memory were investigated in 24 patients with schizophrenia and 24 normal subjects using an autobiographical fluency task and an autobiographical memory inquiry. Autobiographical memory scores and the proportion of specific memories were lower in patients with schizophrenia than in normal subjects. The deficit of personal episodic and semantic memory, as assessed by the autobiographical memory inquiry and the autobiographical fluency task, respectively, was most apparent after the onset of clinical symptoms. Schizophrenia is associated with an impairment of both personal episodic and semantic memory and with a reduction of specific autobiographical memories. Those impairments are consistent with the existence of an abnormal personal identity in patients with schizophrenia.     

Brain death impairs coronary endothelial function

BACKGROUND: To characterize the impact of brain death (BD) on endothelial dysfunction after cardiac transplantation we investigated coronary circulation and vasomotor function in a canine model. METHODS: Left ventricular pressure-volume data (conductance catheter) and coronary blood flow (CBF) were monitored continuously. Endothelium-dependent vasodilatation after acetylcholine and endothelium-independent vasodilation after sodium nitroprusside were assessed before and 3 hr after BD induction (inflation of a subdural balloon). RESULTS: BD led to an initial hyperdynamic reaction with significant (P<0.05) increase of CBF. After 3 hr, CBF decreased significantly (P<0.05). Although before BD, application of acetylcholine led to a monophasic vasodilatative response, after BD a short mild vasodilatation was followed by a longer vasoconstriction. Endothelium-independent vasodilatation remained unchanged. CONCLUSIONS: BD affects coronary circulation by two means: (1) impairment of CBF to decrease in parallel in afterload with consecutive hemodynamic deterioration and (2) severe endothelial dysfunction that may be a contributing factor to posttransplant outcome.     

Modulation of catecholamine responsiveness and beta-adrenergic receptor/adenylyl cyclase pathway during cardiac allograft rejection1 2

BACKGROUND: This study investigated the changes of catecholamine responsiveness and beta-adrenergic receptor/adenylyl cyclase pathway during acute cardiac transplant rejection. METHODS: Isogeneic Lewis to Lewis and allogeneic Dark Agouti (DA) to Lewis rat cardiac transplants were studied 3 and 5 days after heterotopic intraabdominal transplantation (n=6/group). Myocardial blood flow (MBF), left ventricular systolic pressure (LVSP), maximum pressure development (+dP/dt), and end-diastolic pressure (LVEDP) were measured using an intraventricular balloon. Contractile response to dobutamine (5 microg/kg/min) was also assessed. In separate groups beta-adrenergic receptor density and adenylyl cyclase activity were measured in the grafts, in the recipients' native hearts and in native hearts of sham-operated controls. RESULTS: During mild to moderate rejection cardiac function indices remained unchanged, although MBF and contractile response to dobutamine decreased significantly (P<0.05) in the allogeneic group. The beta-adrenergic receptor density was significantly (P<0.05) increased in both isografts and allografts and in the native hearts of allografted recipients in comparison to native hearts of controls. Adenylyl cyclase activity showed a significant decrease (P<0.05) only in allografts. During severe rejection, LVSP and +dP/dt decreased and LVEDP increased in allografts in comparison to isografts (P<0.05). This was accompanied by a significant decrease in MBF, contractile response to dobutamine, beta-adrenergic receptor density, and adenylyl cyclase activity (P<0.05). CONCLUSIONS: Both microcirculatory disturbances and primary alteration in adenylyl cyclase activity may contribute to decreased contractile reserve in mild to moderate cardiac allograft rejection, whereas beta-adrenergic receptor density seems to be also influenced by cardiac denervation. Severe rejection leads to systolic and diastolic heart failure with complex dysregulation of the beta-adrenergic receptor/adenylyl cyclase pathway and impaired microcirculation.     

Selective removal of circulating immune complexes from patient plasma

The principle of a patient-specific immunoadsorber (PsIA) is demonstrated. Studies with model systems (HSA/anti-HSA) on immobilization, stability, and leakage form the basis for the presented fast-performance liquid chromatography (FPLC) and batch experiments, which were conducted using two different protein A adsorbers and autologous and heterologous PsIA systems. Experiments to determine the binding capacity of protein A adsorbers and PsIAs are described. In all experiments, the adsorption of plasma IgG, total protein, and C1q and C3d circulating immune complexes were measured. Plasma of patients with autoimmune diseases (rheumatoid arthritis, systemic lupus erythematosus) was investigated. Analysis was performed in both the initial plasma and the flow-through or supernatant. Results of the investigations using FPLC and batch experiments were compared. Autologous PsIA systems are suitable for the selective removal of elevated levels of circulating immune complexes in the plasma.     

Combined effects of eptifibatide and anticoagulants: differences between LMWH and UH or rH in thrombin generation inhibition but not in platelet aggregation inhibition

Aim of our study was to investigate effects of eptifibatide and anticoagulants on platelet aggregation and thrombin generation under low and high coagulant challenge in tissue factor-activated platelet rich plasma using a model allowing simultaneous determination of the time course of platelet aggregation and thrombin generation. Eptifibatide exerted a dose-dependent anti-aggregating effect under both high and significantly stronger under low coagulant challenge. Combination of eptifibatide and anticoagulants resulted in significant additive prolongation of the lag phase until the onset of platelet aggregation, more pronounced under low coagulant challenge. Under high, but not under low coagulant challenge combination of eptifibatide and anticoagulants had a significant synergistic inhibitory effect on platelet aggregation. Under low coagulant challenge combination of eptifibatide with LMWH, but not with UH, or rH, resulted in significantly reduced thrombin potential, F 1+2 generation, and FXa formation compared to measurements in the absence of eptifibatide. We demonstrate a synergistic effect of eptifibatide and anticoagulants on platelet aggregation inhibition and an additional inhibitory effect of LMWH and eptifibatide on thrombin generation. Our results support the notion that combination of eptifibatide and anticoagulants might be beneficial in atherosclerotic disease to palliate the thrombogenic potency of ruptured atherosclerotic plaques.