Calmodulin and cyclic nucleotide phosphodiesterase activities in erythrocytes from normal and schizophrenic subjects

Calmodulin and cyclic nucleotide phosphodiesterase activities were measured in hemolysates prepared from 18 normal and 17 schizophrenic subjects. No significant difference between groups was found for either activity. The results suggest that calmodulin is present in normal amounts in patients with schizophrenia. This is compatible with the idea that the interaction of calmodulin with antipsychotic agents is structurally non-specific.     

Resonance energy transfer studies of the mechanisms of microclustering of lentil lectin membrane receptors on HeLa cells

The association and dissociation mechanisms of lectin membrane receptor microclustering on HeLa cells have been studied by measuring resonance energy transfer between fluoresceinated and rhodaminated lentil lectin. Compounds known to affect membrane receptor mobility, such as Ca²⁺ ions, methylamine, cytochalasin D and nocodazole, did not modify the association kinetics nor the maximal energy transfer values at 4 and 37 °C. Dissociation of the membrane receptor microclusters was followed by measuring the temporal decrease in energy transfer values at 4 °C after preincubation for different time intervals at 37 °C. The rate of dissociation of the lectin receptors decreased in the presence of Ca²⁺ ions (10⁻³ M) and after cross-linking with anti-lectin antibodies. An increase was observed in the presence of cytochalasin D (10⁻⁶ M) and, to a lesser extent, of methylamine (10⁻² M). When cytochalasin D and methylamine were combined at subliminal concentrations, a partial synergistic effect was observed. Nocodazole (10⁻⁶ M) had no effect. The results suggest that the association of lectin membrane receptors in microclusters is mediated only by physicochemical parameters. Ca²⁺ ions, cytochalasin D (microfilaments) and methylamine (transglutaminase)-sensitive components appear, however, to play an important role in the stabilization of the receptor microclusters.     

Reversible uncoupling of energy transfer between phycobilins and chlorophyll in Anacystis nidulans. Light stimulation of cold-induced phycobilisome detachment

Phycobilin fluorescence of Anacystis nidulans grown at 28°C increases substantially upon cooling below 10°C. A maximal increase is found around ?5°C and amounts to 300%, with almost complete reversibility upon re-warming. Illumination with actinic light leads to considerable stimulation of the cold-induced phycobilin fluorescence increase. Analysis of the light stimulation phenomenon reveals: (1) Actinic illumination shifts the fluorescence-temperature characteristic by about 3°C upwards on the T-axis. At temperatures below 5°C the light stimulating effect becomes smaller again and fluorescence-temperature characteristics measured at high and low light intensity converge around ?5°C. (2) In the 13-8°C region a large (up to 100%) light-induced phycobilin fluorescence increase is observed, while only negligible changes occur in the dark. (3) 3-(3,4-Dichlorophenyl)-1,1-dimethyl urea (DCMU) as well as uncouplers inhibit the light stimulation, which hence depends on coupled electron transport.In agreement with previous work (Schreiber, U. (1979) FEBS Lett. 107, 4–9) it is concluded that illumination enhances cold-induced phycobilisome detachment by increasing the net negative charge at the outer surface of the thylakoid membrane. The possible role of a fluid → ordered transition of membrane lipids (Murata, N. and Fork, D.C. (1975) Plant Physiol. 56, 791–796) is discussed.     

Discriminant analysis on cells from developing squamous cancer of the respiratory tract

Cytologic preparations made from the tracheobronchial tree taken by the Schreiber catheter have been scanned by three color microphotometry. The digitized cell images were processed by the analytical cytodiagnostic programs of the TICAS system. Cells were sorted into two control groups and five groups of increasing atypia ranging from normal epithelium to invasive squamous cell carcinoma. Standard statistical tests, including Wilk's Lambda, Rao's V, and the Kruskal-Wallis tests are performed on these subsets of cell image features. This study demonstrates that discriminant analyses permit differentiation between normal cells and those from marked atypia or carcinoma and that the classification achieves a high degree of agreement with visual assignment.     

Heat-induced changes of chlorophyll fluorescence in isolated chloroplasts and related heat-damage at the pigment level

The heat-induced changes of chlorophyll fluorescence excitation and emission properties were studied in isolated chloroplasts of Larrea divaricata Cav. An analysis of the temperature dependency of fluorescence, under Fo and Fmax conditions, of temperature-jump fluorescence induction kinetics, and of 77 degrees K emission spectra of preheated chloroplasts revealed two major components in the heat-induced fluorescence changes: (1) a fluorescence rise, reflecting the block of Photosystem II reaction centers; and (2) a fluorescence decrease, caused by the functional separation of light-harvesting pigment protein complex from the rest of the pigment system. Preferential excitation of chlorophyll a around 420 nm, produced a predominant fluorescence rise. Preferential excitation of chlorophyll b, at 480 nm, gives a predominant fluorescence decrease. It is proposed that the overlapping of the fluorescence decrease on the somewhat faster fluorescence rise, results in the biphasic fluorescence rise kinetics observed in isolated chloroplasts. Both the rise component and the decay component are affected by the thermal stability of the chloroplasts, acquired during growth of the plants in different thermal environments. Mg2+ enhances the stability against heat-damage expressed in the decrease component, but has no effect on the rise component. Heat pretreatment leads to a decrease of the variable fluorescence in the light-induced 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) rise curve, but no change in half-rise time is observed. It is concluded that the block of Photosystem II reaction centers precedes the loss of the light-harvesting pigment protein complex. However, the approximately antiparallel heat-induced Fmax decrease and Fo increase suggest a common cause for the two events. A heat-induced perturbation of the thylakoid membrane is discussed.     

Extracellular vesicle‐mediated crosstalk between NPCE cells and TM cells result in modulation of Wnt signalling pathway and ECM remodelling

Primary open‐angle glaucoma is a leading cause of irreversible blindness, often associated with increased intraocular pressure. Extracellular vesicles (EVs) carry a specific composition of proteins, lipids and nucleotides have been considered as essential mediators of cell‐cell communication. Their potential impact for crosstalk between tissues responsible for aqueous humour production and out‐flow is largely unknown. The study objective was to investigate the effects of EVs derived from non‐pigmented ciliary epithelium (NPCE) primary cells on the expression of Wnt proteins in a human primary trabecular meshwork (TM) cells and define the mechanism underlying exosome‐mediated regulation that signalling pathway. Consistent with the results in TM cell line, EVs released by both primary NPCE cells and NPCE cell line showed diminished pGSK3β phosphorylation and decreased cytosolic levels of β‐catenin in primary TM cells. At the molecular level, we showed that NPCE exosome treatment downregulated the expression of positive GSKβ regulator‐AKT protein but increased the levels of GSKβ negative regulator‐PP2A protein in TM cells. NPCE exosome protein analysis revealed 584 miRNAs and 182 proteins involved in the regulation of TM cellular processes, including WNT/β‐catenin signalling pathway, cell adhesion and extracellular matrix deposition. We found that negative modulator of Wnt signalling miR‐29b was abundant in NPCE exosomal samples and treatment of TM cells with NPCE EVs significantly decreased COL3A1 expression. Suggesting that miR‐29b can be responsible for decreased levels of WNT/β‐catenin pathway. Overall, this study highlights a potential role of EVs derived from NPCE cells in modulating ECM proteins and TM canonical Wnt signalling.