Line tension at lipid phase boundaries regulates formation of membrane vesicles in living cells.

Ternary lipid compositions in model membranes segregate into large-scale liquid-ordered (L(o)) and liquid-disordered (L(d)) phases. Here, we show mum-sized lipid domain separation leading to vesicle formation in unperturbed human HaCaT keratinocytes. Budding vesicles in the apical portion of the plasma membrane were predominantly labelled with L(d) markers 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate, 1,1'-dilinoleyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate, 1,1'-didodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate and weakly stained by L(o) marker fluorescein-labeled cholera toxin B subunit which labels ganglioside GM(1) enriched plasma membrane rafts.

Lipid rafts in anticancer therapy: Theory and practice (Review).

To enlarge and stabilize transient raft patterns, proteins interact with membrane lipids. By lateral movements, these small lipid rafts (LRs) may coalesce into large platforms (raft clusters), allowing the alignment of transmembrane proteins that easily crosslink. The formation of raft clusters permits TNFR superfamily membrane receptors of the Lo phase to subsequently cooperate with transducer and adaptor proteins to create receptosomes and/or signalosomes, even without external ligand binding.

Focal junctions retard lateral movement and disrupt fluid phase connectivity in the plasma membrane.

In cultured keratinocytes, focal junctions are enriched in major constituents of lipid rafts, such as GM1 ganglioside, phosphoinositides, caveolins and flotillins. We have therefore speculated that focal junctions represent superrafts formed by coalescence of microdomains into large areas containing liquid-ordered (L(o)) lipids. Indeed, values of maximal fluorescence recovery after photobleaching revealed that the long-range mobility of cholera toxin B subunit (CTB, marker of L(o)) was approximately 1.

Chemokines, cytokines, and growth factors in keratinocytes and dermal endothelial cells in the margin of chronic diabetic foot ulcers.

Keratinocytes and dermal endothelial cells, excluding leukocytes that infiltrate wounds, are the main source of soluble factors regulating healing of skin ulcers. We used immunohistochemistry to analyze the expression of various chemotactic and growth factors and their receptors in the margin of diabetic foot ulcers and in normal nondiabetic foot skin. Our study found significantly elevated expression of transforming growth factor-beta1 (TGF-beta1) and type I TGF-beta receptors (TGFbetaR1), granulocyte macrophage colony-stimulating factor (GM-CSF), and epidermal growth factor (EGF) in keratinocytes in the ulcer margin (p < 0.

Growth hormone cell phagocytosis in adenohypophysis of mosaic mice: morphological and immunocytochemical electron microscopy study.

An electron microscopy immunocytochemical study was performed to determine the expression pattern of growth hormone (GH) in mosaic mutant mice adenohypophysis. In normal condition GH was restricted to the secretory granules of all growth hormone cells. Mosaic mice adenohypophysis contained growth hormone cells which have distinctive GH labeled secretory granules at the level seen in control animals.

Neurogenic factors in the impaired healing of diabetic foot ulcers.

Background: We hypothesize that the reduced innervation of skin can be observed both in clinically neuropathic and non-neuropathic diabetic foot ulcers and can contribute to low inflammatory cell infiltration.

Materials And Methods: Twenty patients with type 2 diabetes and active foot ulcers, without clinical evidence of peripheral sensory neuropathy (n = 12) and with sensory neuropathy (n = 8) were involved in this study. Biopsies from ulcer margin were examined immunohistochemically.

Expression of natural antimicrobial peptide beta-defensin-2 and Langerhans cell accumulation in epidermis from human non-healing leg ulcers.

Chronic wounds like venous calf and diabetic foot ulcers are frequently contaminated and colonized by bacteria and it remains unclear whether there is sufficient expression of defensins and recruitment of epidermal Langerhans cells in the margin of ulcer compared to normal skin. The aim of this study was to examine immunohistochemically the expression of beta-defensin-2 (hBD2), GM-CSF, VEGF growth factors and accumulation of CD1a+ Langerhans cells (LC) in epidermis from chronic skin ulcers and to compare it to normal skin from the corresponding areas. Studies were carried out in 10 patients with diabetic foot, 10 patients with varicous ulcers of the calf and 10 patients undergoing orthopedic surgery (normal skin for control).

Low recruitment of immune cells with increased expression of endothelial adhesion molecules in margins of the chronic diabetic foot ulcers.

Diabetic foot skin close to an ulcer shows only a few infiltrating cells compared to nondiabetic inflamed tissues. Diabetes is characterized by thickened basement membrane of the blood arterioles and capillaries. This may affect the transcapillary transport of immune humoral factors and cells to the extravascular space.

Transient cerebral ischemia induces delayed proapoptotic Bad translocation to mitochondria in CA1 sector of hippocampus.

Delayed ischemic brain damage is associated with mitochondrial dysfunction, but the underlying mechanisms are not known in detail. Recent data suggest that the process is associated with multidirectional changes in the activities of various proteins located in mitochondria. Of these, the stress-activated kinase JNK is delay-activated postischemia.

Keratinocyte and dermal vascular endothelial cell capacities remain unimpaired in the margin of chronic venous ulcer.

The role of endogenously produced cytokines and growth factors in the impaired healing of chronic leg ulcers remains uncertain. The aim of this study was to determine the functional capacity of skin cells in ulcer bed tissue compared to those in the edge of ulcers and skin distal to ulcers. Biopsies from leg ulcers of ten randomly selected patients were examined immunohistochemically for cytokines and growth factors produced by keratinocytes (KC) and vascular endothelial cells (EC).

Kinetics of Smac/DIABLO release from mitochondria during apoptosis of MCF-7 breast cancer cells.

Smac/DIABLO, a pro-apoptotic protein released from mitochondrial intermembrane space during apoptosis, promotes caspase activation by IAPs neutralization. The kinetics and molecular mechanism of Smac/DIABLO release from mitochondria has remained obscure. Homeostatic confocal microscopy, for the first time, showed the precise kinetics of Smac/DIABLO release from mitochondria during CPT-induced apoptosis in living MCF-7 cells.

Translocation of Bax and Bid to mitochondria, endoplasmic reticulum and nuclear envelope: possible control points in apoptosis.

The cross-talk between endoplasmic reticulum (ER) and mitochondria was investigated during apoptosis in a breast cancer cell line (MCF-7) in culture. The effect of camptothecin, an inducer of apoptosis and a specific inhibitor of topoisomerase I, was investigated by morphological, immunocytochemical and histochemical techniques for electron microscopy. Our ultrastructural morphological data demonstrate alterations in ER configuration and communication with neighbouring mitochondria early after stimulation by camptothecin.

Impaired somatostatin accumulation within the median eminence in mice with mosaic mutation.

Objectives: The mosaic mutation (Atp7a(mo-ms)) linked to X-chromosome is caused by changes in the Atp7a gene encoding CPx-type protein responsible for the ATP-dependent copper transport across cell membranes. Mosaic mutant males represent an animal model for Menkes disease in humans. Starting from the eighth day of life the mosaic males exhibit a progressive decrease in body weight with poor viability and progressive paresis of the hind limbs.

A new look at the cellular scaffold by embedment-free electron microscopy method.

The basic scaffold of most cells is afforded by the cytoskeleton (comprising microfilaments, intermediate filaments and the microtubules). The conventional methods of electron microscopy fail to visualize filamentous cell structure. They can show only these filaments lying at the section surface.

Expression of apoptosis- and cell cycle-related proteins in epidermis of venous leg and diabetic foot ulcers.

Background: Epithelialization of cutaneous ulcers is a long-lasting process. To study the pathomechanism of impaired epithelialization, we evaluated the role of cell cycle- and apoptosis-related proteins in the regenerating epidermis. We characterized immunohistochemically the expression of cell cycle regulators p63, CD29, PCNA, p53, pro- and antiapoptotic proteins bcl2, bax, caspase 3 and DNA breaks, as well as keratin 10, 16 and 17.