Ultrastructural analysis and electron microscopic localization of Nox4 in healthy and atherosclerotic human aorta.

During diverse pathological conditions, vascular smooth muscle cells (SMCs) characteristically change from a quiescent, contractile phenotype to a proliferative, synthetic state, migrate toward the intima, and synthesize excess extracellular matrix. Although reactive oxygen species (ROS) are generally considered to be toxic to cells, recent evidence suggests that they may also modulate multiple signaling pathways. The vascular system contains several sources of ROS, among which NADPH oxidases (NOXes) have been shown to take an important part in the regulation of cell function, with effects on growth and proliferation. In the present study, the authors investigate the ultrastructural features of SMCs and the expression profile of Nox4 in healthy and atherosclerotic human aorta to explore the possibility of a relationship between Nox4 and SMCs differentiation state. The data extend at the level of immunoelectron microscopy previous observations, demonstrating for the first time the precise distribution and the differential expression of Nox4 in the morphologically distinct SMC types of healthy and diseased human aorta.

Ultrastructure of popliteal vein aneurysm.

The term aneurysm is used to indicate a permanent and irreversible localized vascular dilatation that involves all three layers of the blood vessel wall. It may develop in any part of the circulatory system, including veins, and its history, presentation, and management differ depending on the location. A venous aneurysm is defined as a solitary area of fusiform or saccular dilatation occurring in the course of a major vein or connected by a single channel to a major venous structure. The lower extremities are the most frequently affected, with the popliteal vein being the most common site. Although different theories have been advanced, the etiology of the disease remains uncertain. Mechanical stress and/or degenerative changes within the vein wall are believed to represent the most likely causes of venous aneurysm. To date, there are only a few publications dealing with the histological appearance of popliteal vein aneurysm, and no studies that specifically describe its ultrastructural details have been published to our knowledge. In an attempt to fill this gap and to provide better insights into the pathophysiological mechanisms possibly involved in aneurysmal venous disease, we describe the fine structure of popliteal vein wall and valve in health and disease using both scanning and transmission electron microscopy.

Transmission electron microscopy study of the effects produced by wide-band low-power millimeter waves on MCF-7 human breast cancer cells in culture.

Our previous work showed that low-power wide-band millimeter waves (MMW) inhibit the growth of the MCF-7 human breast carcinoma cell line, also causing a marked reduction of the density of microvilli at the apical membrane of the MCF-7 cells, as revealed by scanning electron microscopy. The aim of the present work was to investigate the ultrastructural changes induced by such electromagnetic radiations on this cell line. A transmission electron microscopy study was performed on MCF-7 cells irradiated under the same experimental conditions previously adopted. Transmission electron microscopy analysis revealed several ultrastructural features of the MMW-irradiated cells pertinent to cells subjected to sublethal injury. The antiproliferative effect of the millimeter radiation was confirmed. MMW, in the 52-78 GHz frequency range, act as stress factor on the cells that survive in a non-steady low-mitogenetic metabolic state.

Selective inhibition of tumoral cells growth by low power millimeter waves.

The effects of low power millimetric wave (MMW) radiation on the growth of tumor and healthy cells were studied. A wide-band frequency range between 53.57-78.33 GHz with a radiation density power of 27 x 10(-17) watt/Hz were used. The radiating energy was low enough not to increase the temperature of the cellular samples (cold irradiation). One hour of radiation treatment given every other day to three tumoral human stable cell lines, produced a noticeable inhibition of the cellular growth. The analogous treatment given to two healthy cell lines gave a weak growth stimulation. A scanning electron microscopy study of MCF-7-and K562-irradiated cells revealed that MMW irradiation induced profound morphological changes of the membrane. Finally, we also provided a mechanistic indication, based on millimeter wave spectroscopy of the cells: water is the primary absorber of these electromagnetic waves. Our work provides interesting evidence that wide band low power MMW irradiation, in the appropriate frequency range, could be used in the future as a cold means to cause selective inhibition of tumor cell growth.

MnSOD expression in human atherosclerotic plaques: an immunohistochemical and ultrastructural study.

BACKGROUND: Over the last decades, the role of oxidative stress in atherosclerosis has become well established, and considerable effort has been made to understand the mechanism of action of free radicals within the cardiovascular system. Conversely, relatively little attention has been directed towards the characterization of the antioxidant status of the arterial wall under disease state. Among the antioxidant enzymes, the manganese-dependent and mitochondria-specific isoform of SOD (MnSOD) represents the first line of defense against superoxide radicals attack. To date, the pathological significance of MnSOD in atherosclerosis is still unclear with conflicting data published. METHODS: In the present study, we used immunohistochemical techniques at the light and electron microscopy level in combination with biochemical assays to localize and characterize the activity and expression profiles of MnSOD in healthy and atherosclerotic human aorta. RESULTS: MnSOD has been found to be highly expressed in the atherosclerotic plaques where specifically localized to the foam cells of the lipid-rich regions but not to other (nonfoamy) cell types. No ultrastructural evidence of apoptosis, such as chromatin condensation and membrane blebbing, has been observed in MnSOD-expressing cells. The up-regulation of MnSOD at the protein level has been associated with a parallel, significant increase of its catalytic activity. CONCLUSIONS: Our data demonstrate that MnSOD is not negatively regulated by the prooxidative and proinflammatory environment of the plaque and evidence a regional and cellular selectivity of MnSOD protein expression under disease state. We suggest that MnSOD induction might represent a protective response against the cytotoxic effect of oxidized low-density lipoprotein.

iNOS induction and PARP-1 activation in human atherosclerotic lesions: an immunohistochemical and ultrastructural approach.

BACKGROUND: Several lines of clinical and experimental evidence have demonstrated that reactive oxygen species and nitrogen species are generated in unregulated amounts during diverse cardiovascular disorders. It has been previously reported by our group and others that augmented expression of nitric oxide synthase isoforms is associated with human atherogenesis and that the activity of the enzymes in an atherosclerotic environment may promote the formation of peroxynitrite. Among the downstream mechanisms triggered by oxidants, poly(ADP-ribose) polymerase-1 activation has recently been implicated in the pathogenesis of acute and chronic myocardial dysfunction, diabetes, hypertension, aging, and various forms of shock. METHODS: Based on these observations, we performed immunohistochemical and immunogold labeling analyses to evaluate the expression profile and the subcellular localization of inducible nitric oxide synthase and poly(ADP-ribose) polymerase-1 in healthy and atherosclerotic human aortae. RESULTS: We have demonstrated that inducible nitric oxide synthase colocalizes with poly(ADP-ribose) polymerase-1 within vascular cells of atherosclerotic human aortae. We have reported for the first time, to our knowledge, the ultrastructural localization of poly(ADP-ribose) polymerase-1 within the nuclei of lesional smooth muscle cells. Finally, we have evidenced that poly(ADP-ribose) polymerase-1 induction within cells of the diseased aorta strongly correlates with alterations in mitochondrial morphology. CONCLUSIONS: Our data imply the possibility of a significant role for cross-talk between inducible nitric oxide synthase and poly(ADP-ribose) polymerase-1 in human atherosclerotic lesions. We conclude that the prooxidant milieu of the plaque might exert damaging effects on mitochondria via a poly(ADP-ribose) polymerase-1-mediated mechanism since the absence of the enzyme results in a corresponding lack of changes in mitochondrial morphology. The present report may open avenues for further researches that could have important therapeutic consequences for the treatment of atherosclerosis and its clinical sequelae.

PEG-templated mesoporous silica nanoparticles exclusively target cancer cells.

Mesoporous silica nanoparticles (MSNs) have been proposed as DNA and drug delivery carriers, as well as efficient tools for fluorescent cell tracking. The major limitation is that MSNs enter cells regardless of a target-specific functionalization. Here we show that non functionalized MSNs, synthesized using a PEG surfactant-based interfacial synthesis procedure, do not enter cells, while a highly specific, receptor mediated, cellular internalization of folic acid (FOL) grafted MSNs (MSN-FOL), occurs exclusively in folate receptor (FR) expressing cells. Neither the classical clathrin pathway nor macropinocytosis is involved in the MSN endocytic process, while fluorescent MSNs (MSN-FITC) enter cells through aspecific, caveolae-mediated, endocytosis. Moreover, internalized particles seem to be mostly exocytosed from cells within 96 h. Finally, cisplatin (Cp) loaded MSN-FOL were tested on cancerous FR-positive (HeLa) or normal FR-negative (HEK293) cells. A strong growth arrest was observed only in HeLa cells treated with MSN-FOL-Cp. The results presented here show that our mesoporous nanoparticles do not enter cells unless opportunely functionalized, suggesting that they could represent a promising vehicle for drug targeting applications.