Helicobacter pylori vacuolating toxin accumulates within the endosomal-vacuolar compartment of cultured gastric cells and potentiates the vacuolating activity of ammonia

This study explored the relationship between vacuolating toxin and ammonia in the genesis of Helicobacter pylori-induced vacuolation in cultured human gastric cells and investigated the intracellular sites of toxin accumulation. Neutral red dye uptake and electron microscopy were used in the investigation of the respective roles of, and of the reciprocal interaction between, toxin and ammonia in cell vacuolation and ultrastructural immunocytochemistry was used for the identification of the intracellular sites of internalized toxin. Toxin was found to cause an expansion of the endosomal compartment, where it accumulates after cellular internalization. However, toxin does not form large, neutral red-positive vacuoles unless combined with ammonia, whose moderate vacuolating activity is markedly potentiated by the toxin. It is concluded that the toxin accumulated within the endosomal compartment alters the morphology and function of this organelle and plays a permissive role towards cell vacuolation, possibly by increasing the accumulation of protonated ammonia within endosomes. In turn, ammonia induces excessive dilatation of the endosomes with reciprocal fusion of their membranes, thus causing cytoplasmic vacuolation. © 1997 John Wiley & Sons, Ltd.     

A quantitative electron microscopic study of the intracellular localization of wheat germ agglutinin in retinal neurons

Previous work has established that, following endocytosis, wheat germ agglutinin, like a number of other plasma membrane bound ligands, is transported to the Golgi apparatus-complex. Previous studies that provided qualitative information about the intracellular distribution of internalized wheat germ agglutinin used techniques that precluded any quantitative conclusions about the relative magnitude of the labeling of endosomes, lysosomes, and the Golgi apparatus-complex. Using quantitative ultrastructural autoradiography, this study compares the time course and relative magnitude of labeling of various intracellular compartments to the labeling in the Golgi area. Fifteen minutes after intraocular injection, wheat germ agglutinin is confined to the inner surface of the retina and the immediate subsurface neuropil with little labeling of the retinal ganglion cell perikarya. Thirty minutes after injection, the plasma membrane (6.97 +/- 1.17), endosomes (10.27 +/- 3.98), smooth vesicles and tubules (1.94 +/- 1.66), and lysosomes (2.42 +/- 1.21) of the retinal ganglion cells are labeled, while the Golgi apparatus-complex is not labeled (0.29 +/- 0.25). (Figures in parentheses are the calculated relative radioactive source density +/- the standard deviation.) The relative labeling density of the plasma membrane and endosomes decreases somewhat during the next 90 minutes (plasma membrane, 4.76 +/- 0.67; endosomes, 7.23 +/- 2.02), while the labeling density of smooth vesicles and tubules and of lysosomes rises (smooth vesicles and tubules, 5.56 +/- 0.94; lysosomes, 7.76 +/- 1.56). The Golgi apparatus-complex, which is unlabeled at 30 minutes, is weakly labeled at 2 hours (1.26 +/- 0.28). The fact that the lysosomes are already labeled at 30 minutes while the Golgi apparatus-complex is unlabeled at that time indicates that the transport of wheat germ agglutinin to the Golgi apparatus-complex is a relatively late phenomenon, and suggests that the bulk of the wheat germ agglutinin destined for lysosomes does not pass through the Golgi apparatus-complex.     

Electron microscopic examination of skin and conjunctival biopsy specimens in neuronal storage diseases.

Skin and conjunctival biopsy specimens from fourteen patients with neuronal storage diseases were investigated using an electron microscope. The diseases were Tay-Sachs disease, ceroid-lipofuscinosis (Jansky-Bielschowsky type), Niemann-Pick disease (type B), highly suspected adrenoleukodystrophy, I-cell disease, mucolipidosis of the beta-galactosidase deficient type, Hurler disease, Hunter disease and Morquio disease. This examination provided valuable diagnostic information on some neuronal storage diseases but not on Morquio disease or highly suspected adrenoleukodystrophy. False negative results may sometimes occur using this examination method. However, this examination suggests the usefulness of skin and conjunctival biopsy specimens as a diagnostic tool in some neuronal storage diseases.     

Ultrastructural Changes of Caudate Nucleus in Mice Chronically Treated with Manganese

Manganese (Mn) is able to cross the blood–brain barrier and induces functional and structural alterations during the intoxication by this metal. Therefore, the effects of chronic administration of Mn in the caudate nucleus of mice were evaluated by electron microscopy. Male albino mice were injected intraperitoneally with MnCl₂ (5?mg/kg/d) 5?d per week during 9 weeks. The control group received only 0.9% of NaCl solution. The caudate nuclei were extracted and subsequently processed to be observed on a conventional transmission electron microscope at 2, 4, 6, and 9 weeks after treatment. A high percentage of vacuolated and swollen mitochondria were found throughout all the analyzed periods. Myelin disarrangement and ultrastructural alterations related to edema were observed increased in Mn-treated mice at week 9. Granular degeneration of myelin at week 9 accompanied with deposition of electron dense granules in the neuropil was also observed. Edema in neuropil and glial cells was detected from week 2 to week 9 accompanied by swollen mitochondria. Neuronal bodies, synaptic terminals, and perivascular cells were found swollen. Decreased electron density in postsynaptic areas and decreased and dispersed synaptic vesicles in presynaptic areas were noted in Mn-treated animals. Some neurons from Mn-treated mice showed cisternae dilation of the Golgi apparatus. These results suggest that Mn-treatment produces structural alterations in the caudate nucleus that could be responsible for some of the neurotoxic effects of this metal.     

'Association of platelet function in hypertensive patients with left ventricular hypertrophy'

The present study has used electron microscopic techniques to rapidly detect the success or failure of bone marrow transplantation in three patients with the Chediak-Higashi Syndrome (CHS). The most rapid procedure was the whole mount technique to determine the presence or absence of dense bodies, which are inherently electron-opaque, serotonin-containing storage organelles in platelets. Dense bodies were present in normal numbers in platelets from two patients with successful transplantation and absent in thrombocytes from another patient in whom the transplant had failed.     

Histological and ultrastructural evaluation of human decellularized matrix as a hernia repair device

Recently, interest has been increasing for human decellularized matrices, due to their ability to reduce numerous side effects related to hernia repair. To date, only animal studies investigated the biological interaction post-implant of human decellularized matrices for soft tissue repair. Therefore, the aim of this study was to evaluate the morphological response one year post implant of human decellularized matrix, through morphological analysis of human biopsies. The histological and ultrastructural results revealed a perfect cellular repopulation and neoangiogenesis, with minimal inflammatory response and a well-organized collagen matrix. The results have indicated that this scaffold can be an effective treatment for hernia.     

Clefts in Intradermal Nevi: An Ultrastructural Study

Six intradermal melanocytic nevi, two with and four without intercellular slitlike spaces, were examined by light microscopy and electron microscopy of deparaf-finized tissue. The spaces were partially lined by elongated nevus cells and contained a few collagen fibrils. Other features in both groups included an abundance of (1) immature elastic tissue comprised of aggregates of microfibrils lacking an amorphous component and (2) collagen fibrils that were finer than in normal dermis and did not form compact bundles. We propose that these elastic tissue and collagen fiber changes diminish the resistance of the dermis to the mechanical stress of the biopsy procedure and histologic processing, resulting in the formation of artifactual clefts.     

Atorvastatin induces autophagy in MDA-MB-231 breast cancer cells

This article explores the effects of atorvastatin on cultured breast cancer cells. Our experiment demonstrated that atorvastatin triggered autophagy and inhibited proliferation in breast cancer cells. A CCK8 assay indicated that atorvastatin can inhibit the activity of MDA-MB-231 breast cancer cells. Western blotting results showed that atorvastatin increased the conversion of light chain 3 (LC3)-I to LC3-phosphatidylethanolamine conjugate (LC3-II). Confocal microscopy was used to reveal the appearance of a punctate structure in the cytoplasm, and electron microscopy was used to reveal the formation of double-membrane autophagosome. In conclusion, our study showed that atorvastatin may affect MDA-MB-231 breast cancer cells by inducing autophagy.     

Ocimum sanctum extracts attenuate hydrogen peroxide induced cytotoxic ultrastructural changes in human lens epithelial cells

Hydrogen peroxide (H₂O₂) is the major oxidant involved in cataract formation. The present study investigated the effect of an aqueous leaf extract of Tulsi (Ocimum sanctum) against H₂O₂ induced cytotoxic changes in human lens epithelial cells (HLEC). Donor eyes of the age range 20–40 years were procured within 5–8 h of death. After several washings with gentamicin (50 mL/L) and betadine (10 mL/L), clear transparent lenses (n = 6 in each group) were incubated in Dulbecco's modified Eagle's medium (DMEM) alone (normal) or in DMEM containing 100 µm of H₂O₂ (control) or in DMEM containing both H₂O₂ (100 µm ) and 150 µg/mL of Ocimum sanctum extract (treated) for 30 min at 37 °C with 5% CO₂ and 95% air. Following incubation, the semi‐hardened epithelium of each lens was carefully removed, fixed and processed for electron microscopic studies. Thin sections (60–70 mm) were contrasted with uranyl acetate and lead citrate and viewed under a transmission electron microscope. Normal epithelial cells showed intact, euchromatic nucleus with few small vacuoles (diameter 0.58 ± 0.6 µm) in well‐demarcated cytoplasm. After treatment with H₂O₂, they showed pyknotic nuclei with clumping of chromatin and ill‐defined edges. The cytoplasm was full of vacuoles (diameter 1.61 ± 0.7 µm). The overall cellular morphology was typical of dying cells. Treatment of cells with Ocimum sanctum extract protected the epithelial cells from H₂O₂ insult and maintained their normal architecture. The mean diameter of the vacuoles was 0.66 ± 0.2 µm. The results indicate that extracts of O. sanctum have an important protective role against H₂O₂ injury in HLEC by maintaining the normal cellular architecture. The protection could be due to its ability to reduce H₂O₂ through its antioxidant property and thus reinforcing the concept that the extracts can penetrate the HLEC membrane. Copyright © 2009 John Wiley & Sons, Ltd.