Abstract: Histological studies in organs like gill, liver, muscle and brain of Cyprinus carpio communis were made to assess tissue damage due to sublethal concentration of heavy metals lead and cadmium after 28 days of exposure. In lead treated gill, disintegration and fusion of primary lamellae, extensive vacuolization with disruption of epithelial lining was observed, whereas on sublethal exposure to cadmium, hyperplasia of branchial arch, vacuolization and congestion of blood vessels were well marked. Metal accumulation was clearly visible in treated liver with degeneration and severe necrosis. Both lead and cadmium treated fish showed marked thickening and separation of muscle bundles with severe intramuscular oedema more pronounced in sublethal treatment of cadmium. Neuronal cell degeneration, swelling of pyramidal cells, vacuolization and dystrophic changes were characteristic features observed in treated brain. Key words : Lead, cadmium, histopathology, Cyprinus carpio communis.

Abstract: During this study, topographic changes in healthy and atretic granulosa cells have been investigated during follicular atresia in goat ovary. Under scanning electron microscopy atresia was marked by asymmetrical shrinkage and vacuolization of cytoplasm. The specific topographical alterations observed in atretic cells were loss of micro extensions, disruption of cell–cell interaction, and smooth-textured membrane with a number of uneven depressions and ruffles. Some portions of the cell membrane were marked by extensive shrinkage due to condensation of cytosol. Irregular membrane at occasions was studded with blunt microextensions. The findings of present investigation will help in understanding the cellular changes in granulosa cells during follicular atresia and will find applications in screening of follicles for in vitro culture, in vitro fertilization and Embryo transfer technology. SCANNING 33: 21–24, 2011. © 2011 Wiley Periodicals, Inc.

Abstract: Indium, a IIIA group element of the periodic chart, has many medical uses for diagnostic and clinical investigations in humans. This element is also used in industry and in nuclear fields where released streams can contaminate environment. Consequently, indium can reach humans mainly by natural ways. In this work, we attempted to study the incidence of this element on the food intake and body and testicle weights of rat, as well as the histological and the ultrastructural consequences of its presence in testicles using conventional transmission electron microscopy. Our study showed that this element induced a significant decrease in the food intake and body and testicles weights and caused necrosis and vacuolization in germinal cells. The ultrastructural observations showed the presence of electron-dense deposits characteristic of indium in the lysosomes of Leydig and Sertoli cells as well as sufferance in mitochondria of indium-treated rats. Despite the role of lysosome in the protection of living cells, by sequestration and concentration of indium in testicle cells under insoluble form, it is probable that this element has noxious effects on food intake and body and testicles weight and induces necrosis on seminal tissues of treated rats. Microsc. Res. Tech., 2010. © 2010 Wiley-Liss, Inc.

Abstract: In this report, we describe an effect of photodynamic therapy (PDT) on membrane trafficking in murine 1c1c7 hepatoma cells. A brief exposure of 1c1c7 cells to a 20 nM concentration of the phosphatidylinositol kinase class-3 antagonist wortmannin led to the rapid appearance of cytoplasmic vacuoles. Fluorescence monitoring of plasma membrane-associated 1-[4-(trimethylamino)phenyl]-6-phenylhexa-1,3,5-triene (TDPH) over time demonstrated that the wortmannin-induced vacuoles were derived from endocytosed plasma membrane. Low-dose photodamage catalyzed by the lysosomal photosensitizer NPe6, prior to the addition of wortmannin, prevented formation of these vacuoles. NPe6 was found to suppress for several hours the normal trafficking of TDPH-labeled plasma membrane to the cytosol, and the formation of punctate TDPH-labeled cytoplasmic vesicles. The ability of NPe6-induced photodamage to suppress wortmannin-induced vacuolization occurred under conditions that did not disrupt lysosomes and were at or below the threshold of cytostatic/cytotoxic effects. Furthermore, the suppressive effects of NPe6-PDT were not prevented by inclusion of an agent that stabilized lysosomal membranes, or by E64d, an inhibitor of lysosomal cathepsin proteases. Mitochondrial photodamage was less effective at preventing wortmannin-induced vacuole formation and PDT directed against the ER had no effect. The role of photodamage to the endocytic pathway may be a hitherto unexplored effect on cells that selectively accumulate photosensitizing agents. These results indicate that photodamage directed against endosomes/lysosomes has effects independent of the release of lysosomal proteases.

Abstract: Inhibiting Hsp90 chaperone roles using 17AAG induces cytostasis or apoptosis in tumor cells through destabilization of several mutated cancer promoting proteins. Although mitochondria are central in deciding the fate of cells, 17AAG induced effects on tumor cell mitochondria were largely unknown. Here, we show that Hsp90 inhibition with 17AAG first affects mitochondrial integrity in different human tumor cells, neuroblastoma, cervical cancer and glial cells. Using human neuroblastoma tumor cells, we found the early effects associated with a change in mitochondrial membrane potential, elongation and engorgement of mitochondria because of an increased matrix vacuolization. These effects are specific to Hsp90 inhibition as other chemotherapeutic drugs did not induce similar mitochondrial deformity. Further, the effects are independent of oxidative damage and cytoarchitecture destabilization since cytoskeletal disruptors and mitochondrial metabolic inhibitors also do not induce similar deformity induced by 17AAG. The 1D PAGE LC MS/MS mitochondrial proteome analysis of 17AAG treated human neuroblastoma cells showed a loss of 61% proteins from membrane, metabolic, chaperone and ribonucleoprotein families. About 31 unmapped protein IDs were identified from proteolytic processing map using Swiss-Prot accession number, and converted to the matching gene name searching the ExPASy proteomics server. Our studies display that Hsp90 inhibition effects at first embark on mitochondria of tumor cells and compromise mitochondrial integrity.

Abstract: Calcineurin inhibitors (CNI) have been commonly used as pivotal immunosuppressive agents to renal transplant recipients and have contributed significantly to improving short-term allograft survival. However, long-term administration of CNI may cause an adverse effect on kidney function, known as chronic nephrotoxicity. Chronic CNI nephrotoxicity (CNI-NT) shows characteristic histopathological findings that involve arteriolar hyalinosis. Recently, the term alternative arteriolar hyalinosis (aah) is used to discriminate CNI-specific arteriolar hyaline deposition from non-specific arteriolar hyalinosis. We studied whether arteriolar vacuolization represents an early lesion of aah as a predictor of CNI-NT. We retrospectively studied the 79 patients under treatment with a CNI immunosuppressant, who underwent living-related renal transplantation (RTx) from January 2007 to March 2009. We examined serial protocol graft biopsies at one h, one, six, and 12 months after RTx. We classified histological findings into two groups on the basis of aah lesion (with or without aah) in serial biopsies for 12 months. Arteriolar vacuolization was more frequently observed in the aah group than in the non-aah group with a significant difference. Arteriolar vacuolization was found even in the one-h biopsy specimens, indicating a non-specific histopathological finding. But in the aah group, arteriolar vacuolization tended to be more frequently observed later on. Aah can be a predictor of CNI-NT.

Abstract: Antisense oligonucleotides (AON) delivered via inhalation are in drug development for respiratory diseases. In rodents and monkeys, repeated exposure to high doses of inhaled phosphorothioate (PS) AON can lead to microscopic changes in the lungs, including accumulation of alveolar macrophages in the lower airway that have a foamy appearance. The functional consequences that result from this morphological change are unclear as there is controversy whether the vacuoles/inclusion bodies reflect normal clearance of the inhaled AON or are early indicators of lung toxicity. The morphological and functional responses of macrophage to PS AON were characterized in vitro using the comparator drug amiodarone, as a known inducer of foamy macrophages. Morphological changes of increased vacuolization with the presence of lamellated structures were observed in macrophages in response to both amiodarone and AON treatment. Functional responses to the drugs clearly differed with amiodarone treatment leading to apoptosis of cells and cell death, release of proinflammatory mediators IL-1RA, MIP-1 α and TNF α , decrease in IP-10, a cytokine shown to be involved in protection against pulmonary fibrosis and altered phagocytosis capacity of the cells. In contrast, AON in concentrations up to 30 μM, had no effect on cell viability or apoptosis, had minimal effects on pro-inflammatory cytokines, increased IP-10 levels and did not alter the phagocytic capacity of the cells. Exposure of macrophages to AON in vitro , led to morphological changes of increased vacuolization, but did not lead to functional consequences which were observed with another vacuolization-inducing drug, suggesting that the in vivo phenotypic changes observed following inhalation of AON may be consistent with a clearance mechanism and not an activation or impairment of macrophages.

Abstract: Mercury is the highly toxic metal that exerts its adverse effect on health of human and animals through air, soil water and food. For the present study, the toxic effect was observed by oral administration of mercuric chloride daily for 4 weeks at doses ranging from of 0 to 8 mg/kg/day. Histopathology was also carried out of both for kidney and liver. Significantl increased in lipid peroxidation and decreased Superoxide dismutase enzymes were observed in treated groups as compared to control . The gross changes were characterized by swellingand paleness of kidney and liver. Microscopic examination has revealed liver revealed hemorrhage, leukocytic infiltration, hepatocyte hypertrophy, necrosis and increased vacuolization incidence and the severity increased with increase treatment dose. In addition kidney showed proteineous cast in the lumen of tubules, narrowing of lumen, infiltration and necrosis.

Abstract: We have previously found that the weak base 4-aminopyridine induces Brownian motion of acidic organelles around which vacuoles are formed, causing organelle traffic disorder in neurons. Our present study investigated the characteristics of vacuoles induced by weak bases (NH(4)Cl, aminopyridines, and chloroquine) using mouse cells. Individual vacuoles included acidic organelles identified by fluorescent protein expression. Mitochondria and actin filaments were extruded outside the vacuoles, composing the vacuole rim. Staining with amine-reactive fluorescence showed no protein/amino acid content in vacuoles. Thus, serous vacuolar contents are probably partitioned by viscous cytosol, other organelles, and cytoskeletons, but not membrane. The weak base (chloroquine) was immunochemically detected in intravacuolar organelles, but not in vacuoles. Early vacuolization was reversible, but long-term vacuolization caused cell death. The vacuolization and cell death were blocked by the vacuolar H(+)-ATPase inhibitor and Cl--free medium. Staining with LysoTracker or LysoSensor indicated that intravacuolar organelles were strongly acidic and vacuoles were slightly acidic. This suggests that vacuolization is caused by accumulation of weak base and H(+) in acidic organelles, driven by vacuolar H(+)-ATPase associated with Cl(-) entering, and probably by subsequent extrusion of H(+) and water from organelles to the surrounding cytoplasm.

Abstract: Nephrosialidosis is a rare subgroup of sialidosis characterized by a progressive and fatal course of nephropathy. The authors report a 2-year-old boy who had suffered from steroid-resistant nephrotic syndrome. Renal biopsy showed diffuse and severe vacuolization of glomerular and tubular epithelial cells. The vacuoles were ultrastructurally membrane bound, most of which were not empty but contained electron-dense material lining the inner surface of the membrane. The pathologic changes were consistent with neuraminidase-deficient disorders, which was later confirmed by biochemical analysis. Although rare, nephrosialidosis should be considered in children with steroid-resistant nephrotic syndrome if renal biopsies exhibit severe vacuolar degeneration of renal epithelial cells.

Abstract: Heterocyclic compounds such as quinazolinones have variety of biological and pharmacological properties (anticancer, antiinflammatory, antimicrobial, antimalaria, etc.). Effects of two new quinazolinones viz., 4(3H)-quinazolinone-2-propyl-2-phenylethyl (QPPE) and 4(3H)quinazolinone-2-ethyl-2-phenylethyl (QEPE) were investigated on Balb/C mice embryos livers-the major organ of metabolism and detoxification of drugs and toxins. Histological and pathological studies demonstrated QPPE and QEPE as producers of toxic metabolites after biotransformation, creating necrosis, fatty changes, increase in the number of band cells, hepatocytes' diameters and alkaline phosphatase, in addition to sinusoid dilation, hemorrhages and hyperemia. Transmission electron micrographs showed lipid droplets in hepatocytes' cytoplasm, necrosis, vacuolization, cytoplasm disintegration, disfigured and swollen mitochondria, irregular and abnormal nuclei, nuclei with heterochromatin, condensed chromatins, myelin figures and autophages in injured hepatocytes. In conclusion, QPPE and QEPE make toxic components after biotransformation injuring membranes and creating inflammatory reactions. They also disturb metabolism of lipids pathways and cause the appearances of lipid droplets in hepatocytes.

Abstract: We examined ultrastructural modifications in cortical neurons of the frontal part of the cerebral hemispheres of rats under conditions of chronic intoxication of the animals by tert-butyl methyl ether (TBME). Everyday doses of intrastomachic introductions of TBME in the experimental groups were 0.5, 5, 50, and 500 mg/kg. Within 60 days of the experiment, the animals preserved their viability even in the case of the highest TBME doses, but significant negative structural changes were observed in neocortical neurons under these conditions. A significant part of the mitochondria demonstrated swelling, decreases in the number of the cristae, destruction of the external membrane, and, finally, transformation into vacuoles. The endoplasmic reticulum underwent comparable modifications (dilation and destruction of the cisterns and intense vacuolization). The osmiophilia of the neuronal nuclei increased, and the latter were also deformed; the integrity of the nuclear envelope was disturbed. Other cellular organelles also underwent spatial redistribution and destruction. All these processes led to death of a considerable part of cortical neurons because of intense apoptosis followed by phagocytation of the apoptotic bodies by microgliocytes. The above-described modifications progradiently increased throughout the experiment (up to the 60th day). Their pattern was most dramatic at the highest TBME dose used (500 mg/kg), but the corresponding manifestations were quite obvious (although mild) even at the lowest doses (0.5 and 5 mg/kg). High polymorphism and variability were characteristic features of TBME-induced ultrastructural modifications in the rat frontal cortex. Therefore, TBME, an agent extensively used in industry and transport, which is believed to demonstrate relatively low integral toxicity, exerts powerful negative neurotropic effects on neuronal systems in the cerebral cortex and dramatically intensifies apoptosis in these structures.

Abstract: There is a still concern about the effects of organotin compounds on freshwater organisms especially on their reproductive system. We investigated the toxicity of TBTCl and its effects on ovary of freshwater prawn, Macrobrachium kistnensis at different concentrations in present study. After exposure of different concentrations as 0.33 ppm, 0.26 ppm, 0.17 ppm and 0.09 ppm LC50 values for 24 h, 48 h, 72 h and 96 h respectively of tributyltin chloride, the ovaries showed different changes in their architecture. There was a destruction of epithelial layer and evidence of degeneration of oocytes and disorganization of nucleus after treatment of 0.26 ppm of TBTCl. Vacuolization at periphery, degenerating oocytes with disintegrated nuclei and vacuolization and alteration in shape was observed for of 0.09 ppm.

Abstract: Our previous studies have shown that taxol, a potent anticancer agent, induces caspase-independent cell death and cytoplasmic vacuolization in human lung cancer cells. However, the mechanisms of taxol-induced cytoplasmic vacuolization are poorly understood. Cytoplasmic vacuolization have been reported to be a characteristic of cell senescence. Here, we employed confocal fluorescence microscopy imaging to study the reversibility of taxol-induced cytoplasmic vacuolization and whether taxol triggers senescence in A549 cells. We found that taxol-induced cytoplasmic vacuolization at 6 or 9 h after treatment with taxol did not decrease but increase at 24 h or 72 h after refreshing the culture medium without taxol, indicating taxol-induced cytoplasmic vacuolization is irreversible. We used SA-β-Gal (senescence-associated β-galactosidase) to assess whether taxol-induced cell death in cytoplasmic vacuolization fashion is senescence, and found that hydrogen peroxide (H 2 O 2 )-treated, but not taxol-treated cells is significantly stained by the SA-β-Gal, a senescence testing kit, indicating that the form of taxol-induced cell death is not senescence.

Abstract: Pompe disease also known as glycogen storage disease type II (GSD type II) is a lysosomal disorder due to alfa-glucosidase deficiency, a key enzyme in glycogen degradation. The juvenile and adult forms, considered late-onset GSDII, are mainly characterized by slowly progressive muscle disorders mimicking limb-girdle dystrophies or inflammatory myopathies, and or by respiratory involvement with diaphragmatic paralysis and restrictive respiratory insufficiency. The histopathological hallmark is revealed by increased muscle fiber vacuolization with vacuoles filled of PAS-positive material and by strong reaction for lysosomal acid phosphatase. The degree of vacuolization is extremely variable in late-onset patients, and seems independent from age of onset, disease duration, or clinical features. In the last few years, major advances in this field have been represented by the development and manufacturing of recombinant human GAA (rhGAA) produced and purified from Chinese hamster ovary cells (CHO) for enzyme replacement therapy (ERT).

Abstract: Midgut apoptosis was investigated in the brown planthopper(Nilaparvata lugens Stl,Hemiptera:Delphacidae) larvae.A terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling(TUNEL) assay revealed that two to five percent of the 1st -5th larval instar midgut cells showed typical fluorescent signals representing genomic DNA cleavage.4',6 -diamidino-2-phenylindole(DAPI) staining revealed apoptotic nuclei presenting characteristic features in apoptosis stage Ⅰ,Ⅱ a and Ⅱ b;including chromatin condensation,margination and nuclear fragmentation.Transmission electron microscopy revealed that early stage apoptotic midgut cells had apoptotic nuclear morphology,such as chromatin condensation and margination,while late stage cells had broken nuclei,apoptotic bodies and vacuolization of the cytoplasm.These findings demonstrate that a small percentage of the midgut cells of N.lugens larvae undergo apoptosis during normal development.It may be possible to regulate N.lugens midgut apoptosis artificially thereby making this process a novel method of control for this species.

Abstract: We have previously found that the weak base 4-aminopyridine induces Brownian motion of acidic organelles around which vacuoles are formed, causing organelle traffic disorder in neurons. Our present study investigated the characteristics of vacuoles induced by weak bases (NH 4 Cl, aminopyridines, and chlo- roquine) using mouse cells. Individual vacuoles included acidic organelles identified by fluorescent protein expression. Mitochondria and actin filaments were extruded outside the vacuoles, composing the vacuole rim. Staining with amine-reactive fluorescence showed no protein/amino acid content in vacuoles. Thus, serous vacuolar contents are probably partitioned by viscous cytosol, other organelles, and cytoskeletons, but not membrane. The weak base (chloroquine) was immunochemically detected in intravacuolar organelles, but not in vacuoles. Early vacuolization was reversible, but long-term vacuolization caused cell death. The vacuolization and cell death were blocked by the vacuolar H + -ATPase inhibitor and Cl - -free medium. Stain- ing with LysoTracker or LysoSensor indicated that intravacuolar organelles were strongly acidic and vacu- oles were slightly acidic. This suggests that vacuolization is caused by accumulation of weak base and H + in acidic organelles, driven by vacuolar H + -ATPase associated with Cl - entering, and probably by subsequent extrusion of H + and water from organelles to the surrounding cytoplasm. (Folia Histochemica et Cytobio- logica 2011; Vol. 49, No. 2, pp. 272-279)

Abstract: The epsilon-toxin of Clostridium perfringens forms a heptamer in the membranes of Madin–Darby canine kidney cells, leading to cell death. Here, we report that it caused the vacuolation of Madin–Darby canine kidney cells. The toxin induced vacuolation in a dose-dependent and time-dependent manner. The monomer of the toxin formed oligomers on lipid rafts in membranes of the cells. Methyl-β-cyclodextrin and poly(ethylene glycol) 4000 inhibited the vacuolation. Epsilon-toxin was internalized into the cells. Confocal microscopy revealed that the internalized toxin was transported from early endosomes (early endosome antigen 1 staining) to late endosomes and lysosomes (lysosomal-associated membrane protein 2 staining) and then distributed to the membranes of vacuoles. Furthermore, the vacuolation was inhibited by bafilomycin A1, a V-type ATPase inhibitor, and colchicine and nocodazole, microtubule-depolymerizing agents. The early endosomal marker green fluorescent protein–Rab5 and early endosome antigen 1 did not localize to vacuolar membranes. In contrast, the vacuolar membranes were specifically stained by the late endosomal and lysosomal marker green fluorescent protein–Rab7 and lysosomal-associated membrane protein 2. The vacuoles in the toxin-treated cells were stained with LysoTracker Red DND-99, a marker for late endosomes and lysosomes. A dominant negative mutant of Rab7 prevented the vacuolization, whereas a mutant form of Rab5 was less effective. These results demonstrate, for the first time, that: (a) oligomers of epsilon-toxin formed in lipid rafts are endocytosed; and (b) the vacuoles originating from late endosomes and lysosomes are formed by an oligomer of epsilon-toxin. Structured digital abstract • Epsilon-toxin colocalizes with Epsilon-toxin by cosedimentationthroughdensitygradient (View interaction)

Abstract: We previously reported that mice deficient in UDP-GlcNAc:lysosomal enzyme GlcNAc-1-phosphotransferase (mucolipidosis type II or Gnptab −/− mice), the enzyme that initiates the addition of the mannose 6-phosphate lysosomal sorting signal on acid hydrolases, exhibited extensive vacuolization of their exocrine gland cells, while the liver, brain, and mus - cle appeared grossly unaffected. Similar pathological findings were observed in several exo - crine glands of patients with mucolipidosis II. To understand the basis for this cell type-specif - ic abnormality, we analyzed these tissues in Gnptab −/− mice using a combined immunoelectron microscopy and biochemical approach. We demonstrate that the vacuoles in the exocrine glands are enlarged autolysosomes containing undigested cytoplasmic material that accumu - late secondary to deficient lysosomal function. Surprisingly, the acid hydrolase levels in these tissues ranged from normal to modestly decreased, in contrast to skin fibroblasts, which ac - cumulate enlarged lysosomes and/or autolysosomes also but exhibit very low levels of acid hydrolases. We propose that the lysosomal defect in the exocrine cells is caused by the com - bination of increased secretion of the acid hydrolases via the constitutive pathway along with their entrapment in secretory granules. Taken together, our results provide new insights into the mechanisms of the tissue-specific abnormalities seen in mucolipidosis type II.

Abstract: Basal vacuolization of renal tubular epithelial cells (so-called Armanni-Ebstein phenomenon) has been attributed to hyperglycemia causing accumulation of cytoplasmic glycogen. Review of 34 autopsy cases with significant hyperglycemia (vitreous glucose ≥ 15 mmol/L/270 mg/dL) was undertaken to determine whether there was any significant association between the degree of hyperglycemia and the severity of this morphological change (graded as 0, 1+, 2+, and 3+). No association was demonstrated. Review of the subgroup of 14 cases with terminal hyperglycemia without ketoacidosis was then undertaken to assess the effect of hyperglycemia in isolation on renal tubular epithelial cells. Vitreous glucose levels in these 14 cases ranged from 17 to 49.7 mmol/L (306-894.6 mg/dL) with a mean of 26.25 mmol/L (472.5 mg/dL) and β-hydroxybutyrate levels ranged from 0.02 to 2.55 mmol/L (0.36-45.9 mg/dL) with a mean 0.79 mmol/L (14.22 mg/dL). Not one of the latter cases displayed basal vacuolization. No relationship between basal vacuolization of renal tubular epithelial cells at autopsy and terminal hyperglycemia could, therefore, be demonstrated.