Mg2+-dependent ATPase activity in Malpighian tubules of Triatoma infestans Klug

Mg2+-dependent ATPases were investigated in Malpighian tubules of the blood-sucking insect, Triatoma infestans, with cytochemical procedures for light and electron microscopy. The aim was to establish patterns of enzyme occurrence in the blood-sucking insect under control rearing conditions for further comparisons with animals subjected to the action of stress factors. Enzyme activity was found in laminated "concretions" present in distal cells, in edges of urate crystals at the lumen of the proximal region of tubules, in the basement membrane of proximal cells, and variously distributed in plasmalemma invaginations of both distal and proximal cells. Presence of ATPases in the "concretions" and urate crystals is presumed to be due to engulfment of other ATPase-containing components during formation of these structures. Cytochemical reactivity in the basement membrane and plasmalemma invaginations is assumed to be involved with active transport of waste molecules from and to hemolymph and differs as a function of the Malpighian tubule region. This paper provides a basic understanding of the enzyme occurrence in the blood sucking insects, and can be used as a pattern for comparative means of the staining patterns among Triatominae species. (C) 2011 Elsevier Ltd. All rights reserved.

Immunocytochemical localization of heparin in secretory granules of rat peritoneal mast cells using a monoclonal anti-heparin antibody (ST-1)

We performed immunogold labeling with an ST-1 monoclonal antibody (IgM), specific for intact heparin, to define the subcellular localization of heparin in mast cells. Rat peritoneal mast cells were fixed by a modified Karnovsky method and embedded in Araldite. Ultrathin sections were first treated with sodium periodate and then sequentially incubated with MAb ST-1, rabbit anti-mouse IgM, and protein A-gold. By transmission electron microscopy, gold particles were localized inside cytoplasmic granules of peritoneal mast cells. In contrast, with the same procedure, no labeling was observed in mast cells from rat intestinal mucosa. Control sections of rat peritoneal or intestinal mucosa mast Mast cells cells treated with an irrelevant MAb (IgM) did not show any labeling. Treatment with nitrous Heparin acid abolished the reactivity of MAb ST-1 with peritoneal mast cells. These results Granules show that different mast cells can be identified regarding their heparin content by immunochemical procedures using MAb ST-1.

Leukocyte and thrombocyte reference values for channel catfish (Ictalurus punctatus Raf), with an assessment of morphologic, cytochemical, and ultrastructural features

Background: Hematology tests are useful to evaluate physiologic disturbances in fish and can provide important information for the diagnosis and prognosis of disease. Objectives: the primary purpose of this study was to define reference intervals for thrombocytes and leukocytes in healthy channel catfish (Ictalurus punctactus). In addition, the morphologic, cytochernical, and ultrastructural features of blood cells were assessed. Methods: Blood samples (0.5 mL were collected into EDTA from 40 clinically healthy catfish on a commercial fish farm in Jaboticabal, Brazil. Thrombocyte, total WBC, and differential WBC counts were determined and reference intervals were calculated as the 25-95th percentiles of data. Thrombocyte and leukocyte morphology was assessed in blood smears stained with May Griinwald-Giemsa-Wright and ultrastructurally by transmission electron microscopy. Cytochemical staining patterns were described using periodic acid-Schiff (PAS), peroxidase, nonspecific esterase, alkaline phosphatase, and toluidine blue. Results: Reference intervals were as follows: thrombocytes 58,802-99,569/mu L; total WBCs 27,460-41,523/mu L; lymphocytes 5380-11,581/mu L; monocytes 2949-7459/mu L; neutrophils 12,529-22,748/mu L, and basophils 736-2003/mu L. Neutrophils were positive for peroxidase and PAS; monocytes were positive for nonspecific esterase; and basophils were positive with toluidine blue. Conclusion: the morphologic and staining features of neutrophils and monocytes of channel catfish are similar to those of mammals, and the presence of basophils in this species was verified. These reference intervals and morphologic findings provide a foundation for future investigations on the functions and alterations of blood cells in channel catfish.

Cytochemical characterization of the endomembranous system during oocyte secondary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Characidae)

The endomembranous system of Serrasalmus spilopleura oocyte secondary growth was analysed using structural and ultrastructural cytochemical techniques. In vitellogenic oocytes, the endoplasmic reticulum components, the nuclear envelope intermembranous space, some Golgi dictiossomes, lysosomes, yolk granules, regions of the egg envelope and sites of the follicle cells react to acid phosphatase detection (AcPase). The cortical alveoli, some heterogeneous cytoplasmic structures, regions of the egg envelope, and sites of the follicle cells are strongly contrasted by osmium tetroxide and zinc iodide impregnation (ZIO). The endoplasmic reticulum components, some vesicles, and sites of the follicle cells also react to osmium tetroxide and potassium iodide impregnation (KI). The biosynthetic pathway of lysosomal proteins, such as acid phosphatase, required for vitellogenesis, involves the endoplasmic reticulum, Golgi complex, vesicles with inactive hydrolytic enzymes, and, finally, lysosomes. In S. spilopleura oocytes at secondary growth, the endomembranous system takes part in the production of the enzymes needed for vitellogenesis, and in the metabolism of yolk exogenous components (AcPase detection). The endomembranous system compartments also show reduction capacity (KI reaction) and are involved in the metabolism of proteins rich in SH-groups (ZIO reaction).

Storaged products and presence of acid phosphatase in fat body cells at pre-pupal worker stage of Apis mellifera Linnaeus, 1758 (Hymenoptera, Apidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Galectin-3 Overexpression in Invasive Laryngeal Carcinoma, Assessed by Computer-assisted Analysis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Glycogen distribution in ameloblasts and odontoblasts of newborn rats of sialoadenectomized dams. Histochemical study

In order to get information about the distribution of glycogen in ameloblasts and odontoblasts, studies were made of newborn rats of sialoadenectomized dams and newborn rats of control dams. Rodent offspring were decapitated on the 1st, 3rd, 5th, 7th, and 9th days after birth. Their heads were fixed in 10% neutral formalin solution, decalcified in sodium citrate-formic acid and embedded in paraffin, and frontal sections were prepared. Sections 6 micrometers thick were stained by specific histochemical reactions to detect glycogen. Based on the results obtained, it was concluded that the amount of glycogen was lower in the cytoplasms of ameloblasts and odontoblasts of experimental animals when compared to controls.

Characterization of myeloid or lymphoid acute leukemia by a chemiluminescence assay: Comparison with immunocytochemistry using an antimyeloperoxidase antibody

A simple and sensitive chemiluminescence assay for the demonstration of the activity of intracellular myeloperoxidase (MPO) is described, which is useful for the distinction between myeloid and lymphoid commitment in blasts from acute leukemia patients. When the cut-off point was settled at 13 mV of chemiluminescence all cases of acute myeloid leukemia (AML) were distinguished from those of acute lymphoid leukemia. In addition, this technique was able to demonstrate MPO activity in AML poorly differentiated (FAB-M0) which usually does not stain for MPO in classical cytochemistry preparations and could be negative also by immunocytochemistry with anti-MPO monoclonal antibody. Therefore the method here described presented a higher sensitivity than the immunocytochemistry procedure with anti-MPO.

Morphology and histochemistry of the hyolingual apparatus in chameleons

We reexamined the morphological and functional properties of the hyoid, the tongue pad, and hyolingual musculature in chameleons. Dissections and histological sections indicated the presence of five distinctly individualized pairs of intrinsic tongue muscles. An analysis of the histochemical properties of the system revealed only two fiber types in the hyolingual muscles: fast glycolytic and fast oxidative glycolytic fibers. In accordance with this observation, motor-endplate staining showed that all endplates are of the en-plaque type. All muscles show relatively short fibers and large numbers of motor endplates, indicating a large potential for fine muscular control. The connective tissue sheet surrounding the entoglossal process contains elastin fibers at its periphery, allowing for elastic recoil of the hyolingual system after prey capture. The connective tissue sheets surrounding the m. accelerator and m. hyoglossus were examined under polarized light. The collagen fibers in the accelerator epimysium are configured in a crossed helical array that will facilitate limited muscle elongation. The microstructure of the tongue pad as revealed by SEM showed decreased adhesive properties, indicating a change in the prey prehension mechanics in chameleons compared to agamid or iguanid lizards. These findings provide the basis for further experimental analysis of the hyolingual system. © 2001 Wiley-Liss, Inc.

Distribution of acid phosphatases in the hypopharyngeal glands from workers, queens and males of a Brazilian stingless bee Scaptotrigona postica Latreille: An ultrastructural cytochemical study

The present study reports the localization of acid phosphatase in the hypopharyngeal gland cells from workers (newly-emerged, nurse and forager), queens (newly-emerged and laying) and males (newly-emerged and mature for mating) of the Brazilian stingless bee, Scaptotrigona postica. The phosphatase activity varied in intensity and localization depending on the individual class, physiological age and the substrate used. In newly-emerged workers, the phosphatase-positive sites suggest the involvement of the enzyme with cellular differentiation that occurs in the presecretory phase, in nurse workers with protein synthesis and in forager workers with changes in cellular activity or glandular regression. In males mature for mating and laying queens, the positive sites are related to secretory activity, showing that the gland maintains some activity in spite of the regressive aspect. Of the substrates used, β-glycerophosphate gave the least specific localization.

Morphology and histoenzymology of eosinophilic granulocytes in the circulating blood of the turtle (Chrysemys dorbignih)

The studies on the characterization of eosinophils and neutrophils/ heterophils of turtles are contradictory. Some authors have pointed out the existence of two distinct cell types: eosinophils and heterophils. Other authors have proposed that eosinophils and heterophils may be the same cells in different stages of maturation. These interpretations are based only on a morphological analysis. In the blood of the turtle (Chrysemys dorbignih), a South American freshwater species, there are two types of granulocytes with eosinophilic staining pattern: the first with round cytoplasmic granules and the second with ellipsoidal cytoplasmic granules. In the present study by using histoenzymological methods for the analyses of enzymological cellular content, we found that the cells with round cytoplasmic granules were positive for nonspecific esterase and the cells with ellipsoidal granules were positives for acid phosphatase, alkaline phosphatase, nonspecific esterase and peroxidase. The results show that these cells are distinct cells and that the cells with ellipsoidal cytoplasmic granules have the same histoenzymological characteristics as the neutrophils/heterophils of mammalians and other vertebrates.

Localization of peroxidase activity in blood mononuclear phagocytes in pacu fish (Piaractus mesopotamicus)

The localization of peroxidase activity in different cell regions is used as a criterion for classifying the stage of maturity of mammalian mononuclear phagocytes, with a positive peroxidase reaction indicating the presence of monoblasts, promonocytes, monocytes, and macrophages. Peroxidase activity was observed ultrastructurally in the circulating blood of pacu fish (Piaractus mesopotamicus), identifying monoblasts, promonocytes, monocytes, and macrophages. These observations suggest that differentiation of mononuclear phagocytes occurs in the blood circulation of fish, whereas in mammals, monoblasts and promonocytes are detected in bone marrow, with only monocytes detected in circulating blood and differentiation into macrophages occurring in other body compartments.

p16INK4A immunohistochemical overexpression in premalignant and malignant oral lesions infected with human papillomavirus

Human papillomavirus (HPV) is believed to promote the oncogenic process, and the correlation between viral oncoproteins and dysfunction of p16 INK4A tumor suppressor protein in oral lesions is controversial. To test the hypothesis that anogenital HPV types participate in disruption of the regulation of p16INK4A suppressor protein in oral lesions, we analyzed 46 oral biopsy specimens for the presence of HPV 6/11 and 16/18 by in situ hybridization (ISH) and for p16INK4A expression by immunohistochemistry (IHC). Eighteen (39%) of the 46 oral lesions were HPV-positive and 28 (61%) were HPV-negative. HPV 6/11 DNA was found in 5 (11%) and HPV 16/18 in 13 (28%) of 46 biopsies. Nine of the 18 HPV-positive oral lesions (50%), assessed by catalyzed signal amplification coupled to ISH (CSA-ISH), gave high-intensity p16INK4A immunostaining. Focal and diffuse patterns were observed in 11/13 (77%) lesions with HPV 16/18, focal immunopositivity in 3/5 (80%) with HPV 6/11, and negative or sporadic p16-labeling in 18/28 (64%) without the presence of HPV DNA. These results showed a strong association between overexpression of p16 protein and malignant oral lesions, mainly those infected by HPV 16/18. We can conclude that high-risk HPV types are associated with p16 overexpression, and p16 may serve as a biomarker in oral cancer related to high-risk HPV infection.

Structural, cytochemical, immunocytochemical and ultrastructural characterization of blood granulocytes of the roadside hawk Buteo magnirostris (Gmelin, 1788) (Avian, Falconiform)

In the peripheral blood of the roadside hawk, Buteo magnirostris, the following types of granulocytic leucocytes were identified: heterophil, eosinophil and basophil. The heterophils presented acidophilic and spindle shaped granules, the eosinophils possess spherical eosinophilic granules and the basophils showed spherical and basophilic granules. The heterophils and eosinophils presented positive cytochemical reaction to glycogen and basic polyaminoacid, while the eosinophils presented sudanophilic granules, which were also positive for myeloperoxidase. The heterophils, alone, presented positivity for acid phosphatase in some granules and immunoreactivity to TGF-β1 was observed only in the cytoplasm of the eosinophils. Electron microscopy demonstrated the heterophil granules as predominantly spindle shaped, being strongly electron-dense, while the eosinophils had numerous uniformly electron-dense spherical granules and the basophils presented three different types of granules identified according to their electron-density and the aspect of their matrix.

Morphological, cytochemical, and ultrastructural study of thrombocytes and leukocytes in neotropical fish, Brycon orbignyanus Valenciennes, 1850 (Characidae, Bryconinae)

Morphological, cytochemical and ultrastructural studies are important to demonstrate the function of the blood cells, which is very little understood in teleosts. In peripheral blood of piracanjuba' Brycon orbignyanus, thrombocytes, lymphocytes, monocytes, neutrophils and heterophils were studied and characterized. Thrombocytes had a fusiform or oval shape with PAS-positive granules. Lymphocytes presented small size with sparse basophilic cytoplasm. Monocytes were large in size, presented basophilic cytoplasm that may be foamy or vacuolated, with non-specific esterase staining. The neutrophils presented lightly neutrophilic granule cytoplasm, with positivity for PAS and peroxidase. The heterophils were large in size, with eosinophilic and basophilic granules cytoplasm and PAS-positive. Transmission electron microscopy study demonstrated that the thrombocytes, lymphocytes and monocytes features were similar to other teleosts. In ultrastructural study only one type of neutrophils was observed. Cytochemical findings indicated that neutrophils and monocytes of B. orbignyanus may be involved in phagocytosis, and neutrophils play an important microbicidal role.