Short-term stromal alterations in the rat ventral prostate following alloxan-induced diabetes and the influence of insulin replacement

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Differentiation of Leydig Cells in the Mongolian Gerbil

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

Localized matrix metalloproteinase (MMP)-2 and MMP-9 activity in the rat ventral prostate during the first week of postnatal development

The initial events in prostatic morphogenesis involve cell proliferation, epithelial canalization and outgrowth toward the stroma. We have hypothesized that stromal rearrangement takes place at the sites of epithelial growth and branching and that this rearrangement involves the action of gelatinases matrix metalloproteinase (MMP)-2 and MMP-9. Thus, the purpose of the present study was to characterize structural aspects of epithelial growth during the first week of postnatal development of the rat ventral prostate and to investigate the expression, localization and activity of MMP-2 and MMP-9 during this period by histological, ultrastructural and immunocytochemical analysis, in addition to gel zymography, in situ zymography and Western blotting. An increasing complexity of prostatic architeture was observed within the first postnatal week. Concurrently, the stroma became more organized and some cells differentiated into smooth muscle cells. Reticulin fibers appeared in a basket-like arrangement around both growing tips and epithelial sprouts, associated with a fainter staining for laminin. MMP-2 and MMP-9 activities were detected. MMP-2/MMP-9 expression decreased during the first week. Developing epithelial cords showed strong and difuse gelatinolytic activity. This activity coincided with the distribution of MMP-2 as determined by immunocytochemistry. on the other hand, MMP-9 was rather concentrated at the epithelial tips. These results suggest that gelatinolytic activity (with contribution of both MMP-2 and MMP-9) in the epithelium and at the epithelium-stroma interface are at least in part responsible for the tissue remodeling that allows epithelial growth and its projection into the surrounding stroma.

Neonatal Gonocyte Differentiation in Mongolian Gerbil Meriones unguiculatus Involves Asynchronous Maturation of Seminiferous Cords and Rapid Formation of Transitional Cell Stage

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Oxidative stress markers and apoptosis in the prostate of diabetic rats and the influence of vitamin C treatment

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Epithelial-stromal transition of MMP-7 immunolocalization in the rat ventral prostate following bilateral orchiectorny

Epithelial cells from involuting rat ventral prostate (VP) express Matrilysin (MMP-7) mRNA. Herein, we investigated by immunohistochemistry the NIMP-7 protein location and its association with tissue changes following castration in the VP. Normal and castrated adult male Wistar rats were sacrificed at different times after surgery. VP was examined by immunocytochemistry and immunoprecipitation. Castration promoted a shrinking of prostate ducts with an extensive stromal remodeling. In the VP from normal rats, MMP-7 immunoreactivity was found in epithelial secretory granules. Three days after castration, immunostaining for MMP-7 was found in both the epithelial secretory granules and in the stroma just below the epithelium, mainly at the distal ductal tips. At seven and 21 days after castration, the immunostaining for MMP-7 was found only in the stromal space. Immunoprecipitation confirmed the specificity of the primary antibody by rescuing a pro-enzyme form (28 kDa) in the prostate extracts. The present results suggest that MMP-7 participates in the epithelial-stromal interface remodeling of the ventral prostate during the involution achieved by castration, probably in the degradation of components of the epithelial basement membrane. (c) 2007 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

Neutrophil interaction with inflamed postcapillary venule endothelium alters annexin 1 expression

Annexin 1 (ANX-A1) exerts antimigratory actions in several models of acute and chronic inflammation, This is related to its ability to mimic the effect of endogenous ANX-A1 that is externalized on neutrophil adhesion to the postcapillary endothelium. In the present study we monitored ANX-A1 expression and localization in intravascular and emigrated neutrophils, using a classical model of rat peritonitis, For this purpose, a pair of antibodies raised against the ANX-A1 N-terminus tie, able to recognize intact ANX-A1) or the whole protein tie, able to interact with all ANX-A1 isoforms) was used by immunofluorescence and immunocytochemistry analyses. The majority (similar to 50%) of ANX-A1 on the plasma membrane of intravascular neutrophils was intact. Extravasation into the subendothelial matrix caused loss of this pool of intact protein (to similar to6%), concomitant with an increase in total amount of the protein; only similar to 25% of the total protein was now recognized by the antibody raised against the N-terminus tie, it was intact). In the cytoplasm of these cells, ANX-A1 was predominantly associated with large vacuoles, possibly endosomes, In situ hybridization confirmed de novo synthesis of ANX-A1 in the extravasated cells. In conclusion, biochemical pathways leading to the externalization, proteolysis, and synthesis of ANX-A1 are activated during the process of neutrophil extravasation.

Testosterone stimulates growth and secretory activity of the female prostate in the adult gerbil (Meriones unguiculatus)

The prostate of the female gerbil (Meriones unguiculatus) is similar to the human female prostate (Skene gland) and, despite its reduced size, it is functional and shows secretory activity. However, virtually nothing is known about its physiological regulation. This study was thus undertaken to evaluate the behavior of the gerbil female prostate in a hyperandrogenic condition. Adult females received subcutaneous injections of testosterone cypionate (1 mg/kg body weight every 48 h) up to 21 days. Circulating levels of testosterone and estradiol were monitored, and the prostate and ovaries subjected to structural and immunocytochemical analyses. The treatment resulted in sustained high levels of circulating testosterone, and caused a transient increase in estradiol. There was an increase in epithelial cell proliferation accompanied by significant reorganization of the epithelium and an apparent reduction in secretory activity, followed by a progressive increase in luminal volume density and accumulation of secretory products. Immunocytochemistry identified the expression of androgen receptor and a prostate-specific antigen (PSA)-related antigen in prostatic epithelial cells. A circulating PSA-related antigen was also found, and its concentration showed strong negative correlation with circulating estrogen. Epithelial dysplasia was detected in the prostate of treated females. Analysis of the ovaries showed the occurrence of a polycystic condition and stromal cell hyperplasia. The results indicate that testosterone has a stimulatory effect on the female prostate, inducing epithelial cell proliferation, differentiation, secretory activity, and dysplasia. The results also suggest that prostatic growth and activity, polycystic ovaries, and ovarian stromal cell hyperplasia are related to a hyperandrogenic condition in females.

Estimation of growth fraction in fine needle aspirates from non-Hodgkin's lymphoma using anti-proliferating cell nuclear antigen - Correlation with the Kiel classification

OBJECTIVE: This study was undertaken to assess whether fine needle aspirates from non-Hodgkins lymphoma (NHL) could be used for growth fraction analysis with proliferating cell nuclear antigen (PCNA) staining and if there was a relationship between the growth fraction and cytomorphologic classification according to the Kiel classification.STUDY DESIGN: the study group consisted of 40 patients with NHL diagnosed by fine needle aspiration (FNA) cytology. The cytologic classification of the lymphomas was made by two cytopathologists on May-Grunwald-Giemsa-stained slides using the Kiel classification. There were 27 cases of low and 13 of high grade lymphoma. The estimation of the growth fraction was made by PCNA immunoreactivity. The PCNA index was quantitated in smears by counting an average of 1,000 cells, and the count teas correlated with the cytomorphologic classification.RESULTS: There was It strong correlation between the PCNA index and lymphoma grading. High grade lymphomas exhibited a mean PCNA positivity of 74.0%, which was significantly higher (P <.001) than that of low grade lymphomas (17.6%).CONCLUSION: Our study showed that PCNA evalua tion is suitable for smears obtained by FNA on NHL, correlates with increasing grades of lymphoma according to the Kiel classification and may offer a method of monitoring treatment of lymphoma.

Identification, content, and distribution of type VI collagen in bovine tendons

Tendon composition changes according to differentiation, mechanical load, and aging. In this study, we attempted to identify, localize, and quantify type VI collagen in bovine tendons. Type VI collagen was identified by the electrophoretic behavior of the alpha chains and Western blotting, and by rotary shadowing. Type VI collagen was extracted from powdered tendon with three sequential 24-h extractions with 4 M guanidine-HCl. The amount of type VI collagen was determined by enzyme-linked immunosorbent assay for purely tensional areas and for the compressive fibrocartilage regions of the deep flexor tendon of the digits, for the corresponding fetal and calf tendons, and for the extensor digital tendon. The distal fibrocartilaginous region of the adult tendon was richer in type VI collagen than the tensional area, reaching as much as 3.3 mg/g (0.33%) of the wet weight. Calf tendons showed an accumulation of type VI at the fibrocartilage site. Immunocytochemistry demonstrated that type VI collagen was evenly distributed in the tensional areas of tendons but was highly concentrated around the fibrochondrocytes in the fibrocartilages. The results demonstrate that tendons are variable with regard to the presence and distribution of type VI collagen. The early accumulation of type VI collagen in the region of calf tendon that will become fibrocartilage in the adult suggests that it is a good marker of fibrocartilage differentiation. Furthermore, the distribution of type VI collagen in tendon fibrocartilage indicates that it organizes the pericellular environment and may represent a survival factor for these cells.

CYTOMORPHOLOGICAL AND IMMUNOCYTOCHEMICAL EVALUATION OF FINE NEEDLE ASPIRATES FROM HIGH-GRADE NON-HODGKINS-LYMPHOMAS

1. Fine needle aspirates from ten patients with high-grade malignant non-Hodgkin's lymphomas were analyzed by cytomorphology and immunocytochemistry.2. The following morphologic diagnoses were made: lymphoblastic lymphoma (3 cases), Burkitt's lymphoma (3 cases), mixed small and large cell lymphomas with predominance of large cells (2 cases), and centroblastic lymphoma (2 cases). Immunocytochemistry showed a B-cell phenotype in five cases and a T-cell phenotype in four. One case of lymphoblastic lymphoma was negative for the T and B cell markers used.3. The results of histological and immunohistochemical analyses performed on surgical biopsies from 8 patients confirmed the morphological diagnosis in all cases. Two cases of Burkitt's lymphoma were submitted only to cytological and immunological diagnosis.4. The high diagnostic accuracy of combined cytomorphology and immunocytochemical assessment of fine needle aspirate samples validates the use of the technique in the diagnostic work-up of high-grade non Hodgkin's lymphomas.

IMMUNOCYTOCHEMICAL DIFFERENTIATION OF REACTIVE HYPERPLASIA FROM FOLLICULAR LYMPHOMA USING MONOCLONAL-ANTIBODIES TO CELL-SURFACE AND PROLIFERATION-RELATED MARKERS

We tested the hypothesis that a panel of antibodies to cell surface, cytoplasmic, and nuclear antigens could reliably distinguish the cells composing reactive germinal centers from those composing follicular lymphoma. Immunocytochemistry was performed on deparaffinized sections of methacarn-fixed lymph node and tonsil (15 cases of reactive hyperplasia and 14 cases of follicular lymphoma) using antibodies to the nerve growth factor receptor (NGFR5), bcl-2 protein (124), proliferating cell nuclear antigen (PCNA; 19A2), and CD45RA (MT2). In 100% of cases of reactive hyperplasia, both MT2 and 124 showed positive immunostaining of mantle zone and scattered interfollicular lymphocytes, but in all cases there was a sharply demarcated absence of immunostaining of germinal center cells. However, diffuse immunostaining of follicular centers with MT2 (64%) and 124 (93%) and scattered intervening cells were seen in follicular lymphoma. The combination of antibodies to CD45RA and bcl-2 yielded positive immunostaining of follicular center cells in 93% of follicular lymphomas. The germinal center cells of reactive hyperplasia showed >75% nuclear positivity with antibodies to PCNA, in contrast to the follicular lymphoma cells, which showed variable PCNA indices ranging from 25 to >75%. A minority of follicular lymphoma cases (29%) showed PCNA indices comparable with those seen in cases of reactive hyperplasia. Antibodies to NGFR were positive in all cases of reactive hyperplasia and in 79% of cases of follicular hyperplasia, although the immunostaining intensity was generally decreased in follicular hyperplasia. In summary, antibodies to bcl-2 appear to be superior to those to CD45RA in distinguishing reactive hyperplasia from follicular lymphoma. Reactive hyperplasia cannot be discriminated from follicular hyperplasia using antibodies to PCNA or to nerve growth factor receptor.

Collagen type VI is a component of the extracellular matrix microfibril network of the prostatic stroma

Stroma-epithelium relationships are of great relevance in prostatic morphogenesis and physiology, However, little knowledge exists about either stromal cells or extracellular matrix composition and arrangement in this system, Ultrastructural analysis revealed the existence of a microfibrillar system which occupies large areas of the rat prostatic stroma, In this work, we have applied immunocytochemistry and an ATP treatment for the ultrastructural identification of collagen type VI microfibrils, aiming at examining its participation in the prostatic microfibrillar network. Immunocytochemistry was also extended to a human case of prostatic nodular hyperplasia, Both methods succeeded in identifying collagen type VI in the rat ventral prostate, Collagen type VI is evenly distributed throughout the stroma but mainly associated with the basal lamina, collagen fibrils, and around the stromal cells, the use of ATP treatment allowed for the discrimination between collagen type VI and elastin-associated microfibrils, and demonstrated that these two classes of microfibrils establish an extended, mixed, and open network. The same aspects of association with the basal lamina, with stromal cells (particularly with smooth muscle cells), and with fibrillar components of the stroma were observed in the human tissue, We suggest that the collagen type VI and elastin-associated microfibril system may be involved in the control of some aspects of cellular behavior and may also play a structural role, maintaining the organ integrity after the deformations occurring under smooth muscle contraction.

Effects of histone hyperacetylation on the preimplantation development of male and female bovine embryos

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

A2 Noradrenergic Lesions Prevent Renal Sympathoinhibition Induced by Hypernatremia in Rats

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)