José Carlos Nogueira

Antiglaucomatous effects of the activation of intrinsic Angiotensin-converting enzyme 2.

PURPOSE To evaluate the effects of the activation of endogenous angiotensin-converting enzyme 2 (ACE2) using the compound diminazene aceturate (DIZE) in an experimental model of glaucoma in Wistar rats. METHODS DIZE (1 mg/kg) was administered daily, either systemically or topically, and the IOP was measured weekly. To examine the role of the Mas receptor in the effects of DIZE, the Ang-(1-7) antagonist A-779 was co-administered. Drainage of the aqueous humor was evaluated by using scintigraphy. The analysis of ACE2 expression by immunohistochemistry and the counting of retinal ganglion cells (RGCs) were performed in histologic sections. Additionally, the nerve fiber structure was evaluated by transmission electron microscopy. RESULTS The systemic administration and topical administration (in the form of eye drops) of DIZE increased the ACE2 expression in the eyes and significantly decreased the IOP of glaucomatous rats without changing the blood pressure. Importantly, this IOP-lowering action of DIZE was similar to the effects of dorzolamide. The antiglaucomatous effects of DIZE were blocked by A-779. Histologic analysis revealed that the reduction in the number of RGCs and the increase in the expression of caspase-3 in the RGC layer in glaucomatous animals were prevented by DIZE. This compound also prevented alterations in the cytoplasm of axons in glaucomatous rats. In addition to these neuroprotective effects, DIZE facilitated the drainage of the aqueous humor. CONCLUSIONS Our results evidence the pathophysiologic relevance of the ocular ACE2/Ang-(1-7)/Mas axis of the renin-angiotensin system and, importantly, indicate that the activation of intrinsic ACE2 is a potential therapeutic strategy to treat glaucoma.

Distribution of estrogen receptors (ERα and ERβ) and androgen receptor in the testis of big fruit-eating bat Artibeus lituratus is cell-and stage-specific and increases during gonadal regression

The testis is a classical target for androgens, especially testosterone, acting via androgen receptor (AR). Alternatively, androgens can be aromatized to produce estrogens which act via specific receptors ERalpha and ERbeta. Although estrogen action is essential for maintenance of male fertility, studies regarding the expression of ERalpha and ERbeta in testis are restricted to a few species of rodent and domestic animals, but rarely in wild species. To our knowledge, there are no studies in Chiroptera species. Chiroptera represent one of the largest and most diversified orders of mammals, which possess several interesting reproductive features, including higher affinity of SHBG for estrogens than androgens. Therefore, we thought that bats would constitute a good model for investigation of the role of estrogens in the male. In this study, the distribution of ERalpha, ERbeta and AR were evaluated in the testis of the big fruit-eating bat Artibeus lituratus and their levels were compared during reproductive and regressive periods. The results showed that ERalpha and AR were restricted to the somatic cells of the testis, whereas ERbeta was widely distributed in both somatic and spermatogenic cells in a cellular and stage-specific fashion. We demonstrated for the first time by immunohistochemistry, and confirmed by Western blotting, that ERbeta and AR increased during regression. The localization of ERalpha, ERbeta and AR in a seasonal, cell and stage-specific fashion in the testis of A. lituratus suggests that these receptors may play important roles in testis function during reproductive and non-reproductive periods.

Sequential order of appearance of ossification centers in the opossum Didelphis albiventris (Didelphidae) skeleton during development in the Marsupium

Pouch-young of the opossum Didelphis albiventris were studied. They were removed from the marsupium at a crown-rump length (CRL) of 9.0 to 100.0 mm and submitted to differential staining of the bone and cartilage. Newborn D. albiventris of 9.0 to 10.0 mm CRL have a cartilaginous skeleton, with no macroscopic evidence of ossification. In 10.5 to 13.0 mm CRL pouch-young, ossification occurs in the bones surrounding the oral cavity, clavicle, ribs, cervical and thoracic vertebrae and thoracic limbs, structures necessary to ensure the survival and maintenance of the newborn in the marsupium. Ossification of most of the skull begins in 15.5 to 16.0 mm CRL pouch-young. The ossification in the pelvis and the pelvic limbs is observed at 18.0 mm CRL and in the sternum and the epipubic bone at 25.0 to 28.0 mm CRL. Most of the skeleton is ossified in 45.0 to 58.0 mm CRL pouch-young, except in the carpus, tarsus and epiphyses of the long bones where ossification occurs in 60.0 to 100.0 mm CRL pouch-young. In the skeleton of D. albiventris the omosternum, haemal arches, prepollex, prehallux and parafibula, are present. These are inconstant or absent elements in eutherian mammals.

Differential expression and seasonal variation on aquaporins 1 and 9 in the male genital system of big fruit-eating bat Artibeus lituratus.

Efferent ductules and epididymis are involved in water and solute transport, which is indispensable for storage and maintenance of the sperm viability. The reabsorption process involves proteins such as aquaporins (AQP), which has been described in the male genital system of limited species, including primate, rodents, cats and dogs. To contribute with information about AQPs in the male system, here we investigated the distribution of AQP1 and AQP9 in the tropical bat Artibeus lituratus, along the annual reproductive cycle. A. lituratus is a seasonal breeder with natural variation in components of the androgen and estrogen responsive system, thus being a good model for exploring the AQPs modulation. AQP1 was found restricted to differentiating spermatids, efferent ductules epithelium and venular endothelia along the male tract. AQP9 was detected throughout the epididymis being more abundant in the cauda and ductus deferens, but was not found in testis, rete testis and efferent ductules. Contrasting with AQP1 which appear to be constitutively expressed, there was seasonal variation in AQP9 expression, which was reduced in regressed epididymis. The AQP9 does not appear to be modulated by estradiol or androgens, but possibly by other factor related to luminal sperm. The establishment of specific function for aquaporins in the male tract remains undetermined; however, the cellular distribution presently found are compatible with the main function of AQP1, as a selective water channel, and AQP9, which is a conduct for water and a plethora of neutral solutes present in the epididymis milieu such as glycerol and urea.

Morphology of the male genital system of Chironectes minimus and comparison to other didelphid marsupials

Abstract The male genital system was examined in 9 male specimens of Chironectes minimus (2 adult and 7 young) from Brazil. Unlike other didelphids, C. minimus lacks a cloaca and has distinct anal and urogenital openings, and the tunica vaginalis lacks black pigmentation. Similar to other American didelphids, C. minimus has a segmented carrot-shaped prostate gland with 3 anatomical segments that can be distinguished by color, length, and diameter in adults. The 2 pairs of bulbourethral glands of C. minimus are similar to those found in species of Caluromys, Gracilinanus, Monodelphis, Thylamys, and Glironia. The tip of each hemi-glans is rounded and bulbous shaped, whereas the urethral grooves end at the ventral aspect of the tip of each half glans.

Ocular Inserts for Sustained Release of the Angiotensin-Converting Enzyme 2 Activator, Diminazene Aceturate, to Treat Glaucoma in Rats

The aim of this study was to develop and evaluate the effects of chitosan inserts for sustained release of the angiotensin-converting enzyme 2 (ACE2) activator, diminazene aceturate (DIZE), in experimental glaucoma. Monolayer DIZE loaded inserts (D+I) were prepared and characterized through swelling, attenuated total reflectance Fourier transformed infrared spectroscopy (ATR-FTIR), differential scanning calorimetry (DSC) and in vitro drug release. Functionally, the effects of D+I were tested in glaucomatous rats. Glaucoma was induced by weekly injections of hyaluronic acid (HA) into the anterior chamber and intraocular pressure (IOP) measurements were performed. Retinal ganglion cells (RGC) and optic nerve head cupping were evaluated in histological sections. Biodistribution of the drug was accessed by scintigraphic images and ex vivo radiation counting. We found that DIZE increased the swelling index of the inserts. Also, it was molecularly dispersed and interspersed in the polymeric matrix as a freebase. DIZE did not lose its chemical integrity and activity when loaded in the inserts. The functional evaluation demonstrated that D+I decreased the IOP and maintained the IOP lowered for up to one month (last week: 11.0±0.7 mmHg). This effect of D+I prevented the loss of RGC and degeneration of the optic nerve. No toxic effects in the eyes related to application of the inserts were observed. Moreover, biodistribution studies showed that D+I prolonged the retention of DIZE in the corneal site. We concluded that D+I provided sustained DIZE delivery in vivo, thereby evidencing the potential application of polymeric-based DIZE inserts for glaucoma management.

Seasonal variation in estrogen receptor ERα, but not ERβ, androgen receptor and aromatase, in the efferent ductules and epididymis of the big fruit-eating bat Artibeus lituratus

The efferent ductules (ED) are a major target for estrogens, which act via the estrogen receptors ERα (ESR1) and ERβ (ESR2). ERα has been found in the ED of all species studied so far. However, in the epididymis (EP), the expression of ERα is controversial, as is data about the occurrence of aromatase in the epithelium lining the excurrent ducts. Therefore, to further investigate this estrogen-responsive system, we used a seasonal breeder, the Neotropical bat, Artibeus lituratus, in which testicular expression of androgen (AR) and estrogen (ER) receptors vary with reproductive phase. The localization of aromatase, ERα, ERβ and AR in the ED and EP of A. lituratus was investigated. The results showed that aromatase, AR and ERβ were distributed throughout the excurrent ducts and did not vary during the annual reproductive cycle. Conversely, ERα was detected primarily in the ED epithelium, had marked seasonal variation and was increased during regression, especially in the EP epithelium. The results suggest that ERα may be involved in preparing the male genital tract for recrudescence. Together, the data obtained under natural conditions emphasize that specific segments of the excurrent ducts downstream of the testis are the primary targets for estrogen action via ERα, which is similar to previous findings in animals lacking functional ERα.

Ultrastructural pattern of glucagon producing-cells in the gastric mucosa of the developing opossum Didelphis albiventris (Marsupialia)

Gastric glucagon-producing cells of the pouch young, weaned and adult opossum Didelphis albiventris were investigated by immunocytochemical and ultrastructural methods. In sections of the oxyntic mucosa stained by immunoperoxidase it was observed that glucagon-immunoreactive cells increased in number during pouch development (from 7.9 +/- 4.7 to 35.4 +/- 5.9 cells/mm2), decreasing progressively to weaned (14.2 +/- 4.8 cells/mm2) and adult animals (10.4 +/- 3.0 cells/mm2). The glucagon-immunoreactive cells presented numerous electrondense, round cytoplasmic granules surrounded by a membrane. Frequently, the dense core of the granules was separated from the enveloping membrane by a delicate electronlucent halo or by a less electrondense peripheral halo. This pattern of the granular component was similar to the ultrastructural morphology observed in pancreatic islet glucagon-producing cells and A-like cells of the gastric mucosa of those eutherian mammals studied so far.

Atividade secretória de tubas uterinas com e sem patologia de porcas adultas nas fases folicular e luteínica

The goal of this study was to describe the anatomopathological and histochemical features of uterine tubes from 62 sexually mature sows, in both follicular and luteal phases. The animals were assigned into four groups as follows: the first (LSP) containing sows in the luteal phase without tube pathology (n=45), the second (LCP) with sows in the luteal phase with tube pathology (n=19), the third (FSP) with sows in the follicular phase without tube pathology (n=45) and the fourth (FCP) with sows in the follicular phase with tube pathology (n=15). PAS, Alcian Blue (pH 2.5 and 0.4), and Ninhidrina-Schiff stains were used for the histochemical study. Grossly, 34 uterine tubes presented morphological lesions. Wall cysts and squamous metaplasia were the changes found more frequently in both phases. Ampulla and infundibulum were the segments affected more frequently. In the follicular phase, the infundibulum had a higher secretion intensity of neutral mucosubstances, whereas the ampulla and the infundibulum had a higher secretion intensity of AB 2.5-positive mucosubstances. The ampulla had a higher intensity of sulphomucins and Ninhidrina-Schiff-positive secretions in the follicular phase. The morphologic changes did not correlate with reduction in secretion of mucosubstance in the adjacent areas of the uterine tube in both follicular and luteal phases. There was a reduction in secretion of neutral and acid mucosubstances in areas affected by squamous metaplasia and wall cysts.

Morphological and pathological aspects of the rete ovarii in sheep (Ovis aries)

Sixty-four ovaries of adult woolly sheep from the Centro de Pesquisa dos Campos Sul Brasileiros - Embrapa, Bage-RS, Brazil, were analyzed histologically. The presence of the rete ovarii was detected in 55 of the 64 ovaries (85.9%), located in the periovarian tissue, in the hilus and in the medullary or cortical regions. As also observed in other mammals, the rete ovarii of sheep is morphologically divided into three portions: intraovarian rete, connecting rete and extraovarian rete. Continuity between the extraovarian rete and the uterine tube was observed in five ovaries (7.8%), suggesting a tuboretial connection, as reported for cows and does. Pathological examination of the rete showed the formation of cysts surrounding the connecting and/or extraovarian rete in seven cases (10.9%) and one case of hyperplasia of the extraovarian rete (1.6%).