Hyaluronan content and distribution in the rat ventral prostate after castration.

Hyaluronan (HA) has been implicated in tissue remodeling, healing, and tumor growth. This study investigated the variation in hyaluronan content, distribution, and metabolism in the rat ventral prostate (VP) in response to androgen deprivation after castration. The mRNA abundance of hyaluronan synthases (Has1-3) and hyaluronidases (Hyal 1-3) were assessed by reverse transcription (RT)-PCR and immunohistochemistry, respectively. The results demonstrated an increased concentration, but an overall reduction in HA content. HA was located in both epithelium and stroma of the prostate of both the noncastrated and castrated animals. Quantitative RT-PCR (qRT-PCR) showed that Has1 and Has2 are major synthases, and that Hyal 1 was the predominant hydrolase expressed in the VP. qRT-PCR also showed that Has1 and Has2 mRNA increased transiently after castration, whereas Has3 mRNA declined markedly. While Hyal 1 mRNA increased slowly up to day 21 after castration, Hyal 2 and Hyal 3 mRNA dropped significantly. CD44 was found in the epithelial cells and in some stromal cells in both hormonal conditions. In conclusion, castration results in increased abundance of Has1 and Has2 mRNA, but is associated with a decrease in the total content of HA, with an increased concentration, and a predominance of short-chain HA molecules.

Angiotensin-(1-7) receptor Mas is an essential modulator of extracellular matrix protein expression in the heart.

In this study we investigated the effects of genetic deletion of the Angiotensin-(1-7) receptor Mas or the Angiotensin II receptor AT(2) on the expression of specific extracellular matrix (ECM) proteins in atria, right ventricles and atrioventricular (AV) valves of neonatal and adult mice. Quantification of collagen types I, III and VI and fibronectin was performed using immunofluorescence-labeling and confocal microscopy. Picrosirius red staining was used for the histological assessment of the overall collagen distribution pattern. ECM proteins, metalloproteinases (MMP), ERK1/2 and p38 levels were quantified by western blot analysis. Gelatin zymography was used to evaluate the activity of MMP-2 and MMP-9. We observed that the relative levels of collagen types I and III and fibronectin are significantly higher in both the right ventricle and AV valves of neonatal Mas(-/-) mouse hearts (e.g., collagen type I: 85.28±6.66 vs 43.50±4.41 arbitrary units in the right ventricles of Mas(+/+) mice). Conversely, the level of collagen type VI was lower in the right ventricle and AV valves of Mas(-/-) mice. Adult Mas(-/-) mouse hearts presented similar patterns as observed in neonates. No significant differences in ECM protein level were detected in atria. Likewise, no changes in ECM levels were observed in AT(2) knockout mouse hearts. Although deletion of Mas induced a significant reduction in the level of the active form of MMP-2 in neonate hearts and a reduction of both MMP-2 and MMP-9 in adult Mas(-/-) mice, no significant differences were observed in MMP enzymatic activities when compared to controls. The levels of the active, phosphorylated forms of ERK1/2 and p38 were higher in hearts of both neonatal and adult Mas(-/-) mice. These observations suggest that Mas is involved in the selective expression of specific ECM proteins within both the ventricular myocardium and AV valves. The changes in the ECM profile may alter the connective tissue framework and contribute to the decreased cardiac performance observed in Mas(-/-) mice.

Membrane-type 4 matrix metalloproteinase (MT4-MMP) modulates water homeostasis in mice.

MT4-MMP is a membrane-type metalloproteinase (MMP) anchored to the membrane by a glycosyl-phosphatidylinositol (GPI) motif. GPI-type MT-MMPs (MT4- and MT6-MMP) are related to other MT-MMPs, but their physiological substrates and functions in vivo have yet to be identified. In this manuscript we show that MT4-MMP is expressed early in kidney development, as well as in the adult kidney, where the highest levels of expression are found in the papilla. MT4-MMP null mice had minimal renal developmental abnormalities, with a minor branching morphogenesis defect in early embryonic kidney development and slightly dysmorphic collecting ducts in adult mice. Interestingly, MT4-MMP null mice had higher baseline urine osmolarities relative to wild type controls, but these animals were able to concentrate and dilute their urines normally. However, MT4-MMP-null mice had decreased daily water intake and daily urine output, consistent with primary hypodipsia. MT4-MMP was shown to be expressed in areas of the hypothalamus considered important for regulating thirst. Thus, our results show that although MT4-MMP is expressed in the kidney, this metalloproteinase does not play a major role in renal development or function; however it does appear to modify the neural stimuli that modulate thirst.

Smooth muscle cell behavior in the ventral prostate of castrated rats.

Smooth muscle cells (SMC) play roles in prostatic development and function. The cells also respond to tissue injury and hormonal variations, alternating between a fully differentiated and contractile phenotype and a dedifferentiated synthetic or secretory phenotype. However, the phenotypic changes in SMC after androgen deprivation have not yet been described. The ventral prostate of control and castrated rats was processed for routine histology, immunocytochemistry, reverse transcriptase polymerase chain reaction (RT-PCR), and scanning electron microscopy (SEM). The maintenance of SMC phenotype was confirmed by immunocytochemistry and by RT-PCR. Stereological analyses were done to define the relative and absolute volume of the SMC. SMC were elongated and flattened against the epithelium. After castration, the cells shortened concomitantly with pleating of the cell surface, leading to a spinous aspect. SEM showed that the smooth surface of SMC became progressively folded. Immunocytochemistry demonstrated both smooth muscle myosin heavy chain and smooth muscle alpha-actin in the prostatic SMC 21 days after castration, whereas RT-PCR amplified the message for smoothelin. Stereological analysis showed an increase in the relative volume of SMC in relation to the whole gland and the stroma. A decrease in the absolute volume of SMC occurred only within the first 7 days after castration and remained unchanged thereafter. The prostatic SMC are affected by the absence of androgens and there is a critical transition point during the first week in which the total volume occupied by SMC diminished. The remaining SMC showed a marked phenotypical change. These findings indicate that ventral prostate SMC maintain their differentiated phenotype after castration. The alterations in SMC behavior correlate with general stromal modifications taking place after castration.