Epithelial-mesenchymal interactions in prostatic development. I. morphological observations of prostatic induction by urogenital sinus mesenchyme in epithelium of the adult rodent urinary bladder

Tissue recombinants of embryonic urogenital sinus mesenchyme (UGM) and epithelium of the urinary bladder (urothelium, BLE) of adult rats and mice were grown for 3-30 d in male syngeneic hosts. Short-term in vivo growth indicated that prostatic morphogenesis is initiated as focal outgrowths from the basal aspect of the adult urothelium. The solid epithelial buds elongate, branch, and subsequently canalize, forming prostatic acini. After 30 d of growth in the male hosts, prostatic acini exhibit secretory activity. The marked changes in urothelial morphology induced by the UGM are accompanied by the expression of fine- structural features indicative of secretory function (rough endoplasmic reticulum, Golgi apparatus, and secretory granules). During this process, urothelial cells express prostatic histochemical markers (alkaline phosphatase, nonspecific esterase, glycosaminoglycans) and prostate-specific antigens. The expression within BLE of prostatic characteristics is associated with the loss of urothelial characteristics. These data indicate that adult urothelial cells retain a responsiveness to embryonic mesenchymal inductors. Furthermore, mesenchyme-induced changes in urothelial cytodifferentiation appear to be coupled to changes in functional activity.     

Enzymatic characterization of arginine esterase from dog seminal plasma

Previously purified arginine esterase from dog seminal plasma was characterized enzymatically. The enzyme was found to have a rather narrow specificity for arginine esters, much less for lysine esters and was practically devoid of activity towards tyrosine esters, casein, albumin and azocoll. It had a broad optimum pH between 8 and 9. It presented no kallikrein-like activities either in the blood pressure test in dog or in the rat uterus contraction test. It was inhibited by bovine pancreas trypsin inhibitor, aprotinin, phenylalanylprolyl arginine chloromethyl ketone, diisopropylfluorophosphate, phenylmethylsulfonyl fluoride, sodium dodecyl sulfate and leupeptin, but not by soybean trypsin inhibitor, tosyllysine chloromethyl ketone, tosylamide-2-phenylethyl chloromethyl ketone, iodoacetamide, Triton X-100 and EDTA. Experiments involving incubation of prostatic cytosol with purified arginine esterase showed that actin was the only important prostatic protein that was extensively hydrolyzed by this enzyme. It is not known presently whether the hydrolysis of actin is related to a true physiological function of the enzyme and whether actin and arginine esterase ever come into contact with each other in vivo. These properties indicate that arginine esterase from dog seminal plasma is different from other known proteinases including classical kallikreins, although it presents many similarities with this class of enzyme.     

Proteolytic activity of arginine esterase from dog seminal plasma towards actin and other structural proteins. Comparison with trypsin and kallikrein

At equimolar ratio of enzyme/substrate, actin, tropomyosin, fibronectin and myosin were extensively hydrolyzed during an incubation of one hour at 37 degrees C. Dog serum albumin, ovalbumin, bovine gamma-globulin and human prostatic acid phosphatase were not hydrolyzed. The activity of arginine esterase towards actin at pHs 6.5, 7.1 and 7.6 was respectively 60, 74 and 84% of the one found at optimum pH 8.2. The cleavage products of actin by arginine esterase and trypsin were similar although trypsin activity was 5000-fold higher. Kallikrein produced a major fragment of actin not observed with arginine esterase and trypsin. It is concluded that arginine esterase has a low trypsin-like activity towards structural proteins and that this activity may have a physiological significance.     

Common causes of male dog infertility

A complete breeding soundness evaluation is essential for assessment of the infertile male dog. Cryptorchidism, a sex-limited autosomal recessive trait, is more common as a unilateral condition. Azoospermia is an ejaculate consisting of seminal plasma but lacking sperm; repeated semen collections in the presence of an estrual bitch will rule out inadequate experience and lack of sexual stimulation. Both carnitine and alkaline phosphatase (AP) are produced in the epididymis; seminal plasma AP concentrations>5000 U/L indicate a normal ejaculate, whereas <5000 U/L is associated with incomplete ejaculation. Benign prostatic hypertrophy (BPH), the most common age-related condition in intact male dogs, is characterized by a sanguineous urethral discharge, hematuria, or hemospermia; diagnosis is based on prostatic enlargement and confirmed by a transabdominal biopsy. Although castration is recommended, valuable breeding dogs can be given finasteride. Prostatitis is more common in older dogs with BPH. Culture of the third fraction of the ejaculate or urine obtained by cystocentesis is indicated. Bacterial prostatitis is treated with antibiotics with high lipid solubility. Some dogs with bacterial prostatitis may develop prostatic abscesses (a medical and surgical emergency). Prostatic cysts are often asymptomatic. Approximately, 5-7% of dogs with prostatic disease have prostatic neoplasia, most commonly adenocarcinoma (it occurs in both intact and castrated dogs), which often metastasizes and has a very poor prognosis. Although a specific diagnosis can be made in many cases of male dog infertility, not all causes are amenable to treatment.     

Characterization and localization of alkaline phosphatase in canine seminal plasma and gonadal tissues

Alkaline phosphatase (AP) is a useful indicator of the presence of the sperm-rich (2nd) fraction in the canine ejaculate. Two AP isoenzymes originating from separate genes have been identified in the dog: tissue nonspecific (TNS) and intestinal. Bone, liver, and corticosteroid-induced AP are different isoforms of the TNS and intestinal isoenzymes. Using gel electrophoresis and levamisole inhibition assays, it was determined that seminal plasma AP (SAP) is a unique isoform of canine TNS AP whose glycosylation is distinct from either of the TNS AP isoforms commonly found in canine serum. Using immunocytochemistry, SAP activity was localized to the epididymal and seminiferous tubular epithelium. The ability to distinguish SAP from bone AP, liver AP and corticosteroid-induced AP could be beneficial to the practitioner in determining the quality of a semen sample.     

Review of acid phosphatase in the diagnosis and prognosis of prostatic cancers

Acid phosphatase is a secretory product frequently utilized as a tumor marker for disseminated, late stage (D2) prostatic cancer. In the 40 years since this association has been recognized, this enzyme has been subjected to extensive biochemical and immunological characterizations. These techniques have also been adapted for rapid and specific determinations of the prostatic isoenzyme levels using a variety of techniques. Since acid phosphatase levels do not become significantly elevated until late stage cancer, newer markers such as prostate-specific antigen have been sought which appear earlier and may be more useful for the screening and monitoring of high risk populations. At this time it is appropriate to review the current and future status of acid phosphatase as a diagnostic aid.     

Phosphomonoesterases in the male genital system of some mammals.

The various male genital organs of the experimental animals used in this investigation (rat, guinea pig, rabbit, cat and dog) showed a widely different localizations of alkaline and acid phosphatases. Alkaline phosphatase was presented as secretory, stromal, nuclear and vascular, while with acid phosphatase, distinction was made only between secretory and nuclear phosphatases. Although the morphological distributions of both enzymes were sometimes overlapping, they were not identical.     

Proteomic analysis of formalin-fixed prostate cancer tissue

Proteomic analysis of formalin-fixed paraffin-embedded (FFPE) tissue would enable retrospective biomarker investigations of this vast archive of pathologically characterized clinical samples that exist worldwide. These FFPE tissues are, however, refractory to proteomic investigations utilizing many state of the art methodologies largely due to the high level of covalently cross-linked proteins arising from formalin fixation. A novel tissue microdissection technique has been developed and combined with a method to extract soluble peptides directly from FFPE tissue for mass spectral analysis of prostate cancer (PCa) and benign prostate hyperplasia (BPH). Hundreds of proteins from PCa and BPH tissue were identified, including several known PCa markers such as prostate-specific antigen, prostatic acid phosphatase, and macrophage inhibitory cytokine-1. Quantitative proteomic profiling utilizing stable isotope labeling confirmed similar expression levels of prostate-specific antigen and prostatic acid phosphatase in BPH and PCa cells, whereas the expression of macrophage inhibitory cytokine-1 was found to be greater in PCa as compared with BPH cells.     

Comparison of the alpha-adrenoceptor characteristics in human and canine prostate

Alpha adrenoceptors, mediating contraction, have been shown to be present in strips of hypertrophic prostate surgically removed from patients with benign prostatic hypertrophy (BPH), providing a rational explanation for the demonstrated effectiveness of alpha antagonists in the symptomatic treatment of this disease. Inasmuch as the dog develops spontaneous and hormonally induced prostatic enlargement, studies were performed to compare the adrenoceptor characteristics of canine and human prostate to determine whether the dog represents a useful model to search for more effective alpha-adrenolytic therapy for human BPH. Norepinephrine produces contraction in isolated strips of canine prostate although it is only one-tenth as potent as previously reported in human tissue. In contrast, several selective alpha 1-adrenoceptor agonists are potent contractile agents in canine prostate, but are nearly inactive in the human tissue. This difference may be a consequence of their partial agonist character. The potency of selective alpha-adrenoceptor antagonists in blocking the norepinephrine-induced contractile response in both canine and human tissue is consistent with an action of norepinephrine on the alpha 1 adrenoceptor. The receptor dissociation constants for these antagonists are similar in prostatic tissue from dogs and humans, and the values in canine tissue correlate well with those obtained in the rabbit ear artery, a standard model for vascular alpha 1 adrenoceptors. Hence the dog may represent a useful model for studies of the potential responsiveness of human prostate to adrenergic agents.     

Fine needle aspiration cytology of prostatic adenocarcinoma metastatic to the lung confirmed by the immunoperoxidase technique

Two cases of prostatic adenocarcinoma metastatic to the lung are presented. The prostatic origin of the metastatic carcinomas was confirmed by immunocytochemical demonstration of prostate-specific antigen and prostatic acid phosphatase in cellular samples obtained by fine needle aspiration of the pulmonary lesions.     

Male Infertility Workup Needs Additional Testing of Expressed Prostatic Secretion and/or Post-Massage Urine

The male factor accounts for almost 50% of infertility cases. Inflammation may reduce semen quality via several pathways, including oxidative stress (OxS). As male infertility routinely is assessed using semen analysis only, the possible presence of non-leukocytospermic asymptomatic inflammatory prostatitis may be overlooked. We compared local and systemic OxS levels in male partners of infertile couples with different inflammation patterns in their genital tract and/or oligospermia.Subjects (n=143) were grouped according to inflammation in their semen, expressed prostatic secretion (EPS), and/or post-massage urine (post-M). Systemic (8-isoprostanes in urine) and local (diene conjugates and total antioxidant capacity in seminal plasma) OxS was measuredThe levels of OxS markers were significantly elevated in both severe inflammation groups – leukocytospermic men and subjects whose inflammation was limited only to EPS and/or post-M. Comparison between oligospermic and non-oligospermic men with genital tract inflammation, and oligozoospermic men with or without inflammation in the genital tract indicated that inflammation but not oligospermia status had significant impact on the measured OxS markers.Hence, a high leukocyte count in prostate-specific materials (EPS, post-M), even in absence of clear leukocytopsermia, is an important source of local and systemic OxS that may be associated with male infertility and affect general health. We suggest including the tests for detection of inflammation of the prostate into the workup of infertile men as was suggested in the WHO 1993 recommendation.     

Prostatic acid phosphatase immunoperoxidase staining of cytologically positive effusions associated with adenocarcinomas of the prostate and neoplasms of undetermined origin

An immunoperoxidase staining technique was employed in an effort to demonstrate prostatic acid phosphatase in sections of the effusion cell blocks in a retrospective investigation of the incidence of malignant prostatic cells in body cavity effusions in 33 patients with histologically confirmed prostatic cancer. An attempt was also made to identify the prostate as a possible anatomic site of origin in 26 patients with an unknown primary but with cytologically positive fluids. Neoplastic cells were identified in the effusion specimens in 21.2% of the patients with confirmed prostatic cancer; the sources, however, were either primary or metastatic carcinomas of nonprostatic origin. None of the cytologic specimens in this study demonstrated a positive prostate-specific acid phosphatase staining reaction, as did the prostatic metastases to the lungs used as controls.     

Characterization of protein-tyrosine kinase activity in the canine prostate.

Following the measurement of the phosphorylation of the substrate poly(Glu80Na,Tyr20) and the analysis of the alkali-resistant phosphorylation of endogenous proteins, the protein-tyrosine kinase of the canine prostate was partially characterized with regard to its subcellular localization, as well as certain kinetic and molecular properties. This kinase was mainly found in the cytosolic fraction (75%); however, its specific activity was similar to that of the residual enzyme present in the particulate fraction. Conditions for optimal activity of both fractions were determined. Under these conditions, several endogenous phosphoproteins (44-63 kilodaltons upon electrophoresis) were alkali resistant and phosphotyrosine was present in all of the major ones (pp63, pp57, pp52, and pp44). The particulate protein-tyrosine kinase activity was partially solubilized (58%) with 0.5% Triton X-100; this percentage was increased to 85% in the presence of 0.25 M KCl. Upon gel filtration, both cytosolic and particulate kinases showed an apparent molecular mass of 44 kilodaltons; these enzymes also phosphorylated similar major alkali-resistant phosphoproteins. The soluble protein-tyrosine kinase, with a sedimentation coefficient of 4.0S and an isoelectric point of 5.5, could be separated from arginine esterase and prostatic acid phosphatase.     

Arginine esterase from isolated dog prostate secretory granules is fully active enzymatically

We have isolated secretory granules from dog prostate homogenates and have determined whether a major portion of arginine esterase was localized in this fraction and if it was enzymatically active. Secretory granules were purified by density gradient centrifugation on sucrose, metrizamide, or Percoll. A major proportion of whole prostate homogenate arginine esterase was found in the granule fractions. Furthermore, the specific enzymatic activity in the granules was similar to the one observed in seminal plasma. No evidence could be found for the existence of significant amount of a zymogen inactive form of arginine esterase. These results suggest that arginine esterase could be active within the secretory granules in vivo and that it could hydrolyze protein substrates contained in this organelle.     

Stem Cell-Associated Marker Expression in Canine Hair Follicles

Functional hair follicle (HF) stem cells (SCs) are crucial to maintain the constant recurring growth of hair. In mice and humans, SC subpopulations with different biomarker expression profiles have been identified in discrete anatomic compartments of the HF. The rare studies investigating canine HF SCs have shown similarities in biomarker expression profiles to that of mouse and human SCs. The aim of our study was to broaden the current repertoire of SC-associated markers and their expression patterns in the dog. We combined analyses on the expression levels of CD34, K15, Sox9, CD200, Nestin, LGR5 and LGR6 in canine skin using RT-qPCR, the corresponding proteins in dog skin lysates, and their expression patterns in canine HFs using immunohistochemistry. Using validated antibodies, we were able to define the location of CD34, Sox9, Keratin15, LGR5 and Nestin in canine HFs and confirm that all tested biomarkers are expressed in canine skin. Our results show similarities between the expression profile of canine, human and mouse HF SC markers. This repertoire of biomarkers will allow us to conduct functional studies and investigate alterations in the canine SC compartment of different diseases, like alopecia or skin cancer with the possibility to extend relevant findings to human patients.     

Dimeric nature and amino acid compositions of homogeneous canine prostatic human liver and rat liver acid phosphatase isoenzymes. Specificity and pH-dependence of the canine enzyme.

Two isoenzymes of rat liver acid phosphatase (orthophosphoric-monoester phosphohydrolase (acid optimum) EC 3.1.3.2) have been purified to homogeneity, at least one of these for the first time. Both of the rat liver isoenzymes have identical specific activities towards p-nitrophenyl phosphate. Molecular weights of the native enzymes are 92 000 for rat liver isoenzyme I and 93 000 for isoenzyme II, while the subunit molecular weights are 51 000 and 52 000 respectively. Data on substrate specificity and pH dependence are presented for the homogeneous canine prostatic enzyme, which is also isolated as a dimeric enzyme of (native) molecular weight 89 000. Carbohydrate analysis data are presented for canine prostatic acid phosphatase and it is further noted that both isoenzymes of rat liver acid phosphatase are also glycoproteins. The amino acid compositions of the two rat liver isoenzymes are presented together with those of the similar dimeric acid phosphatase of human liver and of canine prostate. Comparison of these results with published data for the amino acid composition of human prostatic acid phosphatase shows substantial similarities. However, significant differences are seen in the amino acid composition of rat liver acid phosphatase isoenzyme I as compared to a previous literature report. Most notably, 17 histidine residues are found per mol of isoenzyme I and 18 for isoenzyme II.     

Characterization of 463 Type I markers suitable for dog genome mapping

In total, 463 canine gene markers were identified and characterized to serve as reagents in canine genome map projects. These markers are distributed over 221 canine gene markers, 139 TOASTs (Traced Orthologous Sequence Tags), 27 canine TOASTs, and 76 huESTs (human Expressed Sequence Tags). Out of 310 canine gene markers, 59%–84% were successfully amplified on dog DNA, the highest rates of success being observed when the exon/intron structure is known. Concerning TOASTs and human ESTs, of the 225 and 300 markers analyzed, 62% and 25% respectively were able to produce a dog positive amplification. As part of an ongoing project to map the canine genome using a dog/hamster radiation hybrid panel, these markers were tested for their specificity on dog versus hamster DNA. Thus 61%, 21%, and 12% of dog gene markers, TOASTs, and huESTs met the criteria required for radiation hybrid mapping, respectively. All of these 463 canine gene markers, however, are available and will be of value to any other mapping strategies. Received: 5 January 1999 / Accepted: 13 April 1999     

Incorporation of androgens at the time of cell differentiation of the prostatic epithelium in the rat

The appearance of androgen binding sites and acid phosphatase and esterase activities in the rat prostatic epithelium during the normal development was studied by means of steroid autoradiography and enzyme histochemistry. The incorporation of androgen into the prostatic epithelium began immediately before the lumen formation in the prostatic buds and the onset of their functional differentiation.     

Canine prostatic disease: a review of anatomy, pathology, diagnosis, and treatment

Disease conditions affecting the canine prostate gland are encountered frequently in small animal practice. The most common conditions affecting the canine prostate include benign prostatic hyperplasia, prostatitis, prostatic cysts, and prostatic neoplasia. Clinical signs associated with each of these conditions often overlap; therefore, it is important to reach a definitive diagnosis prior to initiating treatment. This paper reviews the diseases associated with the prostate gland of the dog, their diagnosis, as well as current treatment options for management of these conditions. Emphasis is placed on proper diagnostic sampling of the prostate gland, its fluid, and interpretation of findings, as well as emerging medical options for treatment of canine prostatic disease.