Evaluation of angiogenic apelin/apelin receptor axis in normal prostate, high grade prostatic intraepithelial neoplasia and prostatic adenocarcinoma.

INTRODUCTION AND OBJECTIVES: Prostate cancer is one of the most commonly diagnosed cancers in American men. Apelin is an endogenous peptide identified as the ligand of the G protein-associated apelin receptor. Apelin and apelin receptor have many tissues distribution and they participate in pathological processes, such as cancer. Apelin stimulates cancer angiogenesis. However, there are insufficient data in the literature regarding the role of apelin/apelin receptor in normal tissue, highgrade prostatic intraepithelial neoplasia, and prostatic adenocarcinoma tissues. Therefore, this study aimed to investigate the apelin and apelin receptor expression levels in tissues of normal prostate tissue, high-grade prostatic intraepithelial neoplasia, and prostatic adenocarcinoma. MATERIALS AND METHODS: In this study, 38 samples of patients undergoing radical prostatectomy were used. Among 38 samples; 20 patients were with prostatic adenocarcinoma, 18 patients were with high-grade prostatic intraepithelial neoplasia and adjacent normal prostatic tissue areas. The immunolocalisation of apelin and apelin receptor in these tissues were determined immunohistochemically. RESULTS: Apelin and apelin receptor expressions were higher in prostatic adenocarcinoma than normal prostate tissue and high-grade prostatic intraepithelial neoplasia. Apelin receptor expression was also increased in high-grade prostatic intraepithelial neoplasia compared to normal tissue. CONCLUSION: Apelin and apelin receptor are increase in the process of prostate carcinogenesis. This increase may adversely affect the clinical course of prostate cancer patients by stimulating angiogenesis, which is important for invasion and metastasis in prostate cancer.

The effect of apelin on the functions of peritoneal macrophages.

Apelin, the endogenous ligand of the G protein-coupled receptor (APJ), plays an important role in the physiological response to homeostatic perturbations. The aim of the present study was to investigate the effect of apelin on the functions of peritoneal macrophages. A double staining immunofluorescence technique was used to determine the expression of APJ in peritoneal macrophages. Rat peritoneal macrophages were randomly divided into three groups: control, apelin and apelin+F13A. A significant decrease in phagocytic and chemotactic activity of peritoneal macrophages resulted when the macrophages were incubated with [Pry(1)]-Apelin-13 (10 ng/ml). Incubation of peritoneal macrophages with the APJ receptor antagonist, F13A (20 ng/ml) prevented the suppressive effect of apelin on phagocytosis and chemotaxis. Peritoneal macrophages incubated with [Pry(1)]-Apelin-13 exhibited a decrease in the production of TNF-alpha and IL-6 compared to the control macrophages. Incubation of peritoneal macrophages with [Pry(1)]-Apelin-13 plus F13A prevented the decrease in the production of proinflammatory cytokines produced by [Pry(1)]-Apelin-13. In conclusion, apelin may be a mediator that inhibits the functions of activated macrophages.

Dynamic Hedgehog signalling pathway activity in germline stem cells.

Although the contribution of Hedgehog (Hh) signalling to stem cell development and oncogenesis is well recognised, its importance for spermatogonial stem cells (SSCs) has not been established. Here we interrogate adult rat SSCs using an established model in which only undifferentiated spermatogonial cells remain in the testis at 15 weeks following irradiation, and spermatogonial differentiation is induced within 4 weeks by gonadotrophin-releasing hormone antagonist (GnRH-ant) administration. Synthesis of Hh pathway components in untreated adult rat testes was compared with that in irradiated testes prior to and after GnRH-ant exposure using in situ hybridization. In adult testes with complete spermatogenesis, the Desert Hedgehog ligand transcript, Dhh, was detected in Sertoli cells, some spermatogonia and in spermatocytes by in situ hybridization. Spermatogenic cells were identified as sites of Hh signalling through detection of transcripts encoding the Hh receptor, Ptc2 transcripts and proteins for the key downstream target of Hh signalling, Gli1 and the Hh transcriptional activator, Gli2. Remarkably, the undifferentiated spermatogonia present in irradiated adult rat testes contained Dhh in addition to Ptc2, Gli1 and Gli2, revealing the potential for an autocrine Hh signalling loop to sustain undifferentiated spermatogonial cells. These transcripts became undetectable by in situ hybridization following GnRH-ant induction of spermatogonial differentiation, however, detection of Gli1 protein in spermatogonia in all groups indicates that Hh signalling is sustained. This is the first evidence of active Hh signalling in mammalian male germline stem cells, as has been documented for some cancer stem cells.

Cell cycle inhibitor p57 expression in normal and diabetic rat placentas during some stages of pregnancy.

Placentomegaly, an abnormal increase in the size of the placenta, is commonly seen in human diabetic pregnancies and diabetic animal experimental models. Proper placental development depends on the proliferation and differentiation of trophoblasts. However, our knowledge about the mitotic regulators that play key roles in synchronizing these events is limited. p57 is a cyclin-dependent kinase (CDK) inhibitor acting in the G1/S transition of the cell cycle. There is no data regarding p57 expression in either rat or human diabetic placentas. The purpose of this study was to investigate p57 expression in control and diabetic rat placentas at different stages of pregnancy. Diabetes was induced by streptozotocin on the first day of pregnancy, and placentas were taken on days 11, 13, 17, and 21 of pregnancy. Our results showed that on day 11, p57 immunostaining intensity was stronger in control group placentas compared to the diabetic group. On day 13, p57 immunostaining intensity increased in both groups, but increased more in the diabetic group. On day 17, p57 immunostaining intensity decreased in both the control and diabetic groups compared to day 13, yet the intensity remained higher in control placentas compared to diabetic placentas. On day 21 of pregnancy, p57 immunostaining intensity increased in the control group and it decreased from the day 17 level in the diabetic group. Western blot results showed consistency with immunohistochemistry results. Our study shows different expression patterns of p57 between control and diabetic rat placentas, which indicate p57 may play a role in abnormal placental formation resulting in placentomegaly arising from diabetes.

The distribution of transglutaminase in the rat oocytes and embryos.

Transglutaminases (TGs) are calcium-dependent enzymes that catalyze the transamidation of glutamine residues of a protein substrate to form intermolecular isopeptide bonds. The zona pellucida (ZP) is an extracellular, glycoprotein matrix that surrounds the oocytes of all Eutherian mammals. We aimed to identify the immunoreactivity of tissue transglutaminase (tTG) and ultrastructural changes occuring in rat oocytes before and after fertilization. Female rats were stimulated to superovulate, then mated with males. Oocytes and embryos were collected and examined by immunohistochemistry and electron microscopy. Before fertilization, tTG was present only in the oolemma and the cortical cytoplasm. After fertilization, tTG reactivity increased in the ZP of the early zygote and the preimplantation embryos, but decreased in the cytoplasm and perivitelline space (PVS). After fertilization, the PVS ultrastructure became asymmetrical and large around the polar bodies with many cortical granule contents. In conclusion, tTG immunoreactivity was found to be spatially and temporarily heterogeneous in the rat oocytes and embryos, especially in the ZP.

The zonal distributions of alkaline phosphatase, adenosine triphosphatase, laminin, fibronectin and chondroitin 4-sulphate in growing rat humerus proximal epiphyseal cartilage: a histochemical and an immunohistochemical study.

Although there are many studies about epiphyseal cartilage extracellular matrix (ECM) macromolecules in bone formation, studies of their distribution and role in the mineralization of these components in growing rat humerus proximal epiphyseal cartilage have not been sufficiently detailed. The aim of this study was to determine the distributions of alkaline phosphatase (ALP), adenosine triphosphatase (ATPase), laminin, fibronectin and chondroitin 4-sulphate in growing rat humerus proximal epiphyseal cartilage. The rats were killed by cervical dislocation, and the humeri were removed, sectioned (6 and 10 microm) on a cryotome or paraffin microtome, and stained using histochemical and immunohistochemical methods. ALP and ATPase were markedly observed in the hypertrophy and calcifying cartilage. In addition, ATPase was found to be very strongly positive in the tangential zone of articular cartilage. Results of immunohistochemical staining for laminin, fibronectin and chondroitin 4-sulphate showed that the immunostaining was the heaviest in the tangential zone of articular cartilage. In growing epiphyseal plates, there were differences in the density of these macromolecules of chondrocytes as a function of the maturation process. In conclusion, these ECM macromolecules of epiphyseal cartilage may regulate the cell-cell and cell-matrix interactions as well as the matrix calcification during the ossification of epiphyseal cartilage.

The effect of testosterone on gastrocnemius muscle fibres in growing and adult male and female rats: a histochemical, morphometric and ultrastructural study.

In this study, the effect of testosterone on gastrocnemius muscle fibres in growing and adult rats (male and female) was examined using histochemical, morphometric and ultrastructural techniques. After physiological saline (PS), olive oil (OvO) or olive oil + testosterone (OvOT) injections on 72 rats (growing and mature, 36 male and 36 female), the sample tissues of fibre types of the gastrocnemius muscle taken were examined by histochemical [alkaline adenosine triphosphatase (alk-ATPase), acid ATPase (ac-ATPase)], morphometric and ultrastructural techniques. In PS-injected control groups, the gastrocnemius muscle of both sexes contained all the fibre types studied [slow-oxidative muscle fibres (type I), fast-oxidative glycolytic muscle fibres (type IIA) and fast-glycolytic muscle fibres (type IIB)]. The type I fibres had the smallest diameter, type IIA had a medium diameter and type IIB fibres had the largest diameter. In OvO-injected groups, it was observed that the OvO had little effect on the gastrocnemius muscles of either sex, although there was significant enlargement of type IIB fibres. After the injection of OvOT, hypertrophy of muscle fibres was determined by morphometric study. The biggest increase in diameter was on type I fibres. In addition, degenerations on some mitochondria, accumulation of lipid droplets on type I and type II fibres, an increase in glycogen particles, bifurcation of myofibrils, an increase in the number and diameter of units resembling T tubules and an increase in ribosomal content were also observed in the same group by transmission electron microscope. Consequently, it was determined that testosterone can induce protein synthesis in gastrocnemius muscle fibres, and induces changes in shape and size, and also can change the appearance and the number of fibres.

A histochemical, morphometric and ultrastructural study of gastrocnemius and soleus muscle fiber type composition in male and female rats.

The fiber type composition of gastrocnemius and soleus muscles of adult male and female rats was examined by histochemical, morphometric and ultrastructural methods. Six male and 6 female, 3.5-month-old rats were used in the study. The fiber types were determined using the adenosine triphosphatase (ATPase) and succinate dehydrogenase (SDHase) staining techniques. The number of fibers of different types and the diameters of these two muscles were examined by morphometric methods. Gastrocnemius muscles were also examined by transmission electron microscopy. Z-line thickness, actin length, sarcomere length and M-line thickness of the fiber types were compared at the ultrastructural level. No significant differences were observed between the diameters of the gastrocnemius muscle (p > 0.05) of male and female rats although the male animals were significantly larger than the females (p > 0.05). In males, type I fibers of the gastrocnemius were larger than in females (p < 0.05). Region II of the gastrocnemius muscles of females contained more type IIA and type IIB fibers than in males. Sarcomere lengths were also longer in females (p < 0.05). The composition of fiber types of gastrocnemius and soleus muscles may be sex dependent.

The fine structure of normal and ectopic (tubal) human placental villi as revealed by scanning and transmission electron microscopy.

The aim of this study was to examine the development of chorionic villous trees during early periods of normal intrauterinal and ectopic (tubal) pregnancies, and to study the structural specializations on the free surface of mature placental villi by scanning and transmission electron microscope (SEM and TEM). In order to study the structures of placental villi between 28 and 34 days old (pc), early, 6-8 week normal and ectopic, and full term human placenta samples were obtained from legal curettage and hysterectomized cases, and spontaneous deliveries, and tissues samples were prepared for SEM and TEM. Three-dimensional configurations of the developing chorionic villous trees were observed as large main villus groups, covered with abundant microvilli of different size and diameters. It appeared that the chorionic villous trees which emerged from the chorionic plate divided gradually into branches of which ramifications originated as buds. These buds gradually grew and were transformed into shoots. The number of developing new villi appeared to increase gradually from 28 days to 9 weeks (pm) of gestation. From the 4th week onwards the massive trophoblastic sprouts were observed on the surface of main chorionic villi which transformed into primary, secondary and tertiary villous trees. When the placental villi formation in ectopic pregnancy was compared with the intra-uterinal pregnancy, an arrested development was remarkable. The configurations of ectopic placental villi seemed to be disparate, such as curved lines or compressed and wrinkled positions so that the three dimensional aspect had been wizened. The ramification and new villi formation seen as in the normal placenta were not only decreased but also infrequent. Some placental villi samples displayed a gradually thinning terminal region. Trophoblastic degenerations were frequently found on the surface of ectopic villi ultrastructurally. According to these results, we comment that in ectopic pregnancy the placental villi formation and development could have been delayed. At term, some specialized structural modifications were observed on the free surface of the mature placental villi. The presence of some dome-like balloonings and many crateriform hollows were the most striking features of the mature intermediate and terminal villi. According to the increasing physiological needs of the growing fetus, these special structures that are related to lung-like and kidney-like functions and named "nephropneumonic-like units", formed in the mature placental barrier. We have observed that these special units were showing a smooth surface similar to an inflated balloon.(ABSTRACT TRUNCATED AT 400 WORDS)

A histochemical study on the enzymatic activity in the proximal epiphysis of the humerus during the prenatal and postnatal periods in rats.

In order to investigate the activity of certain enzymes [Alkaline phosphatase (ALP), Acid phosphatase (ACP), Adenosine triphosphatase (ATPase), 5'-Nucleotidase (5'-N)] in the proximal epiphysis of the humerus, tissue specimens were obtained from pregnant rats on the 15th, 17th, 19th and 20th days of gestation and on the 1st, 10th, 20th, 30th, 40th and 60th days of postnatal development. Enzymatic activity in the chondral ossification, in the perichondral areas of the epiphysis, was first seen on the 15th day of gestation. ALP and ATPase could also be observed for the first time in fetuses aged 15 days, whereas ACP and 5'-N could not be detected. These latter enzymes were observed for the first time in the proximal humeral epiphysis of fetuses aged 17 days. ALP, a marker for hypertrophic and calcifying cartilage, was observed extensively in the central hypertrophic part of the cartilaginous perichondral zones, which showed calcification during the development of the epiphysis. ALP, ATP and 5'-N activity was very marked in the cytoplasm of osteoblasts and in the periosteal matrix, but strong ACP activity was found in the cells of the chondrolysis zone. In conclusion; according to our observations, heterogeneity of the proximal epiphysis of the humerus exhibits intrinsic differences between the cells of different zones. The activity of all enzymes showed an increase according to the developmental age. This suggests that all of these enzymes play a role during developmental ossification.