Expression of placental growth factor and a disintegrin and metalloprotease with a thrombospondin type motifs 1-4-8 during the three trimesters of rat pregnancy at the maternal-fetal interface.

The functional roles of the a disintegrin and metalloprotease with a thrombospondin type motifs (ADAMTS) gene family in reproductive physiology, reproductive organs developments and adult reproductive health are still under investigation. The expression of the anti-angiogenic proteases ADAMTS-1, ADAMTS-4 and ADAMTS-8 in placental angiogenesis at various stages of pregnancy also remains unclear. The purpose of this study was therefore to determine the localization and expression of the ADAMTS-1, ADAMTS-4 and ADAMTS-8 proteins during the three stages of pregnancy in rats. Maternal-fetal tissue samples were collected on Days 5, 12 and 19 of each trimester, corresponding to the first, second and third trimesters. The expression of placental growth factor (PlGF) and ADAMTS-1, ADAMTS-4 and ADAMTS-8 at the maternal-fetal interface was examined using immunohistochemistry and western blot at three distinct phases of pregnancy. ADAMTS-1, ADAMTS-4 and ADAMTS-8 were detected in all three trimesters of pregnancy. The relative amount of PIGF increased in the first trimester and decreased significantly in the third trimester (p < 0.05). The expression of ADAMTS-1 and ADAMTS-4 was significantly higher in the second (p < 0.05) and third trimesters (p < 0.01) compared to the first trimester. However, no statistically significant change was observed in ADAMTS-8 expression between trimesters. The ADAMTS exhibiting the highest expression during the first trimester was ADAMTS8. These findings indicate that the expression of ADAMTS-1, ADAMTS-4 and ADAMTS-8 in the three different stages of rat pregnancy may be involved in the modulation of decidualization, morphogenesis and angiogenesis. Periodic changes in ADAMTS expression are thought to be regulated by gonadal steroids.

Expression of ADAMTS 1-4-8 and placental growth factor in ovary and oviduct during pregnancy in the first trimester.

A Disintegrin and Metalloprotease Domains with Thrombospondins Motifs (ADAMTS), proteinases responsible for the destruction of extracellular matrix structures, have essential roles in the physiological and pathological processes of the female reproductive system, which is a dynamic structure. This study aimed to evaluate the immunoreactivity of placental growth factor (PLGF) and ADAMTS' (1, -4, and -8) in the ovary and oviduct during pregnancy in the first trimester. Our findings suggest a predominant role of ADAMTS-4 and ADAMTS-8 as proteoglycan-degrading enzymes from the ADAMTS-1 in the first trimester. As an angiogenic factor, PLGF showed more immunoreactivity than ADAMTS-1 in the ovary. This study provides the first evidence that ADAMTS-4 and ADAMTS-8 are more expressed in ovarian cells and follicles at different developmental stages during the first trimester of pregnancy than ADAMTS-1. Consequently, we suggest that ADAMTSs and PLGF act together and may exert specific effects on the formation, stabilisation, and function (or a combination thereof) of the matrix surrounding and protecting the follicles.

Distributions of PCNA and Cas-3 in rat uterus during early pregnancy.

The aim of present study was to determine the distributions of proliferating cell nuclear agent (PCNA) and Cas-pase-3 (Cas-3) and their possible roles in implantation and decidualization during early pregnancy at immunohistochemical level. The tissue samples from pregnant animals between gestational days 1-5 were incubated by PCNA and Cas-3 antibodies and the obtained results were evaluated quantitatively. It was observed that PCNA immunoreactivity in uterine luminal epithelium and glandular epithelium reduced as from day 2 of gestation and disappeared as from day 4 of gestation. PCNA staining intensity in stromal cells and myometrium increased gradually with progressing gestation. While Cas-3 immunoreactivity was strongly detected in luminal and glandular epithelium throughout the whole gestational period, its reactivity markedly increased as from day 3 of gestation. In conclusion, it may suggest that the blastocyst implantation induces the uterine luminal epithelial cell death and stromal cell proliferation around the embryo in the uterus.

The effect of pinealectomy and leptin hormone on the proliferation and apoptosis activation in Syrian hamster testis in different photoperiods.

The effects of pinealectomy and leptin hormone on proliferative and apoptotic processes in the epithelia of testicular seminiferous tubules of Syrian hamsters have been investigated. Proliferative and apoptotic processes were assessed semi-quantitatively by proliferating cell nuclear antigen (PCNA) and caspase-3 immune stainings. Animals used in the study were divided into four groups; control, pinealectomy (PinX), leptin-treated (10 microg/mL/day/kg body weight, intraperitoneally) and pinealectomy + leptin groups. Half of the hamsters in each group were exposed to short and the other half to long photoperiods for 8 weeks. In short photoperiod, PCNA activity especially in spermatogonia was significantly higher in the pinealectomy and leptin-treated groups compared with the control group. Histological score (HSCORE) value of PCNA in the PinX + leptin group was lower than those of PinX and leptin-treated groups. HSCORE value of caspase-3 in PinX and PinX + leptin groups was increased. In the long photoperiod, PCNA activation in the PinX group was significantly lower than the control group while the differences between the controls and other groups were not significant. The difference between the increases in caspase-3 activity in the PinX and control groups was significant. Thus, it was observed that photoperiods had no effect on the proliferation activity in the control groups. The inhibiting effect of short photoperiod on testis was not observed throughout 8 weeks. PinX eliminated the inhibiting effect of short photoperiod but did not alter the stimulating effect of long photoperiod. Leptin did not show any effect in long photoperiod but decreased proliferation by stimulating melatonin in short photoperiod.

Melatonin is as effective as testosterone in the prevention of soleus muscle atrophy induced by castration in rats.

The purpose of this experiment was to compare the weight, insulin-like growth factor-I (IGF-I) expression, and ultrastructure of the soleus muscle in growing castrated rats treated with testosterone or melatonin. In this study, adult male Wistar albino rats were used. The groups were arranged as sham, castrated, and testosterone- or melatonin-injected groups after castration. The soleus muscle samples were fixed in Bouin's solution for immunohistochemistry, and in 2.5% gluteraldehyde in 0.1 M phosphate buffer (pH 7.4). Whereas castration reduced the soleus weight and fiber diameter, testosterone and melatonin administration increased them. IGF-I immunostaining observed in the satellite cells and periphery of the myofibers was least intense in the castrated group. Strong staining of IGF-I was observed in the testosterone- and melatonin-administered groups. The ultrastructure of the soleus muscle in castrated animals showed the important ultrastructural modifications related to degeneration. In these groups, degenerative mitochondria, glycogen clusters under the sarcolemma, irregular Z lines, and loss of lamina externa were observed. The ultrastructure of myofibrils in the testosterone- and melatonin-injected groups was similar to that in sham groups in view of structure. In conclusion, we suggest that melatonin is as effective as testosterone in the prevention of atrophy induced by castration through the IGF-I axis.

Immunodetection of thyroid hormone receptor (alpha1/alpha2) in the rat uterus and oviduct.

The aim of this study was to investigate the immunolocalization and the existence of thyroid hormone receptors (THR) (alpha1/alpha2) in rat uterus and oviduct. For this purpose 6 female Wistar albino rats found in estrous period were used. Tissue samples fixed in 10% neutral formalin were examined immunohistochemically. Sections were incubated with primary mouse-monoclonal THR (alpha1/alpha2) antibody. In uterus, THR (alpha1/alpha2) immunoreacted strongly with uterine luminal epithelium, endometrial gland epithelium and endometrial stromal cells and, moderately with myometrial smooth muscle. In oviduct, they were observed moderately in the epithelium of the tube and the smooth muscle cells of the muscular layer. In conclusion, the presence of THR in uterus and oviduct suggests that these organs are an active site of thyroid hormones.

Immunolocalization of insulin-like growth factor I (IGF-I) during preimplantation in rat uterus.

OBJECTIVE: The aim of this study was to determine the insulin-like growth factor (IGF-I) immunolocalization in rat uterus at preimplantation period. DESIGN: The tissue samples were examined from pregnant animals between gestational day 1 and 5 using immunocytochemistry. RESULTS: While both the uterine luminal epithelium and the glandular epithelium were stained strongly from gestational day 1 - 3 by IGF-I antibody, the IGF-I immunoreactivity was moderate in myometrium and capillary endothelium. At this period, the IGF-I immunoreactivity was weakly present in a few endometrial stromal cells. At day 4, while IGF-I immunostaining intensity was particularly decreased in the basal domains of uterine luminal epithelium and glandular epithelium, it was similar to the first 3 days of gestation in myometrium and capillary endothelium. The endometrial vascularization and the number of stromal cells immunoreacting with IGF-I increased. The differentiation of stromal cells into decidual cells was seen at gestational day 5 and IGF-I was strongly expressed in the decidualizing cells. The IGF-I immunoreactivity in uterine luminal epithelium and glandular epithelium was similar to gestational day 4. IGF-I immunoreactivity was strongly detected in all of endometrial stromal cells. CONCLUSION: These results indicate that IGF-I expression in the rat uterus changes in the early pregnancy process and increase toward to day when implantation will be initiated.

The effects of triiodothyronine on rat testis: a morphometric and immunohistochemical study.

The aim of present study was to investigate the effects of 3,3',5-triiodothyronine (T(3)) on rat testis both morphometrically and immunohistochemically with determining of insulin-like growth factor I (IGF-I) expression. Adult male Wistar-albino rats used in the study were divided into two groups; control and T(3)-treated groups. After T(3) treatment there was observed to be a decrease in testicular weights, diameters of seminiferous tubules and the number of sertoli cells, and an increase in the number of leydig cells (P<0.05). Some of the seminiferous tubule lumens of T(3) administrated rats had cellular debris. IGF-I was localized in sertoli cells, late spermatids and leydig cells of all groups. IGF-I immunoreactivity in T(3) treated rats was higher than in controls in all stages of the cycle of rat seminiferous epithelium, but the staining intensity of leydig cells were similar in both groups. In conclusion, the present results suggest that T(3) may modulate the testicular function by affecting IGF-I activity at the gonadal level.

Expression of nidogens in rat uterus and embryo during decidualization and implantation.

The purpose of this study was to demonstrate the expression of nidogen-1 and nidogen-2 and their possible role in decidualization and implantation events during early pregnancy in rats. The tissue samples were examined from pregnant animals between gestational days 1-8 using immunocytochemistry. The uterine luminal epithelium, the glandular epithelium, and the myometrial smooth muscle cells stained strongly from gestational days 1-8 with both nidogen antibodies. At day 4 the decidual reaction areas began to appear in the stromal matrix and immunostaining of both nidogens revealed that the basement membrane of the surface epithelium was discontinuous. The differentiation of stromal cells into decidual cells was seen at gestational day 5 and both nidogens were weakly expressed in the decidualizing cells. At day 6, nidogen-2 immunoreactivity was higher in the primary decidual cells close to the embryo than nidogen-1, and during development of the decidual tissue both nidogens appeared in the endometrial stromal cells. At day 7, while expression of both nidogens declined in the primary decidual cells, their expression was markedly observed in the secondary decidual cells close to the myometrium. At day 8, expression of both nidogens was also observed to increase in the primary decidual cells. While nidogen-2 expression was seen in the parietal endoderm and primary ectoderm of the rat embryos at this developmental stage, nidogen-1 expression was only detected in the parietal endoderm. These results indicate that nidogen-1 and nidogen-2 could play important roles during embryogenesis, decidualization, and implantation in the endometrium of rat uterus.

Effects on rat testes of the thiosemicarbazone derivative Schiff base (4-(1-phenylmethylcyclobutane-3-yl)-2-(2-hydroxybenzylidenehydrazino)thiazole) and its cadmium(II) complex.

The aim of this study was to investigate structural and biochemical changes in testes of rats treated with the thiosemicarbazone derivative thiazole ring Schiff base, (4-(1-phenyl-methylcyclobutane-3-yl)-2-(2-hydroxybenzylidene-hydrazino) thiazole (L), and its Cd(II) complex (CdL(2)). The animals were divided into three groups. Group I was designated as control. The rats in groups II and III were injected subcutaneously with L or CdL(2) respectively at 150-mg kg(-1) doses at 3-day intervals for 15 days. At the end of the study, blood samples were collected for biochemical analysis, and testes were removed for histological examinations. Serum levels of vitamin A, E and MDA of the L-injected group were similar to the control group. While CdL(2) treatment decreased serum vitamin A and E levels, it increased the MDA level compared to other groups. Histologically, the testes structures of L-treated animals were similar to the control. Spermatogenic cells in seminiferous tubules of CdL(2)-treated animals displayed necrosis. Nuclei of spermatogonia and primary spermatocytes were pyknotic and heterochromatic. Homogenous pink particles were present in place of the spermatids. The interstitial areas were oedematous and intertubular vessels were plugged. In conclusion, the present results indicate that L does not cause biochemical and morphological alterations, but its Cd(II) complex has degenerative effects in normal rat testes.

Effects of high dose retinoic acid on TGF-beta2 expression during pancreatic organogenesis.

The aim of this study was to investigate the effects of excess all-trans retinoic acid, a vitamin A metabolite, on pancreatic organogenesis and TGF-beta2 expression during prenatal development in rats. First group of animals used as control while a single dose of 60 mg/kg all-trans retinoic acid was ingested by the mothers, at day 8 of gestation (before the neurulation period) in group II and at day 12 of gestation (after the neurulation period) in group III, and all embryos were sacrificed at day 18 of gestation. TGF-beta2 expression was detected in the capsule, acini and Langerhans islets in the control group. In the pancreas of group II, dilatation and congestion of interlobular vessels were observed. Langerhans islet structures were completely absent. Moreover acinar TGF-beta2 immune reactivity was not determined. In group III, acinar expression of TGF-beta2 in acid was similar to that in the controls but their Langerhans islets TGF-beta2 immune reactivity was significantly less than the controls. In view of the present findings we suggest that TGF-beta2 plays important role in pancreatic morphogenesis and administration of excess all-trans retinoic acid before neurulation inhibit TGF-beta2 expression disrupted pancreatic morphogenesis particularly Langerhans islets. However, its administration after neurulation had less adverse affect on pancreatic organogenesis and TGF-beta2 immune reactivity.

Reduction of carbon tetrachloride-induced nephropathy by melatonin administration.

The aim of this study was to investigate possible protective effects of melatonin on carbon tetrachloride (CCl4)-induced renal damage in rats. A total of 24 animals were divided into three equal groups: the control rats received pure olive oil subcutaneously, rats in the second group were injected with CCl4 (0.5 ml kg-1, s.c. in olive oil) and rats in the third group were injected with CCl4 (0.5 ml kg-1) plus melatonin (25 mg kg-1, s.c. in 10% ethanol) every other day for 1 month. At the end of the experimental period, the animals were sacrificed and blood samples were collected. The kidneys were removed and weighed. Urea and creatinine levels were determined in blood samples. Histopathological examination of the kidney was performed using light microscopic methods. Administration of CCl4 significantly increased relative kidney weight (g per 100 g body weight) and decreased serum urea levels compared to controls (p<0.01). Melatonin treatment significantly (p<0.01) reduced relative kidney weight, and it produced a statistically equal (p=0.268) relative weight with the kidneys of control rats. CCl4 administration alone also caused histopathologically prominent damage in the kidney compared to the control group. Glomerular and tubular degeneration, interstitial mononuclear cell infiltration and fibrosis, vascular congestion around the tubules, and interstitial haemorrhage in perivascular areas were observed in the renal cortex and cortico-medullary border. However, the affect of CCl4 on the medulla was limited. Melatonin provided protection against CCl4-induced renal toxicity as was evident by histopathological evaluation. In view of the present findings, it is suggested that melatonin protects kidneys against CCl4 toxicity.

Possible effects of melatonin on thymus gland after pinealectomy in rats.

OBJECTIVES: The purpose of this study was to investigate the effects of pinealectomy and pinealectomy plus melatonin administration on thymus weight and histology in adult Wistar-albino rats. METHODS: The animals were divided into three groups. Group I and Group II were designated as control (sham-pinealectomized) and pinealectomized rats, respectively. They received 10% ethanol (0.1 ml per day s.c.) alone. The rats in Group III were pinealectomized and daily injected with melatonin (3 mg/kg/0.1 ml 10% ethanol per day s.c.) commencing on the day seven after surgical operation. Injections were applied for two months. RESULTS: The thymus atrophied and its weight decreased after pinealectomy (p<0.001). The cortico-medullary boundary could not be distinguished and in the thymus induced a loss of lymphoid elements, increased number of phagocytic macrophages and enlarged blood vessels. Melatonin prevented the thymic involution. CONCLUSIONS: These results suggest that pinealectomy decreases thymus weight and that long-term administration of melatonin restores thymus weight to normal levels.

Effects of melatonin on skeletal muscle of rats with experimental hyperthyroidism.

The aim of this study was to investigate structural changes that occurred in the skeletal muscle of rats with experimental hyperthyroidism and the effect of melatonin on these changes. Groups of animals were designated as controls, 3,3',5-triiodothyronine (T3) injected and T3 + melatonin injected group. At the end of the study the tissue specimens were harvested and their structure examined. In the skeletal muscle of T3 injected rats a decrease was observed in muscle fiber diameter, splitting of fiber, collections of adipose tissue in perimysium, and gathering of nuclei in central compared to the control. Electron microscopic examination showed that mitochondria were dilated and the I band was less clear. In the T3 + melatonin injected group, the structure of fibers was similar to control. In conclusion, this study showed that T3 injection caused structural changes in the skeletal muscle and that melatonin had a positive effect on these changes.

Effect of vitamin E on follicular cell proliferation and expression of apoptosis-associated factors in rats with 6-N-propyl-2-thiouracil-induced goitrogenesis.

We have investigated immunohistochemically the effect of dl-alpha-tocopherol (vitamin E) on thyroid gland with 6-n-propyl-2-thiouracil (PTU)-induced hypothyroidism in rats. The animals were divided into four groups. Rats in group I were designated as control, rats in group II were treated with injections of PTU (10 mg/kg) for 15 days, rats in group III were treated with injections of PTU+vitamin E (10 mg/100 g) for 15 days. Rats in group IV were treated with injections PTU for 15 days and kept for 15 next days after cessation of PTU treatment. At the end of experiment, the animals were killed by decapitation, blood samples were obtained, thyroid tissues were collected and processed for quantitative evaluation of immunohistochemical PCNA (marker of cell proliferation), Bax (pro-apoptotic marker) and Bcl-2 (anti-apoptotic marker) staining. There was an increase in the number of PCNA-immunopositive cells in follicular epithelial cells of group II rats compared with other groups (p<0.05). After vitamin E treatment, the number of PCNA-immunopositive cells decreased (p<0.05) while the number of Bax-immunopositive cells increased (p<0.05). The number of Bcl-2-positive follicular epithelial cells of group IV rats was higher than in those of other groups (p<0.05). The results of this study indicate that hypothyroidism induces cell proliferation in the thyroid gland and vitamin E may promote involution of the gland.