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Στην παρούσα εργασία γίνεται μία ανασκόπηση της βιβλιογραφίας που αφορά στη συμμετοχή του ενζυμικού συστήματος «ενεργοποιοί του πλασμινογόνου/πλασμίνη» (ινωδολυτικό σύστημα) σε φυσιολογικές διεργασίες στο γεννητικό σύστημα των θηλαστικών. Στο αρσενικό, οι ενεργοποιοί του πλασμινογόνου και οι αδρανοποιοί τους διαδραματίζουν σημαντικό ρόλο στη σπερματογένεση, στην ωρίμανση και στη γονιμοποιητική ικανότητα των σπερματοζωαρίων. Στο θηλυκό, η τοπικά ελεγχόμενη πρωτεολυτική δραστηριότητα των ενεργοποιών του πλασμινογόνου από τους αδρανοποιούς τους συμβάλλει σημαντικά στην πραγματοποίηση της ωοθυλακιορρηξίας, της γονιμοποίησης στον ωαγωγό, της εμφύτευσης του εμβρύου στη μήτρα, στη διατήρηση της κυοφορίας και στην παλινδρόμηση της μήτρας μετά τον τοκετό. Η δραστηριότητα του πρωτεολυτικού αυτού συστήματος μπορεί να επηρεαστεί από ποικίλους παράγοντες, όπως ορμόνες, φάρμακα, βιταμίνες, αλλά και φυσιολογικές και παθολογικές διεργασίες, με ποικίλα αποτελέσματα στη φυσιολογία του αναπαραγωγικού συστήματος., The current knowledge of the role of local and directed fibrinolysis controlled by plasminogen activators (PAs) and regulated by plasminogen activator inhibitors (PAls) in reproduction is summarized. The PA system has been found to play an important role in spermatogenesis in testis and modulation of sperm maturation in epididymis while a lot of studies indicate a role for sperm or seminal plasma PAs in sperm hyperactivation and/or capacitation. Hormoneinduced expression of tissue-type PA (tPA) and PAI-1 in the ovary is involved in the processes of ovulation and luteal regression; increases of urokinase-type PA (uPA) and PAI-1 in the early stage of luteinized follicles may be responsible for ovarian tissue remodeling and angiogenesis. The targeted proteolytic activity plays an essential role in the processes of the cyclic uterine angiogenesis, implantation and placentation as well as in the parturition. As the PA system is involved in multiple phases of mammalian fertilization specific regulatory molecules of this system provide opportunities for pharmacological intervention.

Plasminogen activators (PA) are proteolytic enzymes present in the spermatozoa and seminal plasma of various species. They play a role in the binding of the spermatozoon and its penetration through the layers surrounding the oocyte. Plasminogen activator activity (PAA) is modulated by hormones that have a seasonal variation, such as testosterone and melatonin. The present study investigates the seasonal variation of PA activity in sperm extracts and seminal plasma of four farm animal species: boar, buck, bull and stallion. Semen samples were collected every second week during a 12-month period and PAA was determined. With respect to sperm enzyme activity, the boar showed a peak from late January until the beginning of April, whereas the activity in the bull was at the highest levels from April until October and gradually declined during autumn and winter period. Plasminogen activator activity of stallion spermatozoa peaked during March and April, and remained low throughout the rest of the year, whereas in the buck sperm, PAA increased from late October until the end of January. No biologically significant variation was detected regarding the seminal PAA activity in any of the species studied. While seasonality of reproduction is typically studied from the female perspective, the present data provide compelling information about a factor that may affect the reproductive ability of the male.

Summary. Plasminogen activator activity (PAA) and plasminogen activator inhibition (PAI), against t-PA (t-PAI) or u-PA (u-PAI), in spermatozoa and seminal plasma as well as testosterone in the blood of Friesland, Chios, and Karagouniki rams all showed a seasonal variation with the highest values during the corresponding breeding season of the ewes (Autumn-Winter). The seasonal variation of PAA and PAI in spermatozoa or seminal plasma as well as blood testosterone was different among the three breeds studied. Increased spermatozoal PAA was observed in November and May in Friesland rams, in October and November in Chios rams, and in October in Karagouniki rams. Spermatozoal t-PAI was increased in December and June in Friesland rams, in November and December in Chios rams, and in November in Karagouniki rams. Spermatozoal u-PAI was increased in December in Friesland rams, in October and December in Chios rams, and in November and December in Karagouniki rams. Plasminogen activator activity and PAI in seminal plasma also showed similar seasonal variations. Plasminogen activator activity and PAI in spermatozoa and seminal plasma showed a positive correlation with blood testosterone. The results of the present study support our previous findings on the possible role of spermatozoal PAA and PAI in the fertilizing ability of spermatozoa.

After a single injection of serum gonadotrophin (PMSG) at the dose of 15 IU/kg, i.m. into rams testosterone in the plasma of blood showed a significant rise between 4th and 7th day post-injection. At the same time (4th-7th day) the plasminogen activator activity (PAA) in seminal plasma was found to be increased, but the plasminogen activator inhibition (PAI) expressed against t-PA (anti-t-PA) showed an increase between 32nd and 46th day. In spermatozoa a marked increase of PAA was revealed between 32nd and 46th day post-injection, while an increase of PAI (anti-t-PA) was exhibited on the 74th day. Plasmin inhibition (PI) in seminal plasma and spermatozoa showed no change compared to controls. A positive correlation has been found between increased concentrations of testosterone and PAA or PAI (anti-t-PA) in spermatozoa and seminal plasma. The induced increase of PAA in spermatozoa under the effect of testosterone might be of physiological importance, since PAA is localized to sperm membranes and might participate in the whole process of fertilization.

The mammalian oviduct is a dynamic tissue, which lies under the influence of ovarian steroids and produces proteins that affect various stages of fertilization and post-fertilization events. In this study, expression of urokinase-type plasminogen activator (u-PA mRNA) and plasminogen activator activity (PAA) were examined in porcine oviducts by reverse transcription and polymerase chain reaction (RT-PCR) and activity assays, respectively. For this purpose, oviducts were collected from Landrace cycling sows and divided into three segments (isthmus, ampulla, infundibulum). Different concentrations of u-PA mRNA were detected in the three segments following the pattern isthmus > ampulla > infundibulum and this pattern was maintained during the oestrous cycle. On the contrary, the highest PAA was measured in the ampulla compared to the isthmus and the infundibulum and the highest ampullary PAA was detected during the first 2 days of the oestrous cycle. The different regulation of u-PA mRNA expression and PAA is probably due to the existence of PA inhibitors. Recent observations suggest that PAI-1, the main inhibitor of PAs, shows greater expression in the isthmus compared to the ampulla and the local generation of plasmin is inhibited. The latter may be related to observations that spermatozoa are quiescent in the isthmus before fertilization. This study supports the suggestion that urokinase-type plasminogen activator has a biological role within the porcine oviduct, especially at or near the time of fertilization.

Acrosin and plasminogen activators are proteolytic enzymes of ram spermatozoa that play an essential role in the induction of the acrosome reaction, as well as the binding of spermatozoa to the oocyte and their penetration through the layers that surround the oocyte. Since vitamin A can alter gene expression in various tissues, testis included, this study was undertaken to evaluate the possible effect of vitamin A intake on acrosin- and plasminogen-activator activity. During a 20-week experiment, 15 rams of the Greek breed Karagouniki, divided to three groups, received different amounts of vitamin Aper osin retinyl acetate capsules (group A, controls, 12 500 iu/animal per day; group B, 50 000 iu/animal per day; group C, 0 iu/animal per day up to the 13th week, then 150 000 iu/animal per day until the end of the experiment). Acrosin- and plasminogen-activator activity were determined by spectrophotometric methods. Vitamin A was determined in blood plasma by HPLC. No statistical differences were detected regarding the body weight of the rams or the qualitative and quantitative parameters of their ejaculate throughout the whole experiment. No statistically significant alterations of enzyme activity were detected in group B. In group C, both enzyme activities started declining in week 9. Compared with controls, maximum reduction for acrosin was 49% on week 11 and for plasminogen activators 51% in week 14. Activities returned to normal rates after vitamin A resupplementation. To date, the main result of vitamin A deficiency was known to be arrest of spermatogenesis and testicular degeneration. A new role for vitamin A may be suggested, since it can influence factors related to male reproductive ability before spermatogenesis is affected.

The aim of this study was to investigate the effect of dexamethasone on acrosin activity of spermatozoa in Chios rams during autumn (breeding season for sheep in Greece), in correlation with possible changes in blood testosterone. Dexamethasone was administered in four equal consecutive intramuscular injections, one every four hours (total dose: 3 mg kg(-1)). Total acrosin activity was determined in semen samples collected 48 h before and on the 4th and 7th day and thereafter once every week until the 77th day after dexamethasone administration. Blood samples for testosterone radioimmunoassays were collected 24 h before, during dexamethasone administration and on the 4th, 7th, 14th and 21st day after administration. Total acrosin activity in spermatozoa was reduced between days 7-28 after dexamethasone administration. Dexamethasone also induced a reduction in mean value and basal level of blood testosterone and inhibited its episodic secretion between 1 and 4 days after administration. As the reduction of acrosin activity appeared relatively soon after dexamethasone administration (7th day), it is likely that the increased amount of dexamethasone did not influence the synthesis of proacrosin in the late spermatids. As glucocorticoid receptors exist in the epididymis and accessory glands in various species, dexamethasone may have a direct influence on the synthesis and/or release of acrosin inhibitors in epididymal fluid or seminal plasma. These changes in acrosin activity in ovine spermatozoa mediated by dexamethasone may be of importance regarding the role of stress in the reduction of sperm fertilizing ability.

The effect of tannic acid, a common flavonoid, on the acrosin and plasminogen activator activity and plasmin activity of human and ram spermatozoa was evaluated. Acrosin and plasminogen activator activity were determined by spectrophotometry using the chromogenic substrates N-alpha-benzoyl-DL-arginine para-nitroanilide-HCl (BAPNA) and H-D-valyl-L-leucyl-L-lysine-p-nitroanilide-2HCl (S-2251), respectively. In extracts from both human and ovine acrosomes, the activities of acrosin and plasminogen activators were susceptible to tannic acid inhibition. The inhibitory effect of tannic acid was observed at concentrations > 50 micromol l(-1) in a dose-dependent manner. In additional experiments, low concentrations of tannic acid significantly inhibited tissue-type plasminogen activator, urokinase-type plasminogen activator and plasmin activity in a concentration-dependent manner over the range 0.25-200 micromol l(-1). Tannic acid reduced the motility of ram spermatozoa at a concentration of 1000 micromol l(-1) after 2 and 3 h co-incubation with spermatozoa. The motility of human spermatozoa remained unchanged over the range 0.1-1000 micromol tannic acid l(-1) during 3 h co-incubation. These results indicate that tannic acid inhibited the activity of both acrosin and plasminogen activator and indicates a possible mechanism by which flavonoids exert their antifertility effects.

The effect of gossypol--a polycyclic compound isolated from cotton seeds--on the plasminogen activator activity of man and ram acrosomal extracts was explored in vitro. The action of gossypol on the plasminogen activator activity was investigated by a spectrophotometric method using the chromogenic substrate S-2251. Gossypol, a known antispermatogenic agent, was found to effectively inhibit human and ovine acrosomal plasminogen activator activity. The inhibition was dose-dependent. Plasminogen activator activity from man and ram extracts was completely inhibited by 350 mumol l-1 and 300 mumol l-1 of gossypol, respectively. In additional experiments, low, non-spermicidal concentrations of gossypol (2.5-40 mumol l-1) were found to significantly inhibit plasmin activity in a dose-dependent manner. The results suggest that inhibition of both acrosomal plasminogen activator and plasmin activity is a possible mechanism by which gossypol exerts its antifertility effect, since the plasminogen activator/plasmin system plays a role in the whole process of ovum fertilization.

The aim of the study was to investigate the effect of dexamethasone on plasminogen activator activity (PAA), t-PA antigen level, plasminogen activator inhibition (PAI) and plasmin inhibition (PI) in ram lymphocytes. The PAA of ram lymphocytes was due to t-PA (tissue-type plasminogen activator), but not to u-PA (urokinase-type plasminogen activator). Dexamethasone at the dose of 3 mg kg ˇ1 increased the PAA and t-PA antigen level of lymphocytes within 2 h after administration, while decreased the PAI 6‐14 h after administration; no change in PI was noted. When dexamethasone was administered at the dose of 0.75 mg kg ˇ1 every 4 h for 12 h, PAA and t-PA antigen level of lymphocytes showed an increase 2‐6 h after the end of the administration, while PAI and PI remained unchanged. These changes in PAA or PAI of lymphocytes, dependent on the dose or the pattern of dexamethasone administration, may be of importance regarding the role of lymphocytes and PAA or PAI in several pathophysiological conditions. # 1999 Elsevier Science B.V. All rights reserved.

Summary The effect on airway pressure of xylazine alone or following the administration of atipamezole or atropine was studied in 31 halothane-anaesthetized sheep. Xylazine produced a significant increase in airway pressure which lasted for at least 30 min. This effect was inhibited by both atipamezole and atropine. The results suggest that the xylazine-induced increase in airway pressure in sheep is α2-adrenergically mediated. Moreover, activation of central α2-adrenoceptors leading to vagal stimulation may be involved.

Plasminogen activator activity (PAA), tissue-type PA (t-PA) antigen level, plasminogen activator inhibition (PAI), and plasmin inhibition (PI) showed no sex-dependent differences in the stomach of the rat. Chronic ethanol consumption induced an increase of PAA and t-PA antigen and a decrease of PAI (urokinase-type PA [u-PAI]) and PI in the body and pyloric region (PI only in the pyloric region). PAA increase and u-PAI decrease in the pyloric region were greater in the male than in the female. Orchidectomy, but not ovariectomy, induced effects similar to those of ethanol. The effects of orchidectomy and chronic ethanol consumption on gastric PAA, t-PA antigen, and PAI were additive. An increased PAA and decreased PAI or PI in the stomach in cases of hypogonadism, particularly if this deficiency is combined with chronic ethanol consumption, might influence the susceptibility to pathological conditions related to or affected by the local fibrinolytic activity (PAA, PAI, or PI).

In normal Friesland rams, plasminogen activator activity (PAA) and plasminogen activator inhibition (PAI) in spermatozoa and seminal plasma, plasmin inhibition (PI) in seminal plasma and testosterone concentration in blood all showed seasonal variation. In a Friesland ram with chromosomal chimaerism XX/XY, however, although PAA, PAI and PI in seminal plasma and testosterone in the blood exhibited the same seasonal variation as in normal rams, in spermatozoa the PAA and PAI showed no seasonal variation. This is the first case in which a chromosomal anomaly has been linked with abnormal patterns in PAA and PAI.

The effect of unilateral and bilateral adrenalectomy, unilateral and bilateral demedullation or unilateral adrenalectomy/unilateral demedullation on tissue plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) was studied in the rat. All the above treatments induced an increase of PAA in lungs. Increased PAA was also noted in brain after bilateral adrenalectomy or demedullation, as well as unilateral adrenalectomy/unilateral demedullation. In kidneys the PAA was decreased after bilateral adrenalectomy or demedullation and unilateral adrenalectomy/unilateral demedullation. In aorta only after bilateral adrenalectomy a decreased PAA was induced. In heart and liver no changes in PAA were noted. A decreased PAI expressed against tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) was found in lungs of all treated rats compared to controls. However, an increased PAI was noted in heart, liver and aorta against t-PA or u-PA after bilateral adrenalectomy and in liver after bilateral demedullation as well as after unilateral adrenalectomy/unilateral demedullation (rats with only one adrenal cortex). A dissociation between anti-t-PA and anti-u-PA response was seen in heart, liver and aorta. A variable change in PI was induced in kidneys, aorta, lungs and brain in all treatments except unilateral demedullation or unilateral adrenalectomy. Therefore, the effect of adrenalectomy or demedullation on tissue PAA, PAI and PI was variable and dependent on the extent of the treatment (unilateral or bilateral), the organ as well as the time after the treatment.

Plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) have been studied with spectrophotometric methods in extracts of human, bovine, ovine and rat kidneys of both sexes. In all species studied, renal PAA (cortex or medulla) was higher in females than in males. The PAA was also higher in the medulla than in the cortex in all species and both sexes. The PAA was due to both types of plasminogen activator; tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA). In the human kidney (cortex or medulla) the measurement of t-PA antigen showed that t-PA is higher in females than in males; t-PA is also higher in the medulla than in the cortex in both sexes. The PAI showed the opposite pattern in all species studied; it was lower in females than in males. It was also lower in the medulla than in the cortex. PAI-1 was identified in the human kidney. Sex-related differences in renal PAA or PAI almost disappeared after bilateral orchidectomy in rats. PI showed no sex or regional differences in the species studied. Sex-related differences in renal PAA and PAI in man and various animal species might be of physiological or pathophysiological importance.

In the normal stomach of male rats marked differences in plasminogen activator activity (PAA) and plasmin inhibition (PI), but not in plasminogen activator inhibition (PAI), were noted among cardiac area, body and pyloric region. Chronic ethanol consumption (for 15 or 30 days) at the concentration of 6% or 12% in the drinking water induced an increase in PAA in the pyloric region and the body of the stomach (the higher concentration after 15 days and both concentrations after 30 days). The response was time- and dose-dependent. At the cardiac area no change of PAA was noted. Ethanol at both concentrations induced after 30 days a decreased PAI in the pyloric region and the body of the stomach, which was expressed against u-PA, but not against t-PA. A decreased PI was noted at both concentrations of ethanol after 30 days only in the pyloric region. Therefore, changes in PAA, PAI and PI after chronic ethanol consumption were dependent on the concentration, the period of the consumption and the area along the gastric wall.

The aim of the present study was to investigate the effect of melatonin on plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) in ram spermatozoa and seminal plasma, in correlation with changes in blood testosterone. Melatonin implants (18 mg) were placed subcutaneously in sixteen Chios rams in autumn and spring. Semen samples for spectrophotometrical assays were collected 36 h before the implantation of melatonin and thereafter once a week, for 17 weeks. Blood samples for testosterone assay (RIA) were collected 8h before implantation (one sample/30 min x 7.5 h) and thereafter every 15 days for 105 days after implantation. For each ram, six parameters of testosterone were estimated: mean value, basal level, number of peaks, peak amplitude, peak duration and mean testosterone concentration during peaks. Melatonin implantation during autumn induced an increase in PAA and t-PAI in spermatozoa; melatonin implantation in spring induced an additional increase in u-PAI and PI; no change in PAA, PAI or PI was found in seminal plasma, during autumn or spring. The melatonin-induced increase of PAA, PAI and PI in spermatozoa was in positive correlation with the increase of testosterone mean value, basal level and number of peaks; the increase of testosterone parameters was greater in autumn compared to spring. Changes of PAA, PAI and PI of spermatozoa, under the influence of melatonin, might indicate changes in the fertilizing ability of spermatozoa, since plasminogen activators and their inhibitors are present on the plasma and the outer acrosomal membrane of spermatozoa and are released during the acrosome reaction.

The aim of the present study was to evaluate plasminogen activator activity (PAA), tissue-type plasminogen activator (t-PA) antigen level and plasminogen activator inhibitor-1 (PAI-1) antigen in normal canine gingival tissue samples, gingivitis as well as in different stages of periodontal disease. Gingival tissue from 141 adult dogs were analysed spectrophotometrically in order to determine PAA. The tissues were also examined histopathologically. The Sulcus Bleeding Index was used to evaluate the active and inactive phase of periodontal disease. T-PA antigen as well as PAI-1 antigen level was measured by ELISA. There was a significant increase of PAA and t-PA antigen in samples from inflamed gingival tissue compared with normal gingival tissue, while PAI-1 antigen was not detected in either normal or inflamed gingiva. As the severity of periodontal disease was increasing, PAA and t-PA antigen values were significantly higher in periodontitis tissue sample groups, according to the pattern: gingivitisearly periodontitismoderate periodontitissevere periodontitis (P0.001). PAA and t-PA antigen were increased in samples from the inflamed gingival tissue with higher Bleeding Index, (heavy bleedingmoderate bleedingslight bleeding, P0.001). In conclusion, this study indicates that PAA and t-PA antigen level may be used to evaluate the evolution of periodontal disease in dog.

In a series of consecutive blood sampling in 15 days intervals over 15 weeks after implantation of melatonin in rams an increased mean value, basal level and number of peaks of testosterone was observed in samples of the third fortnight (45th day). This increase was greater in the autumn (breeding season) than in spring (non-breeding season). Total acrosin activity in spermatozoa was increased between days 35-56 (autumn) and days 49-70 (spring) after implantation and the relative increase was higher in autumn than in spring. The increase of acrosin activity was independent of the changes of testosterone. An increase of acrosin activity by melatonin, in cases of low activity, might improve fertilization rates in sheep not only during the breeding season, but also during the non-breeding season (after oestrus induction).

The objective of this study was to evaluate the effect of alpha-tocopherol on blood testosterone and specific proteolytic enzymes in spermatozoa and seminal plasma, with final aim the increase of sperm fertilizing ability with a nutritional supplement. The effect of alpha-tocopherol supplementation on testosterone parameters (mean value, basal level, peak number, mean concentration value during peaks, peak amplitude, peak duration) and plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) of spermatozoa and seminal plasma was studied in the ram during autumn (estrous period for the sheep in Greece) and spring (anestrous period). Treated animals showed a marked increase in serum alpha-tocopherol. Testosterone parameters were not affected by the alpha-tocopherol in either autumn or spring, however, the spermatozoal PAA and PAI (anti-tPA) were increased in the spring but not in autumn. These enzymes and their inhibitors are normally increased in autumn (the breeding season) and low in spring. If PAA can improve the fertilizing ability of spermatozoa in the spring, our finding may mean that a nutritional supplement, such as alpha-tocopherol, could provide rams for an accelerated onset of the breeding season in the ewe.

Dexamethasone in low and high doses affects blood fibrinolytic activity both in animals and humans. In this study the effect of a high dose of dexamethasone on plasminogen activator activity (PAA), t-PA antigen level, plasminogen activator inhibition (PAI) and plasmin inhibition (Pl) in rat heart, brain, liver, lungs and kidneys was investigated in both sexes.Twenty male and twenty female adult Wistar rats were used. Dexamethasone was administered as a single intraperitoneal injection (3mg/kg/day) in rats, once daily, for a period of 5 consecutive days. t-PA antigen level was assayed by an enzyme-linked immunoabsorbant assay method. PAA, PAI and Pl were determined by spectrophotometric methods. The plasminogen used was isolated from rat plasma.Dexamethasone induced variable changes in the fibrinolytic parameters in rat heart, brain and liver of both sexes; in lungs and kidneys dexamethasone had no effect.These changes of PAA, PAI and t-PA antigen level in heart, brain and liver induced by dexamethasone might be of importance regarding the involvement of glucocorticoids and plasminogen activators/plasmin in many pathophysiological conditions.

The effect of aspirin or endotoxin or aspirin plus endotoxin on the circadian variations of plasminogen activator activity (PAA), tissue-type plasminogen activator (t-PA) antigen level, plasminogen activator inhibition (PAI) and plasmin inhibition (PI) in aorta, heart and brain of the rat was studied. In aorta the circadian variations of PAA, t-PA antigen and tissue-type PAI (t-PAI) were blunted by endotoxin, while the circadian variation of PI was reversed; t-PAI variation was blunted by aspirin plus endotoxin, while PI variation was reversed. Various disturbances of the circadian variations of PAA, t-PA antigen, PAI or PI in heart and brain were also observed. In conclusion, the circadian variations of fibrinolytic parameters in key organs are influenced by aspirin or endotoxin or aspirin plus endotoxin in a varying way or degree depending on the treatment, the parameter studied or the organ.

1. In ileal segments obtained from 1, 15, 30, 45 and 60-day-old guinea pigs, histamine produced a dose-dependent significant contractile response from a threshold concentration of 3.2 x 10(-8) M. 2. The mean contractile responses (at 10(-5) M) of the ileum from the 1, 15, 30 and 45-day-old guinea pigs compared to that from the 60-day-old ones were 73.79, 92.30, 94.07 and 94.41% respectively. 3. Dimaprit (at 10(-5) M) had no effect on the ileum of the 1-day-old guinea pigs, while it produced a significant contractile response on the ileum of the guinea pigs at all other ages. 4. The mean contractile responses of the ileum from the 15, 30 and 45-day-old guinea pigs compared to that from the 60-day-old ones were 74.44, 66.66 and 76.94% respectively. 5. Cimetidine (at 10(-5), 3.2 x 10(-5) and 10(-4) M) significantly potentiated the histamine-induced contraction on the ileum of the 1-day (from 10(-7) to 3.2 x 10(-6) M) and the 15-day-old guinea pigs (at 10(-8) M). 6. On the contrary, cimetidine (at 3.2 x 10(-5) M) significantly inhibited the contractile response of the ileum to dimaprit. 7. In conclusion, the guinea pig ileum appears increasingly sensitive to histamine from the newborn to the 60-day-old ones, but its sensitivity to dimaprit, not detectable in the newborn, fluctuates during development. 8. Cimetidine produces a potentiating effect, which declines with age, on the on the histamine-induced contraction, while it inhibits the dimaprit contractile response on the ileum.

Plasminogen activator activity (PAA), t-PA antigen level, plasminogen activator inhibition (PAI) and plasmin inhibition (PI) showed a circadian variation in rat aorta, heart, brain and lungs of both sexes, but in a different way depending on the organ and the fibrinolytic parameter studied and the sex as well. In general, the lowest values of PAA and t-PA antigen were found in the early morning and the highest values in late afternoon, while PAI or PI showed the opposite pattern (with the exception of lungs). In kidneys and liver the fibrinolytic parameters studied showed no circadian variation. Sex-related differences, mostly quantitative, were noted in aorta, heart, brain and kidneys, with lower PAA and t-PA antigen and higher PAI or PI in the male than in the female.

We have recently shown that spermatozoa of various species contain both types of plasminogen activator, the tissue-type (t-PA) and the urokinase-type (u-PA). In the present study, the localization of t-PA and u-PA in plasma membrane and outer acrosomal membrane of human and boar spermatozoa has been investigated. The identification of the type of the plasminogen activator (t-PA or u-PA) was made immunologically. In human spermatozoa, the outer acrosomal membrane and plasma membrane contained both types of plasminogen activator (t-PA and u-PA); in addition, t-PA antigen was measured. In boar spermatozoa, the outer acrosomal membrane contained only t-PA, whereas plasma membrane contained both types of plasminogen activator (t-PA and u-PA). Plasminogen activator inhibition (PAI) has also been demonstrated in plasma and outer acrosomal membranes of both species and identified as PAI-1 in membranes of human spermatozoa.

Unilateral or bilateral ovariectomy was performed in young and adult rats. Mainly bilateral ovariectomy induced variable changes of plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) in key organs (brain, lungs, heart, aorta and kidneys). The most remarkable changes were induced after bilateral ovariectomy performed in young animals and mostly after two and three months of ovariectomy. Therefore, the effect of ovariectomy on tissue PAA, PAI or PI was variable and dependent on the extent of the ovariectomy (unilateral or bilateral), the age of the animal at ovariectomy (young or adult), the time after ovariectomy, and the organ. An additional interesting finding was the dissociation in the response of tissue anti-t-PA and anti-u-PA activities to ovariectomy in some of the organs studied.

The transplacental effect of lead compounds (lead acetate and tetraethyl lead) on the tissue plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) was studied in the rat. The concentration of lead in organs of the newborn showed a great variation; the distribution of lead in the organs studied depended on the dose and the stage of gestation at injection. In each organ the concentration of lead was dose-dependent. In control specimens no lead could be detected. The tissue response of PAA, PAI and PI to the lead compounds also showed a great variation; however, there was no correlation between lead concentrations and PAA, PAI or PI responses. Changes of one or more of the parameters studied (PAA, PAI or PI) were noticed in lungs, liver, heart, brain and kidneys. The PAA was due to the tissue type plasminogen activator in all organs studied; in kidneys and lungs the urokinase type of plasminogen activator was also detected. Therefore, fetal tissue PAA, PAI and PI can be affected transplacentally by lead compounds.

The effects of the proteolytic enzyme plasmin and the plasmin inhibitor ϵ-aminocaproic acid (EACA) on the rupture of the follicular wall were studied in the sheep. Injections into mature follicles of the same size and morphology located at the same ovary showed that plasmin increases, while EACA decreases, significantly the incidence of follicle rupture compared to this incidence after intrafollicular injections of the buffered diluent of these agents (controls). The present in vivo study provides evidence that plasmin plays a significant role in the process of the rupture of the follicular wall during ovulation as has been previously shown for plasminogen activator activity.

The effect of endotoxin, aspirin and endotoxin after administration of aspirin on the tissue plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) was studied in the rat. PAA, PAI and PI were determined in key organs (brain, heart, lungs, kidneys, liver and aorta) spectrophotometrically by procedures involving hydrolysis of the chromogenic substrate S-2251. Aspirin at three different doses had not any significant effect on tissue PAA; PAI and PI were affected in several organs. Four hours after a sustained infusion of endotoxin PAA was found to be increased in brain, kidneys and aorta, decreased in heart and lungs, while in liver the PAA was unchanged compared to controls. Changes in PAI and PI showed also a tissue variation. In endotoxin-infused rats pretreated with aspirin the PAA changes induced by endotoxin were prevented or modified; PAI and PI were affected in most organs studied. However, this effect of aspirin was varying and depending on the tissue, the parameter studied and the dose of aspirin. In some organs, as the heart and lungs, changes in PAI or PI were noticed, while neither aspirin nor endotoxin separately induced such changes in these organs. Also, a differential response of PAI and PI to the same stimulus in the same tissue was a noteworthy finding. The results of the present study show that the response of PAA, PAI and PI to aspirin depends on the tissue, the physiological or pathophysiological condition of the tissue and the dose of the aspirin.

Marked changes of the plasminogen activator activity, plasmin inhibition, and plasminogen activator inhibition were found in fatty streaks and fibrous plaques compared to the normal intima of the human aorta. Plasminogen activator activity and plasmin inhibition showed the following pattern: fibrous plaques greater than fatty streaks greater than normal intima. Plasminogen activator inhibition showed the opposite pattern: normal intima greater than fatty streaks greater than fibrous plaques. The tissue-type, but not the urokinase-type, plasminogen activator was detected immunologically.

Plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) were demonstrated in spermatozoa and seminal plasma of man, bull, ram, buck, boar and stallion. There was a great species variation in the PAA, PAI and PI. In spermatozoa and seminal plasma both types of plasminogen activator, namely of the tissue type (t-PA) and urokinase type (u-PA), were detected immunologically. The effect of plasmin on sperm motility was studied in the semen of bull. The addition of plasmin to the semen induced in certain concentrations a marked increase in sperm motility. The effect of plasmin on sperm motility seems to be direct.

The wall of the follicle in the sow (theca externa, theca interna-membrana granulosa) as well as the follicular fluid showed plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI). The follicular wall exhibited the highest PAA at the time of ovulation. The PAA was much higher at the area of the rupture than in the remaining outstanding part of the follicular wall. In the wall of ruptured follicles, with their openings more or less sealed, the PAA was markedly decreased. In mature follicles, the PAA was higher in theca interna-membrana granulosa than in theca externa. Follicular fluid showed an increasing PAA with the growth of the follicle. In the follicular wall and fluid both types of plasminogen activator, the tissue-type plasminogen activator and the urokinase-type plasminogen activator, were detected with the use of monoclonal antibodies. The PI was lower in theca externa than in theca interna-membrana granulosa of mature follicles, while the PAI was of the same level. In ruptured follicles, the PAI was lower than in mature and growing follicles; the PI showed the opposite pattern. The above findings support the suggestion that PAA plays a significant role in the process of the ovulation.

Chronic hypertension was induced in rats after partial nephrectomy. The systolic blood pressure was significantly elevated from the first week after nephrectomy to the end of the experimentation (8th week).Plasminogen activator activity (PAA) and plasminogen activator inhibition (PAI) showed a tissue- and time-dependent pattern of changes in some key organs compared to controls (sham-operated rats). Two weeks after nephrectomy (one week after the induction of hypertension) the PAA was markedly increased in lungs, heart and aorta. In aorta the PAA continued to be enhanced until the end of the experimentation (the 8th week after nephrectomy), while in heart and lungs the PAA returned to the normal eight weeks after nephrectomy. In vena cava, brain and liver no change in PAA was noticed compared to controls. Tissue PAI was mostly increased or unchanged, while tissue plasmin inhibition (PI) was unchanged. The differential response of PAA and PAI was varying not only from one organ to another or in the same organ at a given time but also in the same organ throughout experimentation. In a number of nephrectomized rats, however, hypertension was not induced. In these rats similar changes in tissue PAA and PAI were noted compared to hypertensive nephrectomized rats. Therefore, all the changes in the parameters studied should be due to the partial nephrectomy itself. In conclusion, experimentally induced chronic hypertension had not any effect on tissue PAA, PAI and PI.

In guinea pig isolated uterine strips, separate addition of adenosine, adenine and NAD was without any effect. Adenosine and NAD when administered after previously applied subthreshold dose of PGE2 exhibited a strong contractile effect. Indomethacin, added to the organ bath before addition of prostaglandin, blocked, partially or totally, the observed synergistic action of PGE2 with adenosine and NAD.From the experimental data of the present work it is postulated that adenosine and NAD stimulate prostaglandin biosynthesis in myometrium as already it has been suggested for ATP, ADP and AMP. The aspect for the existence of a functional link between adenyl compounds and prostaglandins, based on these findings, is reinforced.

The hydroxylation rate and rate of tyrosine catabolism are measured by injection of ring-deuterated L-phenylalanine and ring-deuterated L-tyrosine and subsequent deuterium determination in the water fraction of the blood. The hydroxylation rate was confirmed as the rate-limiting step. Whereas tyrosine catabolism yields normal Michaelis-Menten kinetics, homotropic activation is demonstrated for the hydroxylation step. In all tumor rats under study, the rates of phenylalanine hydroxylation and tyrosine catabolism are decreased. The delta-values to control increase with tumor age. In tumor rats, the hydroxylation step remains the rate-limiting one. Kinetic data indicate that the decrease in hydroxylation is due to a decreased turnover rate of the enzyme, whereas the increase in tyrosine catabolism is caused by the branching off of tyrosine or an intermediate on the route to D2O formation. The results are discussed with respect to altered levels of tetrahydrobiopterin, phenylalanine and tyrosine found during parallel investigations.

The levels of free phenylalanine and free tyrosine (and m-tyrosine) as well as of protein-bound tyrosine were determined by means of automated aminoacid analysis and by reaction with 1-nitrosonaphthol(2). Their absolute values as well as their percentages relative to total aminoacids of the blood were markedly increased in all tumor patients tested, as well as in experimental tumor rats. Although tetrahydrobiopterin could scarcely be detected by fluorometric methods in the blood of control persons, it is accumulated blood of all tumor patients tested. The major part was found in the erythrocyte fraction. The results are discussed with respect to decreased phenylalanine hydroxylation rate and to depressed tyrosine catabolism recently found by means of in vivo experiments in tumor-bearing rats.

The plasminogen activator activity (PAA) in extracts of the intima, media, and adventitia of the normal human aorta and other large arteries (carotid artery, renal artery and iliac artery) was studied with a sensitive, quantitative spectrophotometric assay using plasminogen and the chromogenic plasmin substrate S-2251. All layers of the arteries showed PAA which was highest in the adventitia, lowest in the media, while in the intima (aorta) PAA was intermediate, but much closer to that of the media. Plasminogen activator inhibition (PAI) was at the same level in all layers of the arteries studied. Plasmin inhibition (PI) was higher in adventitia than in intima (aorta), while in media the PI was intermediate. The PAA was due to the tissue-type plasminogen activator (t-PA), but not to the urokinase-type (u-PA), as judged by addition of respective antibodies. The relatively low PAA found in the intima of large arteries is therefore due to a low plasminogen activator and not a high plasminogen activator inhibitor activity or plasmin inhibitor level.

Summary By means of paper chromatography and the Feulgen reaction the effects of reserpine and amphetamine on the pteridine pattern and nuclear morphology were studied in the liver of Triturus cristatus . It was demonstrated that the Feulgen stainability and state of aggregation of chromatin, as well as the pteridine pattern can be altered by the influence of these drugs. From the results obtained it is suggested that the observed changes in nuclear morphology are due to the interaction of the drugs with the pteridine metabolism.