NMR structural studies of human cystatin C dimers and monomers.

Human cystatin C undergoes dimerization before unfolding. Dimerization leads to a complete loss of its activity as a cysteine proteinase inhibitor. A similar process of dimerization has been observed in cells, and may be related to the amyloid formation seen for the L68Q variant of the protein. Dimerization is barrier controlled, and no dimer/monomer interconversion can be observed at physiological conditions. As a consequence, very stable, "trapped" dimers can be easily separated from monomers. A study of the structural aspects of cystatin C dimer formation was undertaken using NMR spectroscopy. The monomer/dimer model was verified by (pulse field gradient NMR) self-diffusion molecular mass measurements. Complete backbone resonance assignments and secondary structure determination were obtained for the monomer using data from triple resonance experiments performed on 13C/15N doubly labeled protein. A marked similarity of the cystatin C secondary structure to that of chicken cystatin was observed. Using uniformly and amino-acid-specific 15N-enriched protein, backbone NH signals were assigned for cystatin C in its dimeric state. Comparison of 1H -15N correlation NMR spectra of the monomer and dimer shows that the three-dimensional structure remains unchanged in the dimer and that only local perturbations occur. These are localized to the amino acid residues comprising the cysteine proteinase binding site. Such a mode of dimerization readily explains the complete loss of the inhibitory activity in the dimer. The NMR results also demonstrate that the dimer is symmetric.

Accuracy and calibration of integrated radiation output indicators in diagnostic radiology: A report of the AAPM Imaging Physics Committee Task Group 190.

Due to the proliferation of disciplines employing fluoroscopy as their primary imaging tool and the prolonged extensive use of fluoroscopy in interventional and cardiovascular angiography procedures, "dose-area-product" (DAP) meters were installed to monitor and record the radiation dose delivered to patients. In some cases, the radiation dose or the output value is calculated, rather than measured, using the pertinent radiological parameters and geometrical information. The AAPM Task Group 190 (TG-190) was established to evaluate the accuracy of the DAP meter in 2008. Since then, the term "DAP-meter" has been revised to air kerma-area product (KAP) meter. The charge of TG 190 (Accuracy and Calibration of Integrated Radiation Output Indicators in Diagnostic Radiology) has also been realigned to investigate the "Accuracy and Calibration of Integrated Radiation Output Indicators" which is reflected in the title of the task group, to include situations where the KAP may be acquired with or without the presence of a physical "meter." To accomplish this goal, validation test protocols were developed to compare the displayed radiation output value to an external measurement. These test protocols were applied to a number of clinical systems to collect information on the accuracy of dose display values in the field.

Interactions of transforming growth factor-alpha and -beta and luteinizing hormone in the regulation of plasminogen activator activity in avian granulosa cells during follicular development.

This study examined the influence of transforming growth factor-alpha (TGF alpha), TGF beta, and LH on progesterone (P4) secretion and plasminogen activator (PA) activity in cultured avian granulosa cells from the first (F1), third (F3), and fifth and sixth (F5-6) preovulatory follicles during a 21-h incubation period. PA activity in the cell (PAc) and the medium (PAm) fractions was measured by fibrinolysis and fibrin overlay methods. P4 was determined by RIA. Basal PAc and PAm activities were highest in cell cultures from the less mature (F5-6) follicles and decreased as follicles matured to the F1 stage of development. PAc activity was greater than PAm activity regardless of the stage of follicular maturation. TGF alpha (0.1-10 ng/ml) increased PA activity in cultures of granulosa cells from F1, F3, and F5-6 follicles in a concentration-dependent manner. TGF alpha-induced PAc and PAm activities were observed by 6 and 15 h of incubation, respectively, and increased rapidly between 15-21 h. LH (100 ng/ml) attenuated TGF alpha-induced PA activity by 15 h in cultures of granulosa cells from F1 and F3, but not F5-6, follicles. Basal PA activities were unaffected by the gonadotropin. TGF beta (2-100 ng/ml) stimulated PAc activity in a dose-dependent manner only in cultures of granulosa cells from F5-6 follicles and significantly enhanced TGF alpha-induced PAc and PAm activities in cell cultures from F3 and F5-6, but not F1, follicles. Basal and growth factor-induced PAc and PAm activities corresponded to a mol wt of about 35 kDa, a value consistent with that of the low mol wt uPA species. TGF alpha and TGF beta, alone or in combination, had no effect on basal P4 secretion at all stages of follicular development. TGF alpha, however, decreased LH-induced P4 secretion in F1 and F3 cultures. These results demonstrate a tightly controlled interaction of TGF alpha, TGF beta, and LH in regulating PA activity and P4 secretion during follicular development in the domestic hen.

Influence of growth factors on the plasminogen activator activity of avian granulosa cells from follicles at different maturational stages of preovulatory development.

Granulosa cells from the first (F1), third (F3) and fifth and sixth (F5-6) preovulatory follicles and the small yellow follicles (SYFs; diameter 6-8 mm) were cultured for 21 h in the absence and presence of murine and human epidermal growth factors, fibroblast growth factor, transforming growth factors alpha and beta-I (TGF alpha, TGF beta), platelet-derived growth factor and insulin-like growth factor-I at concentrations of 0.1-100 ng/ml. Plasminogen activator (PA) activities in the cell (PAc) and in the medium (PAm) were measured by fibrinolysis and fibrin overlay methods. Basal PAc and PAm activities were highest in cell cultures from the less mature follicles (F5-6 and SYF) and decreased as the follicles matured (F3 > F1). PAc activity was greater than PAm activity, irrespective of the stage of follicular development. All growth factors examined at the 100 ng/ml level were effective in increasing PAc and PAm activities in cultures of granulosa cells from F1 follicles. However, only TGF alpha was able to increase PA activities at lower concentrations. The stimulation of the PA activities of granulosa cells from F3 follicles was inconsistent. None of the growth factors significantly increased PA activities in granulosa cells from F5-6 follicles and SYFs, as determined by fibrinolysis. The major PAc and PAm species (characterized by fibrin overlay) had a molecular mass of about 35 kDa, which is characteristic of the urokinase type. Both assay methods detected a stimulatory effect of the growth factors on PA activities in the granulosa cells from F1 follicles. However, an increase in PA activities in cells from F3 and F5-6 follicles and SYFs was indicated only after fibrin overlay analysis. Tritiated thymidine was incorporated into the DNA of granulosa cells at all stages of follicular development and was enhanced by all growth factors, although TGF alpha and TGF beta were the most effective and had a ranked order of activity: F3, F5-6 > F1, SYF. The present findings show that, of the growth factors examined, TGF alpha may be an effective regulator of PA activity in avian granulosa cells during follicular development, in addition to its observed mitogenic action.

Novel living skin replacement biotherapy approach for wounded skin tissues.

A novel living skin replacement (LSR) biotherapy concept, addressing the challenging problems related to tissue regeneration and wound healing, is presented for the treatment of skin burns, traumatic injuries and ulcerations. LSR combines elements of cell therapy along with those of tissue engineering to allow for the regeneration of wounded skin. It takes advantage of biodegradable microspheres onto which donor skin epidermal and dermal cells can be attached and expanded in vitro for subsequent direct application down to the deepest recesses of the wound bed. The key element of the biotherapy is the ability of the skin cells to migrate freely from the microspheres into the wound for regeneration of the tissues. The large surface to volume ratio of the microspheres allows for the delivery of appropriate cell numbers while minimizing the amount of biomaterial to be resorbed. This novel approach presents a number of advantages over existing therapies including facilitated cell manipulations, ease of storage and transportation, rapid clinical intervention due to the elimination of any surgical suturing or stapling, and a more natural three-dimensional tissue remodeling and anatomical compliance. Preliminary in vitro and in vivo evidence of the LSR functionality and its potential benefits is presented.

Arginine vasopressin stimulation of prostaglandin F2 alpha release in heifers during the estrous cycle.

The effect of arginine vasopressin on the stimulation of prostaglandin F2 alpha (PGF2 alpha) release has been examined in vivo. Fifty-eight heifers received one intravenous injection of 10 IU arginine vasopressin on either Day 0 (n = 14), Day 6 (n = 12), Day 13 (n = 14) and Day 18 or 19 or 20 (Day 18-20, n = 18) after the onset of oestrus (Day 0) to determine the effect of arginine vasopressin at different times of the oestrous cycle. Frequent blood samples were taken before and after arginine vasopressin injection for the measurement of 13,14-dihydro-15-keto-PGF2 alpha (PGFM) by radioimmunoassay (RIA). Blood samples for progesterone determinations were taken 2 hr before and 24 hr after arginine vasopressin to monitor luteal function. The data show that arginine vasopressin causes an increase (P less than 0.005) in PGFM concentrations only at Day 18-20 of the cycle in 67% of the experimental heifers.

Failure to maintain luteal function: a possible cause of early embryonic loss in a cow.

The effect of early pregnancy failure on the release of prostaglandin F2 alpha (PGF2 alpha) in response to oxytocin (Ot) was examined in an abnormal breeder (AB) heifer that was not able to maintain a pregnancy beyond 21 days. This animal was used in three experiments: 1) She received one intravenous injection of 100 IU Ot 17 days after the onset of oestrus (Day 0). Frequent blood samples were taken for the measurement of 15-keto-13,14-dihydro-PGF2 alpha (PGFM) by radioimmunoassay. Daily samples for progesterone (P4) determinations were taken to monitor luteal function. This was then repeated using the same animal at either day 17 or 18 or 19 (day 17-19) of pregnancy. 2) Embryos from superovulated normal breeder (NB) donors were transferred at day 7 to the AB heifer as well as to NB control animals. 3) Seven day old embryos from the superovulated AB heifer were transferred to NB recipient animals. At day 17-19 of pregnancy all the recipient heifers (experiments 2 and 3) were subjected to the same protocol as in experiment 1. The results showed that the ability of Ot to stimulate PGF2 alpha release was reduced in the NB recipients bearing viable embryos when compared to cyclic animals. However, for the AB heifer, Ot stimulated PGF2 alpha release to the same extent whether the animal was cyclic or pregnant. Furthermore, the AB animal did not have the extended luteal function associated with removal of viable embryos on day 17-19. The data suggest that the embryonic loss might have been caused by failure of the embryos to prevent the luteolytic release of PGF2 alpha.

[Prior use of a telephone-nursing triage service by patients of emergency services]. / Le recours préalable à un service de triage téléphonique infirmier chez les usagers des services d'urgence.

OBJECTIVES: A province-wide telephone-nursing triage service was implemented in Quebec, Canada, in order, among other objectives, to decrease overcrowding in hospital emergency rooms. This study analyses prior use of Info-Sante CLSC telephone service by patients of emergency services. METHODS: Structured interviews were conducted with 850 patients of a general hospital emergency room and of walk-in medical clinics. Patients were recruited while waiting to see the physician between 8h and 23 h, 7 days a week, from November 1997 to June 1998. Information was collected on knowledge and use of the telephone-nursing triage service, health problem and perceived health status, information-seeking behaviour on health services, utilisation habits, social support, and socio-demographic characteristics. Interval estimates and multiple logistic regressions were performed. RESULTS: Of the patients who were aware of this service, 17,4% (CI(95)=0,14 - 0,20) had used it prior to their medical visit. Among these, 85,1% had received a recommendation to consult a doctor. Among the users who were at the hospital emergency room at the time of the study, 56,4% were advised to consult a walk-in clinic or a CLSC, 28,2% their family doctor and only 12,8% a hospital emergency room. The probability of prior recourse to the telephone nursing service is influenced significantly by the duration of the health problem (2-4 days versus<2 days: OR(adjusted)=2,03), new health problem (OR(adjusted)=1,98) and by the frequenting of walk-in clinics rather than hospital emergency rooms (OR(adjusted)=0,31). CONCLUSION: Despite a heightened awareness of the telephone-nursing triage service, few users of emergency services make use of it and, when they do, follow rather loosely the recommendation towards the type of service judged appropriate for their needs.

Involvement of NTS2 receptors in stress-induced analgesia.

Stress activates multiple neural systems that suppress pain sensation. This adaptive phenomenon referred as stress-induced analgesia (SIA) is mediated by the activation of endogenous pain inhibitory systems. Both opioid and non-opioid forms of SIA have been elicited in rodents according to stressor parameters and duration. There is accumulating evidence that the endogenous neurotensin (NT) system plays an important role in SIA. Especially, NT-deficient mice were shown to exhibit reduced SIA following water avoidance or restraint stress. Since central NT produces naloxone-insensitive analgesic effects by acting on spinal and supraspinal NTS2 receptors, we hypothesized that NT might mediate non-opioid SIA through NTS2 activation. Here, we evaluated the influence of an opioid-independent severe stress produced by a cold-water swim for 3 min at 15 degrees C on rodent offspring's pain perception. Our results demonstrated that mice lacking NTS2 exhibit significantly reduced SIA following cold-water swim stress. Indeed, NTS2 knockout mice submitted to both acute (plantar test) and tonic (formalin test) pain stimuli show a greater sensitivity to pain in comparison to wild-type littermates. Accordingly, pretreatment with the NT receptor antagonist SR142948A results in a hyperalgesic response to stress induced by cold-water swim. Endogenous NT regulates hypothalamic-pituitary-adrenal axis activity in stress condition by increasing corticosterone plasma levels. Accordingly, the plasma levels of corticosterone measured by radioimmunoassay are significantly reduced in non-stressed and stressed NTS2-deficient mice in comparison with wild-type mice. To further investigate the site of action of NT in mediating SIA, we microinjected NTS2 agonists in lumbar spinal cord and quantified post-stress sensitivity to pain in rats using the plantar test. Exogenously administered NTS2 analogs, JMV-431, beta-lactotensin and NT69L markedly enhance the magnitude and duration of stress antinociception in both 25- and 60-day-old rats. In sum, by using genetic and pharmacological approaches, we demonstrated here that NTS2 receptors mediate non-opioid SIA. Our results also revealed that the release of endogenous NT in response to stress requires the presence of NTS2 to stimulate corticotropin-releasing factor (CRF)-induced elevation of plasma corticosterone, and that NTS2 receptors localized at the lumbar spinal cord participate to the disinhibition of descending pain control pathways. Therefore, these data highlight the significance of NTS2 as a novel target for the treatment of pain and stress-related disorders.

Altered morphine-induced analgesia in neurotensin type 1 receptor null mice.

Both neurotensin (NT) and opioid agonists have been shown to induce antinociception in rodents after central administration. Besides, previous studies have revealed the existence of functional interactions between NT and opioid systems in the regulation of pain processing. We recently demonstrated that NTS1 receptors play a key role in the mediation of the analgesic effects of NT in long-lasting pain. In the present study, we therefore investigated whether NTS1 gene deletion affected the antinociceptive action of mu opioid drugs. To this end, pain behavioral responses to formalin were determined following systemic administration of morphine in both male and female NTS1 knockout mice. Acute injection of morphine (2 or 5 mg/kg) produced strong antinociceptive effects in both male and female wild-type littermates, with no significant sex differences. On the other hand, morphine analgesia was considerably reduced in NTS1-deficient mice of both sexes compared to their respective controls, indicating that the NTS1 receptor actively participates in mu opioid alleviating pain. By examining specifically the flinching, licking and biting nociceptive behaviors, we also showed that the functional crosstalk between NTS1 and mu opioid receptors influences the supraspinally-mediated behaviors. Interestingly, sexual dimorphic action of morphine-induced pain inhibition was found in NTS1 null mice in the formalin test, suggesting that the endogenous NT system interacts differently with the opioid network in male and female mice. Altogether, these results demonstrated that NTS1 receptor activation operates downstream to the opioidergic transmission and that NTS1-selective agonists combined with morphine may act synergistically to reduce persistent pain.

Confessions of the flesh and biopedagogies: discursive constructions of obesity on Nip/Tuck.

Informed by the work of Michel Foucault, the authors discuss the Nip/Tuck episode entitled "Momma Boone" and how it discursively constructs "obesity". They show how this popular media text can be understood as a crystallisation of the dominant discourse surrounding fat bodies. In the process, how the episode can be seen as a "biopedagogy" that instructs its viewers in how to think and feel about the fat body is examined. Foucault's formulation of the confessional is seen to be useful to theorise the ways in which biopedagogy leads subjects to believe and ultimately take part in processes leading to salvation. It is argued that in this Nip/Tuck episode, biopedagogy functions in and through Momma Boone's "confessions of the flesh", that is, confessions aimed at revealing her obese body so that it can be rescued, rehabilitated and saved. Momma Boone's salvation is shown to require three stages: first, the "confession" of obesity; second, the conversion to the "truth" of the "dominant obesity discourse"; and third, the codification of a "new life" for the obese subject. In the end, it is argued that since it is represented as abject, monstrous and out of control, Momma Boone's body is made to inspire fear and panic in so far as it provides constructed "evidence" regarding the consequences of the obese subject's failure to convert to the truth of obesity discourse.

Role of arachidonic acid and its metabolites in the regulation of progesterone and oxytocin release from the bovine corpus luteum.

We have examined the effects of arachidonic acid (AA) and some of its metabolites on progesterone (P4) and oxytocin (OT) release by corpora lutea obtained from Holstein heifers at day 8 of the estrous cycle (Day 0 = estrus). The luteal cells were dispersed with collagenase and small and large cells were separated by unit gravity sedimentation and flow cytometry. After an 18-hr preincubation period, the cells were incubated in the presence of various treatments for 1 hr, followed by a 23-hr incubation period with no treatment. OT was secreted by the large, but not by the small, luteal cells into the incubation medium. AA elicited a significant (P less than 0.05) release of OT from the large cells and P4 from both the large and small cells within 1 hr of incubation, having a specific effect at a concentration of 10 microM. Larger doses (25 and 100 microM) of AA adversely affected the cell viability. Phospholipases A2 (0.5 unit/ml) and C (0.05 unit/ml) and calcium ionophore A23187 (0.1 microM) stimulated OT release from the large cells to the same extent as AA (10 microM). Inhibition of the AA cyclooxygenase metabolic pathway by indomethacin did not affect AA-induced release of OT and P4, although exogenous prostaglandins F2 alpha and I2 (5-25 ng/ml) stimulated the release of OT. Lipoxygenase products of AA (hydroxyeicosatetraenoic acid and leukotrienes; 25 ng/ml) also stimulated OT release. Inhibition of the lipoxygenase metabolic pathway by nordihydroguaiaretic acid abolished AA-induced release of both OT and P4. These results suggest that intracellular accumulation of free AA may modulate secretory functions in the bovine corpora lutea, including OT and P4 release.

Effects of progesterone and oestradiol-17 beta on oxytocin-induced release of prostaglandin F-2 alpha in heifers.

Seven bilaterally ovariectomized heifers were used in 4 experiments and received: (1) saline injections, as control; (2) one injection of oestradiol (3 mg; i.v.); (3) two i.v. injections of oxytocin (100 i.u.) 6 h apart; or (4) one oestradiol injection 30 min after the first oxytocin injection and a second oxytocin injection 6 h later. All experiments were performed without progesterone and then after 7, 14 and 21 days of progesterone treatment. Frequent blood samples were taken for 1 h before and 7 h after the first injection of oxytocin or oestradiol for the measurement of 13,14-dihydro-15-keto-PGF-2 alpha (PGFM) by radioimmunoassay. After 7, 14 and 21 days of progesterone priming, oestradiol caused a significant increase (P less than 0.001) in plasma PGFM after 6 h but not before. After 7, 14 and 21 days of progesterone, there was a significant increase (P less than 0.005) in PGFM after the first oxytocin injection and a similar increase following the second. The oxytocin-induced increase in PGFM after 14 and 21 days of progesterone was significantly higher (P less than 0.001) 6 h after oestradiol injection than before the oestradiol injection. There was no significant effect of oestradiol on the response to oxytocin in animals that received no progesterone or in those animals that received progesterone for only 7 days. These results show that, under the influence of progesterone, oestradiol enhances the oxytocin-induced release of PGF-2 alpha, and suggest a possible synergistic action of these hormones for the induction of luteolysis in heifers.

Effect of pregnancy on oxytocin-induced release of prostaglandin F2 alpha in heifers.

The effect of pregnancy on the release of prostaglandin F2 alpha (PGF2 alpha) in response to oxytocin (OT) has been examined. Fourteen cyclic heifers received one intravenous injection of 1 IU OT (n = 6) or 100 IU OT (n = 8) 17, 18, or 19 days (Day 17-19) after the onset of estrus (Day 0). Five of these animals also received 100 IU OT at Days 6 and 13 to determine the effect of OT at different times of the cycle. Frequent blood samples were taken for 60 min before and for 90 min after OT injection for the measurement of 15-keto-13,14-dihydro-PGF2 alpha (PGFM) by radioimmunoassay. The experiment was then repeated using the same animals at Day 17-19 of pregnancy (confirmed by the recovery of an embryo the day after OT injection). Following the injection of 1 IU OT, plasma PGFM reached its peak within 30 min with the increase significantly lower (P less than 0.05) in pregnant (1.13 +/- 0.10-fold) than in nonpregnant animals (2.07 +/- 0.27-fold). However, because only 3 of the 6 cyclic animals showed a response to 1 IU OT, the dose was increased to 100 IU in subsequent experiments. The animals that received 100 IU at Days 6 and 13 had no significant increase in PGFM concentrations (1.18 +/- 0.05-fold and 1.01 +/- 0.04-fold, respectively). At Day 17-19 the increase in plasma PGFM reached its peak 5-15 min after 100 IU OT and the increase was significantly greater in nonpregnant (3.23 +/- 0.17-fold) than in pregnant (1.21 +/- 0.02-fold; P = 0.003) heifers. Six of 11 animals injected at Day 17-19 of the cycle showed a decrease in progesterone (P4) the day after OT administration. These data show that the release of PGF2 alpha in response to OT is suppressed in pregnant animals in vivo, suggesting an antiluteolytic role for the embryo in luteostasis.

Control of bovine uterine prostaglandin F2 alpha release in vitro.

Prostaglandin F2 alpha (PGF2 alpha) release from the uterus causes luteolysis in ruminants, and oxytocin is thought to be a regulator of this release. In the present study, we have examined the mechanisms involved in oxytocin stimulation of PGF2 alpha secretion by bovine endometrium in vitro. Endometrial tissue explants, obtained from heifers at Day 19 or 20 (n = 3) and Day 0 (estrus, n = 5) of the estrous cycle, were incubated for 2 h and 6 h, and PGF2 alpha concentration in the medium was determined by radioimmunoassay (RIA). Basal PGF2 alpha release increased for up to 6 h and was significantly stimulated after 2 h of incubation with 100 microU and 1000 microU of oxytocin at Day 0 but not at Day 19 or 20. Secretion of PGF2 alpha was not affected by cholera toxin (10 ng/ml) or the cyclic nucleotide analogs dibutyryl cyclic adenosine 3',5'-monophosphate and dibutyryl cyclic guanosine 3',5'-monophosphate at a concentration of 1 mM. A protein kinase A inhibitor (500 microM) had no effect on the oxytocin-induced release of PGF2 alpha. Both the phorbol ester, 12-myristate-13-acetate (100 mM), and the non-phorbol stimulator of protein kinase C, 1-octanoyl-2-acetylglycerol (500 microM), significantly stimulated PGF2 alpha secretion to the same extent as oxytocin. Neither basal nor stimulated PGF2 alpha release was affected by the calcium ionophore A23187 (0.1-5.0 microM). However, PGF2 alpha secretion was sensitive to cycloheximide (1 microgram/ml) suggesting that protein synthesis may be involved. In conclusion, these data suggest that the stimulation of PGF2 alpha by oxytocin is via the protein kinase C effector pathway.

On the interdependency between attentional biases and visual field effects with bilateral presentations of stimuli.

Previous research indicates that bilateral presentations of visual stimuli yield greater visual field differences than unilateral ones. Although this fact has generally been interpreted by reference to functional models of hemispheric specialization, an alternative explanation has been offered, which suggests that it is artifactually caused by an attentional bias (AB). Indeed, consistent positive correlations between the direction of ABs and that of visual field effects have been reported. However, this relationship remains uncertain since different tasks, presented at different moments, have been used in order to document it. We report on a bilateral presentation task which controlled this potential discrepancy by mixing trials measuring ABs with those assessing visual field effects. We showed that, in a group of dextrals, the direction of the visual field effect is intimately correlated with that of the AB; subjects with a right AB showed a right visual field advantage, whereas subjects with a left AB showed a left visual field advantage; moreover, subjects with no overall AB did not show any visual field advantage. The results of this experiment confirm the fact that the formally reported correlations between the direction of ABs and visual field effects are determined by individual differences in ABs rather than by the task being used to assess them. This observation points to the necessity for further research on variables affecting the between-subjects variability in ABs with the use of bilateral presentations of visual stimuli.

Avian granulosa cell prostaglandin secretion is regulated by transforming growth factor alpha and beta and does not control plasminogen activator activity during follicular development.

The aim of the present study was to determine the role of transforming factor alpha (TGF alpha) and beta (TGF beta) in the regulation of prostaglandin (PG) secretion, and the relationships between PG and plasminogen activator (PA) activity in hen granulosa cells during ovarian follicular development. Cells from the first (F1), third (F3), and fifth and sixth (F5-6) largest preovulatory follicles were cultured for up to 21 h in the presence of TGF alpha (0.1-10 ng/ml) and/or TGF beta (4-20 ng/ml) or TGF alpha together with a cyclooxygenase inhibitor, indomethacin (0.05-0.5 microM). The release of PG into the incubation medium was determined by RIA. Cell-associated (PAc) and secreted (PAs) PA activities were measured by a fibrinolysis assay and characterized by zymography. Basal PGF secretion from F1, F3, and F5-6 cells was 2.2 +/- 0.3, 2.2 +/- 0.5, and 1.1 +/- 0.3 ng/micrograms DNA, respectively, and was higher than that of PGE. Basal total PA (PAc+PAs) activity from F1, F3, and F5-6 cells was 41 +/- 13,261 +/- 68, and 958 +/- 268 x 10(3) cpm/micrograms DNA, respectively. TGF alpha stimulated PG secretion and PA activity in a dose-dependent manner. The TGF alpha-induced PA activity was predominantly associated with a molecular mass of 30-35 kDa, corresponding to that of urokinase PA. The stimulation of PG secretion by TGF alpha was maximal in F3 and F1 granulosa cells whereas PA activity in the presence of TGF alpha was highest in cells from F5-6 follicles.(ABSTRACT TRUNCATED AT 250 WORDS)