RESUMEN
The process of embryo implantation requires extensive remodelling of the endometrial extracellular matrix, a function largely performed by matrix-degrading metalloproteinases (MMPs). In the present study, we used trophoblast cells isolated from human term placentas to study the regulation of MMPs by nitric oxide (NO). Using a combination of zymography, Western blot and indirect immunofluorescence, we showed that MMP-2 and MMP-9 are increased during the conversion from low-motile cytotrophoblast cells to the highly motile and differentiated syncytiotrophoblast multinucleated cells. We also observed an increase in NO production and NO synthase (NOS) expression during this cellular differentiation process. In addition, we demonstrated a positive regulatory role of NO on the activity and protein expression of MMP-2 and MMP-9, because NO donors (NOC-18 and spermine-NONOate) or the NOS substrate (L-arginine) stimulate, whereas NOS inhibitors (N(G)-nitro-L-arginine methyl ester and N(G)-monomethyl-L-arginine) reduce the expression and gelatinolytic activity of MMP-2 and MMP-9 in isolated trophoblast cells. Taken together, these results suggest that, in differentiating trophoblasts, NO regulates the induction of matrix-degrading proteases required for invasion during embryo implantation.
Asunto(s)
Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Óxido Nítrico/farmacología , Trofoblastos/enzimología , Arginina/farmacología , Western Blotting , Diferenciación Celular , Movimiento Celular , Células Cultivadas , Implantación del Embrión , Inhibidores Enzimáticos/farmacología , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , NG-Nitroarginina Metil Éster/farmacología , Donantes de Óxido Nítrico/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Nitroarginina/farmacología , Embarazo , Trofoblastos/citologíaRESUMEN
Previous work described an increase in prostaglandin E (PGE) production by oocyte-cumulus complexes (OVA) obtained from non-insulin-dependent diabetic rats. More recently, it has been found that in control OVA nitric oxide (NO) mediates hCG-induced PGE secretion. To determine whether increases in PGE secretion by diabetic OVA are mediated by NO, the present study has evaluated the secretion of PGE by diabetic OVA, cultured in the absence or presence of hCG, NO donors (sodium nitroprusside (NP) and 3-morpholino-sydnonimine-hydrochloride (SIN-1)), and a NO synthase inhibitor (N(G)monomethyl-L-arginine; L-NMMA). hCG, NP and SIN-1 increased PGE secretion by diabetic OVA. L-NMMA did not modify basal secretion of PGE by control OVA but lowered PGE production in diabetic OVA to control values. L-NMMA prevented the hCG-induced PGE accumulation in control and diabetic OVA, and the quantities of PGE produced were similar to those of control OVA but lower than in diabetic OVA incubated in the absence of hCG. The effect of L-NMMA seems to be specific since N(G)monomethyl-D-arginine had no effect. NO synthase activity was higher in diabetic ovaries than in controls. The present results suggest that NO mediates the increased PGE production by diabetic OVA, probably a result of overproduction of NO.
Asunto(s)
Diabetes Mellitus Tipo 2/enzimología , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico/metabolismo , Oocitos/metabolismo , Prostaglandinas E/biosíntesis , Animales , Gonadotropina Coriónica/farmacología , Diabetes Mellitus Tipo 2/metabolismo , Inhibidores Enzimáticos/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo III , Oocitos/efectos de los fármacos , Ratas , Ratas Wistar , omega-N-Metilarginina/farmacologíaRESUMEN
Embryonic development, prostaglandin E (PGE) generation and nitric oxide synthase (NOS) activity during organogenesis were evaluated in an experimental rat model of non-insulin-dependent diabetes (NIDD) generated by neonatal administration of streptozotocin. Gross malformations were detected in 5% of NIDD embryos and these embryos were all non-viable; in the other 95%, growth was retarded but no congenital abnormalities were found. Control embryos were all alive and not malformed. The NIDD 11-day embryos secreted more PGE into the incubation medium than did controls. The NO donor SIN-1 increased PGE production in both control and NIDD embryos. A NOS inhibitor (L-NMMA) reduced PGE generation in both experimental groups, suggesting a modulatory role of NO on embryonic PGE production. Activity of NOS was higher in NIDD 11-day embryos than in controls. Treatment in vivo of control and NIDD rats (Days 7-11 of gestation) with a NOS inhibitor (L-NAME; 5 mg kg(-1) i.p.) reduced embryonic PGE production and induced a higher resorption rate and an increase in neural-tube defects. The results suggest that NO modulates PGE generation in the organogenetic embryo. In the NIDD model, overproduction of NO is observed, this NO probably enhancing embryonic PGE production. The relationship between PGE generation and the appearance of congenital abnormalities is discussed.
Asunto(s)
Diabetes Mellitus Tipo 2/enzimología , Desarrollo Embrionario y Fetal , Óxido Nítrico Sintasa/metabolismo , Embarazo en Diabéticas/enzimología , Prostaglandinas E/biosíntesis , Animales , Inhibidores Enzimáticos/farmacología , Femenino , Masculino , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo III , Embarazo , Ratas , Ratas WistarRESUMEN
Severe reproductive dysfunction has been described in non-insulin-dependent diabetes mellitus (NIDDM), correlated with high glucose levels in the plasma. We have characterized an abnormal prostanoid profile in tissues from NIDDM rats, and a tight correlation between nitric oxide (NO) levels and prostaglandin production. Likewise, we have determined that parturition is delayed in NIDDM rats compared to control animals. In order to characterize the events which precede delayed parturition in NIDDM rats, we evaluate (a) the arachidonic acid (AA) conversion in placental tissue obtained from control (day 21 and 22) and NIDDM (day 21, 22 and 23) late pregnant rats into prostaglandin E2 (PGE2) and F2alpha (PGF2alpha), thromboxane B2 (TXB2) and 6-keto-prostaglandin F1alpha (6-keto-PGF1alpha), and (b) NO synthase (NOS) activity in control and NIDDM late pregnant animals. Placental arachidonate conversion from control rats into different prostanoids, namely PGE2, PGF2alpha, and TXB2, is higher in day 22 than in day 21, and radioconversion from diabetic rats into PGE2, PGF22, TXB2 and 6-keto-PGF1alpha on day 23 is higher than in day 21 and 22. 6-keto-PGF1alpha is lower and TXB2 is higher in diabetic tissues than in control. Placental AA conversion of control diabetic tissues on the day of delivery is decreased by N(G) monomethyl-L-arginine (LNMMA) (600 mM), a well known NOS inhibitor, while prostanoid production remains unaltered on previous days. NOS activity is higher in control on day 22 when compared to day 21, and in diabetic on day 23 when compared to day 22 of pregnancy. We conclude that elevated NO placental levels are observed in control (day 22) and NIDDM (day 23) rats, and may increase placental prostaglandin production on the day of delivery.
Asunto(s)
Diabetes Mellitus Tipo 2/metabolismo , Óxido Nítrico/metabolismo , Placenta/metabolismo , Embarazo en Diabéticas/metabolismo , Prostaglandinas/biosíntesis , 6-Cetoprostaglandina F1 alfa/metabolismo , Animales , Ácido Araquidónico/metabolismo , Dinoprost/metabolismo , Dinoprostona/metabolismo , Inhibidores Enzimáticos/farmacología , Femenino , Edad Gestacional , Masculino , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Embarazo , Ratas , Ratas Wistar , Valores de Referencia , Tromboxano B2/metabolismo , omega-N-Metilarginina/farmacologíaRESUMEN
Severe uterine and placental disturbances have been described in diabetes pathology. The relative severity of these changes appears to correlate with high glucose levels in the plasma and incubating environment. In order to characterize changes in eicosanoid production we compared uterine and placental arachidonic acid conversion from control and non-insulin-dependent diabetes mellitus (NIDDM) rats on day 21 of pregnancy, into different prostanoids, namely PGE2, PGF22alpha, TXB2 (indicating the production of TXA2) and 6-keto-PGF1 (indicating the generation of PGI2). PGE2, PGF2alpha and TXB2 production was higher and 6-keto-PGF1alpha was similar in diabetic compared to control uteri. PLA2 activity was found diminished in the NIDDM uteri in comparison to control. A role for PLA2 diminution as a protective mechanism to avoid prostaglandin overproduction in uterine tissue from NIDDM rats is discussed. Placental tissues showed an increment in TXB2 generation and a decrease in 6-keto PGF1alpha level in diabetic rats when compared to control animals. Moreover, when control uterine tissue was incubated in the presence of elevated glucose concentrations (22 mM), similar generation of 6-keto PGF1alpha and elevated production of PGE2, PGF2alpha and TXB2 were found when compared to those incubated with glucose 11 mM. Placental TXB2 production was higher and 6-keto PGF1alpha was lower when control tissues were incubated in the presence of high glucose concentrations. However, high glucose was unable to modify uterine or placental prostanoid production in diabetic rats. We conclude that elevated glucose levels induced an abnormal prostanoid profile in control uteri and placenta, similar to those observed in non-insulin-dependent diabetic tissues.
Asunto(s)
Glucemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eicosanoides/biosíntesis , Placenta/metabolismo , Embarazo en Diabéticas/metabolismo , Preñez , Útero/metabolismo , 6-Cetoprostaglandina F1 alfa/metabolismo , Animales , Ácidos Araquidónicos/metabolismo , Dinoprost/metabolismo , Dinoprostona/metabolismo , Femenino , Intercambio Materno-Fetal , Embarazo , Prostaglandinas A/metabolismo , Ratas , Ratas Wistar , Tromboxano A2/metabolismo , Tromboxano B2/metabolismoRESUMEN
In the process of embryo implantation in the rat, both nitric oxide and prostaglandins act as vascular and myometrial regulators. The aim of the present work was to evaluate the effect of diabetes on the synthesis of both agents during embryo implantation. In diabetic rats, uterine activity of the enzyme nitric oxide synthase and prostaglandin E production were increased during peri-implantation compared to the control group (P < 0.05 and P < 0.001, respectively). Both parameters showed a prolonged increase in temporal profile during peri-implantation days. Local production of nitric oxide and prostaglandin E in the implantation sites was higher in diabetic rats (P < 0.05), but the intersite:site ratio was similar to that of the control group. On the other hand, the implantation rate and the timing of the beginning of this process were not altered in the diabetic group. These results suggest that the vasoactive modulators of the implantation process, nitric oxide and prostaglandins, are increased in this diabetic pathology, and that this increase is probably functioning as a compensatory mechanism, so as to allow an unaltered rate of embryo implantation in this model.
Asunto(s)
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Implantación del Embrión , Óxido Nítrico/biosíntesis , Prostaglandinas E/biosíntesis , Útero/metabolismo , Animales , Femenino , Embarazo , Embarazo en Diabéticas/metabolismo , Ratas , Ratas WistarRESUMEN
Eicosanoid production by intrauterine tissues from control and neonatal-streptozotocin induced diabetic rats during late pregnancy was evaluated. In diabetic placenta the release of 6-keto-PGF1alpha was found diminished when compared to controls. In addition, LTB4 generation was increased in diabetic placenta. No alterations in the production of TXA2, PGE2, PGE1 and PGF2alpha was found when diabetic and control placenta were compared. In amnion tissue a decreased generation of 6-keto-PGF1alpha was observed in the diabetic group, but no alteration in any other eicosanoid evaluated was found. Oxytocin (5 mU/ml, in vitro), which increases prostaglandin synthesis in rabbit and human amnion tissues, did not modify eicosanoid generation in control rat amnion. In contrast, in diabetic amnion the presence of oxytocin further decreased the release of 6-keto-PGF1alpha and diminished PGE1 generation. The present results suggest that this mildly diabetic state induces alterations in eicosanoid production in intrauterine tissues, abnormalities probably enhanced during parturition, when endogenous concentrations of oxytocin are elevated.
Asunto(s)
Amnios/efectos de los fármacos , Amnios/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eicosanoides/biosíntesis , Oxitocina/farmacología , Placenta/efectos de los fármacos , Placenta/metabolismo , Embarazo en Diabéticas/metabolismo , 6-Cetoprostaglandina F1 alfa/biosíntesis , Alprostadil/biosíntesis , Animales , Medios de Cultivo , Diabetes Mellitus Experimental/metabolismo , Dinoprost/biosíntesis , Dinoprostona/biosíntesis , Femenino , Humanos , Técnicas In Vitro , Embarazo , Conejos , Ratas , Ratas Wistar , Tromboxano A2/biosíntesisRESUMEN
In vivo administration of HOE 140 (a new bradykinin receptor antagonist) and L-NAME (nitric oxide synthase inhibitor) was performed in chronic streptozotocin-diabetic rats. Basal increases (in umol.g dw-1) in liver (45.0 +/- 3.4.1) and uterine (40.0 +/- 2.95) triglyceride levels in diabetic animals vs control (liver: 34.0 +/- 3.87; uterus: 30.2 +/- 4.01) were partially prevented by L-NAME (p < 0.01), HOE 140 (p < 0.01) and L-NAME + HOE 140 (p < 0.01). High glycogen levels (in mg.g dw-1) observed in diabetic uterine tissue (3.07 +/- 0.90), and decreased glycogen content detected in diabetic liver (11.64 +/- 1.50) vs. control (uterus: 1.59 +/- 0.15, liver: 17.25 +/- 0.87) were unaffected. Uterine 14CO2 production from 14C-U-Glucose (in uCi.mg dw), which is lower in diabetic (35.0 +/- 5.12) than in control (50.12 +/- 4.54) tissues, was improved by HOE 140 (p < 0.05) and L-NAME+HOE 140 (p < 0.05), while hepatic glucose oxidation was not increased by the drugs. Glycemia levels were decreased in diabetic rats injected with L-NAME and L-NAME plus HOE 140. Pancreatic 6-Keto-prostaglandin F1 alpha to Thromboxane B2 ratio was lower in diabetic animals than in controls, and L-NAME and/or HOE 140 treatment prevented the decrement. These findings suggest that vasoactive compounds might prevent streptozotocin-induced damage in pancreatic tissue from chronic diabetic rats.
Asunto(s)
Bradiquinina/análogos & derivados , Diabetes Mellitus Experimental/metabolismo , NG-Nitroarginina Metil Éster/farmacología , 6-Cetoprostaglandina F1 alfa/metabolismo , Animales , Glucemia/metabolismo , Bradiquinina/farmacología , Antagonistas de los Receptores de Bradiquinina , Dióxido de Carbono/metabolismo , Femenino , Glucosa/metabolismo , Glucógeno/metabolismo , Hígado/metabolismo , Nitratos/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Nitritos/metabolismo , Páncreas/metabolismo , Ratas , Ratas Wistar , Tromboxano B2/metabolismo , Triglicéridos/metabolismo , Útero/metabolismoRESUMEN
To determine whether nitric oxide (NO) generation mediates human chorionic gonadotrophin (hCG)-induced prostaglandin E (PGE) secretion by oocyte-cumulus complexes (OCC), the secretion of PGE by cultured rat OCC in the presence of NO donors and NO synthase (NOS) inhibitors was characterized. NO donors (sodium nitroprusside and 3-morpholino-sydnonimine-hydrochloride) increased PGE accumulation in OCC to values similar to those obtained in the presence of hCG. The three NOS inhibitors tested (NG-nitro-L-arginine methyl ester, NG-monomethyl-L-arginine and aminoguanidine) prevented the hCG-induced PGE accumulation in cultured OCC. This effect appears to be specific since D-enantiomers NG-nitro-D-arginine methyl ester and NG-monomethyl-D-arginine had no effect. The present results suggest that NO mediates the hCG-induced accumulation of PGE in rat OCC, a process which may occur in vivo in preovulatory follicles prior to ovulation.
Asunto(s)
Gonadotropina Coriónica/farmacología , Hormona Luteinizante/farmacología , Óxido Nítrico/fisiología , Oocitos/efectos de los fármacos , Ovario/efectos de los fármacos , Prostaglandinas E/biosíntesis , Animales , Células Cultivadas , Inhibidores Enzimáticos/farmacología , Femenino , Humanos , Óxido Nítrico Sintasa/antagonistas & inhibidores , Oocitos/metabolismo , Ovario/citología , Ovario/metabolismo , Ratas , Ratas WistarRESUMEN
It has previously been demonstrated that uterine nitric oxide synthase (NOS) activity increases before embryonic implantation in rats. The aim of the present work was to investigate the regulation and the physiological relevance of the nitric oxide (NO) system in ovoimplantation. The increase in NOS activity in early pregnancy was found to be independent of the presence of embryos in the uterus. Whereas the Ca2+-dependent isoform of NOS increased gradually in the preimplantation days, the Ca2+-independent isoform increased just at the beginning of implantation (Day 5, 1800 hours); then the activity of both isoforms declined. Oestradiol, whose concentration peaks before implantation, might be regulating NOS activity in the uterus, since treatment of rats with tamoxifen, a receptor antagonist, reduces the activity of both isoforms to preimplantation levels. Intraluminal injections of L-NAME (0.5 mg kg[-1]), a competitive inhibitor of NOS, reduced by 50% the number of implanted embryos; this suggests that the NO system plays a role during implantation. The data suggest that oestradiol might be a modulator of NOS activity during nidation and that NO production is necessary to achieve a successful embryo implantation.
Asunto(s)
Implantación del Embrión/fisiología , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico/fisiología , Útero/enzimología , Animales , Calcio/farmacología , Implantación del Embrión/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Antagonistas de Estrógenos/farmacología , Femenino , Isoenzimas/antagonistas & inhibidores , Isoenzimas/química , Isoenzimas/metabolismo , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/química , Embarazo , Ratas , Ratas Wistar , Tamoxifeno/farmacología , Factores de Tiempo , Útero/efectos de los fármacos , Útero/metabolismo , Útero/patologíaRESUMEN
In spite of the large quantities of epoxyeicosatrienoic acids (EEts) released by reproductive tissues, their function has not yet been determined. In order to analyze the influence of epoxygenase products on isolated uterine function, Clotrimazole, a cytochrome P450 inhibitor was used. The drug decreased isolated rat uterine isometric developed tension (IDT) and frequency (FC). 14,15 EEt induced a contractile response when added at 10(11) M, 8,9 EEt and 11,12 EEt produced an increment of IDT when added to 10(-7) M and 5,6 EEt did not modify IDT values. A contractile stimulatory effect was observed when 14,15 EEt (10(-7) M) was added to a tissue bath preparation containing Clotrimazole (20 microM). On the other hand, uterine contractile response to 14,15 EEt addition was partially abolished by indomethacin (10(-6) M), a well known cyclooxygenase inhibitor. Uterine response to 5,6; 8,9 and 11,12 EEts was not modified by indomethacin. This is the first evidence of 14-15 EEt uterotonic properties, possibly exerted in part through the cyclooxygenase pathway.
Asunto(s)
Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido 8,11,14-Eicosatrienoico/farmacología , Contracción Uterina/efectos de los fármacos , Animales , Clotrimazol/farmacología , Inhibidores de la Ciclooxigenasa/farmacología , Inhibidores Enzimáticos del Citocromo P-450 , Relación Dosis-Respuesta a Droga , Estradiol/farmacología , Femenino , Técnicas In Vitro , Indometacina/farmacología , Contracción Isométrica/efectos de los fármacos , Ovariectomía , Prostaglandina-Endoperóxido Sintasas/metabolismo , Ratas , Ratas WistarRESUMEN
Ovulation, oocyte maturation and PGE and PGF2 alpha production by oocyte-cumulus complexes were evaluated in rats with non-insulin-dependent diabetes induced by neonatal streptozotocin. Diabetic rats had normal estrous cycles, but ovulated a lower number of oocytes at estrus. When oocytes from control and diabetic rats obtained at proestrus were matured "in vitro" during 1, 2 or 4 hours (hr) of culture, differences were not found in the percent of germinal vesicle breakdown between both experimental groups. PGE and PGF2 alpha accumulation was higher in ovulated oocyte-cumulus complexes when compared to immature or "in vitro"-matured oocyte-cumulus complexes in both normal and diabetic rats. When control and diabetic rats are compared, more PGE and PGF2 alpha accumulation was observed in immature, "in vitro"-matured and in ovulated oocyte-cumulus complexes. A lower number of oocytes ovulated and increased oocyte-cumulus complexes prostaglandin production has been observed in this mildly diabetic experimental model. These abnormalities are similar to those previously found when 10 day embryos were evaluated in non-insulin-dependent diabetic rats.
Asunto(s)
Diabetes Mellitus Tipo 2/metabolismo , Dinoprost/biosíntesis , Oocitos/fisiología , Prostaglandinas E/biosíntesis , Animales , Modelos Animales de Enfermedad , Estro/metabolismo , Femenino , Meiosis , Ratas , Ratas WistarRESUMEN
Embryonic implantation is a complex process in which both maternal and embryonic signals are involved. In the present study, we evaluated changes in uterine prostaglandins production and nitric oxide synthase (NOS) activity during the course of early pregnancy and their interaction during implantation in rats. Uterine phospholipase A2 (PLA2) activity is increased on days 5 (day of ovoimplantation) and 6, compared to preimplantation days (3 and 4). This enhanced activity might be responsible for the observed increase in uterine PGE and PGF2 alpha production observed on day 5 of pregnancy, which induces endometrial vascular permeability and decidualization. When embryo access to the uterus is impaired, the increase of PG production is suppressed. During postimplantation, PGE levels return to preimplantation values, while PGF2 alpha decreased with respect to preimplantation values. Uterine NOS activity is also increased on day 4 and reaches a maximum on day 5, with a profile similar to PGE and PGF2 alpha. Dexamethasone administered in vivo decreased uterine NOS activity on day 4 of pregnancy but not on day 5, suggesting the presence of at least two types of NOS enzymes in the early days of pregnancy. A competitive inhibitor of NOS, L-NAME (600 and 1000 microM) induced a decrease in PGE and PGF2 alpha production in uterine tissue on day 5 of pregnancy. These results suggest the existence of a physiologically relevant nitridergic system which modulates prostaglandin production in the rat uterus during embryonic implantation.
Asunto(s)
Dinoprost/metabolismo , Implantación del Embrión/fisiología , Fosfolipasas A/metabolismo , Prostaglandinas E/metabolismo , Útero/metabolismo , Administración Tópica , Animales , Antiinflamatorios/farmacología , Dexametasona/farmacología , Eicosanoides/biosíntesis , Desarrollo Embrionario/fisiología , Femenino , Glucocorticoides , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Fosfolipasas A2 , Embarazo , Prostaglandinas/biosíntesis , Ratas , Ratas Wistar , Útero/efectos de los fármacosRESUMEN
Eicosanoid production, glucose (Glu), glycogen (Gly) and triglyceride (TG) metabolism, spontaneous contractile activity, PGF2 alpha and oxytocin-induced contractions have been studied in uterine tissue obtained from control (C) and non-insulin-dependent diabetic (D) rats prior to parturition. Parturition occurs on day 22 of gestation in control animals, whereas a 24 hr delay was observed in diabetic rats. Production of PGE2, PGE1, 6-keto-PGF1 alpha, PGF2 alpha, TXB2 and LTB4 was similar in uterine tissue obtained from control and diabetic rats on day 21 of pregnancy. Uterine metabolism, on day 21 of pregnancy, based on the production of 14CO2 from U14C-glucose was lower in tissues obtained from diabetic rats than in controls. Levels of TG were similar at 0 hr and after 60 min incubation in Glu or Glu-free medium in both experimental groups. Initially Gly levels in diabetic and control uteri were similar. After 60 minutes of incubation, levels of Gly in control tissue decreased only in the absence of Glu in the incubation medium. In contrast, in diabetic uterine strips, levels of Gly decreased after 60 minutes of incubation either in Glu or Glu-free medium. "In vitro" isometric-developed tension (IDT) evaluated on day 21 (C and D) and 22 (D) of pregnancy was similar at 0 hr in control and diabetic uterine preparations, but IDT in both diabetic groups was decreased after a 40 minute incubation when compared to controls. Alterations in PGF2 alpha-induced uterine responses were not seen in 21 or 22 days pregnant diabetic uterine tissue when compared to controls. In contrast, impaired oxytocin responses were observed in diabetic uteri on day 21 of gestation, but they were similar to control responses of uterine tissue from day 22 diabetic rats. We conclude that in the non-insulin-dependent late pregnant rat, there are no alterations in uterine tissue eicosanoid production, but metabolic and contractile abnormalities are present. Involvement of these alterations in the delayed initiation of parturition is discussed.
Asunto(s)
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eicosanoides/metabolismo , Contracción Uterina , Útero/metabolismo , Animales , Dinoprost/farmacología , Relación Dosis-Respuesta a Droga , Femenino , Glucosa/metabolismo , Glucógeno/análisis , Técnicas In Vitro , Contracción Isométrica , Oxitocina/farmacología , Embarazo , Ratas , Ratas Wistar , Estreptozocina , Factores de Tiempo , Triglicéridos/análisis , Útero/química , Útero/efectos de los fármacosRESUMEN
3-O-14C-methyl-D-Glucose (3-O-MG) transport and 14C-saccharose incorporation were measured in isolated uterine strips from ovariectomized-estrogenized diabetic rats. Glucose transport was decreased in uterine strips from diabetic rats compared with control animals. PGE1 and PGE2 (10(-7) M) stimulated 3-O-MG transport, PGF2 alpha failed to modify this parameter at the same concentration, while insulin (0.5 U/ml) evoked an improvement 30% greater than PGs. In spite of the negative influence exerted by TXA2 over glucose metabolism in the isolated rat uterus, U46619, 10(-5) M (a TXA2 analogue), and OKY064, 10(-7) M (an inhibitor of TXA2 synthesis), failed to modify basal or insulin-treated hexose transport. Neither additive or synergistic interactions between PGE1 or PGE2 (10(-7) M) and insulin at 0.5 U/ml and 0.25 U/ml were detected. We conclude that the stimulatory action of PGE1 and PGE2 on glucose metabolism that has been previously described by us, involves enhancement of glucose transport.
Asunto(s)
3-O-Metilglucosa/metabolismo , Alprostadil/farmacología , Diabetes Mellitus Experimental/metabolismo , Dinoprostona/farmacología , Útero/efectos de los fármacos , Animales , Transporte Biológico/efectos de los fármacos , Dinoprost/farmacología , Interacciones Farmacológicas , Femenino , Técnicas In Vitro , Insulina/farmacología , Ovariectomía , Ratas , Ratas Wistar , Estreptozocina , Sacarosa/metabolismo , Tromboxano B2/antagonistas & inhibidores , Regulación hacia Arriba , Útero/metabolismoRESUMEN
In uterine tissue obtained from castrated control and non-insulin dependent diabetic (NIDDM) rats, eicosanoid production and its regulation by glucose levels and by the activity of phospholipase A2 (PLA2) was assessed. Basal outputs of prostaglandins (PGs) PGE2, PGE1, PGF2 alpha, 6-keto-PGF1 alpha (indicating the production of prostacyclin), thromboxane B2 (TXB2) (indicating the generation of TXA2) and leukotriene B4 (LTB4) were similar in control and NIDDM uterine preparations as assessed by RIA. When uterine conversion of labelled arachidonate into different prostanoids was evaluated, generation of 6-keto-PGF1 alpha, PGE2 and PGF2 alpha was similar in control and NIDDM uterine tissue, while TXB2 production was higher in the diabetic group. Moreover, when control tissue was incubated in the presence of elevated concentrations of glucose (22 mM) and compared to control tissue incubated in concentrations of glucose 11 mM, similar generation of 6-keto-PGF1 alpha, PGE2 and PGF2 alpha was observed, and higher concentrations of TXB2 were found, similar to those observed in diabetic uterine tissue. When NIDDM uterine tissue was incubated in the presence of glucose 22 mM, no difference in any prostanoid evaluated was observed when compared to values obtained in the presence of glucose 11 mM. In this work we have observed in NIDDM uterine tissue a normal TXA2 production when evaluated by RIA from endogenous arachidonic acid (AA) and a higher TXA2 generation from exogenous labelled AA. In addition PLA2 activity was found diminished in the NIDDM uteri in comparison to control uteri. A role of the diminished PLA2 as a protective mechanism that avoids TXA2 overproduction in uterine tissue from NIDDM rats is discussed.
Asunto(s)
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eicosanoides/biosíntesis , Ovariectomía , Fosfolipasas A/metabolismo , Útero/metabolismo , 6-Cetoprostaglandina F1 alfa/biosíntesis , Alprostadil/biosíntesis , Animales , Ácido Araquidónico/metabolismo , Dinoprost/biosíntesis , Dinoprostona/biosíntesis , Femenino , Leucotrieno B4/biosíntesis , Fosfolipasas A2 , Ratas , Ratas Wistar , Tromboxano B2/biosíntesisRESUMEN
The influence of nitric oxide (NO) on the production of 14CO2 from labeled glucose in uteri isolated from ovariectomized-estrogenized rats was studied. Nitroprusside, an NO donor (NP), 200 microM increased the formation of labeled CO2 from [U-14C]glucose. This effect was blunted by hemoglobin (Hb) 20 micrograms/mL, an NO scavenger. The addition of N-monomethyl arginine (NMMA), an inhibitor of NO synthase decreased the stimulatory action of NP at 400 mM. Incubation of uterine strips in the presence of NP plus acetylsalicylic acid (ASA) 10(-4) M (a cyclooxygenase inhibitor), inhibited the stimulatory action of NP on glucose metabolism. PGE2 (10(-7) M) added to the incubation medium containing NP and ASA reversed the effect of the inhibitor. Neither NP nor Hb nor NMMA modified the 14CO2 production from labeled glucose in uterine strips from ovariectomized rats. The addition of NP to the incubating medium increased PGE accumulation by uterine strips from rats treated with estradiol, but not in ovariectomized animals. These results suggest that NO exerts a positive influence on glucose metabolism and PGE synthesis in isolated rat uteri from estrogenized animals.
Asunto(s)
Estradiol/farmacología , Glucosa/metabolismo , Óxido Nítrico/fisiología , Útero/metabolismo , Animales , Arginina/análogos & derivados , Arginina/farmacología , Dióxido de Carbono/metabolismo , Radioisótopos de Carbono , Inhibidores Enzimáticos/farmacología , Femenino , Hemoglobinas/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Nitroprusiato/farmacología , Ovariectomía , Ratas , Ratas Wistar , Útero/efectos de los fármacos , omega-N-MetilargininaRESUMEN
The relationship between high glucose concentrations and arachidonic acid metabolism in uterine tissue from control and diabetic ovariectomized rats was evaluated. Uterine tissue from diabetic rats produced amounts of PGE2 and PGF2 alpha similar to controls, while a lower production of 6-keto-PGF1 alpha (indicating the production of prostacyclin) and a higher production of TXB2 (indicating the generation of TXA2) was found in the diabetic group. A group of diabetic rats was treated with phlorizin to diminish plasma glucose levels. Phlorizin treatment did not alter production of PGE2, PGF2 alpha, and 6-keto-PGF1 alpha in the diabetic group. A diminished production of TXB2 was found in the treated diabetic uteri when compared to the non-treated diabetic group. Moreover, a positive correlation between plasma glucose levels and uterine TXB2 generation was observed. When control uterine tissue was exposed in vitro to high concentrations of glucose (22 mM) and compared to control tissue incubated in the presence of glucose 11 mM alterations in the generation of PGE2, PGF2 alpha, and 6-keto-PGF1 alpha were not found, but a higher production of TXB2 was observed and values were similar to those obtained in the diabetic tissue. Alteration in the production of the prostanoids evaluated were not observed when diabetic tissue was incubated in the presence of high concentrations of glucose. These results provide evidence of a direct relationship between plasma glucose levels and uterine production of TXA2.
Asunto(s)
Ácido Araquidónico/metabolismo , Hiperglucemia/metabolismo , Tromboxano A2/metabolismo , Útero/metabolismo , Animales , Glucemia/análisis , Dinoprost/metabolismo , Dinoprostona/metabolismo , Femenino , Glucosa/metabolismo , Glucosa/farmacología , Hiperglucemia/patología , Técnicas In Vitro , Ovariectomía , Florizina/farmacología , Ratas , Tromboxano B2/metabolismo , Útero/patologíaRESUMEN
Glucose transport by uterine strips from ovariectomized estrogenized rats was explored. Sugar transport was significantly different from saccharose values (non-specific diffusion) only after 60 min of incubation. The addition of cytochalasin B demonstrated that we are measuring a specific mechanism for glucose transport. Insulin-enhanced sugar transport only at 0.5 or 0.25 U/ml prostaglandin E1 (PGE1), PGE2 and PGF2 alpha (10(-7) M) significantly improved glucose transport, but indomethacin (10(-6) M) failed in modifying this parameter in either control nor insulin-treated tissues. We did not observe an additive or synergistic action between PGE2 (10(-7) M) and insulin (used at maximal or submaximal concentration).
Asunto(s)
Alprostadil/farmacología , Dinoprostona/farmacología , Glucosa/metabolismo , Prostaglandinas F/farmacología , Útero/efectos de los fármacos , Animales , Transporte Biológico/efectos de los fármacos , Transporte Biológico Activo/efectos de los fármacos , Citocalasina B/farmacología , Difusión , Interacciones Farmacológicas , Estradiol/farmacología , Femenino , Indometacina/farmacología , Insulina/farmacología , Ovariectomía , Ratas , Ratas Wistar , Estimulación Química , Sacarosa/metabolismo , Útero/metabolismoRESUMEN
Spontaneous contractile activity, glucose (Glu), glycogen (GLY), triglyceride (TG) metabolism and eicosanoid production, was evaluated in isolated uterine strips from control and non-insulin-dependent diabetic rats on day 10 of pregnancy. Metabolism of Glu, levels of GLY and TG and eicosanoid production were also studied in day 10 embryos obtained from both experimental groups. "In vitro" isometric developed tension (IDT), was similar at 0 hr in control and diabetic uterine preparations, but IDT was decreased after a 60 min incubation in the diabetic group. The frequency of contractions (FC) was similar at 0 hr and after 60 min incubation in both experimental groups. On the other hand, the production of 14CO2 from U14C-glucose was lower in isolated uteri and embryos obtained from diabetic rats than in controls. Initial TG levels were similar in uteri isolated from control and diabetic rats, and higher in embryos obtained from diabetic mothers than in controls. Levels of TG in uterine strips suspended in Glu or Glu-free medium did not differ at 0 hr or at 60 min either in controls or in diabetic rats. On the contrary GLY levels in uterine strips from diabetic animals were higher than in controls, whereas in embryos from diabetic mothers GLY levels were similar to controls. Levels of GLY in uterine strips from controls and diabetic animals decreased after 60 min incubation only in the absence of Glu in the incubation medium. Production of PGE2, PGE1, 6-keto-PGF1 alpha, PGF2 alpha, TXB2 and LTB4 was studied in uterine strips and embryos obtained from control and diabetic rats. No differences were found between control and diabetic uterine prostanoid production, but lower production of LTB4 was observed in diabetic uteri. However production of PGE2 and PGF2 alpha was greater in embryos obtained from diabetic mothers than in controls. In this study, we observed lower uterine metabolic alterations than in the pancreatectomized diabetic rat model studied previously, but important anomalies in the embryos obtained from non-insulin-dependent diabetic mother were found.