Differential expression of upstream stimulatory factor (USF) 2 variants in eutopic endometria from women with endometriosis: estradiol regulation.

BACKGROUND: Endometriosis, pro-inflammatory and invasive benign disease estrogen dependent, abnormally express in endometria the enzyme P450Arom, positively regulated by steroid factor-1 (SF-1). Our objective was to study the nuclear protein contents of upstream stimulating factor 2 (USF2a and USF2b), a positive regulator of SF-1, throughout the menstrual cycle in eutopic endometria from women with and without (control) endometriosis and the involvement of nuclear estrogen receptors (ER) and G-coupled protein estrogen receptor (GPER)-1. RESULTS: Upstream stimulating factor 2 protein contents were higher in mid (USF2b) and late (USF2a and USF2b) secretory phase in eutopic endometria from endometriosis than control (p < 0.05). In isolated control epithelial cells incubated with E2 and PGE2, to resemble the endometriosis condition, the data showed: (a) significant increase of USF2a and USF2b nuclear protein contents when treated with E2, PPT (specific agonist for ERα) or G1 (specific agonist for GPER1); (b) no increase in USF2 binding to SF-1 E-Box/DNA consensus sequence in E2-treated cells; (c) USF2 variants protein contents were not modified by PGE2; (d) SF-1 nuclear protein content was significantly higher than basal when treated with PGE2, E2 or G1, stimulation unaffected by ICI (nuclear ER antagonist); and (e) increased (p < 0.05) cytosolic protein contents of P450Arom when treated with PGE2, E2, PPT or G1 compared to basal, effect that was additive with E2 + PGE2 together. Nevertheless, in endometriosis cells, the high USF2, SF-1 and P450Arom protein contents in basal condition were unmodified. CONCLUSION: These data strongly suggest that USF2 variants and P450Arom are regulated by E2 through ERα and GPER1, whereas SF-1 through GPER1, visualized by the response of the cells obtained from control endometria, being unaffected the endogenously stimulated cells from endometriosis origin. The lack of E2 stimulation on USF2/SF-1 E-Box/DNA-sequence binding and the absence of PGE2 effect on USF2 variants opposite to the strong induction that they exert on SF1 and P450 proteins suggest different mechanisms and indirect regulations. The sustained USF2 variants protein expression during the secretory phase in eutopic endometria from women with endometriosis may participate in the pathophysiology of this disease strongly associated with infertility and its characteristic endometrial invasion to ectopic sites in the pelvic cavity.

Differential expression of upstream stimulatory factor (USF) 2 variants in eutopic endometria from women with endometriosis: estradiol regulation

BACKGROUND: Endometriosis, pro-inflammatory and invasive benign disease estrogen dependent, abnormally express in endometria the enzyme P450Arom, positively regulated by steroid factor-1 (SF-1). Our objective was to study the nuclear protein contents of upstream stimulating factor 2 (USF2a and USF2b), a positive regulator of SF-1, throughout the menstrual cycle in eutopic endometria from women with and without (control) endometriosis and the involvement of nuclear estrogen receptors (ER) and G-coupled protein estrogen receptor (GPER)-1. RESULTS: Upstream stimulating factor 2 protein contents were higher in mid (USF2b) and late (USF2a and USF2b) secretory phase in eutopic endometria from endometriosis than control (p < 0.05). In isolated control epithelial cells incubated with E2 and PGE2, to resemble the endometriosis condition, the data showed: (a) significant increase of USF2a and USF2b nuclear protein contents when treated with E2, PPT (specific agonist for ERa) or G1 (specific agonist for GPER1); (b) no increase in USF2 binding to SF-1 E-Box/DNA consensus sequence in E2-treated cells; (c) USF2 variants protein contents were not modified by PGE2; (d) SF-1 nuclear protein content was significantly higher than basal when treated with PGE2, E2 or G1, stimulation unaffected by ICI (nuclear ER antagonist); and (e) increased (p < 0.05) cytosolic protein contents of P450Arom when treated with PGE2, E2, PPT or G1 compared to basal, effect that was additive with E2 + PGE2 together. Nevertheless, in endometriosis cells, the high USF2, SF-1 and P450Arom protein contents in basal condition were unmodified. CONCLUSION: These data strongly suggest that USF2 variants and P450Arom are regulated by E2 through ERa and GPER1, whereas SF-1 through GPER1, visualized by the response of the cells obtained from control endometria, being unaffected the endogenously stimulated cells from endometriosis origin. The lack of E2 stimulation on USF2/SF-1 E-Box/DNA-sequence binding and the absence of PGE2 effect on USF2 variants opposite to the strong induction that they exert on SF1 and P450 proteins suggest different mechanisms and indirect regulations. The sustained USF2 variants protein expression during the secretory phase in eutopic endometria from women with endometriosis may participate in the pathophysiology of this disease strongly associated with infertility and its characteristic endometrial invasion to ectopic sites in the pelvic cavity.

Extended transvaginal sonography in deep infiltrating endometriosis: use of bowel preparation and an acoustic window with intravaginal gel: preliminary results.

OBJECTIVES: The purpose of this study was to assess the diagnostic performance of extended transvaginal sonography for diagnosing deep infiltrating endometriosis. METHODS: A prospective study was conducted comprising 51 women (mean age, 32.9 years; range, 23-43 years) with suspected deep infiltrating endometriosis based on clinical symptoms. All women underwent extended transvaginal sonography, which included assessment of 2 pelvic compartments (anterior compartment: bladder and distal ureters; and posterior compartment: posterior vaginal fornix, retrocervical area, pouch of Douglas, and rectosigmoid). The sliding sign for detecting pouch of Douglas obliteration was also assessed. All patients received bowel preparation before sonographic examinations. A single examiner performed all examinations. All women underwent laparoscopic surgery, and histologic confirmation of endometriosis was done. The sensitivity, specificity, positive likelihood ratio (LR+) and negative likelihood ratio (LR-) were calculated. RESULTS: Some women had more than 1 lesion, giving a total of 55 histologically confirmed lesions (rectosigmoid, n = 13; vagina, n = 5; retrocervical, n = 32; bladder, n = 5). The sensitivity, specificity, and LR+ for rectosigmoid involvement were 100%, 93%, and 14.0, respectively. The sensitivity, specificity, LR+, and LR- for vaginal involvement were 60%, 98%, 30.0, and 0.41. The sensitivity, specificity, LR+, and LR- for retrocervical involvement were 84%, 96%, 19.4, and 0.16. The sensitivity, specificity, and LR- for bladder involvement were 20%, 100%, and 0.80. The sensitivity, specificity, LR+, and LR- of the sliding sign for diagnosing pouch of Douglas obliteration were 89%, 92%, 10.7, and 0.12. CONCLUSIONS: Except for bladder involvement, extended transvaginal sonography has good diagnostic performance for deep infiltrating endometriosis.

Physiologic activation of nuclear factor kappa-B in the endometrium during the menstrual cycle is altered in endometriosis patients.

OBJECTIVE: To evaluate nuclear factor kappaB (NF-κB) activation and NF-κB-p65 subunit activation, immunolocalization, and expression in the endometrium of healthy women and endometriosis patients throughout the menstrual cycle. DESIGN: Prospective observational study. SETTING: Affiliated hospital and university research laboratory. PATIENT(S): Twenty-four healthy women and 24 endometriosis patients. INTERVENTION(S): Menstrual, proliferative, and secretory endometrial biopsies. MAIN OUTCOME MEASURE(S): Assessment of NF-κB and p65 activation by protein-DNA binding assays and p65 localization and expression by immunohistochemistry. RESULT(S): Total NF-κB-DNA binding was constitutive and variable in human endometrium accross the menstrual cycle. Healthy women (physiologic conditions) showed higher p65-DNA binding in proliferative than in menstrual and secretory endometrium. Conversely, in endometriosis patients, p65-DNA binding was higher in proliferative and secretory endometrium than in menstrual endometrium. Endometrial epithelial cells showed higher p65 expression level score than endometrial stromal cells. CONCLUSION(S): NF-κB activity is constitutive, physiologic, and variable in human endometrium. The physiologic cyclic p65 activation pattern was altered in endometriosis patients, showing no cyclic variation between the proliferative and secretory phase of the menstrual cycle. The absence of decreased p65 activity in secretory endometrium from endometriosis patients is concurrent with progesterone resistance and could participate in endometrial biologic alterations during the implantation window in endometriosis patients.

P450Arom induction in isolated control endometrial cells by peritoneal fluid from women with endometriosis.

OBJECTIVE: To study the effect of peritoneal fluid from women with (PF-E) and without (PF-C) endometriosis on P(450)Arom expression in endometrial cells. DESIGN: Experimental study. SETTING: University research unit. PATIENT(S): Forty women of reproductive age with (n = 22) or without (control; n = 18) endometriosis. INTERVENTION(S): Peritoneal fluid and eutopic endometrial samples were obtained during surgery from women with (n = 13 and 9, respectively) and without (n = 4 and 14, respectively) endometriosis. MAIN OUTCOME MEASURE(S): Expression study for P(450)Arom, steroid factor 1 (SF-1), chicken ovalbumin upstream transcription factor I (COUP-TFI), and COUP-TFII messenger RNA (reverse transcriptase-polymerase chain reacion) and/or protein (immunoblot) in isolated endometrial epithelial cells transfected or not with expression vector containing SF-1, COUP-TFI, or COUP-TFII complementary DNAs. RESULT(S): Basal messenger RNA and/or protein expression of P(450)Arom and SF-1 were augmented in endometriosis, and that of COUP-TF was diminished. In control cells, (Bu)(2)cAMP and PF-E increased P(450)Arom and SF-1 expression (but not COUP-TF expression) in a dose-dependent way, an effect not observed with PF-C, adsorbed PF-E, or 10(-5) M indomethacin. Transfected cells confirmed these results. Any treatments modified the studied molecules in endometriosis cells. CONCLUSION(S): These data indicate that molecules contained in PF-E favor an estrogenic microenvironment, suggesting a role in the etiopathogenesis of endometriosis enabling the survival, maintenance, and growth of endometrial implants in the ectopic locations.

Nuclear factor kappaB pathway and interleukin-6 are affected in eutopic endometrium of women with endometriosis.

In order to investigate the role of the nuclear factor kappaB (NFKB) pathway on gene expression in the eutopic endometrium in endometriosis, and in particular of interleukin-6 (IL6), we evaluated RELA, IkappaB kinase (CHUK), NFKBIA and IL6 expressions and NFKB DNA binding in eutopic endometrium from women with endometriosis. Eutopic endometrium was obtained from 37 women with endometriosis and 42 fertile women during laparoscopy. We analysed RELA, CHUK, NFKBIA and IL6 mRNA levels (RT-PCR); RELA, CHUK and NFKBIA proteins and p-NFKBIA/NFKBIA ratio (western blot); and NFKB binding (DNA shift assay) and IL6 concentration (ELISA) in endometrial explants. Our results indicate that mRNA and cytoplasmic proteins of RELA and CHUK exhibit constant levels in normal endometrium during the menstrual cycle. A dramatic increase (P<0.05) in NFKBIA mRNA expression, RELA nuclear presence and the mRNA and the protein of IL6 during late secretory phase was also observed in this tissue. By contrast, in eutopic endometrium from endometriosis patients, a decrease (P<0.05) in IL6 mRNA and protein (61%), NFKBIA mRNA (46%), p-NFKBIA/NFKBIA ratio (42%), RELA nuclear stromal (68%) and CHUK (48%) proteins were found exclusively during the late secretory phase compared with normal endometrium. In conclusion, the canonical activation of NFKB pathway is deregulated and may have reduced transcriptional function affecting NFKBIA and IL6 expression, genes related local proinflammatory processes. These molecular alterations observed during the late secretory phase in eutopic endometrium from endometriosis patients constitute a NFKB system dysfunction, suggesting that NFKB could be an important factor in endometriosis aetiology.

Ovarian function during puberty in girls with type 1 diabetes mellitus: response to leuprolide.

CONTEXT: An increased prevalence of polycystic ovary syndrome (PCOS) has been reported in adult women with type 1 diabetes mellitus (DM1). We investigated whether these hormonal abnormalities begin during puberty by evaluating the ovarian steroidogenic response to leuprolide acetate. METHODS: We studied 56 adolescent girls with DM1 (aged 12.3 +/- 0.2 yr) and 64 healthy girls (C) (aged 11.9 +/- 0.2 yr) up to 2 yr post menarche, matched by age, body mass index, and pubertal development. We evaluated anthropometrical data and Ferriman-Gallway score and performed a leuprolide test (500 microg sc) to study ovarian function. Ovarian volume was determined by transabdominal ultrasonography. RESULTS: We found five DM1 but no C girls with abnormally located terminal hair (Fisher's exact, P < 0.05). Free androgen index increased throughout puberty in girls with DM1 (ANOVA, P < 0.0001), which was associated with a decrease in SHBG levels in girls with DM1 (ANOVA, P < 0.0001). Stimulated 17OH progesterone (17OHProg) increased throughout puberty only in girls with DM1 (ANOVA, P < 0.01). Girls with DM1 at Tanner stage 5 had higher stimulated LH to FSH ratio, testosterone, and 17OHProg levels than girls at Tanner stage 4. In contrast, in C girls the stimulated testosterone, 17OHProg, and LH to FSH ratio were similar at Tanner stages 4 and 5. Ovarian volumes and uterine length were larger in girls with DM1 (analysis of covariance, P < 0.05). CONCLUSIONS: These data suggest that patients with DM1 have differences in ovarian steroidogenic response to leuprolide, compared with C girls during puberty. Future studies in young women should clarify whether these findings are related to the pathogenesis of hyperandrogenism later in life.

Criopreservacion de pronucleos: rol en el programa de fertilizacion asistida

Objetivo: Determinar el rol de la criopreservación (CP) de pronúcleos (PN), como una herramienta para disminuir la incidencia de embarazos múltiples y dar otra oportunidad de transferencia embrionaria, sin requerir estimulación ovárica nuevamente, en parejas con infertilidad que requieren como tratamiento algún tipo de procedimiento de Fertilización Asistida (FA). Material y Método: Se analizaron los resultados de 545 procedimientos de FA entre marzo del 2000 y junio del 2003. Resultados: La incorporación de la CP se logra en diciembre del 2001, criopreservando hasta la fecha el 51,3% de las parejas que tiene más de 6 folículos a aspirar. Se han criopreservado 623 PN dando un promedio 6,4 PN por pareja. Se han descongelado 166 PN, sobreviviendo 134 PN, lo que implica un 80,7%. Ciento catorce PN clivaron a embrión de 4 células, equivalente a un 85,1%. Se han transferido 114 PN en 39 ciclos con un promedio de 2,9 embriones por pareja, dando origen a 12 embarazos clínicos (30,7%) con una tasa implantacional del 11,4%. Se observa una reducción del número de embarazos múltiples en dobles del 23% al 15,5%, en triples del 7,6% al 2,2% y la no ocurrencia de cuádruples o más, al comparar el período 2000-diciembre 2001 (sin CP de PN), con diciembre de 2001-junio del 2003.

Clomiphene citrate and ovulation induction.

Clomiphene can be used to treat anovulation due to hypothalamus or pituitary gland dysfunction, and it normalizes the luteal phase in stimulated patients. It can be used to estimate ovarian follicle reserve, and may be predictive of ovulation in women aged >/=35 years or with failed IVF. Contraindications include risk of congenital anomalies, chronic liver disease and visual disorders. Clomiphene may impair fertility through its effects on cervical mucus and in causing various endometrial dysfunctions. However, if clomiphene is administered in 50 mg doses, side-effects are avoided and efficacy is similar to that of a 100 mg dose, although daily dosages of 200 mg/day over 5 days can induce ovulation in approximately 70% of treated patients. Gonadotrophin concentrations increase up to days 5-9 when follicles are selected, and clomiphene is effective in patients with polycystic ovary syndrome (PCOS). Fifty percent of normal patients conceive, a value perhaps biased by the antagonistic effects of clomiphene on cervical mucus in some women. Clomiphene is valuable for IVF, and is used by some clinics in combination with HMG or recombinant FSH. Resistance to clomiphene can develop, and human chorionic gonadotrophin may be needed to induce ovulation in clomiphene cycles. Corticosteroids and human menopausal gonadotrophin (HMG) can be combined with clomiphene for stimulation, its combination with HMG long having been a standard protocol in assisted reproduction. PCOS patients may become insulin resistant, a condition improved by the administration of metformin. Other adverse effects include multiple pregnancies, an increase in the rate of multiple births, ovarian hyperstimulation and unsubstantiated claims of ovarian cancer.

Ecografía tridimensional / Tridimensional echography

Se realiza una revisión bibliográfica de la ecografía tridimensional, a objeto de dar a conocer los principios físicos, las eventuales aplicaciones en obstetricia y ginecología, así como las ventajas y desventajas de la técnica. El principio físico es la sumatoria de un conjunto de ecografías convencionales, orientadas en diferentes ejes de corte de una determinada estructura, las que son integradas por un computador, obteniéndose una imagen de volumen, o tridimensional. La limitación del método está en la obtención de las imágenes ecográficas convencionales, en los tres planos espaciales, los cuales deben ser exactamente perpendiculares entre si, para obtener una buena imagen tridimensional. Las aplicaciones abarcan todas las áreas de la medicina, dondo sea posible obtener imágenes, vale decir: cirugía vascular, con el uso de transductores intravasculares; imágenes de lesiones vasculares en cardiología, así como imágenes de tumores en diversos órganos. Se ve un campo promisorio en obstetricia con la evaluación de malformaciones fetales, lo que permite un mejor diagnóstico al poder mover en la pantalla las imágenes obtenidas, para observarlas en diferentes ángulos; además, éstas pueder ser almacenadas, interpretadas y comparadas posteriormente. Al ser una técnica no invasiva se puede utilizar en forma rutinaria en la embarazada

Leptina y sistema reproductivo / Leptin and the reproductive system

La leptina es una hormona proteica recientemente descubierta, la cual es producida por los adipocitos. Esta actúa regulando el apetito, el peso corporal y la termogénesis. Su posible rol como modulador en el eje reproductivo se ha inferido de una cepa de ratas carentes de leptina, que se caracterizan por ser obesas, diabéticas e infértiles y en las que el administrar esta hormona se producen cambios como disminución del apetito, aumento del gasto energético y recuperación de la fertilidad. Estos fenómenos han motivado una gran cantidad de estudios para establecer el rol la leptina en el sistema reproductivo. Se revisa la literatura existente sobre leptina y su acción sobre los cambios de peso corporal, pubertad, sistema nervioso central y periférico, ovarios y embarazo

Mioma uterino y embarazo / Uterine myomas and pregnancy

Se analizan los principales aspectos del embarazo asociado a mioma uterino: incidencia, diagnóstico, evolución, complicaciones más frecuentes y sus alternativas terapéuticas, en base a una revisión bibliográfica del tema. Se comunican, además, dos casos clínicos atendidos en el Servicio de Ginecología y Obstetricia del Hospital San Juan de Dios en 1997 y se señala la frecuencia de esta patología en los último dos años