Cynthia Dela Cruz

Serum antimüllerian hormone measurements with second generation assay at two distinct menstrual cycle phases for prediction of cycle cancellation, pregnancy and live birth after in vitro fertilization

PurposeThis study investigated the usefulness of serum antimüllerian hormone (AMH) measurements at two distinct menstrual cycle phases to predict in vitro fertilization (IVF) outcomes.MethodsThis was a prospective observational study enrolling 135 consecutive patients referred for conventional IVF or ICSI in a university hospital. Blood samples were obtained for serum AMH measurements on days 3 and 18–20, while transvaginal ultrasound was performed for antral follicle count (AFC) at day 3 of the menstrual cycle immediately before treatment. AMH was measured with the new Beckman Coulter Generation II (GenII) assay. The main outcome measures were cycle cancellation due to poor ovarian response, clinical pregnancy, and live birth.ResultsThere was a strong correlation between AMH levels measured at day 3 and day 18–20 of the menstrual cycle (r = 0.837; P < 0.0001). Day 18–20 serum AMH was comparable to day 3 serum AMH and AFC for the prediction of cycle cancellation (areas under the ROC curve were 0.84 for day 3 AMH, 0.89 for day 18–20 AMH, and 0.80 for AFC). Day 18–20 AMH had a modest predictive value for pregnancy or live birth (area under ROC curve 0.71 for both), which was comparable to that of day 3 AMH; however, AFC had no predictive value for these outcomes.ConclusionsDay 18–20 AMH was comparable to day 3 AMH for the prediction of cycle cancellation, clinical pregnancy, and live birth after IVF. Both AMH measurements were accurate for the prediction of cancellation but were significantly less useful for the prediction of pregnancy or live birth.

The role of TGFβ superfamily members in the pathophysiology of endometriosis.

Abstract The transforming growth factor-beta (TGFβ) superfamily comprises over 30 dimeric proteins with conserved structures, which play important roles in the control of cellular proliferation, differentiation and apoptosis. These proteins are expressed and finely regulated in human endometrium during the menstrual cycle, which is consistent with their effects on endometrial cell proliferation and tissue remodeling. This review is focused on summarizing the role of key members of the TGFβ superfamily in the pathophysiology of endometriosis. Evidence suggests that TGFβ, activins, inhibins, nodal, bone morphogenetic proteins, growth differentiation factors, and anti-Müllerian hormone are produced by endometriotic lesions and could be involved in the establishment and progression of the disease. Their receptors and signaling pathways may also be altered in the presence of endometriosis and may be potential targets to the development of therapeutic agents.

Expression of Nodal, Cripto, SMAD3, phosphorylated SMAD3, and SMAD4 in the proliferative endometrium of women with endometriosis.

Background: Nodal is a growth factor of the transforming growth factor β superfamily that is expressed in high turnover tissues, such as the human endometrium, and in several malignancies. The effects of Nodal are modulated by the coreceptor Cripto and mediated by SMAD proteins. This study evaluated the gene and protein expression of Nodal, Cripto, total and phosphorylated (p) SMAD3, and SMAD4 in the proliferative endometrium of women with and without endometriosis. Method: Total RNA was isolated and complementary DNA synthesized from eutopic endometrium of women with (n = 15) and without (n = 12) endometriosis, followed by quantitative real-time polymerase chain reaction (PCR) to evaluate the gene expression of Nodal, Cripto, SMAD3, and SMAD4. Western blot was used to evaluate the protein levels of Nodal and Cripto, and immunohistochemistry was performed to localize SMAD3, pSMAD3, and SMAD4. Results: Although Nodal expression was unchanged in women with endometriosis, real-time PCR indicated lower gene expression of Cripto (fold change 0.27, P < .05) in the endometriosis group. This difference, however, was not maintained at protein expression level as assessed by Western blot. The immunostaining of total SMAD3 was reduced in the endometriosis group (P < .01), but the localization of pSMAD3 and the nuclear staining of SMAD4 were unchanged. Conclusion: These findings suggest that the Nodal signaling pathway has subtle changes in the endometrium of women with endometriosis, but this imbalance may not cause functional damage as it seems not to affect the nuclear expression of SMAD4.

Deep Infiltrating Endometriosis and Endometrial Adenocarcinoma Express High Levels of Myostatin and Its Receptors Messenger RNAs.

Myostatin is a growth factor member of the transforming growth factor β superfamily, which is known to play major roles in cell proliferation and differentiation. The present study investigated the messenger RNA (mRNA) expression of myostatin and myostatin receptors (activin receptor-like kinase 4 [ALK4], transforming growth factor (TGF)-β type I receptor kinase [ALK5] and activin receptor type IIB [ActRIIB]) in endometrium of healthy women during menstrual cycle as well as in benign (endometriosis, polyps) and malignant (endometrial adenocarcinoma) conditions. Endometrial specimens were collected by hysteroscopy, whereas endometriotic lesions were collected by laparoscopy, and adenocarcinomas were sampled after hysterectomy. Total RNA was extracted from tissue homogenates, and gene expression was assessed by quantitative real-time polymerase chain reaction. Myostatin and myostatin receptors mRNAs were expressed by healthy endometrium throughout the menstrual cycle, with no differences between the proliferative and secretory phase. The highest myostatin mRNA expression was found in patients with deep infiltrating endometriosis (DIE) and in endometrial carcinoma; expression was also found in ovarian endometrioma (OMA ) and endometrial polyps. Myostatin receptors mRNA expression was higher in DIE and adenocarcinomas compared to control endometrium. The expression of ALK5 and ActRIIB in OMA was higher than in controls, whereas polyps had an increased expression of ALK5 mRNA. In conclusion, the present data showed for the first time the expression of myostatin in healthy endometrium and a higher expression in endometriosis and endometrial cancer, suggesting myostatin involvement in human endometrial physiology and related pathologies.

Urocortin and corticotrophin-releasing hormone receptor type 2 mRNA are highly expressed in deep infiltrating endometriotic lesions

Ovarian endometrioma (OMA) and deep infiltrating endometriosis (DIE) are the most severe forms of endometriosis, but different pathogenetic mechanisms and clinical symptoms distinguish these two forms. Corticotrophin-releasing hormone (CRH) and urocortin (Ucn) are endometrial neuropeptides involved in tissue differentiation and inflammation. The expression of CRH, Ucn, Ucn2, CRH-receptors (type-1 and type-2) and inflammatory enzymes phospholipase-A2 group IIA (PLA2G2A) and cycloxygenase-2 (COX2) were evaluated in OMA (n = 22) and DIE (n = 26). The effect of CRH or Ucn on COX2 mRNA expression was evaluated in cultured human endometrial stromal cells. In DIE lesions, CRH, Ucn and CRH-R2 mRNA levels were significantly higher than in OMA (P < 0.01, P < 0.001 and P < 0.05, respectively); DIE lesions showed a higher expression of COX2 (P < 0.01) and PLA2G2A (P < 0.05) mRNA than OMA, which was positively correlated with CRH-R2 mRNA expression (P < 0.05). Intense immunostaining for CRH and Ucn was shown in DIE. Treatment of cultured endometrial stromal cells with Ucn significantly increased COX2 mRNA expression (P < 0.01); this effect was reversed by the CRH-R2 antagonist astressin-2B. In DIE, DIE lesions highly express neuropeptide and enzyme mRNAs, supporting a strong activation of inflammatory pathways.

Follistatin Expression in Human Invasive Breast Tumors: Pathologic and Clinical Associations

Follistatin is a potent native activin antagonist that is expressed in the normal mammary gland and in different breast proliferative diseases. Despite experimental evidence that follistatin can modulate the breast cancer cell cycle, the clinical significance of follistatin expression in these tumors is unknown. The aim of this study was to correlate the intensity of follistatin expression in invasive breast cancer with some of its clinical and pathologic features, such as the disease stage and the hormonal receptor status. Paraffin blocks of tumor samples that had been fixed in buffered formalin were obtained from 154 women subjected to surgery for breast cancer between 2008 and 2012. Sections from all paraffin blocks were cut and processed together by immunohistochemistry using a commercial monoclonal antibody to human follistatin. The intensity of follistatin staining was unrelated to the menopausal status, the disease stage, the grade, progesterone receptor expression, and local or systemic recurrence. However, follistatin immunoreactivity was significantly stronger in estrogen receptor (ER)-negative tumors than in ER-positive tumors. These findings suggest that follistatin expression in invasive breast cancer is unrelated to the disease severity and the risk of recurrence, but is more intense in ER-negative tumors.

Urocortin 1 expression and secretion by human umbilical vein endothelial cells: In vitro effects of interleukin 8, interferon γ, lipopolysaccharide, endothelin 1, prostaglandin F-2α, estradiol, progesterone and dexamethasone.

Urocortin 1 (Ucn1) is a 40-amino-acid peptide that has vasodilatory activity and displays immunomodulatory and antioxidant properties. Maternal and cord plasma Ucn1 levels are increased in preeclampsia and preterm labor, but the mechanisms of such increase are poorly known. Thus, we investigated Ucn1 localization in human umbilical cord and assessed some potential stimuli to Ucn1 release by human umbilical vein endothelial cells (HUVEC). Human umbilical cords were obtained at uncomplicated term pregnancy (n=11). Ucn1 localization was assessed by immunohistochemistry and quantified. HUVEC were grown in vitro to confluence, then incubated with serial concentrations of interleukin (IL)-8, interferon (INF)-γ, lipopolysaccharide (LPS), endothelin (ET)-1, prostaglandin (PG)F-2α, estradiol, progesterone and dexamethasone and Ucn1 concentrations were measured in the supernatants. Ucn1 was immunolocalized with similar intensity in umbilical cord arteries, vein and Whartons jelly. Ucn1 mRNA was detected in all HUVEC cultures and Ucn1 peptide was detectable in culture medium from untreated cells at different time points. Incubation with IFN-γ increased Ucn1 secretion in a dose-dependent manner. Treatments with IL-8, LPS, ET-1 and dexamethasone were able to increase three to fourfold Ucn1 release from cultured endothelial cells. In conclusion, umbilical vessels express Ucn1 and may be a contributive source of Ucn1 release into fetal-placental circulation. IL-8, IFN-γ, LPS, ET-1 and dexamethasone promote Ucn1 secretion from cultured HUVEC.

Expression, localization and control of activin A release from human umbilical vein endothelial cells.

Abstract Activin-A is a member of the TGFβ superfamily found in maternal and umbilical cord blood throughout gestation. We investigated whether human umbilical vein endothelial cells (HUVEC) express activin-A in vivo and tested the effects of vasoactive (endothelin-1), pro-inflammatory (interferon-γ, interleukin-8) and anti-inflammatory (dexamethasone, urocortin) factors on activin-A release by isolated HUVEC in vitro. Activin βA subunit protein and mRNA were strongly localized in the endothelial cells of umbilical veins and were also detectable in scattered cells of the cord connective tissue. Dimeric activin-A was detected in the HUVEC culture medium at picomolar concentrations. Activin-A release by HUVEC decreased after cell incubation with urocortin (p < 0.01), whereas no effect was observed with interleukin-8, interferon-γ, endothelin-1 or dexamethasone. In summary, activin-A is present in the human umbilical vein endothelium in vivo and is produced and released by isolated HUVEC. Activin-A secretion is inhibited in vitro by urocortin, a neuropeptide with predominantly anti-inflammatory action.

Caspase-3 gene expression in human luteinized granulosa cells is inversely correlated with the number of oocytes retrieved after controlled ovarian stimulation

Abstract Granulosa cells control oocyte maturation through paracrine signalling and changes to the microenvironment around the oocyte. Apoptosis occurs as a physiological mechanism of granulosa cell renewal, but how it relates with the ovarian response to induced ovulation is still unclear. Therefore, this study evaluated apoptosis-related gene expression levels in granulosa cells of patients undergoing controlled ovarian stimulation. We enrolled prospectively 59 consecutive IVF patients referred to a tertiary academic hospital for couple infertility treatment. Luteinized granulosa cells were isolated from follicular fluid and the RNA was extracted, reverse-transcribed and the gene expression of apoptosis inducers (caspase-3, caspase-8 and bax) and inhibitor (Bcl-2) was quantified by real-time polymerase chain reaction. Caspase-3 gene expression correlated negatively with the number of pre-ovulatory follicles (Spearman’s r =  −0.308), the number of collected oocytes (r =  −0.451), the number of mature oocytes (r =  −0.526), the number of fertilized oocytes (r =  −0.439) and the number of viable embryos (r =  −0.443, all statistically significant at p < 0.02 level). No such associations were found with caspase-8, bax or bcl-2. These preliminary findings suggest that increased caspase-3 gene expression in granulosa cells is associated with a worse ovulatory response in humans.

Effects of Nandrolone Decanoate and Resistance Exercise on Skeletal Muscle in Adult Male Rats

El objetivo de este estudio fue comparar los efectos del ejercicio de resistencia con o sin decanoato de nandrolona (DN) en el musculo esqueletico y la masa corporal de ratas macho adultas. El protocolo de entrenamiento consistio en 15 sesiones durante 6 semanas de saltos. DN 5mg/kg se administro dos veces durante la semana. El ejercicio fue efectivo para inducir un aumento en el area de las fibras de los musculos extensor largo de los dedos y soleo. El DN asociado con el ejercicio fue capaz de inducir un aumento en el area de las fibras de los musculos. En el grupo de DN sin entrenamiento, se observo un aumento en los parametros musculares evaluados. El ejercicio de resistencia sin DN fue capaz de promover un aumento en el area de las fibras de los musculos, lo que indica que despues de un periodo de adaptacion al ejercicio, el efecto en los musculos causada por la DN se logro por el ejercicio, sin una gestion DN y los consiguientes peligros para la salud.