Linah Al-Alem

Endothelin-2 induces oviductal contraction via endothelin receptor subtype A in rats

Proper function of the oviduct is critical to reproductive success with regulated contraction and relaxation facilitating transportation of the germ cells to the site of fertilization. Endothelin-2 (EDN2) is a potent vasoconstrictor produced by granulosa cells of the preovulatory follicle at the time of ovulation; however, whether this gonadotropin surge-induced peptide played a role in facilitating germ cell transportation by inducing oviductal contraction was unknown. The objectives of these experiments were (1) to determine whether the endothelin receptor system was present in the oviduct, (2) to test the hypothesis that EDN2 induces oviductal contraction via a specific endothelin receptor subtype, (3) to determine, as a possible alternate source of the ligand, whether mRNA for EDN2 was expressed in cumulus-oocyte complexes (COCs) within the oviduct, and (4) to determine whether EDN2 could overcome prostaglandin E(2) (PGE(2))-induced oviductal relaxation. Microarray and real-time PCR analysis indicated that mRNA for both the endothelin receptor subtypes (ET(A) and ET(B)) was present in the oviduct, whereas immunohistochemical examination revealed that ET(A) protein was the dominant isoform, present in the luminal epithelial cells of the oviduct. Real-time PCR analysis demonstrated that mRNA for EDN2 was expressed in COCs after ovulation. Isometric tension analysis indicated that EDN2 was a potent oviductal constrictor and that the contractile effect of EDN2 was mediated by the ET(A) and not the ET(B) receptor subtype. The oviductal contraction induced by EDN2 also reversed oviductal relaxation induced by PGE(2). In summary, ET(A) receptor-specific EDN2-induced contraction as a facilitator of oviductal function suggests a novel pathway involved in germ cell transport and hence mammalian fertility.

Specific Thiazolidinediones Inhibit Ovarian Cancer Cell Line Proliferation and Cause Cell Cycle Arrest in a PPARγ Independent Manner

Background Peroxisome Proliferator Activated Receptor gamma (PPARγ) agonists, such as the thiazolinediones (TZDs), have been studied for their potential use as cancer therapeutic agents. We investigated the effect of four TZDs—Rosiglitazone (Rosi), Ciglitazone (CGZ), Troglitazone (TGZ), and Pioglitazone (Pio)—on ovarian cancer cell proliferation, PPARγ expression and PPAR luciferase reporter activity. We explored whether TZDs act in a PPARγ dependent or independent manner by utilizing molecular approaches to inhibit or overexpress PPARγ activity. Principal Findings Treatment with CGZ or TGZ for 24 hours decreased proliferation in three ovarian cancer cell lines, Ovcar3, CaOv3, and Skov3, whereas Rosi and Pio had no effect. This decrease in Ovcar3 cell proliferation was due to a higher fraction of cells in the G0/G1 stage of the cell cycle. CGZ and TGZ treatment increased apoptosis after 4 hours of treatment but not after 8 or 12 hours. Treatment with TGZ or CGZ increased PPARγ mRNA expression in Ovcar3 cells; however, protein levels were unchanged. Surprisingly, luciferase promoter assays revealed that none of the TZDs increased PPARγ activity. Overexpression of wild type PPARγ increased reporter activity. This was further augmented by TGZ, Rosi, and Pio indicating that these cells have the endogenous capacity to mediate PPARγ transactivation. To determine whether PPARγ mediates the TZD-induced decrease in proliferation, cells were treated with CGZ or TGZ in the absence or presence of a dominant negative (DN) or wild type overexpression PPARγ construct. Neither vector changed the TZD-mediated cell proliferation suggesting this effect of TZDs on ovarian cancer cells may be PPARγ independent. Conclusions CGZ and TGZ cause a decrease in ovarian cancer cell proliferation that is PPARγ independent. This concept is supported by the finding that a DN or overexpression of the wild type PPARγ did not affect the changes in cell proliferation and cell cycle.

OVARIAN CANCER: INVOLVEMENT OF THE MATRIX METALLOPROTEINASES

Ovarian cancer is the leading cause of death from gynecologic malignancies. One of the reasons for the high mortality rate associated with ovarian cancer is its late diagnosis, which often occurs after the cancer has metastasized throughout the peritoneal cavity. Cancer metastasis is facilitated by the remodeling of the extracellular tumor matrix by a family of proteolytic enzymes known as the matrix metalloproteinases (MMPs). There are 23 members of the MMP family, many of which have been reported to be associated with ovarian cancer. In the current paradigm, ovarian tumor cells and the surrounding stromal cells stimulate the synthesis and/or activation of various MMPs to aid in tumor growth, invasion, and eventual metastasis. The present review sheds light on the different MMPs in the various types of ovarian cancer and on their impact on the progression of this gynecologic malignancy.

Activation of the PKC pathway stimulates ovarian cancer cell proliferation, migration, and expression of MMP7 and MMP10.

ABSTRACT Postmenopausal women are at a higher risk of ovarian cancer due, in part, to increased levels of gonadotropins such as luteinizing hormone (LH). Gonadotropins and other stimuli are capable of activating two pathways, PKA and PKC, that are altered in ovarian cancer. To determine the role of LH on ovarian cancer, we explored the effects of human chorionic gonadotropin (hCG), an LH mimic, and an activator of the PKC pathway, phorbol-12-myristate 13-acetate (PMA), on ovarian cancer cell-cycle kinetics and apoptosis in Ovcar3 cells. PMA treatment increased cells in the S phase of the cell cycle and initially increased apoptosis after 4 h before diminishing apoptosis after 8 h. Treatment of ovarian cancer cells with hCG had no effect on these parameters. The PKC pathway is known to differentially regulate matrix metalloproteinase (MMP) expression. Results showed that ovarian cancer cells treated with PMA increased MMP7 and MMP10 mRNA levels after 8 h of treatment, and expression remained high after 12 h before decreasing at 24 h. The mRNA expression of extracellular matrix metalloproteinase inducer (BSG), an activator of MMPs, was unaffected by PMA. Due to the role that MMPs play in migration, we investigated the effect of PMA activation of MMPs on ovarian cancer cell migration. The use of the MMP inhibitor GM6001 blocked the increased migratory effects of PMA on ovarian cancer cells. Together, these studies show that activating the PKC pathway causes significant changes in cell cycle kinetics and selective expression of MMPs that are involved in enhancing ovarian cancer cell proliferation and migration.

Chemokine Ligand 20: A Signal for Leukocyte Recruitment During Human Ovulation?

Ovulation is one of the cornerstones of female fertility. Disruption of the ovulatory process results in infertility, which affects approximately 10% of couples. Using a unique model in which the dominant follicle is collected across the periovulatory period in women, we have identified a leukocyte chemoattractant, chemokine ligand 20 (CCL20), in the human ovary. CCL20 mRNA is massively induced after an in vivo human chorionic gonadotropin (hCG) stimulus in granulosa (>10 000-fold) and theca (>4000-fold) cells collected during the early ovulatory (12-18 h) and late ovulatory (18-34 h) periods after hCG administration. Because the LH surge sets in motion an inflammatory reaction characterized by an influx of leukocytes and CCL20 is known to recruit leukocytes in other systems, the composition of ovarian leukocytes (CD45+) containing the CCL20 receptor CCR6 was determined immediately prior to ovulation. CD45+/CCR6+ cells were primarily natural killer cells (41%) along with B cells (12%), T cells (11%), neutrophils (10%), and monocytes (9%). Importantly, exogenous CCL20 stimulated ovarian leukocyte migration 59% within 90 minutes. Due to the difficulties in obtaining human follicles, an in vitro model was developed using granulosa-lutein cells to explore CCL20 regulation. CCL20 expression increased 40-fold within 6 hours after hCG, was regulated partially by the epithelial growth factor pathway, and was positively correlated with progesterone production. These results demonstrate that hCG dramatically increases CCL20 expression in the human ovary, that ovarian leukocytes contain the CCL20 receptor, and that CCL20 stimulates leukocyte migration. Our findings raise the prospect that CCL20 may aid in the final ovulatory events and contribute to fertility in women.

Induction of proteinases in the human preovulatory follicle of the menstrual cycle by human chorionic gonadotropin

OBJECTIVE To explore the temporal expression in granulosa and theca cells of key members of the MMP and ADAMTS families across the periovulatory period in women to gain insight into their possible roles during ovulation and early luteinization. DESIGN Experimental prospective clinical study and laboratory-based investigation. SETTING University medical center and private IVF center. ANIMAL AND PATIENT(S) Thirty-eight premenopausal women undergoing surgery for tubal ligation and six premenopausal women undergoing assisted reproductive techniques. INTERVENTION(S) Administration of hCG and harvesting of follicles by laparoscopy and collection of granulosa-lutein cells at oocyte retrieval. MAIN OUTCOME MEASURE(S) Expression of mRNA for matrix metalloproteinase (MMPs) and the A disintegrin and metalloproteinase with thrombospondin-like motifs (ADAMTS) in human granulosa cells and theca cells collected across the periovulatory period of the menstrual cycle and in cultured granulosa-lutein cells after hCG. Localization of MMPs and ADAMTSs by immunohistochemistry. RESULT(S) Expression of MMP1 and MMP19 mRNA increased in both granulosa and theca cells after hCG administration. ADAMTS1 and ADAMTS9 mRNA increased in granulosa cells after hCG treatment, however, thecal cell expression for ADAMTS1 was unchanged, while ADAMTS9 expression was decreased. Expression of MMP8 and MMP13 mRNA was unchanged. Immunohistochemistry confirmed the localization of MMP1, MMP19, ADAMTS1, and ADAMTS9 to the granulosa and thecal cell layers. CONCLUSION(S) The collection of the dominant follicle throughout the periovulatory period has allowed the identification of proteolytic remodeling enzymes in the granulosa and theca compartments that may be critically involved in human ovulation. These proteinases may work in concert to regulate breakdown of the follicular wall and release of the oocyte.

Ovarian Membrane-Type Matrix Metalloproteinases: Induction of MMP14 and MMP16 During the Periovulatory Period in the Rat, Macaque, and Human

ABSTRACT An intrafollicular increase in proteolytic activity drives ovulatory events. Surprisingly, the periovulatory expression profile of the membrane-type matrix metalloproteinases (MT-MMPs), unique proteases anchored to the cell surface, has not been extensively examined. Expression profiles of the MT-MMPs were investigated in ovarian tissue from well-characterized rat and macaque periovulatory models and naturally cycling women across the periovulatory period. Among the six known MT-MMPs, mRNA expression of Mmp14, Mmp16, and Mmp25 was increased after human chorionic gonadotropin (hCG) administration in rats. In human granulosa cells, mRNA expression of MMP14 and MMP16 increased following hCG treatment. In contrast, mRNA levels of MMP16 and MMP25 in human theca cells were unchanged before ovulation but declined by the postovulatory stage. In macaque granulosa cells, hCG increased mRNA for MMP16 but not MMP14. Immunoblotting showed that protein levels of MMP14 and MMP16 in rats increased, similar to their mRNA expression. In macaque granulosa cells, only the active form of the MMP14 protein increased after hCG, unlike its mRNA or the proprotein. By immunohistochemistry, both MMP14 and MMP16 localized to the different ovarian cell types in rats and humans. Treatment with hCG resulted in intense immunoreactivity of MMP14 and MMP16 proteins in the granulosa and theca cells. The present study shows that MMP14 and MMP16 are increased by hCG administration in the ovulating follicle, demonstrating that these MMPs are conserved among rats, macaques, and humans. These findings suggest that MT-MMPs could have an important role in promoting ovulation and remodeling of the ovulated follicle into the corpus luteum.

Borderline Personality Disorder: An Overview of History, Diagnosis and Treatment in Adolescents

Borderline personality disorder (BPD) is a cluster B personality disorder. It is characterized by erratic behaviors, emotional instability and one of its hallmarks is self injurious behavior, which starts in adolescence. Patients with BPD are difficult to treat, most have a history of child sexual abuse, about a quarter present with sexual abuse from a caretaker. Although personality disorders are diagnosed only in adults, BPD manifests itself in adolescence in the form of uncontrollable anger, self mutilations, dissociation and other such behaviors. Hence, there is a growing number of scientists discussing the possibility of diagnosing BPD in adolescents. Here, we give an overview of the history and development of BPD in general; and in the adolescent population in particular. We also touch upon pharmacological and clinical interventions available for patients being diagnosed and/or possessing traits of BPD.

Identification of Hepsin and Protein Disulfide Isomerase A3 as Targets of Gelatinolytic Action in Rat Ovarian Granulosa Cells During the Periovulatory Period

The matrix metalloproteinase (MMP) family is believed to play a role in the ovulatory process because MMP inhibitors block oocyte release. However, little is known about the mechanisms by which the MMPs affect ovulation. The present study investigated the degradomic actions of the gelatinases, MMP2 and MMP9, by identifying gelatinolytic targets in periovulatory granulosa cells. Granulosa cells were collected from immature rats 48 h after equine chorionic gonadotropin treatment and were cultured with human chorionic gonadotropin (hCG) in the absence or presence of a specific MMP2/9 inhibitor ((2R)-2-[(4-biphenylylsulfonyl)amino]-3-phenylpropionic acid) for an additional 24 h. The conditioned media was analyzed for gelatinolytic activity, progesterone, and peptide profiles. Gelatinolytic activity and progesterone were induced in response to hCG; however, there was no difference in progesterone between cells treated with or without the inhibitor. Peptide fragments of proteins altered in the presence of the gelatinase inhibitor were identified by two-dimensional gel electrophoresis and mass spectrometry. Protein disulfide isomerase A3 (PDIA3), which plays a role in protein folding, was identified as a peptide that decreased in the presence of inhibitor while the serine protease hepsin, was found to increase with inhibitor treatment. Subsequent experiments established that PDIA3 and hepsin were targets of MMP2/9 action by cleavage with MMP2 and Western blot analysis, respectively. Additionally, hepsin was identified as a gelatinolytic target in ovarian cancer cells. In the present study, proteomics has identified proteins that may be involved in novel ways in the complex cascades that are mediated by gelatinolytic MMPs during the periovulatory period.

Trends in mental health of an adolescent medicine clinic patient population.

The correlation between exposure to forms of violence and development of mental disorders in victims is well established. The purpose of this paper was to identify mental health problems in an adolescent medicine clinic population in Lexington, KY and to investigate potential correlation of mental disorders with psychosocial factors. Data were gathered from the charts of 169 adolescent clinic patients (age 10-22) seen in the clinic for mental health care and analyzed using Excel. Of the patient population, 68% were urban, whereas 32% were rural. In terms of gender, 40% of the patients were male and 60% were female, 80% were white, 13% black, and 7% had other racial background(s). The most prevalent mental disorders in this group were depression 32.12%, 13% with generalized anxiety disorder, 8.2% with an attention deficit disorder (including ADHD), and 5.76% with an adjustment disorder. The abovementioned demographic trends showed that depression continues to be the most common mental health problem in this population regardless of gender, ethnic origin, or economic status. This finding highlights the need for availability of mental health support to this patient population. Further work is needed to spotlight the most significant psychosocial factors and root causes of mental health conditions in this age group.