Effects of Progesterone, Dydrogesterone and Estrogen on the Production of Th1/Th2/Th17 Cytokines by Lymphocytes from Women with Recurrent Spontaneous Miscarriage.

Anti-inflammatory Th2 cytokines have been shown to be associated with healthy, successful pregnancy while pro-inflammatory Th1 and Th17 cytokines are associated with pregnancy loss due to recurrent spontaneous miscarriage. This nexus between unexplained recurrent spontaneous miscarriage (uRSM) and maternal inflammatory has led to the possibility of using pregnancy-related hormones to modify the maternal cytokine bias in a manner that is conducive to successful pregnancy. We investigated the ability of progesterone, dydrogesterone and estrogen to modulate cytokine production by peripheral blood lymphocytes from women undergoing uRSM. Peripheral blood mononuclear cells (PBMC) from females with uRSM were stimulated in vitro with phytohemagglutinin (PHA) in the presence and absence of progesterone or dydrogesterone or 17ß-estradiol. Culture supernatants were assayed for IFN-α, TNF-γ, IL-2, IL-6, IL-10, IL-13, IL-17A, and IL-23 by ELISA. Progesterone and dydrogesterone significantly down-regulated the secretion of the Th1 cytokines IFN-α and TNF-γ, and the Th17 cytokine IL-17A, and IL-23. Additionally, the secretion of the Th2 cytokine IL-6 was up-regulated. Estrogen, on the other hand, decreased the production of IFN-α and IL-2, increased the production of IL-6 but did not affect IL-17A and IL-23 secretion. Progestogens and estrogen can decrease the production of some Th1/Th17 inflammatory cytokines secreted by lymphocytes from uRSM and upregulate the production of anti-inflammatory cytokines. These data support the notion that progestogens can be used for altering maternal cytokine profiles to manage pregnancy complications.

Redirection of cytokine production by lymphocytes from women with pre-term delivery by dydrogesterone.

PROBLEM: To study the ability of dydrogesterone to modulate the production of pro-inflammatory and anti-inflammatory cytokines by lymphocytes from women undergoing pre-term delivery (PTD). METHOD OF STUDY: Peripheral blood mononuclear cells (PBMC) from 18 subjects undergoing PTD were stimulated with the mitogen phytohemagglutinin in the presence and absence of progesterone and dydrogesterone. The levels of interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, interleukin (IL)-4, and IL-10 in culture supernatants were then estimated by enzyme-linked immunoabsorbant assay. Cytokine production in the presence and absence of progesterone and dydrogesterone were compared. RESULTS: The exposure of PBMC to dydrogesterone resulted in a significant inhibition in the production of the pro-inflammatory cytokines IFN-gamma and TNF-alpha and a significant increase in the levels of the anti-inflammatory cytokine IL-4, resulting in a substantial shift in the ratio of Th1/Th2 cytokines. CONCLUSION: Dydrogesterone induces a shift in cytokine bias, by inhibiting pro-inflammatory cytokine production and increasing anti-inflammatory cytokine production.

Modulation of cytokine production by dydrogesterone in lymphocytes from women with recurrent miscarriage.

OBJECTIVE: To examine the effects of dydrogesterone on the production of Th1 and Th2 cytokines by lymphocytes from women undergoing unexplained recurrent spontaneous miscarriage (RSM). DESIGN: Controlled prospective, clinical study conducted in a maternity hospital and a university-based immunology laboratory. SETTING: Faculty of Medicine, Kuwait University and Kuwait Maternity Hospital. SAMPLE: Thirty women with unexplained RSM. METHODS: Peripheral blood mononuclear cells (PBMC) from women with unexplained RSM were isolated from venous blood by density gradient sedimentation and stimulated with phytohaemagglutinin (PHA). Culture supernatants assayed for interferon (IFN)-gamma, tumour necrosis factor (TNF)-alpha, interleukin (IL)-4, IL-6 and IL-10 by ELISA. Levels of the progesterone-induced blocking factor (PIBF) were also measured. MAIN OUTCOME MEASURES: Cytokine production in the presence and absence of progesterone and dydrogesterone. RESULTS: Dydrogesterone significantly inhibited the production of the Th1 cytokines IFN-gamma (P= 0.0001) and TNF-alpha (P= 0.005) and induced an increase in the levels of the Th2 cytokines IL-4 (P= 0.03) and IL-6 (P= 0.017) resulting in a substantial shift in the ratio of Th1/Th2 cytokines. The effect of dydrogesterone was blocked by the addition of the progesterone-receptor antagonist mifepristone, indicating that dydrogesterone was acting via the progesterone receptor. Dydrogesterone induced the production of PIBF. CONCLUSION: Dydrogesterone inhibits the production of the Th1 cytokines IFN-gamma and TNF-alpha from lymphocytes and up-regulates the production of the Th2 cytokines IL-4 and IL-6, inducing a Th1 to Th2 cytokine shift.

Maternal cytokine production patterns in women with pre-eclampsia.

PROBLEM: To determine the levels of cytokines produced upon mitogenic or antigenic stimulation of maternal peripheral blood mononuclear cells (PBMC) from women with pre-eclampsia. METHOD OF STUDY: PBMC from 54 women with a history of successful pregnancy and 32 women undergoing pre-eclamptic delivery were stimulated with a mitogen or with autologous placental cells or with trophoblast antigens, and the levels of cytokines released into the culture supernatants then assessed by enzyme-linked immunosorbent assay. RESULTS: Significantly higher levels of the Th1 cytokines, interferon-gamma, and tumor necrosis factor-alpha were produced by the pre-eclamptic group than by the normal pregnancy group, which on the contrary showed significantly greater production of the Th2 cytokines, interleukin (IL)-4, IL-5, IL-6 and IL-10. A comparison of the ratios of Th2 to Th1 cytokines indicates a higher Th1 cytokine bias in pre-eclampsia as compared with normal pregnancy. CONCLUSIONS: These data are suggestive of a maternal pro-inflammatory cytokine bias in pre-eclampsia.

Maternal-fetal status of copper, iron, molybdenum, selenium and zinc in insulin-dependent diabetic pregnancies.

OBJECTIVE: The objective was to assess the status of essential trace elements such as copper, iron, molybdenum, selenium and zinc in insulin-dependent diabetic pregnancies at term and to compare the data with a control group. Fetal-maternal ratios of the elements and copper:zinc ratio were also computed in the control and study populations. METHODOLOGY: Samples from maternal vein, umbilical artery and umbilical vein of diabetic and control women were collected at the time of spontaneous delivery or cesarean section and activities of trace elements evaluated by atomic absorption spectrophotometry. RESULTS: Cu, Fe, Mo, Se and Zn concentrations in maternal venous blood averaged 2,156, 2,020, 13, 102 and 656 microg/l in control women (n=17) while in the diabetic group (n=14), the corresponding values for the trace elements averaged 3,135, 3,675, 15, 85 and 628 microg/l respectively. Values for copper and molybdenum were significantly higher (p<0.05) in the study group compared to control while those of zinc, iron and selenium were not significantly different (p>0.05). Iron and molybdenum values were significantly higher (p<0.05) and that of zinc significantly lower (p<0.05) in umbilical arterial samples of diabetic group compared to controls. In the case of molybdenum, copper the values were significantly higher (p<0.05) in umbilical venous samples of diabetic group compared to that of control. Significant differences in Cu:Zn ratio of maternal venous and umbilical samples and fetal-maternal ratios of some elements were noted between control and study group as well. CONCLUSION: We speculate that altered status of some essential trace elements and altered antioxidant mineral ratio observed in insulin dependent diabetic patients could have deleterious influences on the health of the mother as well as the fetus and newborn.

Impaired gestational glucose tolerance. Its effect on placental pathology.

OBJECTIVE: The aim of this study was to investigate several macroscopic and microscopic features of placenta in cases with impaired gestational glucose tolerance. METHODS: Seventy-five gm World Health Organization criteria for the diagnosis of gestational diabetes and impaired gestational glucose tolerance were followed during the period June 1999 through to June 2000, at the Maternity Hospital of Kuwait. Macroscopic and microscopic examinations of 95 placentas were carried out. Sixty-five were from the control patients and 30 were from cases with impaired gestational glucose tolerance. RESULTS: Mean maternal age, maternal weight and parity was significantly higher in the impaired gestational glucose tolerance (IGGT) group compared to the control group. Mean birth weight of the baby was significantly higher in the IGGT group compared to the control group. Mean placental weight and the percentage of the cesarean delivery was higher in the IGGT group but did not reach the level of significance. There was no significant association between the microscopic features of the placenta in the control and IGGT groups. CONCLUSION: Impaired gestational glucose tolerance is related to increased neonatal and placental weight, which may lead to a higher number of cesarean deliveries, stressing the similarity between impaired gestational glucose tolerance and gestational diabetes mellitus. More stringent criteria may be necessary to define gestational diabetes. Microscopic features of placenta both in the control and IGGT groups did not show any significant difference.

Increased expression of pro-inflammatory cytokines in placentas of women undergoing spontaneous preterm delivery or premature rupture of membranes.

PROBLEM: The objective of this study was to determine the levels of cytokines in the placentas of women undergoing preterm delivery (PTD) or premature rupture of membranes (PROM) as compared with women undergoing normal delivery at term. METHOD OF STUDY: Placentas were obtained from 30 subjects with spontaneous PTD, 30 women with PROM and 30 women with a history of normal delivery at term. Levels of interleukin (IL)-2, interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, TNF-beta, IL-4, IL-5, IL-6 and IL-10 and IL-12 were estimated by ELISA in detergent lysates of placentas from the subjects. RESULTS: We found significantly increased levels of the Th1 cytokines IL-2 and IFN gamma and of the Th1-inducing cytokine IL-12 in placentas from the PTD and PROM groups as compared with those delivering at term. In contrast, the levels of the Th2 cytokines IL-4, IL-6 and IL-10 were significantly higher in placentas from term pregnancy. CONCLUSIONS: These data support our observation of a pro-inflammatory cytokine bias in women with PTD and PROM.

Measurement of inhibin A and inhibin pro-alphaC in early human pregnancy and their role in the prediction of pregnancy outcome in patients with recurrent pregnancy loss.

OBJECTIVE: To examine the temporal relationship among inhibin A, beta-hCG, and pro-alphaC in early pregnancy and to determine whether the measurement of these hormones has any role in prediction of pregnancy outcome in patients with recurrent spontaneous miscarriage. DESIGN: Prospective descriptive study. SETTING: A tertiary referral center for recurrent miscarriage. PATIENT(S): Thirty-six pregnant women with previous history of recurrent spontaneous pregnancy loss. INTERVENTION(S): Serial blood samples were collected prospectively at 6, 8, 10, and 12 weeks of gestation and were analyzed for inhibin A and inhibin pro-alphac using a two-site enzyme-linked immunosorbent assay as well as for beta-hCG using the microparticle enzyme immunoassay. MAIN OUTCOME MEASURE(S): Serum levels of inhibin A, inhibin pro-alphac, and beta-hCG. RESULT(S): The patients were allocated to two groups according to the pregnancy outcome: group 1 consisted of patients whose pregnancy continued beyond 20 weeks (control group); and group 2 consisted of patients who spontaneously aborted (aborted group). There was a significant difference in inhibin A concentrations between the control and aborted groups at 8, 10, and 12 weeks' gestation. Significant differences in beta-hCG concentrations between the two groups is evident only at 10 weeks' gestation. There were no significant differences in inhibin pro-alphac concentrations between the two groups at any gestational age. Assessment of the trend in the control group over the study period showed a significant increase in inhibin A and beta-hCG but not inhibin pro-alphac levels. CONCLUSION(S): Low serum levels of inhibin A at early gestational age in pregnancies destined to miscarry suggest a role for this glycoprotein as a marker for early pregnancy viability. Its measurement at the time of the first pregnancy test might be able to predict pregnancy outcome.

Differential expression of T-helper cytokines in the peritoneal fluid of women with normal ovarian cycle compared with women with chronic anovulation.

BACKGROUND: Estrogen and progesterone immunoregulate the genital environment by expression of cytokines and growth factors. OBJECTIVE: To investigate the pattern of expression of T-helper cytokines during the ovarian cycle compared with women with chronic anovulation resistant to clomiphene citrate. HYPOTHESIS: Expression of T-helper cytokines in women with chronic anovulation may be different from the pattern in women with a normal ovarian cycle. METHODS: We evaluated 31 infertile women having laparoscopy for evaluation of tubal patency and evidence of ovulation in two groups during (a) the luteal phase (17 women) and (b) the follicular phase (14 women). A third group was composed of 14 women with polycystic ovarian syndrome, but they were resistant to clomiphene citrate for induction of ovulation and had laparoscopic ovarian cautery. Peritoneal fluid was collected during laparoscopy. Estimation of T-helper cytokine interleukin (IL)-2, tumor necrosis factor (TNF)-alpha, IL-4 and IL-6 in serum, peritoneal fluid and culture of the peritoneal mononuclear cells was performed by ELISA. Serum luteinizing hormone (LH), follicle stimulating hormone (FSH), prolactin, estradiol and progesterone were evaluated by the Vidas Parametric System. RESULTS: The LH : FSH ratio was significantly higher in the women with polycystic ovaries than in the ovulatory groups. IL-2 and IFN-gamma were more highly expressed in the follicular phase but the T-helper 2 cytokines IL-4 and IL-6 predominated in the luteal phase, serum, peritoneal fluid and culture of the peritoneal mononuclear cells. From the follicular to the mid-luteal phase, IL-6 increased three to fivefold in the serum and peritoneal fluid, but there was low expression with anovulation. CONCLUSIONS: The peritoneal fluid levels of IL-4 and IL-6 are higher in the luteal phase. Low IL-6 levels in chronic anovulation may be a marker of resistance to clomiphene citrate.

Percutaneous testicular sperm aspiration and intracytoplasmic sperm injection in obstructive and non-obstructive azoospermia: an easy alternative to TESE and MESA.

OBJECTIVE: To evaluate the recovery rate of sperm from the testis using percutaneous testicular aspiration with a 22-gauge hypodermic needle followed by evaluation of the fertilization rate and pregnancy rate after intracytoplasmic sperm injection. MATERIALS AND METHODS: This is a prospective observational study performed in a private in vitro fertilization setting in Kuwait. Fifteen patients with obstructive and non-obstructive azoospermia were included in the study. Thirteen of them had previous microepididymal sperm aspiration, percutaneous epididymal sperm aspiration or testicular sperm extraction. The sperm were retrieved using percutaneous testicular aspiration under local analgesia. This was followed by intracytoplasmic sperm injection. A total of 146 eggs were collected and 112 were injected. RESULTS: Normal fertilization occurred in 91 oocytes (87.5%) and the total number of embryos cleaved was 83 (91%). Embryo transfer was performed in 13 with pregnancy rate of 33.3 per treatment cycle and 38.5 per embryo transfer. Failure to retrieve sperm was encountered in 2 cases both in the hypospermatogenesis group. CONCLUSIONS: Percutaneous testicular sperm aspiration using hypodermic needles under local analgesic is an easy and cheap method with high patient acceptability, minimal complications and no need of special training. In this small group, it seems to have an acceptable success rate in terms of sperm retrieval and pregnancy in the obstructive type as well as hypospermatogenesis, but to lesser extent.