Nabil Aziz

Pathogenic mechanisms in endometriosis-associated infertility

OBJECTIVE To review the mechanisms by which endometriosis may affect reproductive function. DESIGN Review of the English literature from 1986 to 2007 after searching Medline, EMBASE, Cochrane, and BIOSIS, as well as relevant meeting abstracts. SETTING Fertility research center and obstetrics and gynecology department in a tertiary care hospital. RESULT(S) There is compelling evidence in the literature that endometriosis has detrimental effects on ovarian and tubal function and uterine receptivity, resulting in female infertility. The mechanisms of infertility associated with endometriosis remain controversial and include abnormal folliculogenesis, elevated oxidative stress, altered immune function, and hormonal milieu in the follicular and peritoneal environments, and reduced endometrial receptivity. These factors lead to poor oocyte quality, impaired fertilization, and implantation. CONCLUSION(S) Through unraveling the mechanisms by which endometriosis leads to infertility, researchers are sure to find a nonsurgical means to diagnose endometriosis, most likely through serum and peritoneal markers. Cytokines, interleukins, oxidative stress markers, and soluble cellular adhesion molecules all show potential to be used as a reliable marker for diagnosing endometriosis. After analyzing the pathogenic mechanisms of endometriosis, it seems that the future treatment of this entity may include cyclo-oxygenase-2 inhibitors, immunomodulators, or hormonal suppressive therapy to eliminate the need for surgical treatment of endometriosis.

Critical Appraisal of World Health Organization's New Reference Values for Human Semen Characteristics and Effect on Diagnosis and Treatment of Subfertile Men

In 2010, the World Health Organization established new reference values for human semen characteristics that are markedly lower than those previously reported. Despite using controlled studies involving couples with a known time to pregnancy to establish the new limits, the reference studies are limited with regard to the population analyzed and the methods used for semen evaluation. The present review discusses concerns related to the new reference values for semen characteristics, including the effect on patient referral, diagnosis, and treatment of recognized conditions, such as varicocele, and on the indications for assisted reproductive technologies.

Evaluation of chemiluminescence and flow cytometry as tools in assessing production of hydrogen peroxide and superoxide anion in human spermatozoa

OBJECTIVE To examine simultaneously the levels of hydrogen peroxide (H(2)O(2)) and superoxide (O(2)(-*)) using chemiluminescence and flow cytometry. DESIGN Prospective laboratory study. SETTING Reproductive research lab in a tertiary hospital. PATIENT(S) Semen samples from 18 healthy male volunteers. INTERVENTION(S) Sperm preparation and measurement of reactive oxygen species (ROS) by chemiluminescence using luminol and lucigenin before and after H(2)O(2) exposure and by flow cytometry using dichlorofluorescin diacetate (DCFH-DA) for H(2)O(2) and dihydroethidium (DHE) for O(2)(-*). MAIN OUTCOME MEASURE(S) Sperm count, motility, viability, and ROS levels. RESULT(S) Immature sperm fractions showed significantly higher levels of ROS measured by either luminol or lucigenin compared with the neat and mature fraction. ROS levels were detectable by flow cytometry in chemiluminescence-negative samples. Both mature and immature sperm fractions had a significantly higher percentage of cells positive for H(2)O(2) compared with neat semen. On the other hand, the percentage of O(2)(-*)-positive cells in neat semen was significantly higher compared with the percentage found in mature fractions but significantly lower than that in the immature sperm fractions. CONCLUSION(S) We recommend ROS measurement by flow cytometry on the basis that it requires a lower sperm count, is comparable to chemiluminescence, and has higher specificity for intracellular ROS in viable spermatozoa. Samples tested negative by chemiluminescence still may have high intracellular H(2)O(2) generation that can be detected by flow cytometry.

Sperm chromosomal abnormalities are linked to sperm morphologic deformities

OBJECTIVE To describe the association between specific sperm morphologic abnormalities and sperm chromosomal abnormalities on multicolor interphase fluorescence in situ hybridization (FISH). DESIGN Case report. Reproductive medicine unit in a tertiary referral center. PATIENT(S) Three infertile men with severe oligoasthenospermia and total teratozoospermia who were referred for IVF treatment. MAIN OUTCOME MEASURE(S) Incidence of spermatozoal chromosomal aneuploidy for chromosome 18 and the sex chromosomes by using FISH. RESULT(S) Morphologic assessment of sperm revealed a high incidence of double heads, multinucleated sperm heads, and multiple tails. Hormone profiles and karyotyping of peripheral lymphocytes were normal in the three men. The proportion of sperm with disomy, trisomy and tetrasomy for chromosome 18, and the sex chromosomes in each patient was 100%, 76%, and 82.5%, respectively. CONCLUSION(S) Specific morphologic abnormalities of sperm may be associated with higher incidence of chromosomal abnormalities. Resolving infertility by offering patients in vitro fertilization/intracytoplasmic sperm injection must be approached with caution because of the significant risk for embryonic aneuploidy and chromosomal abnormalities in any subsequent offspring.

Sperm viability, apoptosis, and intracellular reactive oxygen species levels in human spermatozoa before and after induction of oxidative stress

OBJECTIVE To investigate sperm viability, incidence of apoptosis, and intracellular basal and induced reactive oxygen species (ROS) in sperm fractions. DESIGN Prospective controlled study. SETTING Center for Reproductive Medicine at a tertiary care hospital. METHOD(S) Liquefied seminal ejaculates (n = 12) prepared by density gradient centrifugation were reconstituted to 2 mL with phosphate-buffered saline. Oxidative stress was induced by hydrogen peroxide (H(2)O(2), 100 muM). Sperm viability, intracellular ROS, and incidence of apoptosis/necrosis in neat, immature, and mature sperm fractions were assessed. RESULT(S) Before H(2)O(2) exposure, mature spermatozoa fractions showed a significantly lower incidence of apoptotic sperm and intracellular O(2)(-*) levels but higher amounts of intracellular H(2)O(2) compared with neat semen. Higher levels of intracellular H(2)O(2) were demonstrated in immature sperm fractions compared with neat or mature fractions. In all sperm fractions, intracellular H(2)O(2) levels correlated with the intracellular concentration of O(2)(-*). After H(2)O(2) exposure, neat semen showed a significantly higher percentage of apoptosis compared with the prepared mature spermatozoa. However, no differences were observed in the incidence of apoptosis between immature and mature sperm fractions. CONCLUSION(S) There is a differential shift of both intracellular H(2)O(2) and O(2)(-*) in each sperm fraction that may affect sperm quality. Sperm apoptosis is related to intracellular H(2)O(2) levels, which in turn are affected by intracellular O(-*) levels. Oxidative stress was not associated with an increased incidence of apoptosis in immature or mature sperm fractions.

Lipid peroxidation and antioxidant status in preeclampsia: a systematic review.

Background: Preeclampsia is characterized by increased lipid peroxidation and diminished antioxidant capacity; however, there is no consensus as to the extent of these conditions. Objective: To assess the association of lipid peroxidation and antioxidant status with preeclampsia quantitatively using meta-analysis. Design: Systematic review and meta-analysis. Search Strategy: Studies were identified by performing an extensive search using BIOSIS (1986–2007), EMBASE (1986–2007), Medline (1986–2007), and the Cochrane database. Data Analysis: Standardized mean differences (SMD) with 95% confidence intervals (CI) were used in the meta-analysis and sources of heterogeneity were examined. Main Results: In the included studies, the overall SMD was a 1.21 nmol/mL increase in serum malondialdehyde in preeclampsia cases compared to controls (95% CI: 0.76, 1.66). Overall, total serum thiobarbituric acid-reactive substances SMD were 1.62 nmol/mL greater in cases than in controls (95% CI: 0.27, 2.96). The overall estimate SMD for serum vitamin E was 1.12 nmol/mL less in cases than controls (95% CI: −1.77, −0.48) and vitamin C SMD overall estimate was −0.53 (95%CI: −1.03, −0.02), significantly lower in cases compared with controls. The overall SMD for erythrocyte superoxide dismutase was −2.37 (95% CI: −4.76, 0.03), a marginally significant decrease in cases versus controls. Conclusions: Established preeclampsia is associated with increased concentrations of oxidative stress markers including lipid peroxidation products, and a reduction in antioxidant concentrations. Target Audience: Obstetricians & Gynecologists, Family Physicians Learning Objectives: After completion of this educational activity, the participant should be better able to describe the pattern of oxidative stress markers associated with preeclampsia, and interpret the available literature as it relates to oxidative stress and preeclampsia.

The sperm deformity index: a reliable predictor of the outcome of oocyte fertilization in vitro *

OBJECTIVE To evaluate a novel expression of sperm morphological parameters, the sperm deformity index, as a predictor of fertilization in vitro. DESIGN Prospective blind clinical trial. SETTING Academic tertiary referral center. INTERVENTION(S) Detailed sperm morphological assessment applying strict morphological criteria and a multiple entry technique for an unselected male population undergoing IVF. The sperm deformity index, defined as the average number of deformities per sperm assessed, was calculated. PATIENT(S) One hundred fifty-eight patients undergoing IVF treatment. Females with conditions negatively influencing fertilization were excluded. MAIN OUTCOME MEASURE(S) Fertilization rates and pregnancy. RESULT(S) Seventy-three percent of patients achieved fertilization. Patients achieving fertilization had a significantly higher median proportion of normal forms and a significantly lower median sperm deformity index than the nonfertilizers. The receiver operator characteristic (ROC) curves identified cutoff points that maximized the sum of sensitivity and specificity at sperm deformity index 1.6 and normal forms 4%. The sperm deformity index had a greater sensitivity (96%), specificity (72%), positive predictive value (90%), and negative predictive value (86%) than the proportion of normal sperm morphology (87%, 69%, 89%, 66%, respectively) at the optimal cutoff points. The area under the ROC curve was greater for the sperm deformity index (0.875) than for the proportion of normal sperm morphology (0.622). Achieving pregnancy did not correlated with sperm morphology. CONCLUSION(S) The sperm deformity index is a more reliable predictor of the outcome of fertilization in vitro than the proportion of normal sperm morphology and can assist to identify patients who require techniques such as intracytoplasmic sperm injection.

The impact of peritoneal fluid from healthy women and from women with endometriosis on sperm DNA and its relationship to the sperm deformity index

OBJECTIVE To study the effect of peritoneal fluid (PF) from healthy women and from women with endometriosis on sperm DNA and its relationship to sperm morphology as assessed by the sperm deformity index (SDI). DESIGN Experimental study. SETTING Research laboratory at an academic hospital. PATIENT(S) Healthy women undergoing laparoscopic tubal ligation and women with endometriosis. INTERVENTION(S) Aliquots of prepared sperm from 10 healthy donors were incubated with PF from healthy women undergoing laparoscopic tubal ligation (treatment 1, n = 10), with PF from patients with endometriosis (treatment 2, n = 10), and with human tubal fluid media with 10% bovine serum albumin (control, n = 10). MAIN OUTCOME MEASURE(S) Sperm DNA fragmentation was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay and SDI. RESULT(S) There was a significant increase in sperm DNA damage at 24 hours compared with at 1.5 and 3 hours. The SDI scores significantly correlated with sperm DNA damage after 1.5 and 24 hours of incubation in all aliquots. Percentage normal morphology did not correlate with DNA damage. CONCLUSION(S) In vitro exposure of sperm to PF from patients with endometriosis is associated with significantly increased DNA damage. There is evidence of interdependence between the sperm morphology as assessed by SDI scores and DNA damage. The significant increase in sperm DNA damage observed after 24 hours of incubation may be clinically relevant.

Role of Oxidative Stress in Polycystic Ovary Syndrome

Polycystic ovary syndrome (PCOS) is a multifactorial disorder affecting many women of reproductive age, typically due to hyperandrogenemia, hyperinsulinemia, and enigmatic genetic factors. The complex nature of PCOS is reflected in the broad spectrum of the disorders clinical presentation, including metabolic and reproductive disorders. As a result, while the European Society for Human Reproduction and Embryology and the American Society for Reproduc- tive Medicine (ESHRE/ASRM) have agreed on a consensus definition of PCOS to help clinical investigators, the condi- tion is recognized to have multiple clinical phenotypes. Oxidative stress (OS) occurs when destructive reactive oxygen species (ROS) outbalance antioxidants, causing DNA damage and/or cell apoptosis. Moreover, reactive nitrogen species (RNS), such as nitrogen oxide (NO) with an unpaired electron also are highly reactive and toxic. In a quest to delineate the role of OS in the pathogenesis of PCOS, investiga- tors have examined patients with the disorder for a wide array of OS biomarkers, including malondialdehyde (MDA), protein carbonyl, total antioxidant capacity (TAC), superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione (GSH).

Human sperm head morphometric distribution and its influence on human fertility

OBJECTIVE To study the distribution of live sperm head size in semen and sperm preparations as a predictor of fertility. DESIGN Prospective blind clinical trial. SETTING Academic tertiary referral center. PATIENT(S) One hundred fifty-five patients undergoing IVF treatment. Females with conditions negatively influencing fertilization were excluded. INTERVENTION(S) Morphometric analysis (head area, major axis, minor axis, and elongation ratio) of video images of sperm in semen and swim-up preparations used for IVF treatment was performed with a Hamilton-Thorne analyzer V 8.1 (Hamilton-Thorn Research, Beverly, MA). MAIN OUTCOME MEASURE(S) Oocyte fertilization. RESULT(S) Seventy-four percent of patients achieved fertilization. Fertilizers and nonfertilizers had different sperm head area distribution. The fertilizers had a significantly smaller interquartile range of sperm head area and of major axis in both semen and sperm preparation compared with the nonfertilizers. A subgroup of men who had fathered a child naturally had a more uniform sperm head area in semen with a significantly smaller median compared with those who failed to father a child naturally with their healthy female partner. We used multiple logistic regression applying forward stepwise selection of variables in building three predictive models of probability of fertilization. CONCLUSION(S) Successful IVF or history of fathering a child was associated with a more uniform sperm head area in semen and sperm preparation.