Dimercapto-1-propanesulfonic acid (DMPS) induces metaphase II mouse oocyte deterioration.

In light of the recent lead contamination of the water in Flint, Michigan and its potential adverse outcomes, much research and media attention has turned towards the safety profile of commonly used chelators. Dimercapto-1-propanesulfonic acid (DMPS) typically used in the treatment of lead, mercury and arsenic poisoning also displays a high affinity towards transition metals such as zinc and copper, essential for biological functioning. It is given in series of dosages (0.2-0.4g/day) over a long period, and has the ability to enter cells. In this work, we investigated the mechanism through which increasing concentrations of DMPS alter oocyte quality as judged by changes in microtubule morphology (MT) and chromosomal alignment (CH) of metaphase II mice oocyte. The oocytes were directly exposed to increasing concentration of DMPS (10, 25, 50, 100 and 300µM) for four hours (time of peak plasma concentration after administration) and reactive oxygen species (mainly hydroxyl radical and superoxide) and zinc content were measured. This data showed DMPS plays an important role in deterioration of oocyte quality through a mechanism involving zinc deficiency and enhancement of reactive oxygen species a major contributor to oocyte damage. Our current work, for the first time, demonstrates the possibility of DMPS to negatively impact fertility. This finding can not only help in counseling reproductive age patients undergoing such treatment but also in the development of potential therapies to alleviate oxidative damage and preserve fertility in people receiving heavy metal chelators.

Sonographic Measurement of Cervical Volume in Pregnant Women at High Risk of Preterm Birth Using a Geometric Formula for a Frustum Versus 3-Dimensional Automated Virtual Organ Computer-Aided Analysis.

OBJECTIVES: To compare cervical volume measurements by 3-dimensional (3D) sonography using Virtual Organ computer-aided analysis (VOCAL; GE Healthcare, Milwaukee, WI) versus a manual method using a geometric formula for a frustum. METHODS: We included 142 asymptomatic pregnant women at 16 to 24 weeks gestation at high risk for preterm birth. With a Voluson 730 Expert system (GE Healthcare), they underwent 2-dimensional (2D) transvaginal sonographic cervical length measurements and 3D cervical volume acquisition. The stored volumes were processed by VOCAL on a surface tablet. Cervical volume was manually calculated from the 2D images by using the formula V = 1/3 × π × h × (r12 + r22 + r1 × r2), where V represents cervical volume; π was approximated as 3.14159; h, cervical length; r1, radius at the internal os; and r2, radius at the external os. RESULTS: Cervical volume was lower when obtained manually than by VOCAL, with a coefficient of variation of 30%, a mean difference of 10.1 ± 14.9 cm3 (P < .0001), and a poor interclass correlation coefficient of 0.62 (95% confidence interval [CI], 0.31 to 0.78). Both methods had good reproducibility; however, VOCAL had wider limits of agreement. A positive correlation was found between both methods (r = 0.63; P < .0001). No correlation was found between cervical length by 2D transvaginal ultrasound and cervical volume by the VOCAL technique (r = 0.06; 95% CI, -0.10 to 0.22) or cervical volume by the manual method (r = 0.2; 95% CI, 0.08 to 0.39). CONCLUSIONS: The cervix represents a frustum (truncated cone, r1 is not equal to r2) in shape rather than a cylinder. Both methods are reproducible; VOCAL is less reliable but provides higher values of cervical volume.

Mesna (2-mercaptoethane sodium sulfonate) functions as a regulator of myeloperoxidase.

Myeloperoxidase (MPO), an abundant protein in neutrophils, monocytes, and macrophages, is thought to play a critical role in the pathogenesis of various disorders ranging from cardiovascular diseases to cancer. We show that mesna (2-mercaptoethanesulfonic acid sodium salt), a detoxifying agent, which inhibits side effects of oxazaphosphorine chemotherapy, functions as a potent inhibitor of MPO; modulating its catalytic activity and function. Using rapid kinetic methods, we examined the interactions of mesna with MPO compounds I and II and ferric forms in the presence and absence of chloride (Cl-), the preferred substrate of MPO. Our results suggest that low mesna concentrations dramatically influenced the build-up, duration, and decay of steady-state levels of Compound I and Compound II, which is the rate-limiting intermediate in the classic peroxidase cycle. Whereas, higher mesna concentrations facilitate the porphyrin-to-adjacent amino acid electron transfer allowing the formation of an unstable transient intermediate, Compound I*, that displays a characteristic spectrum similar to Compound I. In the absence of plasma level of chloride, mesna not only accelerated the formation and decay of Compound II but also reduced its stability in a dose depend manner. Mesna competes with Cl-, inhibiting MPO's chlorinating activity with an IC50 of 5µM, and switches the reaction from a 2e- to a 1e- pathway allowing the enzyme to function only with catalase-like activity. A kinetic model which shows the dual regulation through which mesna interacts with MPO and regulates its downstream inflammatory pathways is presented further validating the repurposing of mesna as an anti-inflammatory drug.

Cyclophosphamide and acrolein induced oxidative stress leading to deterioration of metaphase II mouse oocyte quality.

Cyclophosphamide (CTX) is a chemotherapeutic agent widely used to treat ovarian, breast, and hematological cancers as well as autoimmune disorders. Such chemotherapy is associated with reproductive failure and premature ovarian insufficiency. The mechanism by which CTX and/or its main metabolite, acrolein, affect female fertility remains unclear, but it is thought to be caused by an overproduction of reactive oxygen species (ROS). Here, we investigated the effect of CTX on metaphase II mouse oocytes obtained from treated animals (120mg/kg, 24h of single treatment), and oocytes directly exposed to increasing concentrations of CTX and acrolein (n=480; 0, 5, 10, 25, 50, and 100µM) with and without cumulus cells (CCs) for 45min which correlates to the time of maximum peak plasma concentrations after administration. Oocytes were fixed and subjected to indirect immunofluorescence and were scored based on microtubule spindle structure (MT) and chromosomal alignment (CH). Generation of ROS was evaluated using the Cellular Reactive Oxygen Species Detection Assay Kit. Deterioration of oocyte quality was noted when oocytes were obtained from CTX treated mice along with CTX and acrolein treated oocytes in a dose-dependent manner as shown by an increase in poor scores. Acrolein had an impact at a significantly lower level as compared to CTX, plateau at 10µM versus 50µM, respectively. These variation is are associated with the higher amount of ROS generated with acrolein exposure as compared to CTX (p<0.05). Utilization of antioxidant therapy and acrolein scavengers may mitigate the damaging effects of these compounds and help women undergoing such treatment.

Galactose and its Metabolites Deteriorate Metaphase II Mouse Oocyte Quality and Subsequent Embryo Development by Disrupting the Spindle Structure.

Premature ovarian insufficiency (POI) is a frequent long-term complication of classic galactosemia. The majority of women with this disorder develop POI, however rare spontaneous pregnancies have been reported. Here, we evaluate the effect of D-galactose and its metabolites, galactitol and galactose 1-phosphate, on oocyte quality as well as embryo development to elucidate the mechanism through which these compounds mediate oocyte deterioration. Metaphase II mouse oocytes (n = 240), with and without cumulus cells (CCs), were exposed for 4 hours to D-galactose (2 µM), galactitol (11 µM) and galactose 1-phosphate (0.1 mM), (corresponding to plasma concentrations in patients on galactose-restricted diet) and compared to controls. The treated oocytes showed decreased quality as a function of significant enhancement in production of reactive oxygen species (ROS) when compared to controls. The presence of CCs offered no protection, as elevated ROS was accompanied by increased apoptosis of CCs. Our results suggested that D-galactose and its metabolites disturbed the spindle structure and chromosomal alignment, which was associated with significant decline in oocyte cleavage and blastocyst development after in-vitro fertilization. The results provide insight into prevention and treatment strategies that may be used to extend the window of fertility in these patients.