Proteomic analysis of sperm proteins in infertile men with high levels of reactive oxygen species.

Oxidative stress is a significant risk factor for male infertility. A pro-oxidant testicular environment may alter the expression profile of functional sperm proteins and result in poor sperm quality. Patients and donors were divided into ROS (-) and ROS (+) groups. Using computational studies, and data mining of available literature on spermatozoa, oxidative stress and proteomics, we identified three core regulatory proteins angiotensin-converting enzyme (ACE), heat-shock protein (Hsp70) family A member 2 (HSPA2) and ribosomal protein subunit 27A (RPS27A) and seven interlink proteins NOS2, SUMO2, UBL4A, FBXO25, MAP3K3, APP and UBC. HSPA2 was validated by Western Blot, while the localisation of ACE, RPS27A, MAP3K3 and APP was identified by immunocytochemistry. The obtained results showed that HSPA2 was 1.2 (ROS+) and 2.1 (ROS-) fold downregulated in spermatozoa from patients with high levels of reactive oxygen species (ROS). ACE and APP were localised in the post-acrosomal region of spermatozoa, whereas RPS27A and MAP3K3 were localised either in the tail or sperm neck area. Our data show that these proteins may play a role in ROS-induced male infertility.

Diagnostic accuracy of haptoglobin within ovarian cyst fluid as a potential point-of-care test for epithelial ovarian cancer: an observational study.

OBJECTIVE: To investigate haptoglobin within ovarian cyst fluid (OCF) as a diagnostic biomarker for epithelial ovarian cancer (EOC) and develop an in vitro diagnostic point-of-care device test (IVDPCT) for use in the operating theatre. DESIGN: Retrospective and prospective cohort study. SETTING: South-East Asia. POPULATION: Women with suspicious ovarian cysts. METHODS: Proteomic, immunohistochemical and ELISA methods measured haptoglobin in OCF to differentiate benign and EOCs. Diagnostic performance of haptoglobin was compared with CA125, risk malignancy indices (RMI) and frozen section. Blinded validation of the IVDPCT was performed. MAIN OUTCOME MEASURES: Prediction of malignancy. RESULTS: Haptoglobin concentration measured by ELISA was 0.70 ± 0.09 mg/ml in patients with benign cysts (n = 87), 6.22 ± 0.53 mg/ml in early stage-EOC (n = 17), and 6.57 ± 0.65 mg/ml in late stage-EOC (n = 20). Haptoglobin in EOCs was significantly higher than in benign cysts (P < 0.0001). Haptoglobin using rapid colorimetric assay (RCA) on a training set had a sensitivity of 97.3% and a specificity 92.0%, comparable to ELISA and frozen sections. The haptoglobin AUROC curve was 0.999 (95% CI 0.997-1.000) compared with 0.895 (95% CI 0.814-0.977, P < 0.05) for CA125. Haptoglobin performed significantly better than all the RMIs (P < 0.01). Blinded validation studies showed a minor drop in average diagnostic performance (sensitivity 85.2% and specificity 90.5%) compared with the training set. However, when compared with frozen section, haptoglobin was no worse in diagnostic accuracy for malignancy. CONCLUSION: Haptoglobin was identified as a biomarker for the detection of EOC with potential as a point-of-care diagnostic tool. TWEETABLE ABSTRACT: Haptoglobin within ovarian cyst fluid: a biomarker for epithelial ovarian cancer and point-of-care diagnostics.

Development of quantitative-fluorescence polymerase chain reaction for the rapid prenatal diagnosis of common chromosomal aneuploidies in 1,000 samples in Singapore.

INTRODUCTION: We aimed to develop a rapid quantitative-fluorescence polymerase chain reaction (QF-PCR) to detect common foetal aneuploidies in the Singapore population within 48 hours of sample collection in order to alleviate parental anxiety. METHODS: DNA from 1,000 foetal samples (978 amniotic fluids, 14 chorion villi and eight foetal blood samples) was analysed using a QF-PCR of 19 microsatellite markers located on chromosomes 13, 18, 21, X and Y. A total of 523 samples were archived before the QF-PCR analysis (archived), while QF-PCR was performed and the results obtained within 48 hours of sample collection in the remaining 477 samples (live). The results were confirmed with their respective karyotypes. RESULTS: In total, 47 autosomal trisomies (T) were found: 30 among the archived (three T13, 12 T18, 15 T21) and 17 among the live (four T18, 13 T21) samples. The QF-PCR results were verified with their respective karyotypes. We achieved 100 percent sensitivity (lower 95 percent confidence interval [CI], 92.8 percent) and specificity (lower 95 percent CI, 99.5 percent), and the time taken from sample collection to the obtaining of results for the 477 live samples was less than 48 hours. CONCLUSION: Prenatal diagnostic results of common chromosomal abnormalities can be released within 48 hours of sample collection using QF-PCR. Parental anxiety is alleviated and clinical management is enhanced with this short waiting time.