Rapid detection and quantitation of neopterin and biopterin in clinical urine by wooden-tip electrospray ionization mass spectrometry.
A high throughput metabolite fingerprinting tool based on wooden-tip electrospray ionization mass spectrometry (WT-ESI-MS) has been established for the serum metabolic profiling study of endometriosis with little sample pre-treatment, no chromatography and instrument cycle times of less than 5 min. Serum samples from endometriosis patients and healthy controls were analyzed by direct WT-ESI-MS with a high resolution ESI-Q-TOF-MS. The resulting data were analyzed by multivariate data analysis. MS/MS experiments were carried out to identify potential biomarkers. Global metabolic profiling and subsequent multivariate analysis clearly distinguished endometriosis patients from healthy controls.A total of ten metabolites, up-regulated or down-regulated, were identified which contribute to the progress of endometriosis. These promising identified biomarkers underpin the metabolic pathway including steroid hormone biosynthesis, glycerophospholipid metabolism, sphingolipid metabolism, pyruvate metabolism, bile acid biosynthesis, and androgen and estrogen metabolisms. Considering that a much higher throughput can be obtained without a chromatographic step, the present WT-ESI-MS method could be developed as a fast prognostic or diagnostic method for endometriosis.
Endometriosis is a complex and heterogeneous pre-malignant inflammatory disease harboring multiple gene mutations. Previous studies have suggested that caspase recruitment domain family member (CARD)10 and CARD11 mutations may exist in endometriosis. In the present study, a collection of endometriotic lesions and paired peripheral blood from 101 patients with ovarian endometriosis were obtained, and the entire coding sequences of the CARD10 and CARD11 genes were sequenced. Evolutionary conservation analysis and online prediction programs were applied to analyze the disease-causing potential of the identified mutations. A total of 4 novel somatic mutations were identified in 4 out of the 101 (4.0%) samples: 2 in-frame deletions in CARD10 (c.785_790delAGGAGA, p.K272_E273delKE; c.785_802delAGGAGAAGGAGAAGGAGA, p.K272_V277delKEPDNV) and 2 heterozygous missense mutations in CARD11 (c.49G>T, p.D17Y; c.160G>C, p.E54Q). The sample with CARD10 p.K272_E273delKE deletion was obtained from a 47-year-old patient who was also diagnosed with uterine leiomyoma, while the CARD10 p.K272_V277delKEPDNV-mutated sample was from a 43-year-old patient exhibiting a decreased blood eosinophil granulocyte ratio (0.3%) and an elevated serum creatine kinase level (314 U/l). The patient with the CARD11 p.D17Y mutation was 38 years old and exhibited an increased level of cancer antigen 125 (45.4 U/ml), while the patient with the CARD11 p.E54Q mutation was 46 years old and exhibited no other gynecological conditions. Evolutionary conservation analysis and online prediction programs suggested that these mutations may be disease-causing. In summary, 4 novel somatic mutations in the CARD10 and CARD11 genes were identified from amongst 101 cases of ovarian endometriosis for the first time, these mutations may serve active roles in the development of ovarian endometriosis.
Abstract.Cancer is caused by multiple genetic alterations within cells. Recently, large-scale sequencing has identified frequent ribonuclease type III (DICER1), CCCTC-binding factor (CTCF), ribosomal protein L22 (RPL22), DNA (cytosine-5-)-methyltransferase 3α (DNMT3A), transformation/transcription domain-associated protein (TRRAP), isocitrate dehydrogenase (IDH)1 and IDH2 hotspot mutations in diverse types of cancer. However, it remains largely unknown whether these mutations also exist in ovarian carcinomas. In the present study, a collection of 251 patients with distinct subtypes of ovarian carcinomas were recruited and sequenced for the presence of these hotspot mutations. However, no mutations in the seven genes were detected in the samples. These negative results, together with certain recent reports, indicate that the hotspot mutations in the CTCF, RPL22, DNMT3A, TRRAP, IDH1 and IDH2 genes may not be actively involved in the carcinogenesis of ovarian carcinoma. Of note, the DICER1 mutation frequency in Sertoli-Leydig cell tumor in the present study was significantly lower compared to prior observation, and therefore, it is speculated that this discrepancy may be mainly due to the small sample size analyzed in the study. In addition, among these samples, frequent polymerase (DNA directed) ε, catalytic subunit (POLE1) and ring finger protein 43 (RNF43) mutations were identified in endometrioid and mucinous ovarian carcinomas, respectively; thus DICER1, CTCF, RPL22, DNMT3A, TRRAP, IDH1 and IDH2 hotspot mutations may not play synergistic roles with POLE1 or RNF43 mutations in the carcinogenesis of endometrioid or mucinous ovarian carcinomas. IntroductionThe current understanding of human malignancy is that it mainly arises due to the accumulation of multiple genetic alterations, transforming normal cells into malignant cells (1,2). Of these genetic alterations, a myriad of genomic mutation data derived from a high-throughput DNA sequencing technique provided a unique opportunity to profile the mutation spectra underlying human cancers and a large number of significant functional mutations in multiple genes were identified in diverse types of cancer (1,3,4). These genes can be defined as oncogenes or tumor suppressor genes and are being used as molecular markers for diagnosis, staging and prognosis of human cancers (5,6).Ovarian carcinoma constitutes a heterogeneous group of malignancies with significantly different clinical expression, pathological characteristics and genetic etiology (7,8). However, the majority of ovarian carcinomas shared certain common genetic alterations, such as frequent tumor protein p53 (TP53) and PIK3CA, catalytic subunit α mutations (9,10), and patients also exhibited subtype-specific mutations (11-13), which are possibly essential for the differential clinical expression and molecular-targeted therapy in ovarian carcinomas (14,15). These observations emphasized the requirement to identify novel subtype-specific molecular genetic aberrations in ovarian carcinomas.Recently, large-scal...
BackgroundGiven the important roles of the receptor-mediated lysophosphatidic acid (LPA) signaling in both reproductive tract function and gynecological cancers, it will be informative to investigate the potential role of LPA in the development of adenomyosis. The objective of this study was to evaluate the levels of LPA in plasma and the expression of six LPA receptors in the endometrial tissue collected from women with and without adenomyosis.MethodsPlasma and endometrial tissue samples were collected form women with and without adenomyosis. The levels of LPA in plasma were determined by using high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Immunohistochemistry was performed to evaluate the expression of six LPA receptors (LPA1–6) in endometrial tissue samples. The effects of LPA on IL-8 production, VEGF production and cell proliferation in human endometrial stromal cells (ESCs) were also assessed.ResultsLPA1 staining was localized to the cytoplasm, membrances of the epithelial cells of the endometrial glands, and there was little staining in the stromal cells. LPA2–5 staining were localized to the nuclei of stromal and glandular cells. Plasma levels of LPA were increased in adenomyosis. LPA1, LPA4 and LPA5 immunoreactivity were significantly higher in the adenomyosis group than in the control group, while LPA2 and LPA3 immunoreactivity were significantly lower in the adenomyosis group than in the control group. LPA6 was undetectable in the endometria. LPA induced the release of IL-8 from ESCs but did not affect cell proliferation and VEGF production.ConclusionThese results indicate that elevated plasma levels of LPA and aberrant expression of LPA receptors in the endometria may be associated with the development of adenomyosis.
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