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- Kushnir, Mark M., et al.
(författare)
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High-Sensitivity Tandem Mass Spectrometry Assay for Serum Estrone and Estradiol
- 2008
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Ingår i: American Journal of Clinical Pathology. - 0002-9173 .- 1943-7722. ; 129:4, s. 530-539
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Tidskriftsartikel (refereegranskat)abstract
- High-sensitivity measurement of serum estrogens is important in adult and pediatric endocrinology and oncology. We developed a high-sensitivity liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for simultaneous measurement of estrone (E-1) and estradiol (E-2). Aliquots of 200 mu L of serum were spiked with internal standard, extracted, derivatized with dansyl chloride, and analyzed by LC-MS/MS using 2-dimensional chromatographic separation. Total imprecision for the method was less than 11%; the limit of quantitation was 1 pg/mL. Reference intervals were established with samples from more than 900 healthy postmenopausal women, men, girls, and boys. Concentrations of estrogens in children reached adult levels by Tanner stage 3. In men and postmenopausal women, the median concentrations of total estrogens (E-1 + E-2) were 39 and 22 pg/mL, and the median E-2/E-1 ratios were 0.98 and 0.55, respectively. The method requires a small sample volume and has adequate sensitivity and specificity for analyzing estrogens in samples from postmenopausal women, men, and children.
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| 6. |
- Kushnir, Mark M., et al.
(författare)
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Liquid chromatography tandem mass spectrometry for analysis of steroids in clinical laboratories
- 2011
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Ingår i: Clinical Biochemistry. - : Elsevier BV. - 0009-9120 .- 1873-2933. ; 44:1, s. 77-88
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Forskningsöversikt (refereegranskat)abstract
- Liquid chromatography tandem mass spectrometry is one of the most specific techniques available in clinical laboratories. In the past, immunoassays were the primary methodology for analysis of steroids in biological samples because they are rapid and easy to perform. However, these methods were shown to suffer from the lack of specificity for measuring many of the diagnostically important steroids. LC-MS/MS overcomes many of the limitations of immunoassays, enhances diagnostic utility of the testing, and expands diagnostic capabilities in endocrinology. In addition to the superior quality of the measurements, LC-MS/MS allows high throughput testing using small sample volume with minimal sample preparation, and frees the laboratory from dependence on suppliers of assay specific reagents. LC-MS/MS is being widely employed for routine measurement of steroids, and the methodology plays an important role in the standardization and harmonization of measurements among clinical laboratories.
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- Bergquist, Maria, et al.
(författare)
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Altered adrenal and gonadal steroids biosynthesis in patients with burn injury
- 2016
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Ingår i: Clinical Mass Spectrometry. - : Elsevier BV. - 2213-8005 .- 2376-9998. ; 1, s. 19-26
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Burn injury inevitably leads to changes in the endogenous production of cytokines, as well as adrenal and gonadal steroids. Previous studies have reported gender-related differences in outcome following burn injury, which suggests that gonadal steroids may play a role. The aim of this study was to assess alterations in concentration of endogenous steroids in patients with burn injury.Methods: For this single-center, prospective descriptive study, high-sensitivity liquid chromatography tandem mass spectrometry (LC-MS/MS)-based steroid quantification was used to determine longitudinal profiles of the concentrations of endogenous steroids in plasma from sixteen adult male patients with burn injury (14.5-72% of total body surface area). Steroids were extracted from plasma samples and analyzed using multiple reaction monitoring acquisition, with electrospray ionization on a triple quadruple mass spectrometer. Total protein concentration was measured in the samples using spectrophotometry.Results: Steroid and total protein concentration distributions were compared to reference intervals characteristic of healthy adult men. Concentrations of the following steroids in plasma of burn injured patients were found to correlate positively to the area of the burn injury: cortisol (r = 0.84), corticosterone (r = 0.73), 11-deoxycortisol (r = 0.72), androstenedione (r = 0.72), 17OH-progesterone (r = 0.68), 17OH-pregnenolone (r = 0.64) and pregnenolone (r = 0.77). Concentrations of testosterone decreased during the acute phase and were up to ten-times lower than reference values for healthy adult men, while concentrations of estrone were elevated. By day 21 after injury, testosterone concentrations were increased in younger, but not older, patients. The highest concentrations of estrone were observed on day 3 after the injury and then declined by day 21 to concentrations comparable to those observed on the day of the injury.Conclusion: Burn injury alters endogenous steroid biosynthesis, with decreased testosterone concentrations and elevated estrone concentrations, during the first 21 days after the injury. Concentrations of glucocorticoids, progestagens and androgen precursors correlated positively with the area of burn injury. The finding of increased estrone following burn injury needs to be confirmed in a larger hypothesis driven study.
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- Kushnir, Mark M., et al.
(författare)
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Exploratory study of the association of steroid profiles in stimulated ovarian follicular fluid with outcomes of IVF treatment
- 2016
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Ingår i: Journal of Steroid Biochemistry and Molecular Biology. - : Elsevier BV. - 0960-0760 .- 1879-1220. ; 162, s. 126-133
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Forskningsöversikt (refereegranskat)abstract
- Steroid concentrations in stimulated follicular fluid (sFF) samples have been linked to the quality of oocytes used in IVF treatments. Most of the published studies focused on evaluating the association of the IVF outcomes with only a few of the steroids, measured by immunoassays (IA). We performed a treatment outcome, prospective cohort study using stimulated FF sampled from 14 infertile women undergoing IVF treatment; single oocyte was used per IVF cycle. Fourteen endogenous steroids were analyzed in 22 ovarian follicle aspirations, which corresponded to the embryos used in the IVF. Ten oocytes were associated with live birth (LB) and 12 with no pregnancy (NP). Steroids were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods. Differences in distribution of concentrations in association with the pregnancy outcome (LB or NP), and receiver operating characteristic (ROC) curves analysis were performed for the entire cohort and for within-women data. The predominant androgen and estrogen in stimulated sFF were androstenedione (A4) and estradiol (E2), respectively. Lower concentrations of pregnenolone (Pr), lower ratios of A4/ dehydroepiandrosterone (DHEA), testosterone (Te)/DHEA, and greater ratios of E2/Te, and estrone/A4 were observed in sFF samples associated with LB. Among the oocytes associated with NP, in four out of 12 samples total concentration of androgens was above the distribution of the concentrations in the oocytes corresponding to the LB group. Observations of the study indicated increased consumption of precursors and increased biosynthesis of estrogens in the follicles associated with LB. Our data suggest that potentially steroid profiles in sFF obtained during oocyte retrieval may serve as biomarkers for selection of the best embryo to transfer after IVF.
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| 11. |
- Kushnir, Mark M., et al.
(författare)
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Liquid chromatography-tandem mass spectrometry applications in endocrinology
- 2010
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Ingår i: Mass spectrometry reviews (Print). - : Wiley. - 0277-7037 .- 1098-2787. ; 29:3, s. 480-502
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Forskningsöversikt (refereegranskat)abstract
- Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been recognized as a primary methodology for the accurate analysis of endogenous steroid hormones in biological samples. This review focuses on the use of LC-MS/MS in clinical laboratories to assist with the diagnosis of diverse groups of endocrine and metabolic diseases. Described analytical methods use on-line and off-line sample preparation and analytical derivatization to enhance analytical sensitivity, specificity, and clinical utility. Advantages of LC-MS/MS as an analytical technique include high specificity, possibility to simultaneously measure multiple analytes, and the ability to assess the specificity of the analysis in every sample. All described analytical methods were extensively validated, utilized in routine diagnostic practice, and were applied in a number of clinical and epidemiological studies, including a study of the steroidogenesis in ovarian follicles.
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| 12. |
- Kushnir, Mark M., 1957-
(författare)
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Mass Spectrometric Applications for Diagnosing Metabolic and Endocrine Diseases
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Disease-specific compounds (biomarkers) are analyzed in clinical laboratories to assist with diagnosing diseases. This thesis describes development and validation of liquid chromatography tandem mass spectrometry (LC-MS/MS) based tests for diagnosing a diverse group of endocrine and metabolic diseases. The analytical methods used on-line and off-line sample extraction and analytical derivatization as means of enhancing the analytical sensitivity, specificity and clinical utility. All developed methods were extensively validated and reference intervals for the biomarker concentrations were established in blood samples of healthy adults and children. Advantages of the LC-MS/MS as an analytical technique include possibility of simultaneous measurement of multiple analytes and ability of confirming their identity. In this thesis we proposed and evaluated approaches for the assessment of the specificity of analysis in the methods that use tandem mass spectrometry detection. To enhance throughput of the LC-MS/MS tests for the biomarkers that have endogenous or exogenous isomers an approach was developed for quantitation of isomers from unresolved chromatographic peaks. Using methods developed in this thesis we performed a study of the steroidogenesis in ovarian follicles of healthy women and women with polycystic ovary syndrome (PCOS). Obtained data on the steroid concentrations and associations between the steroid metabolites in the pathway would be helpful for better understanding of the ovarian pathophysiology. Potential biomarkers of PCOS were identified in the thesis; further studies will be necessary to confirm their clinical utility.
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| 13. |
- Kushnir, Mark M., et al.
(författare)
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Protein and Steroid Profiles in Follicular Fluid after Ovarian Hyperstimulation as Potential Biomarkers of IVF Outcome
- 2012
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Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 11:10, s. 5090-5100
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Tidskriftsartikel (refereegranskat)abstract
- Controlled ovarian hyperstimulation is performed to assist with generation of multiple mature oocytes for use in in vitro fertilization (IVF). The goal of our study was to evaluate differences in protein and steroid profiles in ovarian follicular fluid (hFF) samples obtained during oocy-te retrieval from women undergoing IVF treatment and to identify physiological pathways associated with the proteins. The hFF samples were depleted of abundant proteins, fractionated by ultrafiltration, digested, and analyzed by nano-LC QTOF. Concentrations of 15 endogenous steroids were determined in the samples using LC-MS/MS methods. The total number of proteins identified in the samples was 75, of which 4, 7, and 2 were unique to the samples from women with viable pregnancy, miscarriage, and no pregnancy, respectively. Identified proteins were associated with the acute response signaling, coagulation system, intrinsic and extrinsic prothrombin activation, complement system, neuroprotective role of THOP1, FXR/RXR activation, role of tissue factor, and growth hormone pathways. A greater number of proteins associated with biosynthesis was found in hFF samples corresponding to the oocytes resulting in pregnancy. The abundance of seven proteins was found to be associated with steroidogenesis. The obtained data will contribute to better understanding of the pathogenesis and development of noninvasive markers for assessment of oocytes viability.
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- Kushnir, Mark M, et al.
(författare)
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Steroid profiles in ovarian follicular fluid from regularly menstruating women and women after ovarian stimulation
- 2009
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Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 55:3, s. 519-526
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Information on the concentrations of steroids in ovarian follicular fluid (FF) from regularly menstruating (RM) women has been limited because of the absence of methods for the simultaneous quantification of multiple steroids in small volumes of FF. We studied steroid profiles in FF during the early follicular phase of the menstrual cycle and after ovarian stimulation for in vitro fertilization (IVF), and compared concentrations with published values obtained by immunoassay (IA). METHODS: We used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to measure 13 steroids in 40-microL aliquots of FF samples from 21 RM women and from 5 women after ovarian stimulation for IVF. Relationships between concentrations of steroids and their ratios (representations of the enzyme activities) were evaluated within and between subgroups. RESULTS: The concentrations of testosterone (Te), androstenedione (A4), and estradiol (E2) measured by LC-MS/MS were lower than those previously reported in studies with IAs. In RM women, androgens were the most abundant class of steroids, with A4 being the major constituent. The concentrations of 17-hydroxyprogesterone (17OHP), total androgens, and estrogens were 200- to 1000-fold greater in FF than in serum. Compared with RM women, FF samples from women undergoing ovarian stimulation had significantly higher concentrations of E2 (P = 0.021), pregnenolone (P = 0.0022), 17OHP (P = 0.0007), and cortisol (F) (P = 0.0016), and significantly higher ratios of F to cortisone (P = 0.0006), E2 to estrone (P = 0.0008), and E2 to Te (P = 0.0013). CONCLUSIONS: The data provide the first MS-based concentration values for 13 steroids in ovarian FF from RM women, from estrogen- and androgen-dominant follicles, and from women after ovarian stimulation for IVF.
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- Lennartsson, Anna-Karin, et al.
(författare)
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Perceived stress at work is associated with attenuated DHEA-S response during acute psychosocial stress
- 2013
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Ingår i: Psychoneuroendocrinology. - : Elsevier BV. - 0306-4530 .- 1873-3360. ; 38:9, s. 1650-1657
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Tidskriftsartikel (refereegranskat)abstract
- Background: Dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEA-S) have been suggested to play a protective role during acute psychosocial stress, because they act as antagonists to the effects of the stress hormone cortisol. This study aims to investigate whether prolonged psychosocial stress, measured as perceived stress at work during the past week, is related to the capacity to produce DHEA and DHEA-S during acute psychosocial stress. It also aims to investigate whether prolonged perceived stress affects the balance between production of cortisol and DHEA-S during acute psychosocial stress. Method: Thirty-six healthy subjects (19 men and 17 women, mean age 37 years, SD 5 years), were included. Perceived stress at work during the past week was measured by using the Stress-Energy (SE) Questionnaire. The participants were divided into three groups based on their mean scores; Low stress, Medium stress and High stress. The participants underwent the Trier Social Stress Test (TSST) and blood samples were collected before, directly after the stress test, and after 30 min of recovery. General Linear Models were used to investigate if the Medium stress group and the High stress group differ regarding stress response compared to the Low stress group. Results: Higher perceived stress at work was associated with attenuated DHEA-S response during acute psychosocial stress. Furthermore, the ratio between the cortisol production and the DHEA-S production during the acute stress test were higher in individuals reporting higher perceived stress at work compared to individuals reporting low perceived stress at work. There was no statistical difference in DHEA response between the groups. Conclusions: This study shows that prolonged stress, measured as perceived stress at work during the past week, seems to negatively affect the capacity to produce DHEA-S during acute stress. Given the protective functions of DHEA-S, attenuated DHEA-S production during acute stress may Lead to higher risk for adverse effects on psychological and physiological health, particularly if stress exposure continues. (C) 2013 Elsevier Ltd. All rights reserved.
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- Naessén, Tord, et al.
(författare)
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Steroid profiles in ovarian follicular fluid in women with and without polycystic ovary syndrome, analyzed by liquid chromatography-tandem mass spectrometry.
- 2010
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Ingår i: Fertility and Sterility. - : Elsevier BV. - 0015-0282 .- 1556-5653. ; 94:6, s. 2228-2233
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE To compare steroid concentrations and steroid product-to-precursor ratios in ovarian follicular fluid (FF) from women with polycystic ovary syndrome (PCOS) and from regularly menstruating women in their early follicular phase, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Polycystic ovary syndrome involves abnormal regulation of the steroidogenic enzymes, leading to arrest of follicle development. DESIGN Case-control study. SETTING University hospital clinic. PATIENT(S) Follicular fluid from size-matched ovarian follicles (5-8 mm) in 27 nonstimulated women with PCOS and in 21 women without PCOS was sampled. Thirteen steroids were quantitated from 40 muL of FF, using LC-MS/MS. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Concentrations of steroids in the FF and product-to-precursor ratios (enzyme activity) were compared between the groups. RESULT(S) In women with PCOS, ovarian FF contained higher concentrations of individual and total androgens, lower individual and total estrogens (E), and a lower total E-to-androgen ratio, compared with regularly menstruating women. The product-to-precursor concentration ratios indicated higher CYP17-linked and lower CYP19-linked (aromatase) enzyme activity. Receiver operating characteristic plots indicated the early CYP17 step (17-OH5P/5P) being highly important for the prevalence of PCOS (c = 0.95). CONCLUSION(S) The women with PCOS had higher ovarian CYP17-linked and lower CYP19-linked (aromatase) enzyme activity, confirming previous data. Multiple steroid assessments from minute volumes including FF from nonstimulated ovaries, using LC-MS/MS, might be useful in research, clinical endocrinology, and in IVF.
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- Tajar, Abdelouahid, et al.
(författare)
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Frailty in Relation to Variations in Hormone Levels of the Hypothalamic-Pituitary-Testicular Axis in Older Men: Results From the European Male Aging Study.
- 2011
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Ingår i: Journal of the American Geriatrics Society. - : Wiley. - 0002-8614. ; 59:5, s. 814-821
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVES: To explore the associations between frailty and reproductive axis hormones (as an important regulatory system) in middle aged and older men. DESIGN: Cross-sectional. SETTING: The European Male Aging Study. PARTICIPANTS: Three thousand two hundred nineteen community-dwelling European men aged 40 to 79. MEASUREMENTS: Interviewer-assisted questionnaires to assess physical activity, health status, and mood were administered. Testosterone (T), luteinizing hormone (LH), follicle-stimulating hormone (FSH), dehydroepiandrosterone sulfate (DHEAS), and sex hormone-binding globulin (SHBG) were measured in a fasting morning blood sample. Frailty was assessed as an index (FI) according to the number (out of 43 possible) of health deficits (symptoms, signs, and functional impairments). Relationships between FI and hormone levels (as outcomes) were explored using regression models. RESULTS: Mean FI was 0.12 ± 0.11 (range 0-0.67) was highest in the oldest group. After adjustment for confounders, higher levels of FI were significantly associated with lower levels of total T, free T, and DHEAS and higher levels of gonadotropins and SHBG; a 1-standard deviation cross-sectional increase in FI was associated with a regression coefficient of -0.30 nmol/L (95% confidence interval (CI)=-0.53 to -0.07) decrease in total T and 0.66 U/L (95% CI=0.48-0.83) increase in LH. CONCLUSIONS: The associations between high FI, high gonadotropins, and well-maintained circulating T suggest that these changes are markers of aging-related disruptions of multiple physiological regulation, of which alterations in pituitary-testicular function represent a sensitive marker rather than an underlying pathogenic mechanism for frailty.
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