Definition of prepartum hyperketonemia in dairy goats.

A prospective cohort study was conducted on 1,081 dairy goats from 10 commercial herds in Québec (Canada) to define prepartum hyperketonemia based on optimal blood ß-hydroxybutyrate acid threshold values for the early prediction of pregnancy toxemia (PT) and mortality in late-gestation dairy goats. All pregnant goats had blood sampled weekly during the last 5wk of pregnancy. The blood was analyzed directly on the farm for ß-hydroxybutyrate acid quantification using a Precision Xtra meter (Abbott Diabetes Care, Saint-Laurent, QC, Canada). Body condition scores on the lumbar region and sternum were noted. Each goat was classified as being at low (n=973) or high risk (n=108) of having PT by producers based on a standardized definition. The optimal threshold for predicting a PT diagnosis or mortality for each week before kidding was determined based on the highest sum of sensitivity and specificity. The association between hyperketonemia and subsequent PT was tested using a multivariable logistic regression model considering hyperketonemia at wk 4 prepartum, litter size, and body condition score at wk 4 prepartum as covariates, and herd and parturition cohort as random effects. The association between mortality and hyperketonemia was also tested using a logistic regression model accounting for the presence or absence of treatment during the last month of pregnancy. The hyperketonemia definition based on PT varied between ≥0.4 and ≥0.9mmol/L during the last 5wk prepartum. Goats affected by hyperketonemia at wk 4 prepartum and with a large litter size (≥3 fetuses) had 2.1 and 40.5 times the odds, respectively, of subsequent PT than other goats. Hyperketonemia definitions based on mortality varied between ≥0.6 and ≥1.4mmol/L during the last 4wk prepartum, and was ≥1.7mmol/L during the first week postpartum. Goats affected by hyperketonemia and treated by producers had 3.4 and 11.8 times the odds, respectively, of subsequent mortality than did other goats. These results showed that prepartum hyperketonemia could be defined in dairy goats using subsequent risks of PT or mortality during the last month of pregnancy.

Preoperative cow-side lactatemia measurement predicts negative outcome in Holstein dairy cattle with right abomasal disorders.

The objectives of the current study were (1) to determine the gain in prognostic accuracy of preoperative l-lactate concentration (LAC) measured on farm on cows with right displaced abomasum (RDA) or abomasal volvulus (AV) for predicting negative outcome; and (2) to suggest clinically relevant thresholds for such use. A cohort of 102 cows with on-farm surgical diagnostic of RDA or AV was obtained from June 2009 through December 2011. Blood was drawn from coccygeal vessels before surgery and plasma LAC was immediately measured by using a portable clinical analyzer. Dairy producers were interviewed by phone 30 d following surgery and the outcome was determined: a positive outcome if the owner was satisfied of the overall evolution 30 d postoperatively, and a negative outcome if the cow was culled, died, or if the owner reported being unsatisfied 30 d postoperatively. The area under the curve of the receiver operating characteristic curve for LAC was 0.92 and was significantly greater than the area under the curve of the receiver operating characteristic curve of heart rate (HR; 0.77), indicating that LAC, in general, performed better than HR to predict a negative outcome. Furthermore, the ability to predict a negative outcome was significantly improved when LAC measurement was considered in addition to the already available HR data (area under the curve: 0.93 and 95% confidence interval: 0.87, 0.99). Important inflection points of the misclassification cost term function were noted at thresholds of 2 and 6 mmol/L, suggesting the potential utility of these cut-points. The 2 and 6 mmol/L thresholds had a sensitivity, specificity, positive predictive value, and negative predictive value for predicting a negative outcome of 76.2, 82.7, 53.3, and 93.1%, and of 28.6, 97.5, 75, and 84%, respectively. In terms of clinical interpretation, LAC ≤2 mmol/L appeared to be a good indicator of positive outcome and could be used to support a surgical treatment decision. The treatment decision for cows with LAC between 2 and 6 mmol/L, however, would depend on the economic context and the owner's attitude to risk in regard to potential return on its investment. Finally, performing a surgical correction on commercial cows with RDA or AV and a LAC ≥6 mmol/L appeared to be unjustified and these animals should be culled based on their high probability of negative outcome.

Short communication: ultrasonographic assessment of the thorax as a fast technique to assess pulmonary lesions in dairy calves with bovine respiratory disease.

The aim of this study was to assess inter- and intraoperator agreement when assessing lung consolidation secondary to bovine respiratory disease (BRD) by thoracic ultrasonography. Ten calves were blindly assessed by 3 operators with varying expertise in thoracic ultrasound to look for lung consolidation and the presence of comet-tail artifacts (COMT). Systematic ultrasonography of the thorax was performed using an 18-site per side assessment with a linear 8.5-MHz probe. The status of the calves [healthy (n=4) vs. treated for BRD (n=6)] was not known by the operators. The interoperator kappa agreement for detecting consolidation was moderate to almost perfect (from 0.6 to 1.0) depending on the operator's experience (diagnosis of consolidation if depth ≥1cm). The intraclass correlation coefficient for consistency was 0.71 for a single measurement and 0.88 for average measurement. The intraclass correlation coefficient for agreement was 0.73 for single measurements and 0.89 for average measurements. These values were considered good for single measurements and excellent for average measurements. Systematic ultrasonography of the thorax can be used routinely to assess lung consolidation in dairy calves and can therefore be of importance, especially for assessment of subclinical BRD.

Short communication: evaluation of the accuracy of an electronic on-farm test to quantify blood ß-hydroxybutyrate concentration in dairy goats.

The objective of this cross-sectional study was to validate the accuracy of a hand-held electronic on-farm test (Precision Xtra) for quantifying the blood ß-hydroxybutyrate (BHBA) concentration in dairy goats. A total of 114 dairy goats from 3 commercial herds were sampled once for blood in the jugular vein between 1mo before and 2mo after parturition. Blood samples were centrifuged to harvest serum and sera were sent to the Animal Health Laboratory of the Université de Montréal for quantification of BHBA concentration (gold standard). Laboratory BHBA values were between 0.1 and 3.7mmol/L. Precision Xtra values were compared with gold standard values; Pearson correlation coefficient was 0.98 and coefficient of determination was 0.95. Overall, these results suggested that Precision Xtra provides excellent accuracy for measuring blood BHBA concentration in dairy goats compared with the gold standard test.

Use of ichip for high-throughput in situ cultivation of "uncultivable" microbial species.

One of the oldest unresolved microbiological phenomena is why only a small fraction of the diverse microbiological population grows on artificial media. The "uncultivable" microbial majority arguably represents our planet's largest unexplored pool of biological and chemical novelty. Previously we showed that species from this pool could be grown inside diffusion chambers incubated in situ, likely because diffusion provides microorganisms with their naturally occurring growth factors. Here we utilize this approach and develop a novel high-throughput platform for parallel cultivation and isolation of previously uncultivated microbial species from a variety of environments. We have designed and tested an isolation chip (ichip) composed of several hundred miniature diffusion chambers, each inoculated with a single environmental cell. We show that microbial recovery in the ichip exceeds manyfold that afforded by standard cultivation, and the grown species are of significant phylogenetic novelty. The new method allows access to a large and diverse array of previously inaccessible microorganisms and is well suited for both fundamental and applied research.

The effect of stimulus type and background noise on hearing abilities of the round goby Neogobius melanostomus.

The auditory abilities of the round goby Neogobius melanostomus were quantified using auditory evoked potential recordings, using tone bursts and conspecific call stimuli. Fish were tested over a range of sizes to assess effects of growth on hearing ability. Tests were also run with and without background noise to assess the potential effects of masking in a natural setting. Neogobius melanostomus detected tone bursts from 100 to 600 Hz with no clear best frequency in the pressure domain but were most sensitive to 100 Hz tone stimuli when examined in terms of particle acceleration. Responses to a portion of the N. melanostomus call occurred at a significantly lower threshold than responses to pure tone stimulation. There was no effect of size on N. melanostomus hearing ability, perhaps due to growth of the otolith keeping pace with growth of the auditory epithelium. Neogobius melanostomus were masked by both ambient noise and white noise, but not until sound pressure levels were relatively high, having a 5-10 dB threshold shift at noise levels of 150 dB re 1 µPa and higher but not at lower noise levels.

Treatment of prostate cancer with gonadotropin-releasing hormone agonists.

It is now well established that chronic treatment with GnRH agonists offers an advantageous alternative to orchiectomy and estrogens for the treatment of prostate cancer. Castration levels of androgens can thus be easily achieved without side effects other than those related to castration levels of serum androgens. However, man is unique among species in having a high secretion rate of precursor adrenal steroids which are converted into active androgens in the normal prostate and prostatic cancer. All the enzymes required for the transformation of dehydroepiandrosterone sulfate, dehydroepiandrosterone, androstenedione, and androst-5-ene-3 beta, 17 beta-diol are present in prostatic tissue. Moreover, as shown in many systems, castration levels of serum testosterone (T) at 0.2-0.4 ng/ml exert significant androgenic activity in target tissues. In order to inhibit the action of androgens of both testicular and adrenal origin, GnRH agonists have been administered in association with the pure antiandrogen Flutamide in patients having clinical stage D2 (bone metastases) prostate cancer. A positive objective response assessed according to the criteria of the United States National Prostatic Cancer Project (USNPCP) has been observed in 84 of the 88 patients who had received no previous treatment (95.4%). After 2 yr of treatment, the probability of continuing response is 70% compared to 0-10% by previous approaches. In addition, the death rate at 2 yr is at 10.9% as compared to approximately 50% after standard hormonal therapy. When the same treatment was applied to patients who had received previous hormonal therapy (orchiectomy, estrogens or GnRH agonists alone) before showing a relapse, the response rate decreased to 62.9% and the death rate at 2 yr was 52%.(ABSTRACT TRUNCATED AT 250 WORDS)

Inter-rater agreement and reliability of thoracic ultrasonographic findings in feedlot calves, with or without naturally occurring bronchopneumonia.

BACKGROUND: Thoracic ultrasonography (TUS) can be used to assess the extent and severity of lung lesions associated with bronchopneumonia (BP) in feedlot cattle. HYPOTHESIS/OBJECTIVES: To assess inter-rater agreement and reliability of TUS findings in feedlot cattle, with or without naturally occurring BP. ANIMALS: Feedlot steers with (n = 210) or without (n = 107) clinical signs of BP that were assessed by TUS in a previous case-control study. METHODS: A random sample of 50 TUS videos (16-s duration) were scored by 6 raters with various levels of TUS expertise. Lung consolidation, comet tail artifacts, pleural irregularity and effusion were scored. Inter-rater agreement was assessed using raw percentage of agreement (Pa), Cohen's and Fleiss' Kappa (κ), and Gwet agreement coefficient (AC1). Intra-class correlation (ICC) was determined for variables with continuous measurements (mixed factorial design). RESULTS: Median (interquartile range [IQR]) Pa were 0.84 (0.80-0.89), 0.82 (0.80-0.87), 0.62 (0.53-0.67), and 0.82 (0.75-0.86) for presence of lung consolidation, comet tails, pleural irregularity, and pleural effusion, respectively. For the same lesions, Fleiss κ (95% confidence intervals [CI]) were 0.67 (0.49-0.86), 0.56 (0.33-0.80), 0.20 (-0.05 to 0.44), and 0.36 (0.10-0.61), respectively. AC1 were 0.68 (0.51-0.86), 0.73 (0.58-0.89), 0.21 (-0.01 to 0.44), and 0.71 (0.51-0.92), respectively. Moderate reliability was found among raters for all quantitative variables (ICC ranged from 0.52 to 0.70). CONCLUSIONS AND CLINICAL IMPORTANCE: Inter-rater agreement was good for presence of lung consolidation, comet tails and pleural effusion (based on Pa and AC1) but was slight to poor for pleural irregularity.

Bioavailability and metabolism of oral and percutaneous dehydroepiandrosterone in postmenopausal women.

To study the bioavailability of dehydroepiandrosterone (DHEA) administered by the oral and percutaneous routes, three groups of 12 postmenopausal women aged 60-70 years received two capsules of 50mg of DHEA orally before breakfast daily for 14 days or applied 4 g of a 10% DHEA cream or gel at the same time of the day on a 30 cm x 30 cm surface area on the thighs. Detailed serial blood sampling over 24h was performed following 1st and 14th DHEA administration for measurement of DHEA and nine of its metabolites by liquid chromatography tandem mass spectrometry (LC-MS/MS) or gas chromatography mass spectrometry (GC-MS). Serum levels of estrone (E1) and estradiol (E2) did not change following DHEA administration by any of the three formulations, while serum androstenedione (4-dione), testosterone, DHEA sulfate (DHEA-S), E(1)-S, androsterone glucuronide (ADT-G) and 3alpha-androstanediol-G (3alpha-diol-G), increased in all cases, the effect on these parameters being more important after oral than percutaneous administration due to the metabolism of DHEA into these metabolites in the gastrointestinal tract and liver. No qualitative differences in DHEA metabolism are observed between the oral and percutaneous routes of DHEA administration while the levels of all steroids remain on a plateau during the 24h period during chronic percutaneous DHEA administration. The present data show that DHEA is transformed into active androgens and estrogens in peripheral intracrine tissues with no or minimal release of the active steroids E(1), E(2) or testosterone in the circulation. Moreover, DHEA is preferentially transformed into androgens rather than into estrogens. Most importantly, the present data show that changes in serum DHEA following oral or percutaneous DHEA administration are not a valid parameter of DHEA action since the increase in serum DHEA is at least 100% greater than the increase in the formation of active androgens and estrogens and thus much higher than the potential physiological effects.

New chemical and radiochemical routes to [18F]Rho6G-DEG-F, a delocalized lipophilic cation for myocardial perfusion imaging with PET.

New chemical and radiochemical syntheses are described for the preparation of [18F]Rho6G-DEG-F, an 18F-labeled analogue of the fluoresecent dye rhodamine 6G, which has shown promise as myocardidal perfusion imaging agent. Tosylated precursors of [18F]Rho6G-DEG-F amenable to 18F-labeling were obtained either through a two-step synthesis from rhodamine 6G lactone (33% yield), or in one step from rhodamine 575 (64% yield), then purified by preparative C18 chromatography. Manual synthesis of [18F]Rho6G-DEG-F was achieved in a single radiochemical step from either the tosylate salt or the tosylate/formate double salt in DMSO under standard nucleophillic aliphatic 18F-fluorination conditions (K[18F]F/K2CO3/Kryptofix 2.2.2.). Incorporation of the [18F]F- was found to be satisfactory (≥34% by TLC), despite the protic character of the precursor molecules. [18F]Rho6G-DEG-F was manually synthesized in final decay-corrected radiochemical yields of 11-26% (tosylate salt) and 9-21% (tosylate/formate double salt). The protocol was transferred to an automated synthesis unit, where the product was obtained in 3-9% radiochemical yield (n=3) decay corrected to start-of-synthesis, >99% radiochemical purity, and a molar activity of 122-267 GBq/µmol (3.3-7.2 Ci/µmol).

Effect of dehydroepiandrosterone and the antiestrogen EM-800 on growth of human ZR-75-1 breast cancer xenografts.

BACKGROUND: In the mammary gland, androgens are formed from the precursor steroid dehydroepiandrosterone (DHEA). Clinical evidence indicates that androgens have inhibitory effects on breast cancer. Estrogens, on the other hand, stimulate the development and growth of breast cancer. We studied the effect of DHEA alone or in combination with the newly described pure antiestrogen EM-800 on the growth of subcutaneous tumor xenografts formed by the human breast cancer cell line ZR-75-1 in ovariectomized nude mice. METHODS: Immediately after ovariectomy, mice received daily subcutaneous injections of 0.5 microg estrone (E1) (an estrogenic hormone). EM-800 (15, 50, or 100 microg) was given orally once daily. DHEA was administered percutaneously twice daily (total dose of 0.3, 1.0, or 3.0 mg) to the dorsal skin either alone or in combination with a 15-microg daily oral dose of EM-800. Changes in tumor size in response to the treatments (in relation to measurements made on the first day of treatment) were assessed periodically. At the end of the experiments, tumors were dissected and weighed. RESULTS: A 9.4-fold increase in tumor size in 9.5 months was observed in ovariectomized mice receiving E1 alone. Administration of 15, 50, or 100 microg EM-800 in E1-supplemented mice led to inhibitions of 87.5%, 93.5%, and 94.0% in tumor size, respectively. DHEA, on the other hand, at doses of 0.3, 1.0, or 3.0 mg inhibited terminal tumor size by 50.4%, 76.8%, and 80.0%, respectively. Comparable inhibitions in tumor size were obtained with a daily 15-microg oral dose of EM-800 with or without different doses of percutaneous DHEA. CONCLUSIONS: DHEA and EM-800 independently suppressed the growth of E1-stimulated ZR-75-1 xenograft tumors in nude mice. Administration of DHEA at the defined doses did not alter the inhibitory effect of EM-800.

Presence of C-19 steroids in mammary Shionogi carcinoma (SC 115) in castrated mice.

Intact and castrated male DD/S mice were inoculated with androgen-dependent cells (SC 115). All intact animals developed tumors after Day 12 of inoculation; however, six of seven castrated animals presented tumors 48 days postinoculation. The levels of steroids in both tumors were then examined. In castrated mice, dehydroepiandrosterone and androst-5-ene-3 beta, 17 beta-diol levels were diminished by 30% and 70%, respectively, while the amounts of testosterone and androstenedione were reduced by more than 90%. Our data also demonstrate that androstane-3 alpha, 17 beta-diol and androstane-3 beta, 17 beta-diol were decreased to 60% and dihydrotestosterone decreased to 6% of their normal value, respectively. This latter level (0.48 nM) was sufficient to still effect a potent androgenic response in the tumor. Besides, a highly significant correlation was found in these tumors between various C-19 steroids (dehydroepiandrosterone and androstane-3 alpha, 17 beta-diol, r = 0.97, P less than 0.01), suggesting a possible conversion of C-19 precursors into potent androgens in the tumors. Determination of the plasma steroid levels in the castrated animals clearly confirmed that potent androgenic steroids and precursors were still in the circulation 3 days after castration. It thus appears that C-19 steroids from adrenal origin may be also involved in "independent" tumor growth.

Comparison of the effects of the antiestrogens EM-800 and tamoxifen on the growth of human breast ZR-75-1 cancer xenografts in nude mice.

Although estrone supplementation in ovariectomized (OVX) nude mice bearing ZR-75-1 xenografts caused a 365% increase in average tumor size during the 4-month treatment period, administration of the antiestrogen EM-800 at the daily oral doses of 50, 150, or 400 microg completely prevented estrogen-stimulated tumor growth. At the same doses of tamoxifen, tumor size was inhibited to 189, 117, and 120% above pretreatment values. However, when EM-800 (150 microg/day) was added to the daily 150- and 400-microg doses of tamoxifen, final tumor size was decreased further to 12 and 38% above pretreatment values, respectively. EM-800 (400 microg daily) administered to estrone-supplemented OVX mice caused complete, partial, and stable responses in 11, 22, and 49% of estrone-stimulated tumors, respectively, whereas 19% (7 of 37) progressed. At the same dose of tamoxifen, the corresponding responses were 3% (complete response), 3% (partial response), and 25% (no change), whereas 69% (22 of 32) of tumors progressed. In the absence of estrone supplementation, tamoxifen (400 microg) alone administered to OVX mice stimulated tumor growth to 161% compared with initial size whereas the same dose of EM-800 reduced tumor size by 55%, a value superimposable to that observed in OVX control animals. The agonistic effect of tamoxifen is thus illustrated by the observation that 73% of tumors progressed when tamoxifen was administered alone to OVX animals whereas no tumor progressed with EM-800. The present data strongly suggest that at least part of the initial lack of response and resistance to tamoxifen during tamoxifen treatment in women is due to the estrogenic activity of this compound, whereas the new antiestrogen EM-800 exerts pure antagonistic action.

The suprabasal expression of alpha 6 beta 4 integrin is associated with a high risk for malignant progression in mouse skin carcinogenesis.

Enhanced expression of the alpha 6 beta 4 integrin complex has been linked to malignant progression in mouse skin carcinogenesis. To determine if alpha 6 beta 4 expression can predict risk for malignant conversion among populations of benign skin tumors, we analyzed the distribution of alpha 6 beta 4 and other markers of progression in papillomas at high and low risk for malignant progression. After initiation with 7,12-dimethylbenz[a]anthracene, mice were promoted with 12-O-tetradecanoylphorbol-13-acetate to induce predominantly low risk tumors or promoted with mezerein to produce predominantly high risk tumors. When tumors first appeared at 8 weeks after promotion, high risk papillomas demonstrated basal and suprabasal alpha 6 beta 4 expression, loss of keratin 1, and aberrant expression of keratin 13. In these tumors alpha 6 beta 4 expression coincided with an expansion of the proliferating compartment as indicated by suprabasal bromodeoxyuridine labeling. In contrast, alpha 6 beta 4 immunostaining was confined to basal cells in low risk tumors, keratin 1 was abundant, and keratin 13 was absent in the majority of this group, while proliferating cells were largely in the basal compartment. By 33 weeks, alpha 6 beta 4 suprabasal expression continued to distinguish groups at higher risk for malignant conversion, but keratin 13 was expressed in all groups. At this time, high risk tumors displayed focal expression of keratin 8 and gamma-glutamyltranspeptidase, markers also found in chemically induced carcinomas. Keratin 8 and gamma-glutamyltranspeptidase were expressed only in alpha 6 beta 4 positive cells. These results indicate that expression of alpha 6 beta 4 integrin in suprabasal strata serves as an early predictive marker to identify benign squamous tumors at high risk for malignant progression.

Protein kinase Cdelta-mediated phosphorylation of alpha6beta4 is associated with reduced integrin localization to the hemidesmosome and decreased keratinocyte attachment.

In mammalian epidermis, expression of the alpha6beta4 integrin is restricted to the hemidesmosome complexes, which connect the proliferative basal cell layer with the underlying basement membrane. Keratinocyte differentiation is associated with down-regulation of alpha6beta4 expression and detachment of keratinocytes from the basement membrane. Neoplastic keratinocytes delay maturation, proliferate suprabasally, and retain the expression of the alpha6beta4 integrin in suprabasal cells disassociated from the hemidesmosomes. We now show that the alpha6beta4 integrin is a substrate for serine phosphorylation by protein kinase C in keratinocytes. Furthermore, protein kinase C-mediated phosphorylation of alpha6beta4 is associated with redistribution of this integrin from the hemidesmosome to the cytosol. Specifically, in vitro kinase assays identified the protein kinase Cdelta as the primary isoform phosphorylating alpha6 and beta4 integrin subunits. Using recombinant protein kinase C adenoviruses, overexpression of protein kinase Cdelta but not protein kinase Calpha in primary keratinocytes increased beta4 serine phosphorylation, decreased alpha6beta4 localization to the hemidesmosome complexes, and reduced keratinocyte attachment. Taken together, these results establish a link between protein kinase Cdelta-mediated serine phosphorylation of alpha6beta4 integrin and its effects on alpha6beta4 subcellular localization and keratinocyte attachment to the laminin underlying matrix.

Blockade of the stimulatory effect of estrogens, OH-tamoxifen, OH-toremifene, droloxifene, and raloxifene on alkaline phosphatase activity by the antiestrogen EM-800 in human endometrial adenocarcinoma Ishikawa cells.

Although temporary benefits of tamoxifen therapy are observed in up to 40% of women with breast cancer, this compound, which is known to possess mixed estrogenic and antiestrogenic activities, has been associated with increased risk of endometrial carcinoma. This study compares the effects of the novel nonsteroidal pure antiestrogen EM-800 and related compounds with those of a series of antiestrogens on the estrogen-sensitive alkaline phosphatase (AP) activity in human endometrial adenocarcinoma Ishikawa cells. Exposure to increasing concentrations of up to 1000 nM EM-800 or its active metabolite EM-652 alone failed to affect basal AP activity. In contrast, incubation with 10 nM (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, droloxifene, or raloxifene increased the value of this estrogen-sensitive parameter by 3.3-, 3.5-, 2.2-, and 1.6-fold, respectively, a stimulatory effect that was completely reversed by simultaneous exposure to 30 nM EM-800. Moreover, the stimulation of AP activity induced by 1 nM 17beta-estradiol was completely reversed by EM-800, EM-652, or ICI-182780, at the IC50 value of 1.98 +/- 0.23, 1.01 +/- 0.16, and 5.64 +/- 0.59 nM, respectively, whereas the partial blockade exerted by (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, or raloxifene was observed at IC50 values of 13.5 +/- 3.80, 41.0 +/- 7.2, and 3.74 +/- 0.43 nM, respectively. Thus, as assessed by their activity in the human Ishikawa endometrial carcinoma cells, EM-800 and EM-652 are the most potent known antiestrogens in Ishikawa cells, and, most importantly, they are devoid of the estrogenic activity observed in these human endometrial cancer cells with (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, droloxifene, and raloxifene.

Assessing vitamin D nutritional status: Is capillary blood adequate?

BACKGROUND: Venous blood is the usual sample for measuring various biomarkers, including 25-hydroxyvitamin D (25OHD). However, it can prove challenging in infants and young children. Hence the finger-prick capillary collection is an alternative, being a relatively simple procedure perceived to be less invasive. We elected to validate the use of capillary blood sampling for 25OHD quantification by liquid chromatography tandem-mass spectrometry (LC/MS-MS). METHODS: Venous and capillary blood samples were simultaneously collected from 15 preschool-aged children with asthma 10days after receiving 100,000IU of vitamin-D3 or placebo and 20 apparently healthy adult volunteers. 25OHD was measured by an in-house LC/MS-MS method. RESULTS: The venous 25OHD values varied between 23 and 255nmol/l. The venous and capillary blood total 25OHD concentrations highly correlated (r(2)=0.9963). The mean difference (bias) of capillary blood 25OHD compared to venous blood was 2.0 (95% CI: -7.5, 11.5) nmol/l. CONCLUSION: Our study demonstrates excellent agreement with no evidence of a clinically important bias between venous and capillary serum 25OHD concentrations measured by LC/MS-MS over a wide range of values. Under those conditions, capillary blood is therefore adequate for the measurement of 25OHD.

Data in support for the measurement of serum 25-hydroxyvitamin D (25OHD) by tandem mass spectrometry.

This article provides data and a method related to a research paper entitled "Assessing vitamin D nutritional status: is capillary blood adequate?" (Jensen et al., 2016) [1]. Circulating 25OHD, the accepted biomarker of the vitamin D nutritional status, is routinely measured by automated immunoassays, that although may be performed in hospital central laboratories, often suffer from a lack of specificity with regards to the different vitamin D metabolites, "Measurement of circulating 25-hydroxyvitamin D: a historical review" (Le Goff et al., 2015) [2]. Mass spectrometry offers this specificity. This article describes the performance of an in-house tandem mass spectrometry method for the individual measurement of 25OHD3, 25OHD2 and 3-épi-25OHD3.

Formation of pregnenolone- and dehydroepiandrosterone-fatty acid esters by lecithin-cholesterol acyltransferase in human plasma high density lipoproteins.

Pregnenolone- (PREG-), and dehydroepiandrosterone- (DHEA-) fatty acid esters (FA) are present in human plasma, where they are associated with lipoproteins. Because plasma has the ability to form PREG-FA and DHEA-FA in vitro from their unconjugated steroid counterparts, we postulated that the LCAT enzyme might be responsible for their formation. Here we show that lecithin-cholesterol acyltransferase (LCAT) has PREG and DHEA esterifying activities. First, VLDL, IDL, LDL, and HDL were isolated by the sequential ultracentrifugation micromethod from the plasma of fasting men and women and tested for their ability to form PREG-FA, DHEA-FA, and cholesteryl esters in vitro from their respective unconjugated counterparts. The results showed that the three steroids were esterified only in HDL subfractions. The rate of tritiated PREG esterification was clearly higher than that of tritiated cholesterol and DHEA, both in total plasma and isolated HDL, and no gender difference was observed. Second, human and guinea pig LCAT were purified and used in phosphatidylcholine-reconstituted vesicles containing human apoAI to show their ability to esterify tritiated cholesterol, PREG, and DHEA in the absence of unlabeled steroid. The amount of cholesteryl ester, PREG-FA, and DHEA-FA increased after incubation as a function of time and amount of purified LCAT, showing that PREG is preferentially acylated by LCAT compared to cholesterol and DHEA. The PREG and DHEA esterifying activities of LCAT were cofactor-dependent, as shown by the absence of acylation without apoAI. Finally, we determined by HPLC the fatty acid moiety of PREG-FA and DHEA-FA formed in human plasma and guinea pig and rat sera in vitro after incubation with unconjugated tritiated PREG and DHEA. We showed that the fatty acid moieties of newly formed tritiated PREG-FA and DHEA-FA were similar to that reported for cholesteryl esters in the plasma of the three species. We conclude that LCAT has a lecithin-steroid acyltransferase activity and that PREG is probably the preferential substrate of this enzyme. In addition, the fact that the differences in the fatty acid moieties of cholesteryl esters of human, guinea pig, and rat plasmas are also observed for PREG-FA and DHEA-FA suggests that the LCAT is the sole circulating enzyme that has PREG and DHEA esterifying activities.