Serum Sulfate Research Articles

Hormonal Regulation of Sodium/Sulfate Co-Transport in Renal Epithelial Cells

Abstract. Serum sulfate concentrations are elevated in infants, young children, and pregnant women due, at least in part, to increased renal sulfate reabsorption. Little is known about the effects of hormones, particularly those involved in growth, development, and pregnancy, on renal sulfate reabsorption. The objective of this investigation was to examine the effects of growth hormone (GH), insulin-like growth factor 1 (IGF-1), progesterone (PG), and 17β-estradiol (EST) on renal sodium/sulfate co-transport. 35S-sulfate uptake was determined in Madin-Darby canine kidney (MDCK)/NaSi-1 cells (MDCK cells that have been stably transfected with rat sodium/sulfate co-transporter (NaSi-1) cDNA) and in opossum kidney (OK) cells. NaSi-1 mRNA was determined by RT-PCR and protein levels by ELISA. GH (0.1 nM) significantly increased the sodium/sulfate co-transport in MDCK/NaSi-1 cells up to 35%. IGF-1 induced a concentration-related stimulation of the sodium/sulfate co-transport with a maximal response observed at 1000 nM (59% increase). Sodium-dependent sulfate uptake was significantly increased when cells were preincubated with 10 nM PG, 10 nM EST, or 10 nM PG/10 nM EST up to 41%, 46%, or 39%, respectively. OK cells exhibited endogenous sodium-dependent sulfate transport; significantly increased sodium/sulfate co-transport was also observed in OK cells that were preincubated with GH, IGF-1, and PG/EST, although not with EST alone. The NaSi-1 mRNA and NaSi-1 protein levels were significantly increased in MDCK/NaSi-1 cells treated with 0.1 nM GH, 100 nM IGF-1, 10 nM PG, and/or 10 nM EST compared with control. These results suggest that the increased renal sulfate reabsorption that occurs in neonates, young and pregnant humans, and animals could be mediated by the increased steady-state levels of NaSi-1 mRNA produced by the higher plasma concentrations of GH, IGF-1, or PG/EST.

Determination of ethyl sulfate in human serum and urine by capillary zone electrophoresis

The use of capillary zone electrophoresis (CZE) with indirect absorbance detection for the analysis of ethyl sulfate (EtS) in serum and urine was investigated. EtS is a direct metabolite of ethanol employed as marker for recent alcohol consumption. Fused-silica capillaries of 60 cm total length were either coated with cetyltrimethylammonium bromide (CTAB, 50 μm I.D. capillary) or poly(diallyldimethylammonium chloride) (PDADMAC, 100 μm I.D. capillary) to allow CZE analyses to be performed with reversed polarity. At pH 2.2 with a maleic acid/phthalic acid background electrolyte, both approaches provided reliable EtS serum levels down to 0.2 mg L −1 (1.6 μM) for the analysis of solid-phase extracts that were prepared after chloride precipitation. Analysis of urines diluted to a conductivity of 5 S m −1 and analyzed in the two capillary formats resulted in limits of quantification (LOQs) of 2 and 1 mg L −1, respectively. With urines adjusted to 10 S m −1 via dilution or condensation, an LOQ of 0.6 mg L −1 (4.8 μM) was obtained in the CTAB coated capillary whereas in the PDADMAC-coated capillary of equal length not all matrix components were resolved from EtS. The developed assays are robust and suitable to monitor EtS in samples of individuals who consumed as little as one standard drink of an alcoholic beverage containing about 14 g of ethanol.

Sequencing of the CHST6 gene in Czech macular corneal dystrophy patients supports the evidence of a founder mutation

Aims:To characterise the role of the carbohydrate sulfotransferase gene (CHST6) in macular corneal dystrophy (MCD) in Czech patients.Methods:The coding region of the CHST6 gene was directly sequenced in 10 affected...

Regulation of the Sodium/Sulfate Co-transporter by Farnesoid X Receptor α

Fxralpha is known to regulate a variety of metabolic processes, including bile acid, cholesterol, and carbohydrate metabolism. In this study, we show direct evidence that Fxralpha is a key player in maintaining sulfate homeostasis. We identified and characterized the sodium/sulfate co-transporter (NaS-1; Slc13a1) as an Fxralpha target gene expressed in the kidney and intestine. Electromobility shift assays, chromatin immunoprecipitation, and promoter reporter studies identified a single functional Fxralpha response element in the second intron of the mouse Slc13a1 gene. Treatment of wild-type mice with GW4064, a synthetic Fxralpha agonist, induced Slc13a1 mRNA in the intestine and kidney. Slc13a1 mRNA was also induced in the kidney and intestine of wild-type, but not Fxralpha-/- mice, after treatment with the hepatotoxin alpha-naphthylisothiocyanate, which is known to result in elevated blood bile acid levels. Finally, we observed a decrease in Slc13a1 mRNA in the kidney and intestine of Fxralpha-/- mice and a corresponding increase in urinary excretion of free sulfates as compared with wild-type mice. These results demonstrate that mouse Slc13a1 is a novel Fxralpha target gene expressed in the kidney and intestine and that in the absence of Fxralpha, mice waste sulfate into the urine. Thus, Fxralpha is necessary for normal sulfate homeostasis in vivo.

Sulfate ions in mammalian physiology: lessons from sulfate transporter null mice

The NaS1 sulfate transporter maintains serum sulfate levels. Several polymorphisms (SNPs) exist in human NaS1, including R12X and N174S. NaS1 null (Nas1–/–) mice exhibit hyposulfatemia and a decreased sulfate content of intestinal mucins. This is relevant to gastrointestinal (GI) physiology because sulfate plays a role in protecting mucins from degradation. Our aims were to functionally characterise human NaS1 SNPs and to determine the effects of hyposulfatemia in Nas1–/– mice on GI physiology. Compared to wild-type NaS1, R12X and N174S led to 100% & 60% loss of function, respectively. Nas1–/– and Nas1+/+ mice were challenged orally with C. jejuni bacteria or dextran sodium sulfate (DSS), a model for inducing colitis. C. jejuni colonised stomach and intestines of Nas1–/– (n=16) and Nas1+/+ (n=16) mice. One Nas1+/+ and 9 Nas1–/– mice developed systemic infection. DSS treatment (2.5% 7 days) led to shorter intestines (by 15% P<0.01) and decreased hematocrit (by 25% P<0.05) in Nas1–/– mice, suggesting hyposulfatemia enhances DSS-induced colitis and intestinal bleeding. In summary, we characterized 2 loss of function SNPs in human NaS1 and showed that Nas1–/– mice have reduced defence from mucosal bacterial infection, and are more susceptible to DSS-induced colitis. These findings underline the importance of sulfated mucins in intestinal barrier function, and prompt studies of GI mucins in humans with NaS1 SNPs.

Maternal nerve growth factor serum levels in the perinatal period

Neurotrophins, such as nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), are potent modulators of neuronal and immune function, and have been implicated recently in diseases associated with pregnancy. In contrast to serum BDNF, which is reportedly suppressed in the perinatal period, regulation of NGF in the perinatal period is unknown. In this study, serum NGF concentrations were measured in 40 pregnant (follow-up: 30th and 37th week of gestation, 1 week and 8 weeks after childbirth) and 40 non-pregnant women. Maternal NGF serum levels did not differ significantly from controls (median: 7.6 pg NGF/ml serum) neither before nor after childbirth, although there was a trend towards increased NGF concentrations at the 37th week of gestation (median: 12.5 pg NGF/ml serum) and 1 week after childbirth (median: 11.6 pg NGF/ml serum). There was no association of maternal NGF with 17β-estradiol, progesterone, dehydroepiandrosterone sulfate (DHEAS) and cortisol concentrations in maternal serum, or maternal depression, as measured by the Edinburgh Postnatal Depression Scale (EPDS). In the non-pregnant control group, NGF serum concentrations were negatively correlated with the number of days since the first day of the menstrual cycle ( r = −0.32, p < 0.05). In conclusion, NGF is not altered during normal pregnancy on a systemic level. In addition, NGF displays a different regulation compared with BDNF during the menstrual cycle.

Pubertal Development Predicts Resistance to Infection and Reinfection with Schistosoma japonicum

In communities where Schistosoma species are endemic, the prevalence and intensity of schistosomiasis is disproportionately high among children, compared with adults. This epidemiologic pattern is consistent with either the slow development of resistance or the requirement of host developmental changes for the expression of resistance. We enrolled 87 individuals aged 7-18 years who did not have Schistosoma japonicum infection and 641 individuals aged 7-30 years with S. japonicum infection, all of whom reside in 3 villages in Leyte, Philippines. At baseline, S. japonicum infection was assessed by Kato-Katz thick-smear stool examination, and the levels of the pubertal hormone dehydroepiandrosterone sulfate (DHEA-S) in serum were determined. Individuals with S. japonicum infection were treated with praziquantel, after which stool examination and DHEA-S level measurement were performed every 3 months for 18 months. In cross-sectional analyses, the intensity of infection among individuals with high DHEA-S levels was 43% lower (28 eggs per g, n = 243), compared with individuals with low DHEA-S levels (50 eggs per g, n = 242), even after adjusting for age, sex, and village (P = .01). Following praziquantel treatment, increased DHEA-S levels were associated with resistance to reinfection (P = .006). The intensity of reinfection among individuals with high DHEA-S levels was 42% lower, compared with individuals with low DHEA-S levels, even after adjusting for age, baseline intensity of S. japonicum infection, village, sex and water contact (P < .001). Increased DHEA-S levels in serum, a marker for adrenal development, is associated with reduced S. japonicum infection and reinfection, even after adjusting for age and, by proxy, cumulative exposure. These data suggest that an intrinsic property of host pubertal development mediates, in part, the resistance to infection observed in older individuals.

Fasting serum sulfate levels before and after development of osteoarthritis in participants of the veterans administration normative aging longitudinal study do not differ from levels in participants in whom osteoarthritis did not develop

To determine whether the development of osteoarthritis (OA) in men over a 33-year period is related to lower sulfate levels in stored serum collected during that time interval. Stored serum samples from participants in the Veterans Administration Normative Aging Study were assayed for sulfate by ion-exchange chromatography. Samples had been obtained every 3-5 years during part or all of a 33-year portion of the study. Sulfate levels were determined in serum from all participants who underwent knee replacement surgery and had evidence of radiographic hand OA, from some of the participants who had evidence of radiographic hand OA but had not undergone knee replacement surgery, from all participants who underwent knee replacement surgery but had no evidence of radiographic hand OA, and from age-matched participants who had no evidence of OA by history, physical examination, or hand radiography. Serum sulfate levels in participants, with or without radiographic hand OA and/or knee replacements, who were ages 34-72 years at the first examination, ranged from 0.21 mM to 0.51 mM over the course of a maximum of 33 years. Both the overall mean and median sulfate levels rose from 0.32 mM at age 40-50 years to 0.38 mM at age 70-80 years, and the overall mean and median for all ages was 0.36 mM. There were no significant differences in sulfate levels between subjects in any of the 4 groups. There was no evidence of a relationship between these serum sulfate levels and the development of OA. However, all samples were collected after overnight fasting, and no participant was younger than age 34 years at the initiation of the study. It remains to be determined whether differences in the time of ingestion of daily dietary protein providing sulfate are related to the development of OA, or whether sulfate levels measured at an earlier age could be a factor.

Marked attenuation of production of collagen type I from cardiac fibroblasts by dehydroepiandrosterone

Dehydroepiandrosterone (DHEA) is a type of adrenal steroid. The concentrations of DHEA and its sulfate (DHEA-S) in serum reach a peak between the ages of 25 and 30 yr and thereafter decline steadily. It was reported that DHEA-S concentration in humans is inversely related to death from cardiovascular diseases. In this study, we examined the effects of DHEA on regulation of collagen mRNA and collagen synthesis in cultured cardiac fibroblasts. Treatment with DHEA (10(-6) M) resulted in a significant decrease in procollagen type I mRNA expression compared with controls. This was accompanied by a significant decrease in procollagen type I protein accumulation in the medium and also a significant decrease in procollagen type I protein synthesis in the cellular matrix. Furthermore, to confirm in vitro results, we administered DHEA to Sprague-Dawley rats, which were treated with angiotensin II for 8 wk to induce cardiac damage. Procollagen type I mRNA expression was significantly decreased and cardiac fibrosis significantly inhibited in DHEA-treated rat hearts without lowering the systolic blood pressure. These results strongly indicate that DHEA can directly attenuate collagen type I synthesis at the transcriptional level in vivo and in vitro in cardiac fibroblasts.

P70 Quantitation and relative chain length determination of hyaluronan (HA) and chondroitin sulfate (CS) in serum and synovial fluid

P70 Quantitation and relative chain length determination of hyaluronan (HA) and chondroitin sulfate (CS) in serum and synovial fluid

Enzyme immunoassays for the analysis of streptomycin in milk, serum and water: development and assessment of a polyclonal antiserum and assay procedures using novel streptomycin derivatives

A polyclonal antiserum to streptomycin was generated in sheep using a streptomycin-bovine serum albumin conjugate as immunogen. Streptomycin was linked to the carrier protein with cyanuric chloride using a new two-step conjugation method. Plate coating antigen conjugates of streptomycin and gelatine were prepared using either cyanuric chloride (homologous bridge) or 1,4-butanediol diglycidyl ether to provide an heterologous complex. The reagents enabled the generation of a specific antiserum with a titre of 1/40,000 and the development of a sensitive ELISA method suitable for the measurement of streptomycin sulfate in milk, serum and water samples. A minimum detection value of 1 ng mL(-1) and a dynamic range of 1 to 200 ng mL(-1) were demonstrated in the three matrices. No detectable cross reactivity with any of the common aminoglycosides was found except the related dihydrostreptomycin which gave a 75% cross reaction value. The details of the preparation of the hapten-protein conjugates, characterisation of the antiserum and assay construction and assessment methods are presented. The introduction of new coupling methods and antibody assessment provide improved basic methodologies necessary for the advancement of immuno-analysis of streptomycin, one of the most widely used antimicrobial substances.

Novel lipoidal derivatives of pregnenolone and dehydroepiandrosterone and absence of their sulfated counterparts in rodent brain

A new sample preparation method coupled to GC-MS analysis was developed and validated for quantification of sulfate esters of pregnenolone (PREG-S) and dehydroepiandrosterone (DHEA-S) in rat brain. Using a solid-phase extraction recycling protocol, the results show that little or no PREG-S and DHEA-S (<1 pmol/g) is present in rat and mouse brain. These data are in agreement with studies in which steroid sulfates were analyzed without deconjugation. We suggest that the discrepancies between analyses with and without deconjugation are caused by internal contamination of brain extract fractions, supposed to contain steroid sulfates, by lipoidal forms of PREG and DHEA (L-PREG and L-DHEA, respectively). These derivatives can be acylated very efficiently with heptafluorobutyric anhydride and triethylamine, and their levels in rodent brain (approximately 1 nmol/g) are much higher than those of their unconjugated counterparts. They are distinct from fatty acid esters, and preliminary data do not favor structures such as sulfolipids or sterol peroxides. Noncovalent interactions between steroids and proteolipidic elements, such as lipoproteins, could account for some experimental data. Given their abundance in rodent brain, the structural characterization and biological functions of L-PREG and L-DHEA in the central nervous system merit considerable attention.

Sulfur- and Selenium-Containing Compounds Derived from Natural and Synthetic Sources in Angiogenesis Modulation.

Increases in serum sulfate may explain some of the therapeutic effects of methylsulfonyl methane (MSM), dimethyl sulfoxide, and glucosamine sulfate. Organic sulfur, as sulfur-containing amino acids, can be used to increase synthesis of S-adenosylmethionine, glutathione, taurine, and N-acetylcysteine. MSM may be effective for the treatment of allergy, pain syndromes, athletic injuries, and bladder disorders. Epidemiological studies have suggested that brassica vegetables are protective against cancers of the lungs and alimentary tract. Cruciferous vegetables are the dietary source of glucosinolates, a large group of sulfur-containing glycosides. However, dosages, mechanisms of action, and rationales for the use of various sulfur-containing compounds from natural or food products need to be better defined. The low toxicological profiles of these sulfur compounds, combined with promising therapeutic effects, warrant continued preclinical and clinical investigations. In this current study the angiostatic (antiangiogenesis) efficacy for individual sulfur-containing compounds derived from natural or synthetic sources was determined using in vitro (endothelial cell [EC] tube formation assay) and in vivo (chick chorioallantoic membrane [CAM] model). In vitro EC tube formation studies demonstrated significant inhibition of fibroblast growth factor-2 (FGF-2) stimulated EC tube formation by the different sulfur- and selenium-containing compounds, with different potency depending on the oxidation state of the sulfur or selenium. Furthermore, utilizing the CAM model, similar inhibitory efficacy of FGF-2 induced angiogenesis. These data suggest that sulfur and selenium compounds derived from natural sources might be a useful therapy for the inhibition of angiogenesis associated with human tumor growth and other pathological angiogenesis-mediated disorders, such as ocular and inflammatory diseases.Antiangiogenesis Efficacy of Sulfone/Sulfoxide and Selenone/Selenoxide in the Chick Chorioallantoic Membrane ModelTreatmentBranch Points ± SEM% Inhibition ± SEMPBS control64.67 ± 3.00-----FGF-2 (1.5 ug/ml)178.20 ± 5.32-----FGF-2 + diphenyl sulfoxide (100 uM)145.65 ± 11.1034.12 ± 4.8FGF-2 + diphenyl sulfone (3 uM)82.56 ± 4.484.25 ± 3.6FGF-2 + dimethyl selenoxide (30 uM)120.86 ± 3.150.74 ± 2.7FGF-2 + dimethyl selenone (3 uM)71.63 ± 3.893.87 ± 3.3FGF = fibroblast growth factor; PBS = phosphate-buffered saline.

Critical role of vitamin D in sulfate homeostasis: regulation of the sodium-sulfate cotransporter by 1,25-dihydroxyvitamin D3.

As the fourth most abundant anion in the body, sulfate plays an essential role in numerous physiological processes. One key protein involved in transcellular transport of sulfate is the sodium-sulfate cotransporter NaSi-1, and previous studies suggest that vitamin D modulates sulfate homeostasis by regulating NaSi-1 expression. In the present study, we found that, in mice lacking the vitamin D receptor (VDR), NaSi-1 expression in the kidney was reduced by 72% but intestinal NaSi-1 levels remained unchanged. In connection with these findings, urinary sulfate excretion was increased by 42% whereas serum sulfate concentration was reduced by 50% in VDR knockout mice. Moreover, levels of hepatic glutathione and skeletal sulfated proteoglycans were also reduced by 18 and 45%, respectively, in the mutant mice. Similar results were observed in VDR knockout mice after their blood ionized calcium levels and rachitic bone phenotype were normalized by dietary means, indicating that vitamin D regulation of NaSi-1 expression and sulfate metabolism is independent of its role in calcium metabolism. Treatment of wild-type mice with 1,25-dihydroxyvitamin D3 or vitamin D analog markedly stimulated renal NaSi-1 mRNA expression. These data provide strong in vivo evidence that vitamin D plays a critical role in sulfate homeostasis. However, the observation that serum sulfate and skeletal proteoglycan levels in normocalcemic VDR knockout mice remained low in the absence of rickets and osteomalacia suggests that the contribution of sulfate deficiency to development of rickets and osteomalacia is minimal.

Hyposulfatemia, growth retardation, reduced fertility, and seizures in mice lacking a functional NaSi-1 gene.

Inorganic sulfate is required for numerous functions in mammalian physiology, and its circulating levels are proposed to be maintained by the Na+-SO42- cotransporter, (NaSi-1). To determine the role of NaSi-1 in sulfate homeostasis and the physiological consequences in its absence, we have generated a mouse lacking a functional NaSi-1 gene, Nas1. Serum sulfate concentration was reduced by >75% in Nas1-/- mice when compared with Nas1+/+ mice. Nas1-/- mice exhibit increased urinary sulfate excretion, reduced renal and intestinal Na+-SO42- cotransport, and a general growth retardation. Nas1-/- mouse body weight was reduced by >20% when compared with Nas1+/+ and Nas1+/- littermates at 2 weeks of age and remained so throughout adulthood. Nas1-/- females had a lowered fertility, with a 60% reduction in litter size. Spontaneous clonic seizures were observed in Nas1-/- mice from 8 months of age. These data demonstrate NaSi-1 is essential for maintaining sulfate homeostasis, and its expression is necessary for a wide range of physiological functions.

Simultaneous determination of androstenediol 3-sulfate and dehydroepiandrosterone sulfate in human serum using isotope diluted liquid chromatography–electrospray ionization-mass spectrometry

A simple method for simultaneous determination of androstenediol 3-sulfate (Adiol-3S) and dehydroepiandrosterone sulfate (DHEA-S) in human serum using isotope diluted liquid chromatography–electrospray ionization-ion trap-mass spectrometry (LC–ESI-ion trap-MS) was developed. After addition of deuterated internal standards ([ 2 H 5 ]Adiol-3S and [ 2 H 4 ]DHEA-S), human serum (100 μl) was deproteinized with acetonitrile and then applied to a solid-phase extraction cartridge, Oasis HLB. The obtained steroid sulfates fraction was washed with hexane and then analyzed by LC–ESI-MS operated in the negative ion mode. The quantification ranges of Adiol-3S and DHEA-S were 10–400 ng/ml and 0.05–8 μg/ml, respectively. The method does not require the chemical or enzymatic hydrolysis of the conjugates and purification with high-performance liquid chromatography, and shows satisfactory reproducibility and accuracy. The concentrations of these sulfates in the sera of healthy male volunteers ( n=14) were 19.2–245.3 mg/ml (Adiol-3S) and 0.175–5.16 μg/ml (DHEA-S), and those of patients with prostate cancer ( n=19) were 15.3–182.7 ng/ml (Adiol-3S; four samples, not detectable) and 0.110–2.421 μg/ml (DHEA-S).

Regulation of the mouse Nas1 promoter by vitamin D and thyroid hormone.

The renal sodium-sulfate cotransporter, NaS(i)-1, a protein implicated to control serum sulfate levels, has been shown to be regulated in vivo by 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) and tri-iodothyronine (T(3)). Recently, we cloned the mouse NaS(i)-1 gene ( Nas1) and in the present study identified a 1,25-(OH)(2)D(3)- and T(3)-responsive element located within the Nas1 promoter. Mutational analysis of the Nas1 promoter resulted in identification of a direct repeat 6-type vitamin-D-responsive element (DR6 VDRE) at -525 to -508 and an imperfect inverted repeat 0-type T(3)-responsive element (IR0 T(3)RE) at -436 to -425 which conferred 1,25-(OH)(2)D(3) and T(3) responsiveness, respectively. In summary, we have identified responsive elements that mediate the enhanced transcription of Nas1 by 1,25-(OH)(2)D(3) and T(3), and these mechanisms may provide important clues to the physiological control of sulfate homeostasis.

Two neuroactive steroids in midpregnancy as measured in maternal and fetal sera and in amniotic fluid.

Allopregnanolone and pregnenolone sulfate, which are neuroactive steroids that differentially modulate the sensitivity of GABA(A) and NMDA receptors, were measured simultaneously in maternal and fetal sera and in amniotic fluid for the first time during the second trimester of gravidity. The study included 33 pregnant women, who underwent cordocentesis due to suspicion of fetal abnormality or alloimmunization. Allopregnanolone concentrations in maternal and fetal sera were similar and close to the previously found levels in healthy controls at 20 weeks of gestation The levels of pregnenolone sulfate in maternal serum were 2-3 times higher and in fetal serum, as much as 10-25 times higher than those found by others in the serum of healthy, non-pregnant women. A positive correlation between maternal and fetal allopregnanolone indicate similar 5alpha-reductase activities or the efficient transport of allopregnanolone between the two subjects. No correlation of pregnenolone sulfate levels between mother and fetus was found. This finding suggests the autonomous production of pregnenolone sulfate in mother and fetus.

Studies on neurosteroids XIV. Levels of dehydroepiandrosterone sulfate in rat brain and serum determined with newly developed enzyme-linked immunosorbent assay.

An enzyme-linked immunosorbent assay (ELISA) of dehydroepiandrosterone sulfate (DHEAS), one of the neurosteroids, has been developed for measuring its brain and serum levels in rats without deconjugation. 11 alpha-Hemiglutaryloxy-DHEAS was newly synthesized, conjugated with bovine serum albumin (BSA), and immunized to rabbits for the production of anti-DHEAS antibodies. A bridge-heterologous ELISA system employing the sequential saturation method exhibited a high sensitivity with a midpoint of 100 pg. Although the antibody significantly cross-reacted with epiandrosterone sulfate, it easily discriminated the unconjugated steroids and pregnenolone sulfate, which is reported to exist in the brain at a much higher level when compared with DHEAS. The brain homogenate or serum was treated with hexane to remove the lipophilic compounds and purified with an OASIS HLB cartridge. The DHEAS levels were then determined by ELISA. The overall recovery rate through the pretreatment was a satisfactory and constant (81.8 +/- 3.4% for brain, 89.3 +/- 3.0% for serum, mean +/- standard deviation). This ELISA afforded a satisfactory serial dilution study and recovery test. The intra- and inter-assay coefficients of variation were lower than 13%, which showed the precision of the proposed method. The applied method showed that DHEAS was not detected in some brain samples and its levels were much lower than those previously reported and than its serum levels.

Prospective measurements of dehydroepiandrosterone sulfate in a cohort of elderly subjects: relationship to gender, subjective health, smoking habits, and 10-year mortality.

The decrease with age of the adrenal-secreted dehydroepiandrosterone sulfate (DHEAS) in serum has suggested that it may be causally related to longevity. For the PAQUID [People (Personnes) Aged (Agées) About What (Quid, in Latin)] cohort of elderly subjects, we have previously reported higher DHEAS in men than in women, a decrease with age and, among men, a negative correlation between the DHEAS level and mortality at 2 and 4 years. Here, with an 8-year followup in 290 subjects, we show a global decrease of 2.3% per year for men and 3.9% per year for women. However, in approximately 30% of cases, there was an increase of DHEAS. We observed no relationship between the evolution of DHEAS level and functional, psychological, and mental status, possibly because of selection by death. In women, no association was found between mortality and DHEAS level. In men, the relative risk (RR) of death was higher for the lowest levels of DHEAS (RR = 1.9, P = 0.007), with RR = 6.5, P = 0.003 for those under 70 years old, a result indicating heterogeneity of the population. There was an effect of subjective health on mortality that disappeared after adjustment of DHEAS levels, suggesting its relation with these DHEAS levels. Death RR was much higher in smokers with a low DHEAS level than in nonsmokers with high DHEAS (RR = 6.7, P = 0.001). We submit that the involvement of DHEAS is possibly different according to gender, that association between low DHEAS level and mortality only for men under 70 years old possibly reflects heterogeneity of the population, and that DHEAS level is a reliable predictor of death in male smokers.