Free Plasma Cortisol Concentrations Research Articles

Assessment of Plasma-Free Cortisol Concentrations by LC-MS/MS in Patients with Autonomous Cortisol Secretion.

Autonomous cortisol secretion (ACS) of an adrenal incidentaloma (AI) is associated with mild cortisol excess that could result in poor metabolic and cardiovascular outcomes. The biological activity of glucocorticoids depends on the unbound, free fraction. We aimed to evaluate plasma free cortisol (FC) concentrations in patients with ACS in this cross-sectional study. One hundred and ten AI patients in 3 groups; non-functioning (NFA, n=33), possible ACS (n=65), ACS (n=12) were enrolled. Following measurements were conducted: Clinical data and total serum cortisol (TC), plasma corticotrophin (ACTH), serum dehydroepiandrosterone sulfate (DHEA-S), cortisol after 1 mg dexamethasone by both immunoassay and LC-MS/MS (DexF), serum corticosteroid binding globulin (CBG), plasma dexamethasone concentration [DEX] and plasma FC by LC-MS/MS. Patients with ACS featured an unfavorable metabolic profile. Plasma [DEX] and serum CBG levels were similar between groups. Plasma FC was significantly higher in ACS when compared to NFA and possible ACS groups p<0.05 and p<0.01, respectively. In multiple regression analysis DexF (beta=0.402, p<0.001) and CBG (beta=-0.257, p=0.03) remained as the independent predictors of plasma FC while age, sex, BMI, smoking habit, and existing cardiovascular disease did not make a significant contribution to the regression model. In conclusion, the magnitude of cortisol excess in ACS could lead to increased plasma FC concentrations. Further studies in AI patients are needed to demonstrate whether any alterations of cortisol affinity for CBG exist and to establish whether plasma FC concentrations predict the unfavorable metabolic profile in ACS.

Physiological and Psychological Stress Responses to Labor and Delivery as Expressed by Salivary Cortisol: A Prospective Study

(Am J Obstet Gynecol. 2019;221:351.e1–351.e7) Pregnant women experience both physiological stress and psychological stress during labor and delivery, but research is limited on measurement of stress levels during labor. Previous studies on physiological stress during labor have found a significant increase of total and free plasma cortisol concentrations; this increase is considered to be crucial for helping labor progression and promoting maternal and fetal well-being. While there are few studies on psychological stress during labor, researchers found epidural anesthesia helps alleviate women’s anxiety. In addition, postpartum stress levels can affect breastfeeding duration and health care service usage. The objective of this prospective, observational study was to accurately record the levels of physiological and psychological stress during labor to determine their effect on obstetric and neonatal outcomes.

Is EIPH a misnomer and could it be hampering our attempts to understand the etiology of the condition?

Salivary cortisol provides information about free plasma cortisol concentration, and salivary sampling is a noninvasive well-tolerated procedure. The aim of this study was to validate a commercial enzyme immunoassay for the determination of salivary cortisol in donkeys. Saliva samples were collected in four donkey stallions on 13 nonconsecutive days at 8:30 AM to avoid circadian variation. Animals were already accustomed to be handled. Saliva was collected using a swab inserted at the angle of the lips, placed onto the tongue for 1 minute and returned into a polypropylene tube. Tubes were centrifuged, and at least 1 mL of saliva was aspirated from each sample and frozen at −20°C until analysis. A commercial enzyme immunoassay kit without extraction was used for determination of cortisol in saliva. Median cortisol concentrations with minimum and maximum value were calculated. Recovery of cortisol standard in donkey saliva was between 97.3% and 99.7%, and serial dilution of donkey saliva samples with assay buffer resulted in changes in optical density parallel to the standard curve. Cross-reactivity of the antiserum was 10.4% with 11-deoxycortisol, 5.2% with corticosterone, 0.4% with 11-deoxycorticosterone, 0.2% with cortisone, and <0.1% with testosterone, progesterone, and estradiol. The intra-assay coefficient of variation was 10.7%, the interassay variation was 8.0%, and the minimal detectable concentration was 0.01 ng/mL. The results of the present study demonstrate the validity of a commercial kit to determine the concentration of cortisol in donkey saliva as already reported in other species.

Determination of Salivary Cortisol in Donkey Stallions

Salivary cortisol provides information about free plasma cortisol concentration, and salivary sampling is a noninvasive well-tolerated procedure. The aim of this study was to validate a commercial enzyme immunoassay for the determination of salivary cortisol in donkeys. Saliva samples were collected in four donkey stallions on 13 nonconsecutive days at 8:30 AM to avoid circadian variation. Animals were already accustomed to be handled. Saliva was collected using a swab inserted at the angle of the lips, placed onto the tongue for 1 minute and returned into a polypropylene tube. Tubes were centrifuged, and at least 1 mL of saliva was aspirated from each sample and frozen at −20°C until analysis. A commercial enzyme immunoassay kit without extraction was used for determination of cortisol in saliva. Median cortisol concentrations with minimum and maximum value were calculated. Recovery of cortisol standard in donkey saliva was between 97.3% and 99.7%, and serial dilution of donkey saliva samples with assay buffer resulted in changes in optical density parallel to the standard curve. Cross-reactivity of the antiserum was 10.4% with 11-deoxycortisol, 5.2% with corticosterone, 0.4% with 11-deoxycorticosterone, 0.2% with cortisone, and <0.1% with testosterone, progesterone, and estradiol. The intra-assay coefficient of variation was 10.7%, the interassay variation was 8.0%, and the minimal detectable concentration was 0.01 ng/mL. The results of the present study demonstrate the validity of a commercial kit to determine the concentration of cortisol in donkey saliva as already reported in other species.

Analysis and validation of a new extended method for estimating plasma free cortisol including neutrophil elastase and competition from other steroids

A non-linear mechanistic model for the distribution of cortisol in plasma on free and bound forms is proposed. The influence of progesterone, testosterone and neutrophil elastase on the cortisol distribution in the blood is investigated.The activity of neutrophil elastase is directly included in the model with the concentration of elastase and the kinetic constants describing the activity of elastase collected in one single input variable. The model is very sensitive towards this input variable and fits data excellently, when it is allowed to be subject specific.The analysis shows that steroids such as testosterone with low affinity for corticosteroid-binding globulin (CBG) do not significantly influence the concentration of free cortisol. Progesterone has a high affinity for CBG, but low plasma concentrations compared to cortisol. Contrary to expectations, progesterone is shown to impact the distribution of cortisol in plasma both under circumstances with high levels as seen in pregnancy and during the normal menstrual cycle of women.Comparing the predictions of our model with predictions made with the equilibrium models by Coolens et al. [1], Dorin et al. [2] and Nguyen et al. [3] shows that the models differ considerably not only in their predictions for free cortisol, but also for cortisol on bound forms; i.e. bound to albumin, intact CBG and elastase-cleaved CBG.Disregarding some of the smallest terms of the model equations a reduced version of the model in form of a fourth order polynomial equation is obtained. The reduced version of the model performs almost identically to the full version and serves as a new formula for calculating the plasma free cortisol concentration.

Social Support Modulates Stress-Related Gene Expression in Various Brain Regions of Piglets.

The presence of an affiliative conspecific may alleviate an individual’s stress response in threatening conditions. However, the mechanisms and neural circuitry underlying the process of social buffering have not yet been elucidated. Using the domestic pig as an animal model, we examined the effect of a 4-h maternal and littermate deprivation on stress hormones and on mRNA expression of the glucocorticoid receptor (GR), mineralocorticoid receptor (MR), 11ß-hydroxysteroid dehydrogenase (11ß-HSD) types 1 and 2 and the immediate early gene c-fos in various brain regions of 7-, 21- and 35-day old piglets. The deprivation occurred either alone or with a familiar or unfamiliar age-matched piglet. Compared to piglets deprived alone, the presence of a conspecific animal significantly reduced free plasma cortisol concentrations and altered the MR/GR balance and 11ß-HSD2 and c-fos mRNA expression in the prefrontal cortex (PFC), amygdala and hypothalamus, but not in the hippocampus. The alterations in brain mRNA expression were particularly found in 21- or 35-day old piglets, which may reflect the species-specific postnatal ontogeny of the investigated brain regions. The buffering effects of social support were most pronounced in the amygdala, indicating its significance both for the assessment of social conspecifics as biologically relevant stimuli and for the processing of emotional states. In conclusion, the present findings provide further evidence for the importance of the cortico-limbic network underlying the abilities of individuals to cope with social stress and strongly emphasize the benefits of social partners in livestock with respect to positive welfare and health.

Reduced nocturnal ACTH-driven cortisol secretion during critical illness

Recently, during critical illness, cortisol metabolism was found to be reduced. We hypothesize that such reduced cortisol breakdown may suppress pulsatile ACTH and cortisol secretion via feedback inhibition. To test this hypothesis, nocturnal ACTH and cortisol secretory profiles were constructed by deconvolution analysis from plasma concentration time series in 40 matched critically ill patients and eight healthy controls, excluding diseases or drugs that affect the hypothalamic-pituitary-adrenal axis. Blood was sampled every 10 min between 2100 and 0600 to quantify plasma concentrations of ACTH and (free) cortisol. Approximate entropy, an estimation of process irregularity, cross-approximate entropy, a measure of ACTH-cortisol asynchrony, and ACTH-cortisol dose-response relationships were calculated. Total and free plasma cortisol concentrations were higher at all times in patients than in controls (all P < 0.04). Pulsatile cortisol secretion was 54% lower in patients than in controls (P = 0.005), explained by reduced cortisol burst mass (P = 0.03), whereas cortisol pulse frequency (P = 0.35) and nonpulsatile cortisol secretion (P = 0.80) were unaltered. Pulsatile ACTH secretion was 31% lower in patients than in controls (P = 0.03), again explained by a lower ACTH burst mass (P = 0.02), whereas ACTH pulse frequency (P = 0.50) and nonpulsatile ACTH secretion (P = 0.80) were unchanged. ACTH-cortisol dose response estimates were similar in patients and controls. ACTH and cortisol approximate entropy were higher in patients (P ≤ 0.03), as was ACTH-cortisol cross-approximate entropy (P ≤ 0.001). We conclude that hypercortisolism during critical illness coincided with suppressed pulsatile ACTH and cortisol secretion and a normal ACTH-cortisol dose response. Increased irregularity and asynchrony of the ACTH and cortisol time series supported non-ACTH-dependent mechanisms driving hypercortisolism during critical illness.

Development of a formula for estimating plasma free cortisol concentration from a measured total cortisol concentration when elastase-cleaved and intact corticosteroid binding globulin coexist

Cortisol bound to corticosteroid binding globulin (CBG) contributes up to 90% of the total cortisol concentration in circulation. Therefore, changes in the binding kinetics of cortisol to CBG can potentially impact on the concentration of free cortisol, the only form that is responsible for the physiological function of the hormone. When CBG is cleaved into elastase-cleaved CBG (eCBG) by the activity of neutrophil elastase, its affinity for cortisol is reduced. Therefore, when eCBG coexists with intact CBG (iCBG) in plasma, the calculation of free cortisol concentration based on the formulae that considers only one CBG pool with the same affinity for cortisol may be inappropriate. In this study, we developed in vivo and in vitro models of cortisol partitioning which considers two CBG pools, iCBG and eCBG, with different affinities for cortisol, and deduce a new formula for calculating plasma free cortisol concentration. The formula provides better estimates of free cortisol concentration than previously used formulae when measurements of the concentrations of the two CBG forms are available. The model can also be used to estimate the affinity of CBG and albumin for cortisol in different clinical groups. We found no significant difference in the estimated affinity of CBG and albumin for cortisol in normal, sepsis and septic shock groups, although free cortisol was higher in sepsis and septic shock groups. The in vivo model also demonstrated that the concentration of interstitial free cortisol is increased locally at a site of inflammation where iCBG is cleaved to form eCBG by the activity of elastase released by neutrophils. This supports the argument that the cleavage of iCBG at sites of inflammation leads to more lower-affinity eCBG and may be a mechanism that permits the local concentration of free cortisol to increase at these sites, while allowing basal free cortisol concentrations at other sites to remain unaffected.

Real-time free cortisol quantification among critically ill children

Ascertainment of adrenal function assessing free rather that total cortisol may be beneficial for the diagnosis of critical illness-related cortisol insufficiency. We hypothesized that centrifugal ultrafiltration would provide timely free cortisol data that highly correlated with the gold standard, but logistically cumbersome, equilibrium dialysis technique when the free cortisol fractions were identically quantified by chemiluminescence immunoassay. We also hypothesized that free cortisol would correlate with illness severity in a large cohort of critically ill children. Prospective, multi-institutional, observational cohort investigation. Seven pediatric intensive care units within the Eunice Kennedy Shriver National Institute of Child Health and Human Development Collaborative Pediatric Critical Care Research Network. One hundred sixty-five critically ill children across the spectrum of illness severity. Blood sampling. Time to derive plasma free cortisol concentrations after centrifugal ultrafiltration or equilibrium dialysis fractionation with chemiluminescence immunoassay was approximately 2 vs. approximately 24 hrs, respectively. Using centrifugal ultrafiltration, mean plasma free cortisol was 4.1 ± 6.7 μg/dL (median, 1.6 μg/dL; range, 0.2-43.6 μg/L), representing an average of 15.2 ± 9.4% of total cortisol. Nearly 60% of subjects exhibited free cortisol <2 and 30% <0.8 μg/dL, previously suggested threshold concentrations for defining critical illness-related cortisol insufficiency. Plasma-free cortisol concentrations comparing centrifugal ultrafiltration vs. equilibrium dialysis fractionation demonstrated a strong correlation (R2 = 0.97). For free cortisol <2 μg/dL, Bland-Altman analysis revealed minimal negative bias for the centrifugal ultrafiltration technique. Illness severity assessed by Pediatric Risk of Mortality III correlated moderately with free cortisol and percent total cortisol as free cortisol. Determination of centrifugal ultrafiltration fractionated free cortisol was fast and results correlated highly with equilibrium dialysis fractionated free cortisol. Many children exhibited free cortisol <2 and <0.8 μg/dL but did not demonstrate clinical evidence of critical illness-related cortisol insufficiency. This study ascertains that real-time free cortisol quantification is feasible to potentially help guide clinical decision-making for cortisol replacement therapy in the pediatric intensive care unit.

Characterising adrenal function using directly measured plasma free cortisol in stable severe liver disease

Adrenal insufficiency (AI) has been reported in patients with advanced liver disease. Diagnosing AI is problematic owing to controversies in using total serum cortisol as a measure of adrenal function. No published data exist on directly measured plasma free cortisol (PFC) in patients with liver disease. This prospective study compared serum total and measured plasma free cortisol to evaluate adrenal function in clinically stable cirrhotic patients and healthy controls. Cortisol levels were measured at baseline and following 250 μg corticotrophin. AI was defined by total cortisol increments (delta cortisol) of less than 250 nmol/L, or a peak total cortisol under 500 nmol/L after cosyntropin. We used a peak plasma free cortisol concentration of 33 nmol/L as the threshold for AI. Forty-three consecutive patients and 10 healthy controls were studied. Cirrhotic patients had significantly lower peak (526 vs. 649 nmol/L, p=0.004) and delta total cortisol (264 vs. 397 nmol/L, p = 0.002) responses compared to healthy controls. However, basal plasma free cortisol was higher in patients (10.9 vs. 6.4 nmol/L, p = 0.03), and there were no differences in peak plasma free cortisol (p = 0.69) between the two groups. The prevalence of AI using total cortisol criteria was 58% compared to 12% using free cortisol (p<0.001). In patients with stable severe liver disease, a significant discrepancy exists between the rates of diagnosis of AI using the total and free cortisol criteria. We would advise caution in the interpretation of adrenal function testing using total cortisol measurements in this group.

Differential effects of chronic ethanol exposure on cytochrome P450 2E1 and the hypothalamic–pituitary–adrenal axis in the maternal–fetal unit of the guinea pig

BackgroundEthanol neurobehavioural teratogenicity is a leading cause of developmental mental deficiency, in which the hippocampus is a target site of injury. The multi-faceted mechanism of ethanol teratogenicity is not completely understood. This study tested the hypothesis that chronic ethanol exposure (CEE), via chronic maternal ethanol administration, increases cytochrome P450 2E1 (CYP2E1) expression and alters hypothalamic–pituitary–adrenal (HPA) axis activity in the maternal–fetal unit during the third-trimester-equivalent of gestation. MethodsPregnant Dunkin–Hartley-strain guinea pigs received daily oral administration of ethanol (4g ethanol/kg maternal body weight) or isocaloric-sucrose/pair-feeding (control) throughout gestation (term, about gestational day (GD) 68). On GD 45, 55 and 65, pregnant animals were euthanized 2h after the last daily dose. Maternal and fetal body weights and fetal hippocampal brain weight were determined. Maternal and fetal samples were collected for the determination of liver CYP2E1 enzymatic activity and plasma free cortisol and ACTH concentrations. ResultsCEE, with maternal blood ethanol concentration of 108–124mg/dl at 2h after the last dose, decreased fetal hippocampal weight only at GD 65 and had no effect on fetal body weight compared with control. CYP2E1 activity increased with gestational age in the fetal liver microsomal and mitochondrial fractions. CEE increased CYP2E1 activity in the microsomal and mitochondrial fractions of maternal liver at the three gestational ages and in both hepatic subcellular fractions of the GD 65 fetus compared with control. There was a gestational-age-dependent increase in maternal and fetal plasma free cortisol concentrations, but no effect of CEE compared with control. Maternal and fetal plasma ACTH concentrations were unaffected by CEE compared with control, and were virtually unchanged during the third-trimester-equivalent that was studied. ConclusionThese data demonstrate that, in the pregnant guinea pig, this CEE regimen increases liver CYP2E1 activity, without affecting HPA axis function, in the maternal–fetal unit during near-term gestation. The CEE-induced increase in liver CYP2E1 activity and potential oxidative stress in the maternal–fetal unit may play a role in the pathogenesis of ethanol teratogenicity.

Measurement of tissue cortisol levels in patients with severe burns: a preliminary investigation

IntroductionThe assessment of adrenal function in critically ill patients is problematic, and there is evidence to suggest that measurement of tissue glucocorticoid activity may be more useful than estimation of plasma cortisol concentrations. Interstitial cortisol concentrations of cortisol represent the available pool of glucocorticoids able to enter the cell and bind to the glucocorticoid receptor. However the concentrations of plasma cortisol may not accurately reflect interstitial concentrations. We elected to perform a preliminary study into the feasibility of measuring interstitial cortisol by microdialysis, and to investigate the relationship between total plasma cortisol, free plasma cortisol and interstitial cortisol in patients with severe burns.MethodsA prospective observational study carried out in a tertiary intensive care unit. Ten adult patients with a mean total burn surface area of 48% were studied. Interstitial cortisol was measured by microdialysis from patient-matched burnt and non-burnt tissue and compared with that of 3 healthy volunteers. Plasma sampling for estimations of total and free cortisol concentrations was performed concurrently.ResultsIn the burn patients, mean total plasma and free plasma cortisol concentrations were 8.8 +/- 3.9, and 1.7 +/- 1.1 mcg/dL, (p < 0.001), respectively. Mean subcutaneous microdialysis cortisol concentrations in the burn and non-burn tissue were 0.80 +/- 0.31 vs 0.74 +/- 0.41 mcg/dL (p = 0.8), respectively, and were significantly elevated over the mean subcutaneous microdialysis cortisol concentrations in the healthy volunteers. There was no significant correlation between total plasma or free plasma and microdialysis cortisol concentrations. Plasma free cortisol was better correlated with total burn surface area than total cortisol.ConclusionsIn this preliminary study, interstitial cortisol concentrations measured by microdialysis in burnt and non-burnt skin from patients with severe thermal injury are significantly elevated over those from healthy volunteers. Plasma estimations of cortisol do not correlate with the microdialysis levels, raising the possibility that plasma cortisol may be an unreliable guide to tissue cortisol activity.

Sarcoidosis in Pregnancy and Postpartum Period

Sarcoidosis is a multisystemic disease of unknown etiology, characterized by granulomatous inflammation. It typically presents between the ages of 20 to 40 years old. An estimated 0.02% to 0.05% of pregnancies occur in patients with sarcoidosis. Although fetal loss has been reported in mothers with sarcoidosis, limited studies do not suggest an increase risk of fetal or neonatal complications. Several reports suggest an improvement of sarcoidosis during pregnancy. Sarcoidosis, a Th1-mediated disease, seems to follow the same course as rheumatoid arthritis during pregnancy and postpartum. Estrogen levels increase during pregnancy, resulting in a decreased Th1-mediated immune response, which can improve active sarcoidosis. Free plasma cortisol concentrations increase in pregnancy, with plasma levels 2- to 3-fold higher than those of non-pregnant controls, suggesting greater tissue exposure to glucocorticoids during pregnancy. This may result in decreased granulomatous inflammation with improvement in symptoms and clinical findings. During the postpartum period, when free cortisol levels return to the prior non-pregnant levels, reactivation of sarcoidosis can occur. The majority of patients with sarcoidosis will either have a stable disease or will experience improvement in their symptoms. A small group of pregnant mother with active sarcoidosis, however, may develop more progressive disease during pregnancy. The manifestations, course of disease, and treatment options during pregnancy and postpartum period are discussed. Keywords: Sarcoidosis, pregnancy, corticosteroids, lung function, Th1 cells

Production of gender-specific morning salivary cortisol reference intervals using internationally accepted procedures

Salivary cortisol concentrations correlate well with biologically active unbound free plasma cortisol concentrations. Despite its practical and analytical advantages, salivary cortisol measurement has been used mainly as a research tool rather than for the routine evaluation of adrenal function. This may be partly explained by the lack of robust reference data in the literature. Using the recommended procedures for the production of reference intervals published by the International Federation of Clinical Chemistry, we aimed to produce morning salivary cortisol reference intervals for males and females. Salivary cortisol was measured in 496 specimens collected from 248 reference individuals (128 males, median age 41 years, range 16-86; and 120 females, median age 44 years, range 16-98) attending an otorhinolaryngology clinic. Reference individuals mailed saliva specimens sampled on two consecutive mornings to our laboratory, where cortisol concentrations were measured. Statistical analysis showed no significant correlation with age or body mass index. The following 95% gender-partitioned reference intervals were produced: males 10.9-40.3 nmol/l; and females 9.3-40.3 nmol/l. Knowledge of these salivary cortisol reference intervals helps us monitor the adrenal function of outpatients using topical intranasal glucocorticoids for rhinosinusitis.

Perturbations in factors that modulate osteoblast functions in vitamin B6 deficiency

It was hypothesized that the widespread structural defect of collagen in connective tissue of vitamin B6 deficient-animals and the consequent alteration in bone biomechanical properties cause an additional stress to their inflamed swollen tibiotarsometatarsal joints. The present study showed a 32% elevation (P < 0.02) in mean plasma free cortisol concentration. Vitamin D metabolism was impaired but without changing plasma calcium homeostasis and bone mineral content. Mean plasma calcitriol [1,25(OH)2D] concentration was significantly reduced (P < 0.001). Because plasma calcidiol concentration did not change, we speculated that either renal 25-hydroxycalciferol-1alpha-hydroxylase activity was reduced or 1,25(OH)2D turnover was increased. Plasma osteocalcin, an index of osteoblast function related to bone formation, was significantly decreased (P < 0.05). This adverse effect on osteoblasts was consistent with the reduction of bone specific alkaline phosphatase activity (another index of bone formation) found in a previous study. The excess of cortisol may have impaired these bone cells functions directly and (or) indirectly via the decline in calcitriol synthesis. Plasma hydroxyproline concentrations in B6-deficient animals were found to be significantly reduced (P < 0.001), suggesting that cortisol in excess had also a suppressive effect on another hydroxylase, namely tissue (mainly bone and liver) prolyl hydroxylase. The bone uncoupling (in formation and resorption) associated with vitamin B6 deficiency seems to be due to secondary hypercortisolism and (or) another unknown factors but not related to a change in bone modulators such as IGF-1 and eicosanoids.

The Effects of Handling on Behavioural and Physiological Responses to Housing in Tether-Stalls Among Pregnant Pigs

AbstractTwenty-four primiparous pregnant pigs were randomly assigned to three handling treatments: Minimal, Positive and Negative. The pigs were moved individually to indoor concrete-floored partial stalls with neck-tethers, 2 days before handling commenced. Positive (stroking and patting on approach to an experimenter) and Negative (brief electric shock of &lt; Is when failing to withdraw from the outstretched hand of an experimenter) handling was imposed for 3min day-1and the amount of physical contact between handler and pig was recorded. The Minimal treatment group was subjected to routine husbandry practices only. After 3 weeks of the handling treatments and tether-housing, all pigs were catheterized under full surgical anaesthesia. The pigs were allowed 4 days of recovery before collecting the following data: daytime plasma Cortisol concentration profiles, behavioural responses to a human in an arena test, Cortisol responses to human proximity, Cortisol responses to an A CTH-challenge and immunological responses to an injected mitogen.In the Positive treatment, the amount of physical contact between pig and handler increased during the course of the experiment, while the amount of physical contact did not change in the Negative treatment. There were no effects of treatment on behavioural responses in the arena test. The average daytime concentration of free plasma Cortisol was lower in the Positive treatment than in the Negative or Minimal treatments. The Positive treatment also showed lower total and free plasma Cortisol concentrations pre- and posthuman proximity when compared with the Negative treatment. No differences were found between treatments in total and free plasma Cortisol concentrations following an ACTH challenge. The immunological response was greater in the Positive treatment compared with the Negative treatment and tended to be greater when compared with the Minimal treatment.It was concluded that the nature of the human-animal relationship affected the physiological stress responses of pregnant pigs to tether-housing. Indications are that a positive human-animal relationship would obviate some of the negative effects of being kept in tether-stalls by lowering the basal cortisol concentration and by increasing immunological responsiveness.

Divergent changes in plasma ACTH and pituitary POMC mRNA after cortisol administration to late-gestation ovine fetus.

Plasma concentrations of cortisol and adrenocorticotropic hormone (ACTH) rise in the late-gestation sheep fetus at approximately the same time as there is an increase in the plasma levels of corticosteroid-binding globulin (CBG). We hypothesized that intrafetal cortisol infusion during late pregnancy would stimulate an increase in fetal plasma CBG, which in turn would bind cortisol and diminish glucocorticoid negative-feedback regulation of the fetal pituitary, leading to an increase in plasma ACTH concentrations. Cortisol was infused into chronically catheterized fetal sheep beginning at 126.1 +/- 0.5 days of gestation and continued for 96 h. Control fetuses were infused with saline. In cortisol-infused fetuses, the plasma cortisol concentrations rose significantly from control levels (4.4 +/- 0.6 ng/ml) to 19.3 +/- 3.1 ng/ml within 24 h and remained significantly elevated throughout the infusion period. Plasma immunoreactive (i.r.) ACTH concentrations were significantly elevated in cortisol-infused fetuses within 24-48 h and remained significantly higher than in controls throughout the 96-h experimental period. Plasma free cortisol concentrations increased 10-fold and remained significantly elevated in cortisol-infused animals, despite a rise in plasma corticosteroid-binding capacity. Levels of pituitary proopiomelanocortin (POMC) mRNA in the fetal pars distalis and pars intermedia were 96 and 38% lower, respectively, after 96 h of cortisol infusion. Therefore physiological elevations of plasma cortisol, in the late-gestation ovine fetus, lead to increases in mean plasma irACTH concentrations, but this is not associated with increases in fetal pituitary POMC mRNA levels.

Effects of Major Depression, Aging and Gender upon Calculated Diurnal Free Plasma Cortisol Concentrations: a Re-Evaluation Study

Depression, aging and female gender are associated with increased diurnal concentrations of total plasma cortisol. For the physical effects of hypercortisolemia, however, it is generally assumed that free rather than total plasma cortisol concentrations are of importance. Herein, we report a mathematical approach to determine free plasma cortisol concentrations on the basis of total cortisol, corticosteroid binding-globulin (CBG) and albumin plasma concentrations. This approach was used to re-evaluate two sets of data in order to estimate the effect of depression as well as the effect of aging and gender upon free plasma cortisol concentrations. Comparing male depressed patients with healthy controls, we found 24-hour free cortisol minima (MIN: 4.1 ± 1.8 vs. 1.6 ± 1.1 nmol/l, p < 0.0001), mean (MEAN: 25.5 ± 6.7 vs. 10.4 ± 2.7 nmol/l, p < 0.0001) and maximal (MAX: 85.3 ± 23.3 vs. 45.2 ± 15.8 nmol/l, p < 0.0001) concentrations to be significantly increased in depressed patients. In general, the impact of depression upon total plasma cortisol were not only maintained, but stronger regarding free plasma cortisol. Also, age was associated with free plasma cortisol MIN (F1.30= 10.8, p < 0.003) and free plasma cortisol MEAN (F1.30 = 8.9, p < 0.006). All effects of age upon total plasma cortisol were generally also found in free plasma cortisol, though with less impact. No effect of gender upon any of the given free plasma cortisol outcome variables was found. Taken together, our re-evalution clearly shows not only depression but also aging to be associated with increases in free plasma cortisol concentrations. This finding is in line with the observation that in both conditions medical problems triggered and/or maintained by glucocorticoids (e.g. osteoporosis) are frequently seen.

Overnight metabolic fuel deficiency in patients treated conventionally for hypopituitarism.

Hormone replacement in hypopituitary adults attempts to reproduce normal physiology. Conventional regimens fail to mimic normal hormone profiles over 24 hours. To investigate the metabolic consequences of conventional hormone replacement in hypopituitary adults by measuring circulating levels of the major fuels, glucose, non-esterified fatty acids (NEFA), glycerol and 3-hydroxybutyrate (3-OHB) over 24 hours in hypopituitary subjects and controls. Ten GH and adrenocorticotrophin deficient hypopituitary adults on conventional replacement and 13 controls matched for age, sex and body mass index were studied. The patients received replacement with hydrocortisone twice daily (at 0730 and 1730 h; mean (range) daily dose 22 (10-30) mg/24 h) but not with GH. Other hormones were replaced as clinically necessary. Circulating glucose, NEFA, glycerol and 3-OHB levels were measured over 24 hours together with concentrations of cortisol (total and free), GH and insulin, and urinary free cortisol. Levels of glucose, NEFA and 3-OHB were lower in patients than controls (mean +/- SEM) (4.3 +/- 0.1 vs 5.3 +/- 0.1 mmol/l, P = 0.0001; 291 +/- 46 vs 448 +/- 48 mumol/l, P = 0.015; 78 +/- 8 vs 136 +/- 24 mumol/l, P = 0.035, respectively) before breakfast. This decrease in glucose, NEFA and 3-OHB was observed in the patient group throughout the night, from midnight to breakfast. For NEFA, the decrease persisted throughout the 24 hours. Glycerol did not differ significantly in patients and controls. Integrated levels of total and free plasma cortisol, and 24-hour urine cortisol excretion, were normal in patients but total and free plasma cortisol concentrations overnight were markedly decreased (overnight area under the curve (AUC) of total cortisol: 440 +/- 154 vs 1593 +/- 267 nmol/l h, P = 0.0024; overnight AUC of free cortisol: 24 +/- 8 vs 161 +/- 26 nmol/l h, P = 0.0001). GH levels were low throughout the whole 24 hours in the patient group (24-hour AUC: 10.6 +/- 5.1 vs 74.6 +/- 19.6 mU/l h, P = 0.008). Hypopituitary adults on conventional hormone replacement regimens have low concentrations of metabolic fuels, glucose, non-esterified fatty acids and 3-hydroxybutyrate throughout the night, possibly related to GH deficiency or to decreased overnight circulating cortisol levels. This overnight fuel deficiency may underlie the mechanism for the non-specific symptoms, such as fatigue and headache in the early morning, which are frequent in this group of patients.

The welfare of pigs in two farrowing/lactation environments: cortisol responses of sows

Plasma free cortisol concentrations were measured in 24 primiparous sows housed from Day 104 after mating up to Day 2 after parturition. Concentrations were measured in 22 of these sows, from Days 2–29 of lactation, housed in either commercial farrowing crates (Crate treatment) or farrowing pens with straw added (Pen treatment). The mortality and growth data of litters from the time of birth to weaning were also recorded. The mean plasma free cortisol concentration was higher in sows housed in the Crate treatment, compared with sows housed in the Pen treatment, on the first day of treatment (8.2 and 5.8 nmol l −1, respectively, P < 0.05) and on Day 28 of lactation (5.6 and 4.0 nmol l −1, respectively, P < 0.05). The data suggest that housing treatment around parturition and during lactation may have only limited effects on the welfare of sows, providing that lactation is no longer than 28 days. On Days 21 and 24, there were no effects of treatment on free cortisol concentrations and no response to the exogenous adrenocorticotrophic hormone (ACTH), respectively. However, on Day 28, there were mean increases in the concentrations of free cortisol of approximately 150% and approximately 60%, respectively; these may have been the result of unavoidable attention of the piglets to the sows.