Determination Of Cortisol Concentrations Research Articles

Effects of weaning and inactivated Lactobacillus helveticus supplementation on dairy calf behavioral and physiological indicators of affective state

The objectives of this study were to determine if weaning would induce behavioral and physiological indicators of a negative affective state, and if supplementation of inactivated Lactobacillus helveticus (ILH) to dairy calves would reduce those indicators of negative affect during weaning. Male Holstein calves (n = 23) were enrolled in the study on d 1 of life. The calves were housed in individual pens in 1 of 4 rooms for the 42 d study. Calves began a stepdown weaning from 9 L/d of milk replacer (MR), at 150 g of MR powder/L, on d 35 and received 6 L/d on d 35 - 36, 3 L/d on d 37 - 38, and 0.4 L/d on d 39 - 42. The MR was divided between 3 meals/d until the last 0.4 L/d phase which was divided between 2 meals/d. Calves had ad libitum water access throughout the study and calf starter from d 28 onwards. Within room, calves were assigned to 1 of 2 treatments: 1) control (CON; n = 11) and 2) 5 g of ILH/d split over and mixed into the 0800 h and 2000 h milk feedings from d 3-42 (ILH; n = 12). Lying behavior was recorded using HOBO data loggers from d 21-41. On d 33, 37 and 41, infrared eye images were taken to determine maximum eye temperature (MET), saliva samples were collected to determine cortisol concentration, and play assessments were conducted to quantify play behavior. On d 34, 38, and 42, blood samples were collected to determine blood serotonin concentration, whereas on d 38 and 39, calves were tested with a cognitive task. A subset of calves (n = 5/treatment) were euthanized to collect gut and brain tissue samples for serotonin concentration on d 43. Weaning resulted in fewer (d 37-41, tendency: d 36), but longer (d 38-41, tendency: d 37), lying bouts and reduced play (d 41), although no changes in lying time, MET, saliva cortisol, nor blood serotonin were detected with initiation of weaning. Supplementation of ILH was associated with lower lying time throughout the study, and reduced play duration and higher salivary cortisol and MET during weaning. No differences in lying bouts, play count, blood and tissue (colon, ileum, prefrontal cortex and brain stem) serotonin concentration, and time to complete the cognitive task were detected between the treatments. Overall, weaning induced behavioral changes indicative of negative affective state, and some behavioral differences were observed with ILH supplementation both before and during weaning, with some physiological changes observed during weaning.

Methods for determining cortisol concentrations in animals

The steroid hormone cortisol is the end product of the hypothalamic-pituitary-adrenal (HPA) axis. Recently, cortisol has been considered as a potential biomarker for identifying stress, because it is directly related to the body›s response. The most common methods for determining endocrine status, including cortisol, are enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA), as they are the most reliable and have high sensitivity. Cortisol levels are determined mainly in blood, saliva, serum and urine. The concentration of this hormone in these samples is related to the circadian rhythm and fluctuates throughout the day. Therefore, samples of this biomaterial allow the analysis of acute dynamic changes in cortisol. The effects of chronic stress require assessment of long-term stress levels. Hair (fur) cortisol concentrations are increasingly used as a biomarker of long-term stress. Hair cortisol analysis has thus been a major methodological advance because it provides retrospective assessment of cumulative levels over long periods of time (from days to months). Over the past decade, the validity of hair (fur) cortisol levels as an indicator of long-term levels has been established. In addition, this material is the most stable over time and resistant to environmental influences. Measuring hair cortisol levels is an innovative method for determining long-term increases in cortisol levels, independent of many methodological difficulties and not associated with other matrices. Interest in hair as an object of endocrine activity is due to various unique features that provide many advantages over other biomaterials.

291 Effect of Early vs. Late Maturing Sire Lines and Creep Feeding on the Stress Response, Intestinal Permeability, and Growth Performance of Nursery and Finishing Pigs

Abstract A total of 21 litters (early or late maturing Duroc × DNA 241) resulting in 247 pigs were used in a 170-day trial to determine the effect of sire lines selected for either early or late maturing growth rates and creep feeding on the stress response, intestinal permeability, and growth performance of nursery and finishing pigs. Treatments were arranged in a 2 x 2 factorial with main effect of Duroc sire line (early or late maturing) and creep feeding (with or without starting 14 d before weaning). In the nursery, there were 4 or 5 pigs per pen and 12 or 13 replications per treatment. Thirty minutes after weaning (initially 6.4 kg BW), 7 gilts per treatment were used to determine cortisol concentrations in response to weaning. Blood cortisol concentrations were increased (P < 0.01; 104.0 v. 83.5 ng/mL) in late maturing pigs compared with early maturing pigs. After almost no feed consumption by either group on d 1, late maturing pigs had decreased daily feed intake (P < 0.001) than early maturing pigs from days 2 to 14 post-weaning. Likewise, a greater percentage (P < 0.001) of late maturing pigs had BW less than their weaning weights on d 3 post-weaning compared with early maturing pigs (Table). Thus, late maturing pigs had decreased (P < 0.001) ADG and ADFI during the first 3 days in the nursery. Creep feeding had no effect on initial performance. No differences were observed in intestinal permeability between treatments. For overall nursery performance, a significant interaction was observed for ADG (P = 0.007) and ADFI (P < 0.001), with creep feeding providing a benefit in late maturing pigs, but not in early maturing pigs. Early maturing pigs had poorer G:F (P < 0.001) than late maturing pigs. At the end of the nursery, 241 pigs were transported to the finisher and randomly placed in pens based on treatment. There were 7 or 8 pigs per pen and 7 or 8 replications per treatment. For overall finishing performance, an interaction was observed for ADG (P = 0.037), with late maturing pigs that received creep feed having increased ADG compared with the other treatments. An interaction was also observed for ADFI (P = 0.007), with late maturing pigs that did not receive creep feed having decreased ADFI compared with late maturing pigs that received creep feed or early maturing pigs whether they received creep feed or not. In summary, early maturing pigs had lower stress response at weaning and improved ADG and ADFI until approximately 100 kg, at which point late maturing pigs began to exhibit greater ADG. Late maturing pigs had improved feed efficiency throughout the experiment. Creep feeding late maturing pigs resulted in improved growth performance compared with providing no creep feed, whereas creep feed did not impact early maturing pigs.

Stress and adaptation to repeated stress changed the expression of two isoforms of 11β-hydroxysteroid dehydrogenase (11β-HSD1 and 11β-HSD2) in the adrenal glands of sheep

The aim of this study was to estimate rate of cortisol release from ewe’s adrenal gland cortex and medulla tissue and the participation 11β-HSD1 and 2 in modulating adrenal glucocorticosteroids release, determine the expression size of these two isoforms in both parts of adrenal glands after single and repeated stress (3 times every 24h), when adaptive processes take place. The experiment was carried on 15 ewes, outside the breeding season. The research material consisted of adrenal glands obtained from control sheep (CON), sheep subjected to one-hour individual isolation from the herd (S1) and three times repeated stress factor at one-day intervals (S2). Immediately after slaughter, the adrenal glands from each sheep were placed on ice in 0.9% NaCl, then they were cut into smaller sections of similar weight (approx. 50 mg), covering the cortical and medullary layers of the adrenal glands. Sections of obtained tissues were weighed and placed in incubation wells (Sigma cell culture) containing 1 ml of incubation Eagle's medium with the addition of bovine albumin (BSA, 50 mg / 100 ml). Tissue incubation was carried out in an atmosphere of carbogen - 95% O2 and 5% CO2 at 38oC in a Sanyo incubator (MCO-18AIC). After 10 minutes of stabilization, the sections of core and adrenal cortex tissue were transferred to subsequent wells containing pure Eagle Medium. The same section was transferred to consecutive incubation wells containing clean incubation medium every 30 minutes. The medium harvested from the wells after 30, 60 and 90 minutes of incubation of adrenal tissue was frozen until the scheduled cortisol concentration de-termination by RIA (Orion Diagnostica) was performed. Another pieces of tissue the cortex and adrenal medulla from each sheep immediately placed in the RNAlater. Measurements of the gene expression of 11β-HSD1 and 2 expression in the adrenal cortex and medulla were performed using Real-time qPCR method. Starters and TaqMan MGB probes were designed by Applied Biosystem. In the conducted in vitro experiment, a significant reduction in the amount of cortisol released from adrenal cortex tissue into the medium after single stress factor was found, from 3.27 ± 0.69 pg･mg-1 tissue after first 30 minutes of experiment to 0.70 ± 0.15 pg･mg-1 of tissue after 90 minutes (P < 0.01). A similar profile of cortisol release from the adrenal cortex tissue was found in control sheep (P > 0.05). Experimental Group 2 (S2) had a similar profile of cortisol release during the all-time of incubation, however the values found were always lower than those observed in the control and single stress sheep Experimental Group 1 (S1) (1.34 ± 0.37 to 0.30 ± 0.08 pg･mg-1 of tissue; P < 0.05). After repeated stress, incubation of adrenal cortex tissue showed a similar profile of changes in cortisol release, however the values were significantly lower than those found in the singly stressed animals (from 1.30 ± 0.37 pg･mg-1 after 30 minutes to 0.27 ± 0.08 pg･mg-1 tissue after 90 minutes; P < 0.01). Incubation of adrenal medulla tissue obtained from singly stressed animals showed cortisol release to the medium much lower in comparison to adrenal cortex layer, however constant during the whole incubation time, value 1.04 ± 0.36 pg/mg-1 tissue noted after first 30 minutes to 0.77 ± 0.06 pg･mg-1 after 90 minutes of experiment; P > 0.05). When the stress factor was repeated three times, the amount of cortisol released was like that observed in the control secretion; P > 0.05). Analysis of 11β-HSD 1 expression in adrenal cortex tissue from singly stressed sheep showed an over threefold increase (350 ± 25 RQ), while measurement after stress factor was repeated three times demonstrated increase, however the value turned out to be two times lower than after a single stress (160 ± 35 RQ). The expression value of 11β-HSD 2 in the adrenal cortex tissue showed a significant reduction after both single (15 ± 7 RQ and repeated stress (22 ± 3.0 RQ). No significant differences were found in the expression value of this enzyme in the sheep adrenal medulla in both experimental groups in comparison to control group (P > 0.05). The obtained results indicate the immanent abilities of the adrenal glands, probably to a certain extent, to control the release of cortisol, conditioned by the presence and variable activity of the enzyme 11β-HSD, particularly isoform 1, in adrenal glands without the need to activate all HPA axis structures during stress and adaptation processes.

Cortisol Concentration in Horsehair and its Relationship to Body Location, Coat Colour, and Gender

The determination of cortisol concentrations in animal hair appears to be a relatively simple and non-invasive method of assessing chronic stress. This method has advantages but it also has some drawbacks. We discovered that there are specific coat properties and individual factors in animlas that can affect corisol levels and skew the interpretation of results. We removed horsehair from 8 different areas of the body (chest, left saddle, right saddle, right thoracic fetlock, and left thoracic fetlock, the mane was also sampled at the withers, just behind the ears, and the tail strand) in 31 horses and found statistically significant differences in cortisol concentrations There were statistically significant differences in the color of black, white, and brown hair based on the amount of cortisol excreted. In contrast, no significant differences in cortisol concentrations were found when the animals' sexes were compared. Samples should be taken from the same region with the same hair colour to obtain accurate results as the literature states that colour and location of the hair sampling effects cortisol concentrations. Studies also show that the gender of the animal has no effect on cortisol concentration.

Адаптация сердечно- сосудистой и эндокринной систем младших школьников к когнитивной нагрузке при выполнении теста на ноутбуке

Adaptation of children’s body to cognitive activity is an important issue for developmental physiology. The purpose of this study was to assess the nature of short-term adaptation of the cardiovascular and endocrine systems of primary school children to cognitive load when performing a test on a laptop. Materials and methods. Using spectral and temporal analysis of heart rate variability, electrocardiography, bipolar rheoencephalography, tonometry, and enzyme immunoassay to determine cortisol concentrations in saliva, we examined 32 primary school children (10–11 years old) performing a cognitive task (Anfimov’s tables) on a laptop. The cardiovascular system parameters were assessed before and during the test, the saliva was collected before and after the cognitive load. Results. In primary school children, the process of short-term adaptation of the cardiovascular and autonomic nervous systems to cognitive load was going favourably, without straining the regulation mechanisms, and was accompanied by an increase in sympathetic influences and heart rate, a decrease in the RR interval, and an increase in the tone of medium and small vessels in the occipital regions of the brain without changes in the indicators characterizing blood flow intensity. Two types of reaction of the endocrine system to cognitive load were revealed: 1) an increase in cortisol concentration in the saliva, observed in 40 % of children; 2) a decrease in the level of cortisol, characteristic of the majority of the examined schoolchildren. The obtained results showed that short-term adaptation of heart rate and haemodynamic parameters to cognitive load in primary school children is a problem-free process. A distinctive feature of children of this age is the manifestation of a proactive response of the endocrine system to the effects of environmental factors, which manifested itself in an increase in the level of the stress hormone cortisol before cognitive load.

Real-Time Cortisol Detection Using Electrocatalytic Polymer-Based Sensor

Cortisol is a steroid hormone produced by the adrenal gland in response to stress or agitate states. Deficiency or excess cortisol level can be triggered by physical or psychological stress, which is considered a biomarker for many diseases, such as Cushing syndrome [1]. In the human body, cortisol concentrations fluctuate in a circadian rhythm, being the highest in the early morning and the lowest in the night. A reliable cortisol test cannot depend on a single or one-day average measurement. Instead, screening a patient's dynamic cortisol levels over the circadian cycle is expected. The existing clinical cortisol measurements require blood, urine, or saliva samples sent to a lab to await results. The detection methods are mainly based on immunoassay that requires labeled antigens for a competitive assay and time-consuming incubation time (15–30 min), which hinders continuous detection needed for real-time stress management. To enable accurate time course measurements, a new detection approach is required to allow real-time determination of cortisol without costly reagents, labeling processes, additional redox reagents, and cumbersome laboratory equipment.Nanomaterials that mimic the catalytic behaviors of enzymes have great potential for continuous monitoring of cortisol levels through the direct electrochemical reading from cortisol redox reaction. There are very few reports on artificial cortisol catalytic sensors which require complicated fabrication or extremely non-neutral pH conditions for sensing [2]. Copper phthalocyanine (CuPc), a macrocycle with transitional copper (Cu) ion, holds electrocatalytic behavior and can be utilized for electrochemical redox reactions of various biomolecules, including H2O2, phenol, dopamine and serotonin. However, the detection of steroid hormones using CuPc has not been demonstrated yet. Compared to similar compounds based on porphyrin, phthalocyanine has N atoms in the meso positions and rich π-electrons that permit the formation of stable metallic phthalocyanine complexes. In this work, we develop copper phthalocyanine tetrasulfonate doped polypyrrole (PPy:CuPcTS) nanocomposite onto disposable screen-printed electrodes (SPEs) and demonstrate its electrocatalytic activity for real-time cortisol sensing. The incorporation of water-soluble CuPcTS into the PPy film forms an electrochemically conducting polymer in the negative potential window and catalyzes the reduction reaction of cortisol. The PPy:CuPcTS/SPE sensor can directly determine cortisol concentrations in real-time.The PPy:CuPcTS film was formed through one-step anodic electropolymerization on SPEs. The morphology and elemental composition of the film were characterized by scanning electron microscopy (SEM) and energy dispersive X-Ray analysis (EDX), respectively. Electrochemical responses were obtained with cyclic voltammetry (CV) and chronoamperometry. Cortisol detection was performed with 1x PBS solution.During the electropolymerization of PPy with CuPcTS, parts of the pyrrole rings hold a positive charge and form a complex with the sulfonated group in the CuPcTS. The CuPcTS and the PPy rings can also bond through π-π stacking. The stacked arrangement creates more conduction pathways due to highly delocalized electrons. The chelated Cu in CuPcTS catalyzes the reduction of ketone group in cortisol to an alcohol group [3], as schematically shown in Fig. 1(a). The SEM images in Fig.1 (b) show that the polymerization of PPy in the presence of CuPcTS results in a rough surface, very different from the smooth surface morphology on an undoped PPy film. The successful doping of CuPcTS in the PPy film is further confirmed by the presence of Cu peaks in the EDX (Fig.1 (b)). The CV curves in Fig. 1(c) indicate that cortisol reduction can be achieved using PPy:CuPcTS/SPE, evidenced by the increase in cathodic current at -1 V vs. Ag/AgCl upon adding cortisol. The phenomenon was not observed with PPy/SPE. The cathodic current increases with cortisol concentrations. The proposed sensor provides a dynamic detection range from 1 nM to 500 nM (Fig. (d)). Fig. 1(e) shows that the cortisol sensor also exhibits a selective detection against common interfering molecules, including glucose, ascorbic acid, and dopamine.PPy:CuPcTS nanocomposite was successfully demonstrated for electrocatalytic cortisol sensing. The sensor was achieved by a facile fabrication method and can operate at neutral pH without redox probes and labeling steps. The new sensor possesses great potential for low-cost, real-time monitoring of stress levels.

Use of a Guided Imagery Mobile App (See Me Serene) to Reduce COVID-19-Related Stress: Pilot Feasibility Study.

BackgroundThe SARS-CoV-2 pandemic has led to concerns about mental health resulting from regional and national lockdowns, social isolation, job loss, and concern about disease exposure.ObjectiveWe describe results of the pilot feasibility study of the See Me Serene mHealth app. The app provides users with immersive, vivid, nature experiences to reduce stress and anxiety related to COVID-19 and other isolation. The goals of the study were to develop the See Me Serene app and test the feasibility and acceptability of study procedures, and explore the potential impact of the app on stress and anxiety.MethodsWe developed and tested the See Me Serene app and our study procedures for feasibility, and gathered preliminary data with a goal of 100 participants. The research was conducted in 2 phases: (1) development and internal testing of the app; and (2) feasibility and pilot testing with participants recruited online through earned media (eg, news stories), presentations at a university campus, and social media (eg, online sharing of earned media and presentations). The feasibility study employed a mixed methods, within-subjects, pre-/posttest design. At baseline and 30-day follow-up, we assessed stress-related variables via validated self-report measures and saliva samples for determination of cortisol concentrations.ResultsWe met or surpassed all our feasibility benchmarks for recruitment (101 participants recruited), retention (91% [90/99] of 30-day assessment completed), and data collection (99 participants completed all baseline data; 85% [84/99] of salivary cortisol samples returned). Participants adhered to the intervention. On average, participants listened to 48.2 audio files over 30 days or approximately 1.6 audio files per day. Participants were satisfied with the app, with 87% (78/90) rating the app as helpful in dealing with stress and anxiety. The app showed the potential to reduce stress, anxiety, loneliness, and worry. We did not find significant differences (P=.41) in cortisol levels over time. Our findings suggest that future research is warranted to test the efficacy of the See Me Serene app with a representative, diverse sample.ConclusionsThere is a need for evidence-based and easily disseminable stress-reduction interventions. See Me Serene is a feasible intervention and has the potential to reduce stress related to COVID-19 and other forms of social isolation. More research on See Me Serene is warranted.

Determination of cortisol in hair using liquid chromatography-tandem mass spectrometry: a short review.

Cortisol is considered a particularly relevant biomarker in the context of stress evaluation. This study aims to review of the available literature on the determination of cortisol in hair using LC-MS/MS. Currently, there is no standardized procedure for the measurement of cortisol concentrations in hair, and different sample preparation, chromatographic separation and mass spectrometric detection conditions were described. Simple methanolic extraction, reversed-phase separation and MRM detection in negative ion mode are the most common employed analytical approaches. Reported assays presented acceptable sensitivity for clinical purposes. The increasing use of mass spectrometry in clinical laboratories may contribute to the establishment of LC-MS/MS as the method of choice for the determination of cortisol concentrations in hair.

Ewes behavioural and physiological reactions to the odour of fox (Lycalopex gymnocercus) and wild boar (Sus scrofa) faeces

Predation of sheep is a production problem that deserves the studies of how this prey species detects and react to predators. In this context, this study determined the behavioural and physiological responses of ewes exposed to the odour of predators’ faeces as fox (Lycalopex gymnocercus) and wild boar (Sus scrofa). Four experimental treatments were applied to 48 Corriedale sheep (12 ewes each): fox faeces (FFO), wild boar faeces (WBFO), unknown sheep wool (SWOC) and no odour (NOC). All ewes were evaluated in a behavioural test for 5 min and then rectal temperature was measured, and a series of blood samples were obtained to determine cortisol concentration, glycaemia, total protein, albumin and globulin. The frequency of kicking the ground, turning their head, going back and stillness behaviours increased significantly in FFO in comparison with NOC (p = 0.01; p < 0.0001; p = 0.005 and p = 0.002, respectively) and kicking the ground and turning the head compared with SWOC (p = 0.03 and p < 0.001 respectively). The FFO group spent significantly more time turning the head and going back than NOC (p < 0.001 and p = 0.01 respectively) and SWOC (p = 0.002 and p = 0.05). For ewes evaluated with WBFO, kicking the ground, turning their head, going back and remaining still occurred significantly more times than with NOC (p = 0.008; p = 0.005; p = 0.01 and p = 0.003), and kicking the ground, turning their head and remaining still occurred significantly more times than with SWOC (p = 0.02; p = 0.02 and p = 0.05). Furthermore, WBFO group spent significantly less time still (p = 0.01) and more time turning their head and going back (p = 0.001 and p = 0.02, respectively) than NOC, that spent significantly more time turning their head than SWOC (p = 0.006). Rectal temperature and haematological variables were not affected by treatments. The increase in the frequency of agonistic behaviours and duration of locomotion behaviours in ewes treated with predators’ faeces, allow us to conclude that ewes respond to fox and wild boar’s faeces exposure with a similar aversive behavioural repertoire. Also, ewes displayed some physiological responses although these responses cannot be attributed to the odour of predators.

Habituation to Transport Helps Reducing Stress-Related Behavior in Donkeys During Loading

Adopting proper animal management strategies, including training, might reduce to a substantial extent the adverse effects of transport-related stress in animals. The aim of this study was to evaluate the effect of habituation to transport on stress-related behaviors and physiological indicators during loading and unloading in donkeys. Fourteen donkeys were recruited and divided in two treatment groups: Habituation (H; M = 5, F = 2) and Control (C; M = 5, F = 2). H donkeys were gradually habituated to be transported, traveling together with their mothers and other adult donkeys well-accustomed to transport, while C donkeys had never been transported before. Loading and unloading phases were video recorded and behavior was analyzed. Saliva samples for cortisol concentration determination were collected at rest and after unloading. Latency time to load was significantly shorter for H donkeys than C donkeys (Mann-Whitney; p = 0.004). C donkeys also showed significantly more stress-related behaviors (Mann-Whitney; p = 0.026) and required a higher but not statistically significant number of human interventions to load. Cortisol concentration increased in both groups, but no differences were found between them (Mann-Whitney; p > 0.05). These results suggest that habituation to transport could mitigate stress during loading procedures in donkeys reducing loading time, frequency of stress-related behaviors and diminishing the need of human intervention.

16Up: Outline of a Study Investigating Wellbeing and Information and Communication Technology Use in Adolescent Twins

The '16Up' study conducted at the QIMR Berghofer Medical Research Institute from January 2014 to December 2018 aimed to examine the physical and mental health of young Australian twins aged 16-18 years (N = 876; 371 twin pairs and 18 triplet sets). Measurements included online questionnaires covering physical and mental health as well as information and communication technology (ICT) use, actigraphy, sleep diaries and hair samples to determine cortisol concentrations. Study participants generally rated themselves as being in good physical (79%) and mental (73%) health and reported lower rates of psychological distress and exposure to alcohol, tobacco products or other substances than previously reported for this age group in the Australian population. Daily or near-daily online activity was almost universal among study participants, with no differences noted between males and females in terms of frequency or duration of internet access. Patterns of ICT use in this sample indicated that the respondents were more likely to use online information sources for researching physical health issues than for mental health or substance use issues, and that they generally reported partial levels of satisfaction with the mental health information they found online. This suggests that internet-based mental health resources can be readily accessed by adolescent Australians, and their computer literacy augurs well for future access to online health resources. In combination with other data collected as part of the ongoing Brisbane Longitudinal Twin Study, the 16Up project provides a valuable resource for the longitudinal investigation of genetic and environmental contributions to phenotypic variation in a variety of human traits.

Hair cortisol as chronic stress parameter in patients with acute ST-segment elevation myocardial infarction

Stress is arisk factor for the development of cardiovascular diseases. New analytical methods enable the determination of cortisol concentrations in hair (HCC) as aretrospective long-term parameter for chronic stress. So far, it is still uncertain whether stress objectively measured by an increase in HCC might be apredictive factor associated with the development of an acute myocardial infarction. Included in this study were 40patients with an acute ST-segment elevation myocardial infarction (STEMI), confirmed by electrocardiography (ECG). The control group consisted of 80 patients without coronary artery disease (CAD). Patients with STEMI did not show significantly higher HCCs (p = 0.846) compared to the control group and there was no correlation with the extent of myocardial infarction (p = 0.701, r = 0.038). Correlations were found between HCC, diabetes mellitus (p = 0.046, odds ratio, OR = 6.346), low high-density lipoprotein (HDL) cholesterol concentration (p = 0.107) and glycated hemoglobin A (HbA1c) concentration (p = 0.083, r = 0.236). Furthermore, there was an association between an increased HCC and chronic heart failure (p = 0.110) and an association to the red blood cell distribution width (RDW) concentration (p = 0.005, r = 0.293). The results of this study showed that patients with acute ST-segment elevation myocardial infarction did not have significantly higher levels of hair cortisol compared to the reference group; however, long-term metabolic changes as well as advanced chronic heart failure appear to be long-acting stress factors for the organism and increase cortisol levels in hair.

The Effect of Long Distance Running on the Pattern of Production of Salivary Cortisol on Rest Days

INTRODUCTIONCortisol is a glucocorticoid synthesized from cholesterol by the zona fasciculate of the adrenal cortex and is controlled by way of a negative feedback pathway within the hypothalamic‐pituitary‐adrenal (HPA) system. HPA axis receives strong circadian input and plays an essential role in the regulation of stress responses. Cortisol spikes in the morning upon waking up, and then decreases throughout the day, but exercise modifies this pattern of production. Physical exercise activates HPA axis to maintain proper blood glucose levels during these times of energy expenditure. According to previous studies endurance‐trained athletes possess a significantly higher level of circulating cortisol than power trained athletes immediately after exercise. Cortisol is known to have debilitating effects on factors related to bone mineral density (BMD). Consequently, the chronic increase in cortisol in long‐distance runners can lead to adverse effects such as decreased BMD and can contribute to stress fractures. The purpose of this study is to evaluate cortisol levels during days of rest to determine the possible change in the pattern of cortisol production stimulated by repetitive prolonged previous exposure to the physical activity rather than actual stimulus.METHODSMissouri Southern State University (MSSU) Institutional Review Board approval was gained before data collection. A participant group was gathered including 12 males from the men’s cross‐country team at MSSU in the fall of 2019. Participants completed a demographic questionnaire stating the number of hours per week actively participating in sport specified workouts, average hours of sleep per night, and a list of supplements being consumed. Participants gave saliva samples within pre‐determined time intervals before, during, and after their workouts as well as during the same times during days of rest. Samples were collected on two training days and one rest day a week on three separate weeks for a total of 27 samples per athlete. In addition to saliva samples, a stress assessment was filled out by every athlete each day of collection. All samples were placed in a freezer at 0 degrees Celsius until collections were completed and analyzed. Samples were evaluated via ELISA by a competitive immunoenzymatic colorimetric method for the quantitative determination of cortisol concentration. ELISA absorptions were analyzed with linear regression to calculate cortisol concentrations.CONCLUSIONThe result of this study will assist in a better understanding of the hypothalamic‐pituitary‐adrenal axis function. Furthermore, while allowing the endurance‐trained athlete to efficiently and properly recover, it will aid in reducing the risk of injury and overtraining syndrome.

Effects of Alfaxalone or Propofol on Giant-Breed Dog Neonates Viability During Elective Caesarean Sections.

Simple SummaryNowadays, thanks to the increased awareness of owners and breeders and to the most recent techniques available to veterinarians, the management of parturition, especially of C-sections, has become a topic of greater importance. Anesthesia is crucial and must be targeted to both the mother and neonates. The present study aimed to evaluate the effect of the induction agent alfaxalone on the vitality of puppies born from elective C-section, in comparison to propofol. After inducing general anesthesia for elective C-section, puppies from the mothers induced with alfaxalone had higher 5-min Apgar scores than those induced with propofol. The concentration of cortisol in fetal fluids collected at birth is neither influenced by the anesthetic protocol used, nor does it differ between amniotic and allantoic fluids. Nevertheless, the cortisol concentration in fetal fluids affects the relationship between anesthesia and the Apgar score: the present study highlights a significant relationship between the anesthetic protocol used and Apgar score in puppies, and fetal fluids cortisol concentration acts as a covariate of this relationship. Alfaxalone is a safe and effective drug for the induction of anesthesia in dogs, and it can be successfully employed for elective C-section, with neonatal wellbeing improvements.Attention must be paid to C-section anesthesia effects on mother and offspring. Alfaxalone induction results in improved puppy viability when compared to propofol. The present study aims to evaluate effects of alfaxalone or propofol induction for elective C-section on newborns, expressed as Apgar score and fetal fluids cortisol concentration. Anesthesia was induced with alfaxalone 3 mg/kg iv in 5 bitches (group A), and propofol 4 mg/kg iv in another 5 (group P), maintained with isoflurane. Amniotic and allantoic fluids were collected to determine cortisol concentration. Apgar score, litter size, newborn gender, birth-weight, maternal age, and parity were recorded. ANOVA, U Mann-Whitney test and ANCOVA assessed the effects of drugs on the Apgar score and fetal fluids cortisol. Thirty-six puppies were randomly selected for the study: 16 from group A and 20 from group P. Only the Apgar score significantly differed between groups. ANCOVA confirmed a significantly higher Apgar score in group A underlining the influence of fetal fluids cortisol concentrations, both resulting in covariates. Present results confirm the effect of anesthesia on the Apgar score of newborns, which is significantly higher for alfaxalone than propofol, suggesting the use of fetal fluids cortisol as a covariate. These findings could be a starting point for further investigations when less viable puppies are detected or expected, such as during an emergency C-section.

Strenuous Exercise Increases the Risk of Oxidative Stress in Ironman Triathlon Participants

Regular physical activity has been linked to greater overall health. Literature review and studies have also defined regular physical activity as a reducer of life-threatening illnesses such as cardiovascular disease, diabetes and obesity. However, long increments of strenuous exercise can produce oxidative stress and muscle fatigue in the human body. The increase in oxygen consumption during strenuous exercise leads to elevated reactive oxygen species (ROS). Cells continuously produce free radicals and reactive oxygen species as part of metabolic processes in the body. These free radicals are neutralized by an antioxidant defense system in the body consisting of enzymes, such as catalase, and non-enzymatic antioxidants. An Ironman Triathlon consists of a 2.4-mile (3.86 km) swim, a 112-mile (180.25 km) bicycle ride and a marathon 26.2-mile (42.2 km) run, raced in that order and without a break. It is widely considered by athletes to be one of the most demanding sporting events in the world. It is hypothesized that a physically challenging event such as the Ironman Triathlon can be linked to elevated cortisol levels, increased occurrence of DNA damage, elevated concentrations of ROS, and consequently increased oxidative stress in humans. In order to derive conclusive results regarding the hypothesis, groups containing athletes who completed the full Ironman race, the half Ironman race, and a control group of moderately active individuals were established and individuals were required to report Garmin Smartwatch health and wellness data. The half Ironman consists of a 1.2-mile (1.93 km) swim, a 56-mile (90.12 km) bicycle ride and a marathon 13.1-mile (21.1 km) run, raced in that order and without a break. Several protocols were then applied to derive data necessary to complete the research. After the participants were selected, their saliva was collected in a non-invasive fashion and was used in the Elisa Saliva Kit to determine cortisol concentration. The saliva samples were also utilized to perform DNA and RNA extraction; and the resulting products were analyzed for quantity and quality of the DNA and RNA. Real time PCR allows scientists to monitor PCR while it is occuring. In this technique, luminescence is produced by reporter molecules as the PCR products increase with every cycle. To determine ROS concentration, the ROS-Glo assay, which provides a light signal that is proportional to the ROS in a given sample, was utilized. An additional marker of oxidative stress is 8-oxo-2-deoxyguanosine(8-oxo-dG). The OxiSelect™ Oxidative DNA Damage ELISA uses antibody and antigen interactions to report the concentration of 8-oxo-dG in a sample. Furthermore, the results indicate an increase in enzymatic indicators of elevated ROS, elevated cortisol levels, and disruption of sleep in the participating athletes after the race. In conclusion, the athletes who completed the full Ironman triathlon experienced increased amounts of oxidative stress than their less active counterparts in the control group, as was denoted by the elevated cortisol levels, increased 8-oxo-dG concentrations, and increased ROS concentrations.Such a rigorous event negatively impacted participants and caused oxidative stress. <strong>Faculty Sponsor</strong>: Noelle Cutter

Evaluation of Intraperitoneal and Incisional Bupivacaine or Levobupivacaine for Postoperative Analgesia in Ovariohysterectomized Dogs

Background: Ovariohysterectomy (OHE) is the most commonly performed elective surgical procedure in companion animals. OHE offers benefits of control of population and decreased risk of potentially life-threatening diseases such as mammarian tumours and pyometra. Traditional OHE intervention causes inflammation and pain due to trauma during organ manipulation. The purpose of this study was to compare the effect of intraperitoneal and incisional administration of bupivacaine (BP) or levobupivacaine (LP) on postoperative analgesia in dogs undergoing the OHE procedure.Materials, Methods &amp; Results: A total of 24 mix-breed bitches aged between 1 - 3 years and weighed 19 - 20 kg were used in this study. The animals were divided into three groups as control (n = 8), BP (n = 8) and LP group (n = 8). The animals were kept under surveillance at the hospitalisation unit of the animal hospital for one day before the elective OHE. The dogs were fasted for 12 h before the surgery, with adlib water consumption. Atropine sulphate 0.045 mg/kg was administered subcutaneously approximately 30 min before general anaesthesia. Midazolam (0.3 mg/kg) was intravenously injected into all dogs for pre-anaesthetic medication. After sedation, anaesthesia was induced with propofol (4 mg/kg, IV) and then the dogs were orotracheally intubated using cuffed endotracheal tubes. General anaesthesia was maintained by administration of 2% isoflurane. The ventral abdomen was prepared aseptically for OHE following the general anaesthesia. All animals were operated on by the same surgeon. During surgery, sprayed bupivacaine 0.5% (4.4 mg/kg) with an equal volume of saline in BP group, levobupivacaine 0.5% (4.4 mg/kg) with an equal volume of saline in LP were then applied over the ovaries, uterine broad ligaments and cervix uteri. After removal of the uterine body, either LP or BP was sprayed to left and right, or cranial and caudal parts of the abdominal cavity. Finally, the incision line was sprayed before closing the skin. The control group received only sprayed saline. The pain evaluation was carried out with modified Melbourne pain scoring scale. To determine cortisol concentrations, blood samples were taken before anaesthesia induction (baseline) and postoperative 0, 0.5, 1, 2, 4, 6, 12 and 24 h. Postoperative pain scores were higher in the control group than BP and LP groups following surgery at 30 min, 1st, 4th, and 6th h. In all groups, cortisol levels increased postoperatively, whereas they decreased more rapidly in BP and LP groups. There was a steady increase at postoperative 0 min, 30 min, 1st, 2nd, 4th and 6th h in the control group and at postoperative 0 min, 30 min and 1 h in BP and LP groups (P &lt; 0.05).Discussion: Intraperitoneal and incisional bupivacaine shows significantly superior postoperative pain management benefits after the closure of skin, compared to untreated dogs and it is also superior to lidocaine treatment at 0.5 h following the visual analogy pain assessment system. In the present study, the observation of lower pain scores in BP group at 0.5, 1, 4 and 6 h as compared to the control group was similar to findings of other studies. LP group also showed decreased pain scores at above-mentioned h without any significant difference. However, only in LP group, pain scores at 24 h were lower than postoperative pain scores. In conclusion, it is suggested that sprayed intraperitoneal and incisional BP and LP are very effective for preventing postoperative pain ovariohysterectomized dogs.

Effect of a single dose of subcutaneous meloxicam prior to band or knife castration in 1-wk-old beef calves: II. Inflammatory response and healing.

The aim of this study was to evaluate the effect of a single dose of subcutaneous (s.c.) meloxicam administered at the time of knife and band castration on inflammatory response and wound healing over 56-d post-castration. Seventy-two Angus crossbred calves (47.3 ± 6.70 kg of body weight [BW] and 7 to 8 d of age) were randomly assigned according to a 3 × 2 factorial design assessing castration method: sham (CT), band (BA), and knife (KN) castration, and pain mitigation: non-medicated (NM) and medicated calves (M) injected s.c. with meloxicam (0.5 mg/kg of BW). Calf BW, rectal temperature, swelling ("0": no swelling; "4": swelling needing intervention) and healing score ("1" to "5" with "5" being completely healed), scrotal circumference, and maximum scrotal temperature were measured on d -1, immediately before castration (d 0), and weekly thereafter over a 56-d period. Blood samples for haptoglobin (Hp), serum amyloid-A (SAA), and complete blood cell count were collected according to the same schedule. Hair samples were collected on d -1, 28, and 56 to determine cortisol concentrations. Standing and lying behaviors were measured using accelerometers that were placed on the calves on d -1 until d 35, and visual observations of behaviors related to pain were recorded once a week for 35 d. Knife-castrated calves achieved swelling scores of "3" and "2" between d 7 and 14, which was sooner (Z< 0.05) than in BA calves (from d 14 to 35). In addition, greater (P = 0.03) concentrations of SAA were observed in BA calves (76.9 ± 0.12 g/liter) compared with CT (57.6 ± 0.12 g/liter) and KN (51.6 ± 0.12 g/liter) from d 7 to 35. Healing scores of "2" and "4" tended to be achieved sooner (Z < 0.10) in KN calves than in BA calves, although healing scores of "3" tended to be achieved sooner (Z < 0.10) in BA calves than KN calves. No differences (P > 0.10) were observed among treatments for hair cortisol on d -1 and 28, but on d 56, hair cortisol concentrations in BA-NM calves were greater (P > 0.05) than for CT-NM, BA-M, KN-NM, and KN-M, and tended to be greater (P = 0.08) than for CT-M calves. Lying duration tended (P = 0.10) to be greater and suckling behavior tended (P = 0.08) to be lower in NM than M calves. A single s.c. injection of meloxicam did not reduce long-term inflammatory responses or improve wound healing; however, it may be useful in reducing pain and stress in band castrated calves as evidenced by reduced hair cortisol concentrations up to 56 d post-castration.

Disposable electrochemical immunosensor for cortisol determination in human saliva

Cortisol is a steroidal hormone and an important stress marker. Free serum cortisol concentration has been identified to correlate well with free salivary cortisol. In this present work an electrochemical immunosensor was developed to determine cortisol concentration within the physiological concentration range found in human saliva. The immunosensor is based on a direct competitive enzyme linked immunoassay using a home-made cortisol-alkaline phosphatase (AP) conjugate synthesized in our laboratory with disposable graphite screen-printed electrodes (SPEs). 1-nalphtyl phosphate (1-NP) was used as an enzymatic substrate and a square wave voltammetry (SWV) for electrochemical detection. To study method suitability for use with saliva samples, calibration curves were performed both in buffer and saliva. In buffer standard samples showed a limit of detection (LOD) of 0.6 ng/ml and working range (WR) of 0.2–44.6 ng/ml with good reproducibility (RSD 10%). Saliva matrix effect was removed effectively with Salivette Cortisol collection device (polyethylene) and a calibration curve showed similar characteristics as in buffer with LOD 1.7 ng/ml and WR 0.5–55.1 ng/ml (RSD 8%) demonstrating the possibility to determine human salivary cortisol within the desired human physiological range. Spiked saliva samples were analyzed with the developed immunosensor presenting excellent 92–114% recovery. Comparison to liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method showed strong 0.90 correlation between methods indicating good accuracy of the developed immunosensor.

Exercise intensity and its impact on relationships between salivary immunoglobulin A, saliva flow rate and plasma cortisol concentration

IntroductionSalivary secretory immunoglobulin A (sIgA), saliva flow rate and plasma cortisol concentrations have been shown to be influenced by exercise, particularly the intensity exercise is performed at, and circadian variation. The autonomic nervous system partly regulates salivary secretion, but it is not yet known whether cortisol also explains some variation in salivary parameters.MethodsTwelve moderately trained male individuals (dot {V}{{text{O}}_2}peak legs: 46.2 ± 6.8 mL·kg−1·min−1) performed three 45-min constant load exercise trials in the morning: arm cranking exercise at 60%dot {V}{{text{O}}_2}peak arms; moderate cycling at 60%dot {V}{{text{O}}_2}peak legs; and easy cycling at 60%dot {V}{{text{O}}_2}peak arms. Timed saliva samples and blood samples for plasma cortisol concentration determination were obtained before, post, 2 h post, and 4 h post-exercise. Saliva was collected in an additional resting trial at the same time points.ResultsAt each time point for each exercise trial, negative correlations between cortisol and saliva flow rate (explaining 25 ± 17% of the variance, R2 = 0.002–0.46) and positive correlations between cortisol and sIgA concentration (explaining 8 ± 8% of the variance R2 = 0.002–0.24) were found. Saliva flow rate increased over time, whereas sIgA concentration and cortisol decreased over time for all trials (P < 0.05), there was no effect of time for sIgA secretion rate (P = 0.16).ConclusionThese results show a relationship between cortisol and saliva flow rate, which directly impacts on the concentration of salivary analytes. This study further confirms circadian variations in salivary parameters which must be acknowledged when standardising salivary data collection.