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Background: Psychological anxiety and depression, as well as memory impairment, are frequently linked to inflammation. Clove essential oil (CEO) administration and hydrogen (H2) inhalation have been proven to have anti-inflammatory and alleviating effects on related psychological disorders in the past. The current study investigated the potential to improve anxiety-like behaviors and cognitive function by a combination of CEO and H2 treatment. Methods: The mice were administered lipopolysaccharide (LPS) to induce inflammation and oxidative stress response and cause psychological disorders. Using this animal model, we conducted experiments to test whether essential oil and H2 inhalation could improve the psychological damage in behavior caused by LPS. Subsequently, elevated plus maze (EPM), forced swimming test (FST), and passive avoidance (PA) test were performed for evaluation of mice anxiety, depression, and response to electric shock, respectively. Furthermore, the biochemical analysis was used to examine the expression levels of inflammatory and oxidative stress markers. Results: Our results showed that CEO administration and H2 inhalation alone or in combination positively improved inflammation-induced anxiety, depression, and cognitive memory deficits in the mice. In the single treatment groups, CEO demonstrated better results than H2 inhalation in the elevated plus maze, forced swimming, and passive avoidance tests, while combined treatment with both provided a further improved enhancement effect. Biochemical analysis of the cerebral cortex revealed that CEO and H2 therapy reversed the LPS-induced inflammation and oxidative stress response. Conclusions: Our results suggest that a combination of CEO and H2 has the potential to treat psychological disorders or neuropsychiatric disorders. [ABSTRACT FROM AUTHOR]

Objective To examine disease and target engagement biomarkers in the RISE-SSc trial of riociguat in early diffuse cutaneous systemic sclerosis and their potential to predict the response to treatment. Methods Patients were randomized to riociguat (n  = 60) or placebo (n  = 61) for 52 weeks. Skin biopsies and plasma/serum samples were obtained at baseline and week 14. Plasma cyclic guanosine monophosphate (cGMP) was assessed using radio-immunoassay. α-Smooth muscle actin (αSMA) and skin thickness were determined by immunohistochemistry, mRNA markers of fibrosis by qRT-PCR in skin biopsies, and serum CXC motif chemokine ligand 4 (CXCL-4) and soluble platelet endothelial cell adhesion molecule-1 (sPECAM-1) by enzyme-linked immunosorbent assay. Results By week 14, cGMP increased by 94 (78)% with riociguat and 10 (39)% with placebo (P  < 0.001, riociguat vs placebo). Serum sPECAM-1 and CXCL-4 decreased with riociguat vs placebo (P  = 0.004 and P  = 0.008, respectively). There were no differences in skin collagen markers between the two groups. Higher baseline serum sPECAM-1 or the detection of αSMA-positive cells in baseline skin biopsies was associated with a larger reduction of modified Rodnan skin score from baseline at week 52 with riociguat vs placebo (interaction P -values 0.004 and 0.02, respectively). Conclusion Plasma cGMP increased with riociguat, suggesting engagement with the nitric oxide–soluble guanylate cyclase–cGMP pathway. Riociguat was associated with a significant reduction in sPECAM-1 (an angiogenic biomarker) vs placebo. Elevated sPECAM-1 and the presence of αSMA-positive skin cells may help to identify patients who could benefit from riociguat in terms of skin fibrosis. Trial registration Clinicaltrials.gov, NCT02283762. [ABSTRACT FROM AUTHOR]

Background: Acetaminophen (APAP)-induced hepatotoxicity is a potentially life-threatening condition. Gardenia jasminoides fruit extract (GJE), which contains geniposide (Gen) as its major active constituent, possesses anti-inflammatory and antioxidant properties and may help address the underlying pathogenesis of APAP-induced hepatotoxicity. This study aimed to evaluate the effects of GJE in a mouse model of APAP-induced hepatotoxicity. Methods: Twenty-four male ICR mice were divided into 4 groups (n = 6/group): [1] Control group, mice were given distilled water; [2] APAP group, mice received a single dose of 600 mg/kg APAP; [3] APAP + low-dose GJE group, mice received APAP followed 30 min later by 2 doses of low-dose GJE (0.44 g/kg/dose, containing Gen 100 mg/kg/dose) 8 h apart; [4] APAP + high-dose GJE group, mice received APAP followed by 2 doses of high-dose GJE (0.88 g/kg/dose, containing Gen 200 mg/kg/dose). All mice were euthanized 24 h after APAP administration. Liver tissue was used for histological examination and to measure glutathione (GSH) and malondialdehyde (MDA) levels. Serum was used to determine levels of ALT and inflammatory cytokines (tumor necrosis factor- α (TNF-α) and interleukin-6 (IL-6)). Results: Liver histopathology showed moderate to severe hepatic necroinflammation in the APAP group, whereas only mild necroinflammation was observed in both treatment groups. Serum ALT levels were significantly elevated in the APAP group compared to the control group but were significantly reduced after low- and high-dose GJE treatment. Serum TNF- α levels were significantly higher in the APAP group than in the control group and were significantly lower after high-dose GJE treatment (135.5 ± 477.2 vs. 35.5 ± 25.8 vs. 74.7 ± 47.2 vs. 41.4 ± 50.8 pg/mL, respectively). Serum IL-6 followed a similar pattern. Hepatic GSH levels were significantly lower in the APAP group compared to the control group but significantly increased after both low- and high-dose GJE treatment (19.9 ± 4.5 vs. 81.5 ± 12.4 vs. 71.4 ± 7.8 vs. 82.6 ± 6.6 nmol/mg protein, respectively). Conversely, hepatic MDA levels were significantly elevated in the APAP group compared with the control group but significantly decreased after high-dose GJE treatment (108.6 ± 201.5 vs. 40.5 ± 18.0 vs. 40.5 ± 16.8 nmol/mg protein, respectively). Conclusions: Treatment with G. jasminoides fruit extract can alleviate APAP-induced hepatotoxicity, likely through its anti-inflammatory and antioxidant properties. [ABSTRACT FROM AUTHOR]

Context Glucagon plays a role in the development of type 2 diabetes, yet its role in prediabetes (preDM) remains uncertain. Objective To evaluate glucagon levels in the fasting state and its response to glucose inhibition in preDM through meta-analysis. Methods A systematic search across Pubmed, Embase, Web of Science, and Cochrane Library identified studies assessing glucagon levels during 75 g oral glucose tolerance test (OGTT) in both preDM and normal glucose tolerance (NGT) cohorts. Data on glucagon, glucose, and insulin were pooled using a random-effect model. Results Although glucagon levels decreased in both preDM and NGT groups upon glucose challenge, glucagon levels at 0 hours, 0.5 hours, 1 hour, and 1.5 hours in preDM were significantly higher compared to NGT, despite higher glucose levels at all time points and higher insulin levels at 0 hours, 1 hour, 1.5 hours, and 2 hours during OGTT. Subgroup analysis revealed that in studies using the radioimmunoassay method, glucagon levels in preDM were higher at 0.5 hours and 1 hour than NGT, while in studies using the ELISA method, glucagon levels were similar to those of the NGT group despite higher glucose in preDM compared to NGT. Fasting glucagon level was inadequately suppressed in both impaired glucose tolerance (IGT) and impaired fasting glucose (IFG). Responsiveness to glucose inhibition was preserved in IFG, while glucagon level in IGT group at 0.5 hours after glucose intake was not suppressed and was higher than NGT. Conclusion Glucagon was not adequately suppressed during OGTT in preDM. Glucagon dysregulation is a contributing mechanism underlying both IFG and IGT. [ABSTRACT FROM AUTHOR]

Objective: This research aimed to probe the expression of long noncoding RNA TMEM147 antisense RNA 1 (TMEM147‐AS1)/micro‐RNA (miR)‐124/signal transducer and activator of transcription 3 (STAT3) axis in estrogen receptor (ER)‐positive breast cancer (BC). Methods: Sixty ER‐positive BC patients undergoing surgical treatment were gathered. TMEM147‐AS1, miR‐124, and STAT3 expression levels in BC cells and tissues were measured. The binding sites of TMEM147‐AS1 and miR‐124, miR‐124, and STAT3 were analyzed and validated. The miR‐124, STAT3 overexpression (oe) sequences, TMEM147‐AS1 oe, and interference sequences and their control sequences were planned and cells were transfected to assess their functions in BC cells biological functions. Results: TMEM147‐AS1, as well as STAT3 was extremely expressed and miR‐124 was lowly expressed in BC cells and tissues. Interference with TMEM147‐AS1 restrained ER‐positive BC cell malignant activities. Mechanistically, TMEM147‐AS1 could competitively bind miR‐124 in refraining miR‐124 expression, and STAT3 was a target gene of miR‐124. Oe of miR‐124 effectively reversed the enhancement of BC cell proliferation and invasion induced by TMEM147‐AS1 upregulation. Oe of STAT3 could reverse the inhibitory effect of miR‐124 on BC cell malignant behaviors. Conclusion: TMEM147‐AS1 has oncogenic activity in ER‐positive BC, which may be a result of the altered miR‐124/STAT3 axis. Therefore, targeting the TMEM147‐AS1/miR‐124/STAT3 axis may be a target for ER‐positive BC therapy. [ABSTRACT FROM AUTHOR]

Background: Two main types of galenic formulation, immediate release and prolonged release, have been developed to optimize melatonin bioavailability. We recently described the kinetic profile of a prolonged-release form generating a peak of plasma melatonin 1 h (Tmax) after intake, followed by a prolonged decay over time. We have developed a new oral form of melatonin with the aim of producing a melatonin peak several hours after intake. Objective: The objective is to investigate melatonin bioavailability after administration of this new delayed-release form (DR form). Methods: In this single-centre open-label study, 12 healthy male volunteers received one tablet of the DR form containing 1.9 mg melatonin, 10 mg zinc and 200 mg lemon balm extract (Melissa officinalis L aerial parts). Blood samples were collected for 12 h, beginning at 8:00 am. Plasma concentrations of melatonin and 6-sulfatoxymelatonin (6-SMT), the main hepatic melatonin metabolite, were determined by radioimmunoassay. Results: A progressive increase in plasma melatonin concentrations was observed from 20 min and a peak about 3 h after intake (Cmax 740 ± 824 pg/mL; Tmax 179 ± 60 min). Concentrations remained high between 140 and 220 min, the concentration remaining physiologically significant (over 100 pg/mL) up to 7 h after intake. The DR form was well tolerated. Conclusions: The melatonin release profile was consistent with what was anticipated for the DR form. The DR form generated a 2 h delayed Tmax compared with a prolonged-release form previously evaluated. This suggests that the DR form is suitable for the treatment of certain sleep disorders such as short sleep duration or early awakening. Trial Registry: Registration number: NCT05419466. [ABSTRACT FROM AUTHOR]

Alpha‐fetoprotein (AFP) is a structural serum glycoprotein that plays vital roles in reproduction and mammalian development. Analysis of serum prolactin (PRL) is considered one of the useful methods for diagnosing pregnancy in Asian elephants. However, the expression profiles of AFP in pregnant and nonpregnant Asian elephants remain unclear, nor is the relationship with PRL. In this study, serum seven gonadal hormones and AFP in three pregnant and seven nonpregnant Asian elephants were analysed by via radioimmunoassay (RIA) and enzyme‐linked immunosorbent (ELISA) assay. We found that the mean (±SD) concentration of prolactin (PRL) in pregnant (136.782 ± 30.987 ng/mL) elephants was significantly higher than that in nonpregnant elephants (52.803 ± 21.070 ng/mL; p ≤ 0.0005). The mean (±SD) concentration of AFP in pregnant elephants (11.598 ± 0.824 ng/mL) was significantly higher than that in nonpregnant elephants (7.200 ± 2.283 ng/mL; p ≤ 0.05). Furthermore, the AFP concentration was positively correlated with the PRL concentration in the 10 Asian elephants studied. In conclusion, our findings suggest that serum AFP concentration is a potential biomarker of pregnancy outcomes in Asian elephants. [ABSTRACT FROM AUTHOR]

Fibroblast growth factor 21 (FGF-21) has been suggested as a potential therapeutic target for insulin resistance in health-related metabolic disorders such as type 2 diabetes. Despite the metabolic effects of resistance (RT) and aerobic training (AT) on diabetes symptoms, uncertainty exists regarding the superiority of effects manifested through these training approaches on FGF-21 and biochemical and physiological variables associated with metabolic disorders in men diagnosed with type 2 diabetes. This study aimed to investigate the impact of a 12-week RT and AT on FGF-21 levels and symptoms associated with metabolic disorders in male individuals diagnosed with type 2 diabetes. Thirty-six sedentary obese diabetic men (40 to 45 years old) were matched based on the level of FGF-1. They and were randomly divided into two training groups (RT, n = 12 and AT, n = 12) performing three days per week of moderate-intensity RT or AT for 12 weeks and an inactive control group (n = 12). Both training interventions significantly improved FGF-21, glucose metabolism, lipid profile, hormonal changes, strength, and aerobic capacity. Subgroup analysis revealed that RT had greater adaptive responses (p < 0.01) in fasting blood sugar (ES = -0.52), HOMA-IR (ES = -0.87), testosterone (ES = 0.52), cortisol (ES = -0.82), FGF-21 (ES = 0.61), and maximal strength (ES = 1.19) compared to AT. Conversely, AT showed greater changes (p < 0.01) in cholesterol (ES = -0.28), triglyceride (ES = -0.64), HDL (ES = 0.46), LDL (ES = -0.73), and aerobic capacity (ES = 1.18) compared to RT. Overall, both RT and AT interventions yielded significant moderate to large ES in FGF-21 levels and enhanced the management of biochemical variables. RT is an effective method for controlling FGF-21 levels and glucose balance, as well as for inducing hormonal changes. On the other hand, AT is more suitable for improving lipid profiles in overweight men with type 2 diabetes mellitus. [ABSTRACT FROM AUTHOR]

Background: The pancreas contains Vitamin D (Vit.-D) and an enzyme that regulates insulin levels. Vitamin D deficiency is associated with several severe health conditions, such as cardiovascular disease, diabetes, various cancers, and multiple sclerosis (MS). Therefore, the objective of the current study is to investigate the correlation between Vitamin D levels and glycosylated hemoglobin (HbA1c) in both patients and controls. Materials and Methods: The study enrolled 163 T2DM (Group I) and 174 healthy (Group II) subjects to estimate the level of Vit.-D and HbA1c in the blood serum of all subjects, using radioimmunoassay and high-performance liquid chromatography, respectively. Results: The results revealed that Group I had a lower level of Vit.-D (17.82 ± 2.23 ng/mL) than Group II. Similarly, they have high HbA1c levels, showing significant (P < 0.001; Student's t -test). Further, in Group I, Vit.-D levels were negatively correlated with HbA1c levels (P < 0.001; r2 = 0.07, linear regression). Group I and II combined analysis revealed an inverse relationship between Vit.-D levels and HbA1c (P < 0.001; r2 = 0.021). Conclusion: The study concluded a negative correlation between Vit.-D levels and HbA1c levels in T2DM. However, Vit.-D therapy can recover this correlation, which helps control glycemic levels. [ABSTRACT FROM AUTHOR]

Abstract The transition from radioimmunoassay (RIA) to chemiluminescent enzyme immunoassay (CLEIA) for plasma aldosterone concentration (PAC) assays has raised concerns over its impact on primary aldosteronism (PA) diagnosis. This study investigated the correlation between PAC and renin values using RIA, CLEIA, and liquid chromatography/mass spectrometry/mass spectrometry (LC–MS/MS), established cutoff values for PA diagnosis using the aldosterone-to-renin ratio (ARR) with PAC_CLEIA, and assessed the differences in PAC values by measuring weak mineralocorticoids (WMs). This retrospective study evaluated 312 serum PAC samples using RIA, CLEIA, and LC–MS/MS, and analyzed 315 plasma renin samples. Method correlations were assessed through Passing-Bablok regression. Receiver operating characteristic curves determined ARR cutoffs for PA diagnosis. WMs were quantified to evaluate their impact on ΔPAC (RIA-LC–MS/MS) through multiple regression analysis. PAC_CLEIA and PAC_LC-MS/MS values were highly correlated. ARRs derived from PAC_RIAs demonstrated more false positives and lower specificity than ARRs using PAC_CLEIA or PAC_LC-MS/MS. WMs significantly influenced ΔPAC in both the PA and non-PA groups. ARRs using PAC_CLEIA are valuable for determining PA cutoffs in clinical practice. The transition to PAC using CLEIA may enhance PA detection rates. WMs were found to interfere with PAC measurements in the RIA method, affecting outcomes.

Cortisol is the primary biomarker associated with the hypothalamic–pituitary–adrenal (HPA) axis. This research aimed at assessing the predictive potential and the relationship of cortisol concentrations in four media (blood, milk, whey and hair) with parity, lactation stage and productive classes. Multiparous (n = 30) and primiparous (n = 38) Italian Mediterranean buffaloes were used and assigned to four productive classes (based on percentiles of mature equivalent milk yield (EMY) and mature equivalent energy corrected milk (eECM)), and cortisol concentrations were measured using a in house radioimmunoassay (RIA) method. Parity did not show a significant effect on cortisol concentrations of the four media. The catabolic stage of lactation (up to 90 d in milk (DIM)) was characterised by higher cortisol concentrations compared to the second anabolic stage (beyond 150 DIM) in milk formulations. The plasmatic concentrations of cortisol were higher at the catabolic and the first anabolic stage (91–150 DIM) compared to the second anabolic (p = 0.022 and p = 0.009, respectively). Buffaloes beyond 150 DIM differed from those below 90 DIM (p

In Japan, the traditional method for measuring plasma aldosterone concentration (PAC) was radioimmunoassay (RIA), which had several challenges, including poor traceability of certified reference materials and reduced detection sensitivity at low concentrations. To overcome these issues, a chemiluminescent enzyme immunoassay (CLEIA) for PAC measurement was introduced in April 2021 and the Japan Endocrine Society published new guidelines for primary aldosteronism (PA). This study aimed to evaluate the impact of the transition from RIA to CLEIA for PAC measurement on PA diagnosis. Data from 190 patients admitted to the Second Department of Internal Medicine, University of the Ryukyus Hospital, between April 2012 and March 2021 were analyzed. Patients who were diagnosed with PA underwent adrenal venous sampling. The PAC measured by RIA (PAC(RIA)) was converted to the estimated PAC measured by CLEIA (ePAC(CLEIA)) using a conversion formula. The present study evaluated the discordance rates in diagnoses based on screening (SC), captopril challenge test (CCT), saline infusion test (SIT), and diagnosis of PA between results judged by PAC(RIA) according to the previous guidelines and those judged by ePAC(CLEIA) according to the new guidelines. The results revealed discordant diagnosis rates of 6.4% for SC and 10.1% for CCT, with no discordance for SIT. The discordant diagnosis rate for PA was 3.7%. Our study reveals the challenges in establishing appropriate diagnostic criteria for PA using PAC(CLEIA) and highlights the demand for further research on provisionally positive categories.

The Pregnancy-Associated Glycoproteins (PAGs) belong to aspartic protease family and are synthesized by trophoblast cells of ruminant placenta and secreted into maternal blood and milk. These biomarkers very closely associated to pregnancy have demonstrated their efficiency and reliability in detection of vital embryos or pregnancy loss. In order to measure the concentration of PAGs in bovine serum, a direct radioimmunoassay (RIA) system was developed and validated. The radioiodinated bPAG tracer was prepared by chloramine-T direct method and the solid phase was used in RIA/PAGs as a separation system. The optimization tests allowed to set the assays procedure as working buffers, time and temperature of the assays incubation. Assay sensitivity was 0.63 ng/mL, the intra and inter-assay CVs were 7.7% and 5.3% respectively and the accuracy of the assays was expressed by a good parallelism and recovery tests. The serums bPAG values determinated by this developed system were consistent with the reproduction status and pregnancy stages of cows. The developed RIA/PAG system can provide the laboratory with an efficient method for the accurate detection of pregnancy, which could offer veterinarians and farmers an important tool for the reproductive management in dairy herds. Moreover, this diagnostic tool can also be developed for other livestock species.

The Pregnancy-Associated Glycoproteins (PAGs) belong to aspartic protease family and are synthesized by trophoblast cells of ruminant placenta and secreted into maternal blood and milk. These biomarkers very closely associated to pregnancy have demonstrated their efficiency and reliability in detection of vital embryos or pregnancy loss. In order to measure the concentration of PAGs in bovine serum, a direct radioimmunoassay (RIA) system was developed and validated. The radioiodinated bPAG tracer was prepared by chloramine-T direct method and the solid phase was used in RIA/PAGs as a separation system. The optimization tests allowed to set the assays procedure as working buffers, time and temperature of the assays incubation. Assay sensitivity was 0.63 ng/mL, the intra and inter-assay CVs were 7.7% and 5.3% respectively and the accuracy of the assays was expressed by a good parallelism and recovery tests. The serums bPAG values determinated by this developed system were consistent with the reproduction status and pregnancy stages of cows. The developed RIA/PAG system can provide the laboratory with an efficient method for the accurate detection of pregnancy, which could offer veterinarians and farmers an important tool for the reproductive management in dairy herds. Moreover, this diagnostic tool can also be developed for other livestock species. [ABSTRACT FROM AUTHOR]

Radioimmunoassay (RIA) for C-reactive protein (CRP) is a clinical tool to quantify CRP, a cardiovascular diseases (CVDs) marker, in human serum. Development of the Radioimmunoassay includes radioiodination of CRP with 125I radioisotope, where radioiodination is conducted following the Chloramine-T method. The present study standardizes the radiolabeling procedure and key reagent concentrations like chloramine-T (oxidizing reagent), sodium metabisulfite, and potassium iodide. The outcome of the standardized radioiodination includes a reaction time of 60 s, iodination analytical parameters like % Radiochemical purity was ~ 97% with specific activity ~ 17 µCi/µg, and the tracer was stable for the 60 days. The optimized radioiodination method is simple, reproducible, and has high tracer stability with high immunoreactivity to develop an RIA procedure to quantify CRP in human serum. [ABSTRACT FROM AUTHOR]

The cervix is an important organ that has to dilate sufficiently at delivery to allow the foetus to transition to extrauterine life. Insufficient dilatation of the cervix (IDC) is a frequent cause of dystocia in cattle. The mechanisms underlying cervical opening and the pathogenesis of IDC are still widely unclear. Systematic studies on the relationship between IDC and steroid hormones have been limited and have yielded inconsistent findings. This study aimed to measure oestrogen and progesterone (P4) concentrations in intrapartum cows presented with dystocia due to IDC and in a comparison (C) group of cows with eutocic delivery. Before any obstetrical procedures, and right after the initial evaluation, blood samples were taken from IDC and C animals. Concentrations of P4, oestradiol‐17β (E2), free total oestrogens (FTE) and conjugated total oestrogens (CTE) were measured by established radioimmunoassays. Concentrations of P4 (p =.538), FTE (p =.065) and CTE (p =.605) were not statistically different between C and IDC groups. However, E2 levels in group C were significantly lower when compared to those in the IDC group (p =.013), which is inconsistent with the function of oestrogens in cervical dilatation. The correlation analysis demonstrated significant positive correlations between the pairs P4 versus FTE, P4 versus E2 and FTE versus E2 in group C and between the pair FTE versus E2 in group IDC. In conclusion, the results suggest that local activities of steroids relevant to the aetiology of IDC are not reflected by concentrations in the systemic circulation or that other factors are clearly more important. [ABSTRACT FROM AUTHOR]

Sociability indicates the variances of affinity and solitary among animals and it is strongly correlated with cortisol levels. This study was conducted in 2020, from September 16th to October 15th, at the Zhuanglang Musk Deer Farm in Gansu Province of China. We recorded behaviours of 41 captive Alpine musk deer (Moschus chrysogaster) (in 12 enclosures) using the integrated methods of focal sampling and all occurrence recording for sociability analysis, while non-invasive sampling methods and radioimmunoassay were used to collect and measure faecal cortisol level. The relationship between the sociability of captive Alpine musk deer and faecal cortisol hormone was analysed. The results showed that the sociability of captive Alpine musk deer was 14.754% (± 3.026%) (n=41). Individuals in high-density enclosures (19.090% ± 4.497%, n=26) were significantly more sociable as compared to those in low-density enclosures (7.979% ± 2.640%, n=15; P=0.011). The cortisol level of Alpine musk deer was 109.215 (± 5.349) ng/g (n=41). The cortisol level of female individuals (120.419 ± 11.103 ng/g, n=14) was significantly higher than that of male individuals (103.406 ± 5.575 ng/g, n=27; P=0.008). The male musk deer in high sociability (44.114% ± 9.364%, n=8) had lower faecal cortisol levels (102.415 ± 14.885 ng/g, n=8) than those in low sociability (5.518% ± 1.369%, n=19) (103.823 ± 5.243 ng/g, n=19), however these differences were not significant (P=0.911). The female musk deer in high sociability (20.556% ± 2.174%, n=3) had significantly lower (P=0.035) faecal cortisol levels (77.018 ± 17.036 ng/g, n=3) than those in low sociability (7.773% ± 1.595%, n=11) (132.256 ± 11.079 ng/g, n=11), which could be related to maternal care and weaning period. It is suggested that the enclosing density should be increased to improve captive Alpine musk deer’s (especially female musk deer) sociability in the domestication of musk deer. [ABSTRACT FROM AUTHOR]

The objective: to study the characteristics of the physical and sexual development of girls conceived with the help of assisted reproductive technologies (ART). Materials and methods. An examination of 16 teenage girls who were born with the help of ART was carried out. The average age of the girls was 12.0±0.45 (10–13) years. All girls underwent a clinical, functional, hormonal examinations, secondary sexual characteristics were evaluated according to the J. Tanner scale. Blood levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), thyroid-stimulating hormone (TSH), dehydroepiandrosterone sulfate (DHEA-S), as well as estradiol (E2), testosterone, levels of thyroid hormones triiodothyronine (T3) and thyroxine (T4) were determined by the radioimmunoassay method on the microspectrophotometer “Stat fax” 303 PLUS USA in the first phase of the menstrual cycle. To study the functional activity of the liver, fasting glucose level and oral glucose tolerance test, insulin, alanine amino-transferase and aspartate aminotransferase levels were determined. In order to compare the concentration of hormones, 25 teenage girls of the appropriate age who were born from a spontaneous pregnancy and have a physiological course of the puberty period were additionally examined. Results. According to the ultrasound examination, the length of the right and left ovaries, as well as all echographic indicators, are significantly increased in girls who were born with the help of ART. A study of hormone levels in teenage girls conceived with ART revealed statistically higher levels of LH (5.05±1.17 mIU/ ml), DHEA-S (91.9±16.93 μg/dL), E2 (78.13±17.98 pg/ml), which indicates hyperandrogenism and hyperestrogenism of adrenal origin. At the same time, the fasting glucose level and its level after the oral glucose tolerance test were statistically lower than similar indicators in girls born from a spontaneous pregnancy and with the physiological course of puberty, but the fasting insulin level was higher (p<0.05). Conclusions. In girls who were born with the help of assisted reproductive technologies (ART), premature development of secondary sexual characteristics (late precocious puberty) and pronounced hair growth have been identified. Carbohydrate metabolism, enzymatic activity of the liver and functional activity of the kidneys in girls who were born with the help of ART do not differ from similar indicators of girls who were born from spontaneous pregnancy and with the physiological course of puberty. [ABSTRACT FROM AUTHOR]

Hypothyroidism is an endocrine disorder due to diminished synthesis and secretion of triiodothyronine (T3) and thyroxine (T4) by the thyroid glands. The present study was carried out at Veterinary Clinical Complex of NTR College of Veterinary Science, Gannavaram (India) during the period from March to November 2023. The hospital prevalence of hypothyroidism was found to be 0.45 % (27/5957). The mean values of TT4 (0.60±0.05 µg/dL) and FT4 (0.32±0.04 ng/dL) in hypothyroid dogs were significantly (p =0.01) lower when compared to healthy dogs. Haemato-biochemical changes included significantly (p<0.01) reduced levels of haemoglobin, packed cell volume, and total erythrocyte count, along with elevated total leukocyte count, hypercholesterolemia, hypertriglyceridemia, and significantly (p<0.05) higher levels of alkaline phosphatase and blood urea nitrogen. These findings emphasize the significant role of thyroid hormones in regulating the body›s metabolic balance and ensuring the integrity of various organs. [ABSTRACT FROM AUTHOR]

BackgroundThe ability to detect bacteriuria in dogs with a point-of-care test might improve medical care and antimicrobial stewardship.Hypothesis and objectiveA rapid immunoassay (RIA; RapidBac) will provide acceptable sensitivity and specificity for diagnosis of bacteriuria.AnimalsForty-four client-owned dogs with a clinical indication for urinalysis and aerobic bacterial urine culture.MethodsProspective study. Urine, collected by cystocentesis, was submitted for urinalysis and culture at a diagnostic laboratory. Owners completed an enrollment questionnaire regarding their dogs' clinical signs. The RIA was performed according to the manufacturer's guidelines. Results were compared to culture.ResultsForty-four urine specimens were evaluated from 44 dogs. The sensitivity and specificity of the RIA test to detect bacteriuria compared to urine culture were 81.8% (95% CI, 65.7%-97.9%) and 95.5% (95% CI, 86.8%-99.9%), respectively. For cultures yielding ≥103  CFU/mL, sensitivity increased to 90.0% (95% CI, 76.9%-100%) and specificity was similar at 95.2% (95% CI, 86.1%-99.9%). Malodorous urine, bacteriuria, and pyuria were more likely to be present in dogs with positive RIA or urine culture results compared to dogs with negative results.Conclusions and clinical importanceThe RIA was easy to perform and had good sensitivity and excellent specificity in this group of dogs. The RIA might be a useful screening test for decision-making regarding antimicrobial therapy in dogs with a clinical indication for urine culture. Consideration could be given to amending the International Society for Companion Animal Infectious Disease definition of bacterial cystitis as the presence of signs of lower urinary tract disease together with positive culture or a positive RIA.

The Wellness Ready Test (WRT) is a lateral flow, stall-side assay that measures equine insulin in whole blood and requires validation before recommending clinical use. We evaluated intra- and inter-assay precision and linearity and compared the WRT with a radioimmunoassay (RIA). Tested concentrations ranged from 695 pmol/L (100 μIU/mL). For 20 replicates at each insulin level, intra-assay CVs of the WRT for insulin were 13.3%, 12.9%, and 15.3% at low (139-278 pmol/L; 20-40 μIU/mL), intermediate (278-417 pmol/L; 40-60 μIU/mL), and high (>417 pmol/L; >60 μIU/mL) concentrations, respectively. For 10 replicates at each level (3 assay lots), inter-assay CVs were 15.9%, 11.0%, and 11.7%, respectively. In the weighted linear regression of 5 measured insulin concentrations against expected concentrations, R2 = 0.98, slope = 1.02, and y-intercept = 14.4 pmol/L (2.08 μIU/mL). The Spearman correlation coefficient (rs) was 0.90 (95% CI: 0.85-0.94) between the WRT and RIA; the WRT = f(RIA) Passing-Bablok regression yielded the fit, y = 1.005x + 24.3 pmol/L (3.50 μIU/mL). The WRT result averaged 10.4% higher than the RIA result, with targeted bias of 25.9, 26.1, and 26.7 pmol/L (3.74, 3.76, and 3.84 μIU/mL) for cutoffs used to diagnose insulin dysregulation of 312, 347, and 451 pmol/L (45, 50, and 65 μIU/mL). Assay clinical sensitivities, specificities, and accuracies determined at the 3 selected clinical cutoffs and using the RIA as gold standard were 87-95%, 92-96%, and 91-95%, respectively (n = 99 samples). Observed total error was 28.4-30.4%. The WRT had acceptable precision, excellent linearity, and good association with the RIA.

Objectives: Anti‐acetylcholine receptor antibodies (AChR Ab) are one of the pathogenic autoantibodies in myasthenia gravis (MG), and they are specifically mentioned in the Japanese clinical guidelines for MG. In the serological diagnosis and measurement of autoantibodies as an adjunct marker for MG, the radioimmunoassay (RIA) method has been conventionally used as the gold standard. However, there have been several worldwide concerns regarding the use of radioisotopes. In this study, we evaluated the performance of the EUROIMMUN AChR Ab enzyme‐linked immunosorbent assay (ELISA) in comparison with RIA. Methods: ELISA reproducibility and dilution linearity were analyzed using AChR Ab‐positive samples. Sera from 50 patients with suspected MG and 50 healthy donors were used to assess the correlation and qualitative agreement between the two methods. Results: The ELISA showed good interassay precision and dilution linearity. The correlation coefficient, sensitivity, specificity and overall agreement of the ELISA in comparison with the conventional RIA amounted to 0.87, 98%, 92% and 95% (kappa = 0.895), respectively. Analysis of the discordant results suggested that the ELISA might be more capable of detecting low AChR Ab reactivity in patients with suspected MG. Conclusions: The AchR Ab ELISA provides a reliable tool for the quantification of AchR Ab, supporting the diagnosis of MG. As a non‐isotopic assay with good precision, shorter handling time and high correlation with the conventional RIA, the AChR ELISA could be a useful alternative in the laboratory routine. [ABSTRACT FROM AUTHOR]

Background: Polygonum multiflorum (PM), a widely used traditional Chinese medicine herb, is divided into two forms, namely raw polygonum multiflorum (RPM) and polygonum multiflorum praeparata (PMP), according to the processing procedure. Emerging data has revealed the differential hepatotoxicity of RPM and PMP, however, its potential mechanism is still unclear. Methods: In our study, we investigated the differential hepatotoxicity of RPM and PMP exerted in C57BL/6 mice. First, sera were collected for biochemical analysis and HE staining was applied to examine the morphological alternation of the liver. Then we treated L02 cells with 5 mg / mL of RPM or PMP. The CCK8 and EdU assays were utilized to observe the viability and proliferation of L02 cells. RNA sequencing was performed to explore the expression profile of L02 cells. Western blotting was performed to detect the expression level of ferroptosis-related protein. Flow cytometry was used to evaluate ROS accumulation. Results: In our study, a significant elevation in serum ALT, AST and TBIL levels was investigated in the RMP group, while no significant differences were observed in the PMP group, compared to that of the CON group. HE staining showed punctate necrosis, inflammatory cell infiltration and structural destruction can be observed in the RPM group, which can be significantly attenuated after processing. In addition, we also found RPM could decrease the viability and proliferation capacity of L02 cells, which can be reversed by ferroptosis inhibitor. RNA sequencing data revealed the adverse effect of PM exerted on the liver is closely associated with ferroptosis. Western blotting assay uncovered the protein level of GPX4, HO-1 and FTL was sharply decreased, while the ROS content was dramatically elevated in L02 cells treated with RPM, which can be partially restored after processing. Conclusions: The hepatotoxicity induced by RPM was significantly lower than the PMP, and its potential mechanism is associated with ferroptosis. [ABSTRACT FROM AUTHOR]

Background. Detection of type 1 diabetes (T1D) at the preclinical stage is possible by detecting islet autoantibodies (IAs) years before the appearance of symptomatic diabetes. The Antibody Detection Israeli Research is a general population screening program searching for children with multiple IAs who are at risk of developing T1D. IAs are measured in capillary or venous whole blood (WB) samples using the novel ultrasensitive antibody detection by agglutination-PCR (ADAP) technology. Objective. To assess the accuracy and reliability of the ADAP assay in venous and capillary WB. Materials and Methods. In total, 50 children with T1D and 50 healthy controls participated in the study. Venous and capillary blood samples were drawn from participants with T1D, while only venous blood was drawn from the controls. The ADAP assay in venous and capillary blood was compared to the currently used assays in their ability to detect glutamic acid decarboxylase (GADA), islet antigen-2 (IA-2A), and insulin autoantibodies (IAAs). Results. The area under the curve using the receiver operating characteristic curves was comparable between the ADAP assay in WB and standard enzyme-linked immunosorbent assay (ELISA)/radioimmunoassay (RIA) for all three IAs GADA 0.946 (95% CI: 0.900–0.991) vs. 0.949 (0.906–0.992), P = 0.873 ; IA-2A 0.747 (0.649–0.844) vs. 0.666 (0.587–0.744), P = 0.106 ; IAA 1.000 (1.000–1.000) vs. 1.000 (1.000–1.000), P = 1.000. The correlation between the levels of IA in venous and capillary WB using ADAP was R2 = 0.958 (P < 0.01), R2 = 0.943 (P < 0.01), and R2 = 0.711 (P < 0.01) for GADA, IA-2A, and IAA, respectively. IA levels in venous and capillary WB using ADAP were comparable without a proportional bias in Bland–Altman's plots of agreement, suggesting the two methods may be used interchangeably. Conclusions. The ADAP assay is reliable in detecting IA in venous and capillary WB samples with comparable performance to standard RIA and ELISA. These findings open avenues for widespread use of the ADAP assay in future general population screening programs to detect children at risk of developing T1D. [ABSTRACT FROM AUTHOR]

Study Objectives: The aim of this cross-sectional study is to explore the association between serum 25-hydroxyvitamin D [25(OH)D] levels, a marker of Vitamin D status, and excessive daytime sleepiness (EDS), expressed as increased scores of the Epworth Sleepiness Scale (ESS), in a group of prospectively enrolled patients with obstructive sleep apnea (OSA). Methods: Newly diagnosed patients with OSA, divided into two groups, those with EDS (ESS > 10) and those without EDS (ESS < 10). All patients underwent night polysomnography. Measurement of serum 25(OH)D vitamin was performed using a radioimmunoassay. Results: In total, 217 patients with OSA (197 males and 20 females) were included. Patients with EDS had higher AHI (p < 0.001) values and lower mean serum 25(OH)D levels, compared with those of non-somnolent patients [17.4 (12.2–25.7) versus 21.1 (15.3–28.8) ng/mL, respectively, p = 0.005]. In patients with EDS, serum 25(OH)D levels correlated with average oxyhemoglobin saturation during sleep (r = 0.194, p = 0.043), and negatively with ESS score (r = −0.285, p = 0.003), AHΙ (r = −0.197, p = 0.040) and arousal index (r = −0.256, p = 0.019). Binary regression analysis identified Vit D serum levels (β = −0.045, OR: 0.956, 95% CI: 0.916–0.997, p = 0.035), total sleep time (β = 0.011, OR: 1.011, 95% CI: 1.002–1.021, p = 0.016) and AHI (β = 0.022, OR: 1.022, 95% CI: 1.003–1.043, p = 0.026) as independent predictors of EDS in patients with OSA. In patients with EDS, multiple regression analysis indicated that ESS score was negatively associated with Vit D serum levels (β = −0.135, p = 0.014) and minimum oxyhemoglobin saturation during sleep (β = −0.137, p = 0.043). Conclusions: In the present study, EDS in patients with OSA is associated with low levels of Vitamin D, while sleep hypoxia may play a role in this process. [ABSTRACT FROM AUTHOR]

Purpose: To our knowledge, there are very few studies evaluating if the levels of folate modify the risk of cervical intraepithelial neoplasia grade 2 and higher (CIN2+ and CIN3+) associated with the levels of HPV genome methylation, two cofactors related to single carbon metabolism and independently associated with cervical cancer in previous studies. We conducted a case–control study nested in a three-arm randomized clinical pragmatic trial (ASCUS-COL trial) to evaluate the risk of CIN3+ associated with methylation levels according to serum folate concentrations. Methods: Cases (n = 155) were women with histologically confirmed CIN2+ (113 CIN2, 38 CIN3, and 4 SCC) and controls were age and follow-up time at diagnosis-matched women with histologically confirmed ≤ CIN1 (n = 155), selected from the 1122 hrHPV + women of this trial. The concentrations of serum folate were determined by the radioimmunoassay SimulTRAC-SNB-VitaminB12/Folate-RIAKit and the methylation levels by the S5 classifier. Stepwise logistic regression models were used to estimate the association between folate or methylation levels and CIN2+ or CIN3+. The joint effect of folate levels and methylation on the risk of CIN3+ was estimated using combinations of categorical stratifications. Results: Folate levels were significantly lower in women with CIN3+ than in other diagnostic groups (p = 0.019). The risk of CIN3+ was eight times higher (OR 8.9, 95% CI 3.4–24.9) in women with folate deficiency and high methylation levels than in women with normal folate and high methylation levels (OR 1.4, 95% CI 0.4–4.6). Conclusion: High methylation and deficient folate independently increased the risk of CIN3+ while deficient folate combined with high methylation was associated with a substantially elevated risk of CIN3+. [ABSTRACT FROM AUTHOR]

Oxytocin (OT) is a peptide hormone synthesized in the hypothalamus and released into systemic circulation or other areas of the brain. Its physiological roles include action as a hormone with stimulation of uterine contractions and that as a neuromodulator with involvement in social behaviors and regulation of mood. Its small size and low levels within biological matrices make it challenging to accurately measure. The goal of this study was to demonstrate the specificity of the antibody, sensitivity, and reproducibility of the Phoenix Pharmaceuticals (PP) OT radioimmunoassay (RIA) for use in human urine, serum, and saliva. Specificity of the antibody was assessed by high pressure liquid chromatography with ultraviolet (HPLC-UV) separation and assay of the fractions. Immunoreactivity was evaluated using the percent OT bound, and the fraction retention times were compared to the retention time of an intact OT standard to determine which fractions contained OT in the extracted samples. Reproducibility was assessed by running replicates of pools of each biomatrix over several assays. Sensitivity was assessed by repeated measurement of physiologically relevant low-concentration specimens. In all tested specimens the greatest reactivity in assay corresponded to the same fraction(s) as the OT standard. Only minimal reactivity was found in the other fractions, suggesting that in an unfractionated sample the antibody reacts mostly with intact OT. Reproducibility was acceptable for all specimens and the coefficient of variation (CV) ranged from 3.72 to 8.04% and 5.89–12.8%, for intra and inter-assay, respectively. The limits of quantitation (LOQ) were sufficient for measurement of normal values in urine (0.643 & 1.43 pg/mL), serum (1.90 pg/mL), and saliva pools (0.485 & 4.42 pg/mL). In conclusion, the PP OT RIA is specific and sensitive enough for reproducible measurement of intact OT in human peripheral biological matrices.

Plasma aldosterone concentration (PAC) was routinely measured using radioimmunoassay (RIA); however, the RIA kit was discontinued in March 2021 in Japan. This study examined PAC conversion in adrenal venous sampling (AVS) and AVS criteria when measured using chemiluminescent enzyme immunoassay (CLEIA). PAC of 415 adrenal venous blood samples from AVS (including segmental AVS) of 63 patients with primary aldosteronism was measured using RIA (Spac-S aldosterone kit; Fujirebio Inc.) and CLEIA (Lumipulse Presto Aldosterone; Fujirebio Inc.). PAC of 70 AVS samples was also measured using liquid chromatography-mass spectrometry (LC-MS/MS, ASKA Pharma Medical Co., Ltd.). PAC conversion formulas were determined for each AVS sample assay. PAC measured using CLEIA was significantly correlated with that measured using RIA (correlation coefficient = 0.971). The PAC conversion formula was PAC (CLEIA) = PAC (RIA) × 0.772 – 1,199 pg/mL. The PAC of 14,000 pg/mL in RIA was equivalent to 9,613 pg/mL in CLEIA. PAC measured using CLEIA was also correlated with that measured using LC-MS/MS, and the PAC conversion formula was PAC (CLEIA, pg/mL) = 0.97 × PAC (LC-MS/MS, pg/mL) + 211. The inter-assay coefficient of variability (CV) was 1.1–1.3% and intra-assay CV was 1.0–1.7%, measured using CLEIA. The PAC conversion formula for AVS samples was obtained using CLEIA and RIA, and the conversion formula was different from that for peripheral blood. PAC values measured by CLEIA showed preferable accuracy and high concordance with those measured by LC-MS/MS, even in AVS samples. The study outcomes are useful for interpreting AVS results using non-RIA measurement methods.

Introduction: Cerebral white matter hyperintensities (WMHs) are commonly found in the aging brain and have been implicated in the initiation and severity of many central nervous system diseases. Furthermore, an increased WMH volume indicates reduced brain health in older adults. This study investigated the association between WMH volume and physical activity in older adults with depressive symptoms (DS) and mild memory impairment (MMI). Factors associated with the WMH volume were also investigated. Methods: A total of 57 individuals aged over 65 years with DS and MMI were included in this study. The participants underwent magnetic resonance imaging to quantify WMH volumes. After WMH volume was accumulated, normalized to the total intracranial volume (TIV), the percentage of WMH volume was calculated. In addition, all participants wore a triaxial accelerometer for 2 weeks, and the average daily physical activity and number of steps were measured. The levels of blood biomarkers including cortisol, interleukin-6 (IL-6), brain-derived insulin-like growth factor-1, and brainderived neurotrophic factor were measured. Motor and cognitive functions were also assessed. Results: Faster maximum walking speed and longer time spent engaged in moderate physical activity were associated with a smaller percent of WMH volume, whereas higher serum IL-6 levels were associated with a larger percent of WMH volume. The number of steps per day, time spent engaged in low levels of physical activity, cognitive function, and all other measured biomarkers were not significantly associated with percent of WMH volume. Discussion: Higher blood inflammatory cytokine levels, shorter duration of moderate physical activity, and lower maximum walking speed were associated with a higher percent of WMH volume. Our results provide useful information for maintaining brain health in older adults at a high risk of developing dementia and may contribute to the development of preventive medicine for brain health. [ABSTRACT FROM AUTHOR]

Objective Preeclampsia (PE) is a pregnancy-specific condition featured by high blood pressure, edema, and proteinuria. Research about the role of microRNA (miR)-203 in PE remains insufficient. This experiment is designed to investigate the specific role of miR-203 in trophoblasts in PE. Study Design miR-203 expression in placenta tissues of normal pregnant women and PE patients was examined to analyze the relevance between miR-203 and PE diagnostic efficiency and between miR-203 and blood pressure (systolic pressure and diastolic pressure) and proteinuria of PE patients. miR-203 expression was downregulated in hypoxia-cultured trophoblasts using miR-203 inhibitor to assess matrix metalloproteinase-9 (MMP-9) level. Then, the angiogenesis of trophoblasts with different treatments was determined. Subsequently, the target relation between miR-203 and insulin-like growth factor receptor 1 (IGF-1R) was predicted and verified. Additionally, the effect of IGF-1R in the mechanism of miR-203 modulating trophoblast vascular remodeling was detected. Results miR-203 was overexpressed in the placenta of PE patients and it acted as a promising diagnostic indicator for PE. Moreover, miR-203 was positively associated with blood pressure (systolic pressure and diastolic pressure) and proteinuria of PE patients. miR-203 silencing in hypoxia-cultured trophoblasts enhanced trophoblast vascular remodeling. Mechanically, miR-203 bound to IGF-1R to suppress its transcription. IGF-1R downregulation counteracted the promotive effect of miR-203 silencing on trophoblast vascular remodeling. Conclusion miR-203 was overexpressed in PE, and it targeted IGF-1R to limit trophoblast vascular remodeling. Key Points miR-203 is overexpressed in the placenta of PE patients. miR-203 acts as a potential diagnostic marker for PE. miR-203 targets IGF-1R to reduce trophoblast vascular remodeling in PE. [ABSTRACT FROM AUTHOR]

Luteinizing hormone (LH) is a useful biomarker for identifying ovulation events in the cows to predict the time of ovulation to achieve a high success rate of conception following artificial insemination. Although antibody-based radioimmunoassay and enzyme-linked immunosorbent assay are being used for LH measurement, these techniques are expensive, time-consuming, and require expertise and sophisticated laboratory facilities. So, there is a need for a field-applicable, affordable, easy-to-use method for LH detection. For developing such a specific, quantitative, and inexpensive system, an aptamer-based smartphone-enabled aptasensor has been investigated. The aptamer was used instead of the antibody as a biorecognition element due to its comparative stability at ambient temperature, ease of synthesis, and cost-effectiveness. Electrochemical impedance spectroscopy has been used to obtain label-free detection of LH within 20 min in ~ 20 μL sample volume. The screen-printed gold electrode is compatible with a smartphone-enabled miniaturized device (Sensit Smart; Palmsens BV, The Netherlands) and was fabricated with the aptamer to detect LH in biological fluids (limit of detection 0.80 and 0.61 ng/mL in buffer and undiluted/unprocessed serum, respectively, with the dynamic range of detection of 0.01 to 50 ng/mL). All the data were obtained in the 10 kHz to 0.10 Hz frequency range at a bias potential of 0.30 V with an alternating potential of 10 mV. The clinical relevance of the sensor was evaluated in 10 serum samples collected from dairy animals which established a high correlation with standard LH-ELISA (κ > 0.87). The aptasensor can be stored at room temperature for 30 days without any significant loss in electrochemical sensing ability. [ABSTRACT FROM AUTHOR]

This article discusses a study that examined potential biomarkers for assessing bone health in apparently healthy adults. The study focused on Vitamin D, Vitamin D binding protein (VDBP), and bone-specific alkaline phosphatase (BALP) as markers for overall bone turnover. The study found that the participants had low levels of serum vitamin D and VDBP, and increased BALP levels. The researchers suggest that these biomarkers could be used to assess bone health and emphasize the importance of public health education on the role of vitamin D in maintaining bone health. [Extracted from the article]

Background: Pseudorabies virus (PRV) is one of the major viral pathogens leading to reproductive disorders in swine. However, little is known about the effects of PRV infection on porcine reproductive system. Ovarian granulosa cells are somatic cells surrounding oocytes in ovary and required for folliculogenesis. The present study aimed to investigate the interference of PRV on functions of porcine ovarian granulosa cells in vitro. Methods: Primary granulosa cells were isolated from porcine ovaries. To investigate the PRV infectivity, transmission electron microscopy (TEM) was used to check the presence of viral particles, and the expression of viral gE gene was detected by quantitative real-time PCR (qPCR) in PRV-inoculated cells. After PRV infection, cell viability was detected by MTS assay, Ki67 for proliferative status was determined by immunofluorescence assay (IFA), cell cycle and apoptosis were detected by flow cytometry, and progesterone (P4) and estradiol (E2) were determined by radioimmunoassay. The checkpoint genes of cell cycle and apoptosis-related proteins were studied by qPCR and western blotting. Results: Virus particles were observed in the nucleus and cytoplasm of PRV-infected granulosa cells by TEM imaging, and the expression of viral gE gene increased in a time-dependent manner post infection. PRV infection inhibited cell viability and blocked cell cycle at S phase in porcine granulosa cells, accompanied by decreases in expression of Ki67 protein and checkpoint genes related to S phase. Radioimmunoassay revealed decreased levels in P4 and E2, and the expressions of key steroidogenic enzymes were also down-regulated post PRV-infection. In addition, PRV induced apoptosis with an increase in Bax expression and activation of caspase 9, and the phosphorylation of JNK, ERK and p38 MAPKs were significantly up-regulated in porcine ovarian granulosa cells post PRV infection. Conclusions: The data indicate that PRV causes infection on porcine ovarian granulosa cells and interferes the cell functions through apoptosis, and the MAPK signaling pathway is involved in the viral pathogenesis. [ABSTRACT FROM AUTHOR]

Simple Summary: The significance of serum beta-2 microglobulin (sβ2m) in Hodgkin lymphoma (HL) is controversial. In an effort to investigate the prognostic significance of sβ2m levels in a large series of patients with HL, we analyzed 915 patients, who were treated with ABVD or equivalent regimens with or without radiotherapy. Sβ2m levels were measured by a radioimmunoassay (upper normal limit 2.4 mg/L). The median sβ2m levels were 2.20 mg/L. Freedom from progression (FFP) was significantly inferior in patients with a higher sβ2m at all tested cutoffs. The best cutoff was 2.0 mg/L (10-year FFP 83% vs. 70%, p = 0.001), which performed better than the 2.4 mg/L cutoff ("normal versus high"). Our data suggest that higher sβ2m is a significant independent predictor of FFP, OS and HLSS in HL but the optimal cutoff appears to lie within the normal limits in this predominantly young patient population. The significance of serum beta-2 microglobulin (sβ2m) in Hodgkin lymphoma (HL) is controversial. We analyzed 915 patients with HL, who were treated with ABVD or equivalent regimens with or without radiotherapy. Sβ2m levels were measured by a radioimmunoassay (upper normal limit 2.4 mg/L). Sequential cutoffs (1.8–3.0 by 0.1 mg/L increments, 3.5 and 4.0 mg/L) were tested along with ROC analysis. The median sβ2m levels were 2.20 mg/L and were elevated (>2.4 mg/L) in 383/915 patients (41.9%). Higher sβ2m was associated with inferior freedom from progression (FFP) at all tested cutoffs. The best cutoff was 2.0 mg/L (10-year FFP 83% vs. 70%, p = 0.001), which performed better than the 2.4 mg/L cutoff ("normal versus high"). In multivariate analysis, sβ2m > 2.0 mg/L was an independent adverse prognostic factor in the whole patient population. In multivariate overall survival analysis, sβ2m levels were predictive at 2.0 mg/L cutoff in the whole patient population and in advanced stages. Similarly, sβ2m > 2.0 mg/L independently predicted inferior HL-specific survival in the whole patient population. Our data suggest that higher sβ2m is an independent predictor of outcome in HL but the optimal cutoff lies within the normal limits (i.e., at 2.0 mg/L) in this predominantly young patient population, performing much better than a "normal versus high" cutoff set at 2.4 mg/L. [ABSTRACT FROM AUTHOR]

Marine turtles exhibit temperature-dependent sex determination (TSD), and there is widespread concern that global warming is raising nest incubation temperatures, resulting in increasingly female-skewed sex ratios in "feminized" populations. We assessed the sex ratio of a mixed-stock aggregation of immature green turtles (Chelonia mydas) at a midocean developmental foraging ground in the Northwest Atlantic from 1975 to 2018. We used plasma testosterone concentrations, measured by radioimmunoassay (RIA) and calibrated by laparoscopy, to determine the sex of 2,724 green turtles captured 3,940 times in Bermuda (32°18′ N, 64°46′ W) waters. A logistic regression model correctly predicted the sex of 99.5% of turtles (189/190) with associated testosterone concentrations whose sex had been verified via laparoscopy. Empirical evaluation of the trend in sex ratio using four related data sets showed a significant increase (2.8–4.0% yr−1) in the percent females, ranging from 62.7 to 68.1% during the most recent years of evaluation. Using growth rates to predict the year of arrival of turtles in Bermuda, we estimated the sex ratio of recruiting cohorts over 4 decades. Mixed-stock analysis of mtDNA sequences of 602 turtles that recruited to Bermuda between 1970 and 2018 suggested that multiple, geographically dispersed rookeries contributed to the Bermuda aggregation making it regionally representative. Changes in rookery contributions and strong population increases at certain rookeries may partly explain the increasing trend in the percent female. But the steady rate of increase over decades and the increasing female percentage of arriving cohorts are consistent with impacts of global warming at source rookeries. [ABSTRACT FROM AUTHOR]

To determine differences in glucose control and cardiovascular disease risk factors following three weeks of added soda, 100% fruit juice, or water in apparently healthy, college-aged adults. Thirty-six adults (18 males; 18 females) between the ages of 18 and 30 years of age. A 3-arm randomized controlled parallel-arm trial; at baseline and after three weeks consuming the assigned beverage, participants completed glucose control and cardiovascular disease risk factor assessments. There were no significant differences between beverage conditions for glucose control or cardiovascular disease risk factors (ps > 0.05). There were no significant changes in caloric intake or differences in caloric intake between conditions, p = 0.17. In healthy, young adults, under free-living conditions, short-term consumption of two commercially packaged servings of SBs did not lead to significant glucose control or cardiovascular disease risk factor changes, indicating potential compensation and/or resilience to negative short-term effects. [ABSTRACT FROM AUTHOR]

The dim light melatonin onset (DLMO) is the current gold standard biomarker of the timing of the central circadian clock in humans and is often assessed from saliva samples. To date, only one commercially available salivary melatonin assay is considered accurate at the low daytime levels required to accurately detect the DLMO (Novolytix RIA RK‐DSM2). The aim of this study was to conduct the first independent evaluation of a newly improved enzyme‐linked immunosorbent assay (ELISA; Novolytix MLTN‐96) and compare it with the recommended radioimmunoassay (RIA)—both in terms of melatonin concentrations and derived DLMOs. Twenty participants (15 females, 18–59 years old) provided saliva samples every 30 min in dim light starting 6 h before their habitual bedtime, yielding a total of 260 saliva samples. Both the RIA and ELISA yielded daytime melatonin concentrations <2 pg/mL, indicating adequate accuracy to detect the DLMO. The melatonin concentrations from the two assays were highly correlated (r =.94, p <.001), although the RIA yielded lower levels of melatonin concentration than the ELISA, on average by 0.70 pg/mL (p =.006). Seventeen DLMOs were calculated from the melatonin profiles and the DLMOs from both assays were not statistically different (p =.36) and were highly correlated (r =.97, p <.001). Two DLMOs derived from the RIA occurred more than 30 min earlier than the DLMO derived from the ELISA. These results indicate that the new Novolytix ELISA is an appropriate assay to use if the Novolytix RIA is not feasible or available. [ABSTRACT FROM AUTHOR]

Background: The thyroid function of the dog has been little studied in comparison with that of the human. This may have resulted, in part, from the apparent lack of reporting of thyroid-related disorders in the Indian canine population. There is, therefore, a need for a more detailed study of thyroid hormone levels (establish reference interval) in normal dogs using a specific and accurate technique like radioimmunoassay (RIA). Methods: Total 192 healthy dogs from the clinical setup of constituent colleges of MAFSU (viz. Mumbai, Nagpur, Parbhani, Shirwal, Udgir and Akola) were ethically enrolled to examine thyroid hormones levels (TT3, TT4 and FT4). Thyroid hormones were estimated using RIA. Result: The results of the present study recorded mean serum TT3, TT4 and FT4 values in healthy dogs as 1.29±0.04 nmol/l, 28.17±1.18 nmol/l and 13.03±0.68 pmol/l respectively. The reference interval (25th to 75th percentile) for TT3, TT4 and FT4 was found to be 0.88-1.51 nmol/l, 15.70-35.29 nmol/l and 7.80-14.75 pmol/l respectively. The median for TT3, TT4 and FT4 was found to be 1.13 nmol/l, 24.54 nmol/l and 10.00 pmol/l respectively. The baseline serum thyroid hormone concentration would be valuable for identifying thyroid dysfunction in dogs. [ABSTRACT FROM AUTHOR]

It is important to understand the physiological stressors in animals especially for threatened species or intensively managed to improve their conservation and optimise their reproduction. We sought to understand changes in stress hormones (faecal glucocorticoid metabolites) in black rhinoceros (Diceros bicornis michaeli) in relation to population density and sex (intrinsic factors) and plant minerals, rainfall and land surface temperature (extrinsic factors). We used non-invasive faecal sampling techniques on animals of known sex, age and dominance in seven populations of contrasting population densities over 1year. We measured variability in faecal corticosterone metabolites through radioimmunoassay and related them to population density, sex and faecal calcium, phosphorus, copper, zinc and potassium as characteristic of plant minerals, rainfall and temperature. We used linear mixed models (LMM) to analyse the data. We did not detect a significant relationship between physiological stress parameters and population density. However, we have indications that stress levels increased as rainfall and temperature increased and were correlated negatively and positively with concentration of faecal phosphorus and copper respectively; we found higher stress levels in females than in males. These results suggest that both intrinsic and extrinsic factors explain the variation in physiological stress observed in black rhinoceros. [ABSTRACT FROM AUTHOR]

Larval cod require live prey as food, and prey concentration (PC) and feeding frequency (FF) often affect their growth and survival. Apart from this, water exchange rates/water flow (WER/WF) and water current (WC) also affect the prey resident time in the tank and larval/early juvenile behaviour, respectively. High water current is also known to induce stress in finfish larvae, and this stress response is believed to be dependent on larval developmental stage. Thus, we conducted a study to evaluate three larval rearing protocols varying in prey concentration, feeding frequency, and water current/exchange rate. Three protocols were used: low prey concentration (PC), low feeding frequency (FF), and low water flow (protocol 1); medium PC, medium FF, and medium WF (protocol 2); and high PC, high FF, and high WF (protocol 3) (see Table 1). Larvae were sampled periodically for growth and cortisol measurements. Cortisol extraction and radioimmunoassay (RIA) were conducted using methods previously validated for cod larvae. Larvae reared using protocols 2 and 3 were significantly bigger and heavier than larvae reared using protocol 1. Rearing protocol had a significant effect on the cortisol level in larval cod. Larvae showed a developmental stage-dependent stress response. Protocol 2 had significantly higher survival than protocols 1 and 3. Our results indicate that an intermediate PC, FF, and WF (protocol 2) is suitable during cod larval rearing. [ABSTRACT FROM AUTHOR]

In 1992, a paper reported that the melatonin content of the rat duodenum was 24 000 ± 2000 pg/g tissue (range: 4000–100 000 pg/g) while the pineal melatonin content was 580 000 ± 36 000 pg/g. The data has been used for the last 30 years to infer that the gut produces 400 hundred times more melatonin than the pineal gland and that it is the source of plasma melatonin during the daytime. No‐one has ever challenged the statement. In this review, evidence is summarised from the literature that pinealectomy eliminates melatonin from the circulation and that studies to the contrary have relied upon poorly validated immunoassays that overstate the levels. Similarly studies that have reported increases in plasma melatonin following tryptophan administration failed to account for cross reactivity of tryptophan and its metabolites in immunoassays. The most extraordinary observation from the literature is that in those studies that have measured melatonin in the gut since 1992, the tissue content is vastly lower than the original report, even when the methodology used could be overestimating the melatonin content due to cross reactivity. Using the more contemporary results we can calculate that rather than a 400:1 ratio of duodenum: pineal melatonin, a ratio of 0.05–0.19: 1 is likely. The gut is not a major extra‐pineal source of melatonin; indeed it may well not produce any. [ABSTRACT FROM AUTHOR]

The salivary protein quantification reflecting the periodontal pathophysiology among patients with different periodontitis severity has not been well explored. This quantification is fundamental as the changes in the quantity of proteins in different severities of periodontitis can reflect the periodontal complex alterations. This pilot study aimed to quantify and compare the salivary proteins by collecting unstimulated saliva from periodontitis patients and healthy individuals. The radioimmunoprecipitation assay (RIPA) protocol was used to extract protein from patients' saliva and quantified using the Nanodrop Spectrophotometer 2000/2000c (A280) set at 280 nm wavelength. The mean concentration of salivary proteins was shown to be 26.48 ± 2.95 mg/mL in healthy patients, 22.00 ± 0.38 mg/mL in Stage 1 periodontitis patients, 27.34 ± 1.61 mg/mL in Stage 2 periodontitis patients, 27.11 ± 0.66 mg/mL in Stage 3 periodontitis patients and 24.89 ± 1.91 mg/mL in Stage 4 periodontitis patients. However, the mean difference between all groups was not statistically significant. Within the limitations of this study, there was no difference in the salivary protein quantity among patients with different stages of periodontitis. [ABSTRACT FROM AUTHOR]

Fluoxetine is a widely used antidepressant that acts as a selective serotonin reuptake inhibitor. This study investigated the effects of fluoxetine hydrochloride on the hypothalamic-pituitary-testicular axis and spermatogenesis in male albino mice. Fifty mice were divided into five distinct groups, comprising a control group, an observation group and three separate experimental cohorts receiving oral fluoxetine at doses of 50, 100, and 150 mg kg-1 for 28 days. Testosterone, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels were measured by radioimmunoassay. Testicular tissue was analyzed for changes in seminiferous tubule sperm count and spermatogenic cell populations. The high-dose fluoxetine group showed decreased testosterone, increased LH and FSH, reduced seminiferous tubule sperm density, as well as fewer spermatogonia and primary spermatocytes versus controls. Histology revealed reduced germinal epithelium layers and increased vacuolization in the fluoxetine-treated mice. These findings demonstrate that chronic high-dose fluoxetine administration inhibits steroidogenesis and spermatogenesis in male mice, likely by increasing serotonin levels which suppress testosterone synthesis. Further research on fluoxetine's effects on male fertility are warranted given its widespread clinical use. This study affirms the need for judicious fluoxetine prescribing in patients with reproductive concerns. [ABSTRACT FROM AUTHOR]

Background: Adherence to traditional 24‐h fasting periods for serum gastrin concentration in dogs can be challenging and may delay the institution of therapies for suspected hypergastrinemia. Peer‐reviewed publications regarding serum gastrin reference intervals (RI) are lacking. Hypercalcemia is associated with hypergastrinemia in people; limited data exist in dogs. Objective: The objective of the study was to generate a RI for a 12‐h fasted serum gastrin concentration in dogs and to investigate whether correlations exist with age, weight, sex, and total calcium concentration. Methods: Fifty‐five healthy adult dogs (>1 year of age). The screening included: medical history, physical examination, CBC (15 dogs), and serum chemistry (55 dogs). Gastrin was measured via a commercial radioimmunoassay. The RI for 12‐h fasted serum gastrin concentration was calculated according to the recommendations of the American Society for Veterinary Clinical Pathology. Additionally, data were evaluated for correlation with selected variables. Results: The RI for serum gastrin following a 12‐h fasting period was 15.1–78.9 ng/L with 90% confidence intervals for the lower and upper limits of 14.0–22.9 and 68.3–83.0 ng/L, respectively. A generalized linear model did not detect significant relationships between gastrin and age (P = 0.48), sex (P = 0.30), weight (P = 0.93), or total calcium concentration (P = 0.84). Conclusions: A 12‐h fasted serum gastrin concentration RI has been established. Given the limited range of serum calcium concentrations in our healthy study population, additional investigations are needed to determine the effects of hypercalcemia on serum gastrin concentrations in dogs and for any potential clinical consequences thereof. [ABSTRACT FROM AUTHOR]

Faecal glucocorticoid metabolites (FGMs) have gained relevance in ecological studies and population monitoring, allowing non-invasive remote sampling without the need to capture and handle animals. Enzyme immunoassays (EIAs) and radioimmunoassays (RIAs) are commonly used for FGM determination. Although these methods should be validated for each species, non-validated tests are still widely used to determine FGMs in wildlife. Near-infrared reflectance spectroscopy (NIRS) is a predictive method requiring calibration against reference methods used to assess FGMs in wildlife. EIAs and RIAs have been utilized to determine FGMs in chamois (Rupicapra spp.), a medium-sized mountain ungulate.This study aims to assess the potential of NIRS to determine FGMs and to evaluate the correlation among analytical methods used to determine FGMs in chamois. Faecal samples from 125 Alpine chamois (Rupicapra rupicapra rupicapra) and 125 Pyrenean chamois (R. pyrenaica pyrenaica) were collected from the field, frozen at − 20 °C, lyophilized, grounded, and scanned using a NIRSystems 5000 monochromator over a 1108–2492 nm wavelength. After this non-destructive NIRS analysis, FGMs were extracted and analysed using four immunoassays previously used in chamois studies: a 125-I-corticosterone RIA, a cortisol EIA, and two 11-oxoetiocholanolone EIAs (72a and 72T). Only the 11-oxoetiocholanolone 72T EIA has been validated for Alpine chamois. NIRS predictions were calibrated and cross-validated for each of the four immunoassays. The correlation among the four immunoassays was assessed using Spearman's rank.The coefficient of determination for NIRS calibration (R²) values ranged from 0.37 to 0.75, and the ratio of performance to deviation values from 1.2 to 1.6. Therefore, NIRS could not predict FGM concentration in chamois faeces. This could be due to the complexity and variability of the FGM detected by the immunoassays as reference methods, and by potential interference of other compounds in the faecal matrix. The correlation among the immunoassays was low overall.As a conclusion, NIRS cannot be recommended for measuring FGMs in chamois. The low correlation among the immunoassays used for FGM determination raises concern about the reliability of previous studies using non-validated methods in chamois. Only biologically validated tests should be used to assess FGMs to avoid incorrect inferences about biological responses in physiological, conservational, and ecological studies or population monitoring. These conclusions are applicable beyond the species studied here.

Recent studies have focused on how sickness behaviours, including lethargy, are coordinated in the brain in response to peripheral infections. Decreased hypocretin (orexin) signalling is associated with lethargy and previous research suggests that hypocretin signalling is downregulated during sickness. However, there are studies that find increases or no change in hypocretin signalling during sickness. It is further unknown whether hypocretin receptor expression changes during sickness. Using lipopolysaccharide (LPS) to induce sickness in female mice, we investigated how LPS‐injection affects gene expression of hypocretin receptors and prepro‐hypocretin as well as hypocretin‐1 peptide concentrations in brain tissue. We found that hypocretin receptor 1 gene expression was downregulated during sickness in the lateral hypothalamus and ventral tegmental area, but not in the dorsal raphe nucleus or locus coeruleus. We found no changes in hypocretin receptor 2 expression. Using a gene expression calculation that accounts for primer efficiencies and multiple endogenous controls, we were unable to detect changes in prepro‐hypocretin expression. Using radioimmunoassay, we found no change in hypocretin‐1 peptide in rostral brain tissue. Our results indicate that hypocretin receptor expression can fluctuate during sickness, adding an additional level of complexity to understanding hypocretin signalling during sickness. [ABSTRACT FROM AUTHOR]

Vitamin D has received significant attention from clinical societies, researchers, and the general population in recent years. While 25-hydroxyvitamin D (25(OH)D) is the most commonly-used biomarker of vitamin D status, 1α,25-dihydroxyvitamin D (1,25(OH)2D), its bioactive form, plays a critical role in regulating calcium and phosphorus homeostasis and is also involved in the immune system and cellular differentiation. Consequently, accurate measurements of 1,25(OH)2D can aid in the differential diagnosis of calcium-related disorders such as hypocalcemia in vitamin D-dependent rickets and hypercalcemia due to inappropriate increase of serum 1,25(OH)2D in granulomatous diseases. However, due to its lipophilicity and very low circulating concentration, the measurement of 1,25(OH)2D is particularly challenging. Over the past several decades, numerous efforts have been made to develop sensitive, specific, and practical laboratory methods for measuring 1,25(OH)2D. Methods using radioreceptor assay, radioimmunoassay, enzyme immunoassay, enzyme-linked immunosorbent assay, automated chemiluminescent immunoassay, and liquid chromatography-tandem mass spectrometry have been described. Each of these methods has unique advantages and limitations, and some are no longer used. Despite the sophisticated methods in use today, substantial variations between methods still exist. A concerted effort toward standardization of 1,25(OH)2D measurement is needed to ensure accurate and reliable results across laboratories and methods. [ABSTRACT FROM AUTHOR]

Background: Although the role of oxytocin in the pathophysiology of sepsis is still unknown, rising preclinical evidence suggests that oxytocin is possibly involved. However, no direct clinical studies have measured the levels of oxytocin during sepsis. In this preliminary study, the serum oxytocin levels were evaluated throughout the duration of sepsis. Method: Twenty-two male patients over 18 years of age with a SOFA score of 2 points or more who were admitted to the ICU were included. Patients with a history of neuroendocrine, psychiatric, and neurologic disorders, cancer, an infection caused by COVID-19, shock due to reasons other than sepsis, a history of psychiatric or neurologic medication use, and those who died during the study were excluded. The main endpoint included the measurement of serum oxytocin levels using radioimmunoassay at 6, 24, and 48 h of the ICU admission. Results: Mean serum oxytocin level was higher at 6 h of ICU admission (41.27 ± 13.14 ng/L) than after 24 and 48 h of ICU admission (22.63 ± 5.75 and 20.97 ± 7.61 ng/L respectively) (P -value <.001). Conclusion: Our study, while reporting increased serum oxytocin levels in the initial phase of sepsis and decline afterward, supports the possible contribution of oxytocin in the pathophysiology of sepsis. Given that oxytocin seems to modulate the innate immune system, future investigations are necessary to assess the potential role of oxytocin in the pathophysiology of sepsis. [ABSTRACT FROM AUTHOR]