The result revealed that among all incident patients with CKD, 28.7% and 30.3% of the users and nonusers demonstrated CKD progression, respectively. The crude odds ratio (OR) of CKD progression was 0.93 [95% confidence interval (CI) 0.78–1.10]. After adjustment for baseline characteristics, the adjusted OR was 0.80 (95% CI 0.63–1.01). The sensitivity analysis results showed consistent OR for CKD progression, stratification by age, sex, Charlson score, and statins use within 1 year before index date. The effect of statins was significant in patients with CKD stage 3B-5 , but not statistically significant in those with CKD stage 1–3A . The effect of statins was significant in patients with proteinuria ≥1000 mg/day , but not statistically significant in those with proteinuria <1000 mg/day .Dyslipidemia is associated with glomerular injury. However, the effect of statins on chronic kidney disease (CKD) progression remains controversial. We aimed to investigate the efficacy of statins for renal protection in patients with CKD. The retrospective cohort study comprised 3441 patients diagnosed with CKD in multiple medical centers. We divided the patients into two cohorts based on statin prescription, and compared proportions and risks of CKD progression events between the two groups. CKD progression event was defined as an average annual decline of eGFR >5 mL/min/1.73 m According to systemic review data and a meta-analysis of observational studies until September 2014, the global prevalence of CKD was approximately 13.4%, and the prevalence of stage 3–5 CKD was 10.6%1. An estimated 5.4 million people will be receiving dialysis due to end-stage renal disease (ESRD) in 2030, and globally, the number of patients receiving dialysis is increasing the fastest in Asia2. According to the 2017 United States Renal Data System (USRDS) report, the highest prevalence and incidence of ESRD among all countries investigated was noted in Taiwan3. Among patients with ESRD in Taiwan, median estimated glomerular filtration rate (eGFR) at initial dialysis was approximately 4.7 mL/min/1.73 m2 4.Chronic kidney disease (CKD) is a global health concern5. CKD is associated with increased CVD risk, severity of which increases as kidney function deteriorates6. Statins are the mainstay of primary and secondary prevention of CVD in the general population7. The Kidney Disease Improving Global Outcomes (KDIGO) lipid management guidelines suggest statin initiation for primary prevention in all patients with CKD above the age of 50 years, and all adult CKD patients with diabetes who are not receiving dialysis8. In Taiwan, in adults with eGFR <60 mL/min/1.73 m2 and without chronic dialysis, statins therapy is recommended only if low-density lipoprotein cholesterol levels are ≥100 mg/dL9.Cardiovascular disease (CVD), which may occur even at the earliest stages of CKD without manifestations of vascular disease, is the leading cause of morbidity and mortality among patients with CKD10. On the basis of the experimental evidence, dyslipidemia is associated with tubulointerstitial and glomerular injuries, which may result in glomerulosclerosis12. Statins inhibit HMG-CoA reductase activity and thus play a beneficial role in dyslipidemia treatment13. Many studies have investigated the renoprotective effects of statins. The ALERT trial and Scandinavian Simvastatin Survival Study have demonstrated that statins slowed CKD progression15; however, other studies such as the SHARP study, ALLHAT study, and ASUCA trial have reported that statins exhibit little to no effects on CKD progression18. Recent systematic reviews and meta-analyses have indicated that statins do not reduce risk of kidney failure events in adults not receiving dialysis, where kidney failure events are defined as 25–50% decrease in eGFR, doubling of serum creatinine level, or advancement to ESRD stage during follow-up period; however, according to the same reports, statins may reduce proteinuria and eGFR decline19. Therefore, we evaluated the renoprotection efficacy of statins in patients with CKD.Patients with CKD are more likely to exhibit elevated triglyceride and low high-density lipoprotein cholesterol levels, which were the risk factors for CVD20, including National Health Insurance claims data more than 99% of 23 million Taiwan residents. The Ministry of Health and Welfare annually releases the Longitudinal Health Insurance Database for research purposes, which contains the insured population’s registration files and medical claims data, including data on basic demographic characteristics, inpatient and ambulatory care, diagnostic codes, medical expenditure, operations, prescriptions, examinations, and procedures. From January 1, 2008, to Dec 31, 2013, a multicenter project was conducted to survey risk factors for CKD in the Taiwan population. This project was supported by the Bureau of Health Promotion, Ministry of Health and Welfare, Taiwan. The present CKD retrospective observational cohort study is based on the data of 7956 patients from the multicenter project. We used national identification numbers to link laboratory data from study hospitals to the NHID. This research project was approved by the Ethics Committee of Taipei Medical University-Shuang Ho Hospital (TMU-JIRB 20124036), Tri-Service General Hospital (TSGHIRB 100-05-197), Cardinal Tien Hospital (TMU-JIRB 201204035), Changhua Christian Hospital (CCHIRB 20405), Kaohsiung Medical University Chung-Ho Memorial Hospital (KMUHIRB 20120019), Kaohsiung Chang Gung Memorial Hospital (101-1096B), National Cheng Kung University Hospital (A-ER-101-117) and China Medical University Hospital (DMR101-IRB2-273(CR-1)). After a complete explanation of the study, written informed consent was obtained from all participants. All clinical and biological samples were collected after patient consent. All the study methods were in accordance with the guidelines approved by the joint institutional review board and aforementioned governmental regulations.The National Health Insurance Database (NHID) is a research database developed by the Ministry of Health and Welfare2 for more than 3 months, with this eGFR being used to determine CKD stage21. The exclusion criteria were as follows: age <20 years, follow-up duration <1 year, absence of major variables , and cancer patients. The multiple imputation method was used to account for missing data on smoking, alcohol use, and betel nut use22. We input missing data on the basis of patients’ sex and CKD stage. After patients with loss to follow-up, with missing data, or with cancer were removed from the database, the final study cohort comprised 3441 CKD patients.The initial study cohort comprised 7956 patients diagnosed with CKD between November 1, 2008, and June 30, 2013, in multiple medical centers  and nonusers . Patients were individually tracked from the index date to December, 2015, to determine CKD progression. The primary outcome was CKD progression, defined as an average annual decline of eGFR >5 mL/min/1.73 mt test. We calculated proportions of CKD progression events; then, a logistic regression model was used to calculate crude odds ratios (ORs) between statin users and nonusers. After additional adjustments for the potential confounding factors including age, sex, comorbidities, smoking, alcohol use, betel nut use, body mass index (BMI), baseline eGFR, baseline urine protein-to-creatinine ratio (UPCR), and statin use within 1 year before the index date, adjusted ORs (aORs) were calculated. We also performed sensitivity analysis after stratification according to age (<65 and ≥65 years), sex, Charlson comorbidity index score (≤3 and >3), baseline CKD stage (1–3 A and 3B–5), proteinuria (<1000 mg/day and ≥1000 mg/day) and stain use within 1 year before the index time. All analyses were performed the SAS system for Windows . P < 0.05 was considered significant.We compared the demographic data between statin users and nonusers by using Pearson’s chi-squared test and 2, respectively. Furthermore, the users had a significantly higher baseline cholesterol, fasting glucose and HbA1c levels than did the nonusers . Baseline levels of serum albumin, electrolytes, uric acid, hemoglobin, hematocrit, and proteinuria did not differ significantly between the groups. The following levels of cholesterol of the users and nonusers was 177.6 ± 43.15 and 177.14 ± 41.06 mg/dl, respectively (p = 0.8049).From November 1, 2008, to June 30, 2013, across multiple medical centers, a total of 7956 patients with CKD, aged 20–85 years, fulfilling the inclusion criteria were identified. Among them, those who had ever received dialysis or kidney transplant were excluded. After the additional exclusion of patients with a follow-up of <1 year (n = 2343), with incomplete or missing data (n = 1809), and with cancer (n = 363), 3441 patients with CKD were enrolled . In the subgroup of patients with CKD stage 3B-5 , and those with proteinuria ≥1000 mg/day , statins exhibited renoprotective effects with statistically significant.24. Many of these effects are potentially arise from small G-protein disruption24. Because of its resultant biological and genetic stress, the activation of G-protein signaling is pivotal in renal pathologies25.Because of their main mechanism of inhibiting HMG-CoA reductase activity, statins are widely used to treat hyperlipidemia. In addition, statins have other therapeutic benefits, such as anti-inflammatory and antioxidant properties, apoptosis induction, vascular smooth muscle proliferation inhibition, platelet activation and aggregation reduction, and increase in atherosclerotic plaque stability19 examined the data of 57 randomized control trials (RCTs), with 143,888 participants and 8,498 kidney failure events and suggested that statin therapy results in mild decreases in proteinuria levels and an eGFR decline of 0.41 mL/min/1.73 m2 per year. Su et al.19 defined kidney failure events as 25–50% decrease in eGFR, doubling of serum creatinine level, or advancement to ESRD stage during the follow-up period. Statin use did not prevent or mitigate these kidney failure events, consistent with the results of a systemic review conducted by Zhong et al.26; in total, 23 RCTs with 39,419 participants were analyzed, and the authors concluded that statins significantly reduced microalbuminuria, proteinuria, and clinical deaths, but did not slow the clinical progression of non-end-stage CKD significantly.The effects of statins on CKD progression remain subject to debate. A 2016 meta-analysis28 suggest that dialysis should be initiated when the eGFR is approximately 5–9 mL/min/1.73 m2. The 2014 Canadian Society of Nephrology clinical practice guidelines suggest that chronic dialysis should be initiated when eGFR decreases to 6 mL/min/1.73 m2, even if no clinical indicators are exhibited. Although data have shown high prevalence and incidence of ESRD in the Taiwan population3, patients in Taiwan begin dialysis with poor clinical conditions  with low residual renal function (eGFR 4.7 mL/min/1.73 m2)4. Hwang et al.4 attributed the delay in dialysis to patient awareness and attitudes toward dialysis treatment; in Taiwan, to initiate dialysis, pre-ESRD patients tend to wait for the presentation of uremic symptoms and other clinical signs, rather than following the eGFR criteria. The effects of dialysis initiation timing remains controversial. Although dialysis initiation timing is relatively late in Taiwan, mortality due to CKD is low4.The 2012 KDIGO guidelines20. In contrast to definitions used in previous studies, we identified CKD progression events as either an annual average eGFR decline >5 mL/min/1.73 m2 or advancement to the dialysis stage. Because observational studies are prone to bias and being confounded, sensitivity analyses were performed, the results of which suggested that statin use benefited in those with CKD stage 3B-5 and proteinuria ≥1000 mg/day. A possible explanation for this finding is that patients may require a higher dosage of statin therapy. Shepherd et al.29 reported that although some eGFR improvement occurred after low-dosage (10 mg/day) atorvastatin treatment, high-dosage atorvastatin led to a more significant eGFR improvement. However, a 2014 meta-analysis30 involving 6 RCTs compared high-intensity statin therapy (atorvastatin 80 mg or rosuvastatin 20 or 40 mg) with moderate-to-mild statin treatment or placebo and noted that the effect of the high-intensity and moderate-to-mild therapies on eGFR improvement was not substantially different; regarding the safety of statin use, the prevalence of adverse events was low, and the pooled results showed no significant differences in adverse event prevalence among patients receiving high-intensity or nonintensive statin therapy or placebo30. Therefore, the effect of high-intensity statin on renal function is difficult to determine conclusively, and more evidence from high-quality studies is required.Our study enrolled patients from multiple medical centers in Taiwan, and national identification numbers were used to link patients with corresponding data in the NHID2) was broader than that of similar studies. It is worth noting that the beneficial effects of statin can be seen in patients with more than one gram of urine protein, because urinary protein itself is an indicator that can lead to deterioration of kidney function.This study has some limitations. First, this was a retrospective cohort study with a short follow-up period (1–6 years). Second, our criterion for defining CKD progression (annual eGFR decline >5 mL/min/1.73 mThis national cohort study on CKD found that statins effectively delay CKD progression in CKD stage 3B-5 patients, particularly among those with proteinuria ≥1000 mg/day, for whom the benefits of treatment are clear. Thus, statin therapy may have a net clinical benefit for preventing CKD progression, particularly considering the high burden of dialysis. The protective effect of CKD on kidneys may differ according to statin dosage, and additional evidence is required to confirm these benefits. Our results suggest that statin usage in CKD with stage 3B-5 and those with proteinuria ≥1000 mg/day is critical, particularly for targeting CKD progression outcomes.


The usefulness of particle therapy for skull base chordoma has not been established. The aim of this retrospective study was to analyse the treatment outcomes of proton therapy (PT) and carbon ion therapy (CIT) in patients with skull base chordoma at a single institution.All patients who underwent PT or CIT with curative intent between 2003 and 2014 at Hyogo Ion Beam Medical Center were included in this study. Twenty-four patients were enrolled. Eleven (46%) received PT and 13 (54%) received CIT. Overall survival (OS), progression-free survival (PFS) and local control (LC) were calculated using the Kaplan–Meier method. Late toxicities were evaluated according to the Common Terminology Criteria for Adverse Events version 4.0.P = 0.048), PFS (P = 0.028) and OS (P = 0.012) were significantly improved in patients who had undergone surgery before particle therapy. No significant differences were observed in the LC rate and the incidence of grade 2 or higher late toxicities between patients who received PT and CIT.The median follow-up was 71.5 months . The five-year LC, PFS and OS rates were 85, 81, and 86%, respectively. The LC (Both PT and CIT appear to be effective and safe treatments and show potential to become the standard treatments for skull base chordoma. To increase the local control, surgery before particle therapy is preferable. Chordoma is a rare tumour of the bone that arises from embryonic remnants of the notochord . ApproxiFor several decades, particle therapy, such as proton therapy (PT) and carbon ion therapy (CIT), have been used for skull base chordomas –18. The The aim of this study was to evaluate treatment outcomes in patients with skull base chordomas who received PT or CIT at our centre and to investigate the clinical role of particle therapy for the disease to add further evidence.We conducted an Institutional Review Board-approved, retrospective analysis of patients with skull base chordoma who received definitive PT or CIT between April 2003 and May 2014 at Hyogo Ion Beam Medical Center. The inclusion criteria for the present study were as follows: 1) histologically confirmed skull base chordoma, 2) no previous radiotherapy, and 3) a duration of follow-up ≥24 months for survivors. Twenty-four patients were enrolled. All eligible patients provided written informed consent before treatment.Patients were immobilised in the supine position with an adequate head angle using a custom-made thermoplastic cast. The target volumes and organs at risk were delineated on computed tomography (CT) and magnetic resonance imaging (MRI) fusion images. The clinical target volume (CTV) that included regions of suspected microscopic spread was generated around the gross tumour volume (GTV) by expanding three-dimensional margins of 5 mm anatomically and then using manual corrections based on anatomic structures. The planning target volume (PTV) was defined as the CTV plus a setup margin of 5 mm for PT and 3 mm for CIT. Radiation treatments were planned on a CT-based three-dimensional treatment planning system .Due to availability constrictions at the time, from April 2003 to April 2005, either PT or CIT was available. After April 2005, both beams were available and radiotherapy treatments with PT and CIT were simultaneously planned for each patient. After comparing dose distributions and the dose volume histogram (DVH), a team of radiation oncologists selected the more appropriate treatment for each patient. When comparing the radiation treatment plans between PT and CIT, the same total dose and fractionations were established. Dose constraints of OARs were considered the most important factor in selecting which treatment plan should be admitted. If doses to the OARs were similar, then the plan that had the favourable minimum dose to the CTV was selected. Both proton and carbon ion beams were administered using a passive delivery system .Radiobiological experiments at Hyogo Ion Beam Medical Center showed that the relative biological effectiveness (RBE) values for PT and CIT were 1.1 and 2 to 3.7, respectively (depending on the depth in the spread-out Bragg peak) . In partThe follow-up period was calculated from the initial date of particle therapy. Patients were evaluated at 3-month intervals for 1 to 3 years after the start of therapy and at 6-month intervals thereafter. Regular follow-up studies included physical examinations, endoscopy, diagnostic imaging , and blood tests. Acute reactions and late toxicities were evaluated according to the Common Terminology Criteria for Adverse Events version 4.0 .Continuous variables are presented as medians with ranges and categorical variables are shown as frequencies with percentages. The LC, progression-free survival (PFS) and overall survival (OS) rates were assessed for all patients using the Kaplan–Meier method. Local recurrence was defined as confirmed radiographic or clinical disease progression/recurrence on CT or MRI. Disease progression was defined as clinical or radiographic evidence of local, regional, or distant recurrence/progression. Survival was identified as the confirmed date of death or last follow-up. All statistical analyses were performed using SPSS Statistics 22 software .P-values less than 0.05 were considered statistically significant.The following covariates were analysed for the relationship to LC, PFS, and OS using the log-rank test: age, gender, tumour status , proximity of the tumour to the brainstem, surgical intervention, ion type, GTV volume, and minimum dose of GTV (GTV Dmin). In this study, tumours in which the PTV overlapped the brainstem were defined as proximal to the brainstem. The GTV Dmin was standardised to 2 Gy per fraction using the linear-quadratic model with α/β = 2 , 23. LatP = 0.902). In the PT group, 10 patients (91%) received 65.0 Gy (RBE) in 26 fractions, and patients in the CIT group received various doses and fractions. A representative dose distribution for a patient treated with 70.4 Gy (RBE) in 32 fractions is shown in Fig. Patient and treatment characteristics are shown in Table Five patients 21%) experienced local recurrence with a median duration of 51 months . All tumours recurred within their original GTV. Before particle therapy, all tumours were adjacent to the brainstem and the GTV Dmin values of these 5 patients were relatively low . After local recurrence, one patient received salvage surgery, and the disease was controlled at the final follow-up. One patient received re-irradiation using XRT after local recurrence; this tumour was not controlled. Three patients received no treatment after their local recurrence. The 5- and 8- year LC rates were 85%  and 71% (95% CI: 33–90%), respectively  experienced distant metastasis with a median duration of 69 months : one with metastasis in the nasal cavity mucosa, one with bone metastasis and one with meningeal dissemination, respectively. The 5- and 8-year PFS rates were 81% (95% CI: 57–93%) and 65% 95% CI: 27–87%), respectively  had died. Two patients died of the recurrent primary tumour and meningeal dissemination. Three patients died of intercurrent disease. The median time to death was 52 months . The 5- and 8-year OS rates were 86% (95% CI: 62–95%) and 76% (95% CI: 46–91%), respectively , PFS (P = 0.028) and OS (P = 0.012) (Table P = 0.752), PFS (P = 0.187) and OS (P = 0.060) between the PT and CIT groups . These symptoms improved with temporal oral corticosteroids. One patient whose tumour had infiltrated the optic canal experienced grade 3 unilateral blindness that had been predicted before particle therapy. One patient experienced grade 4 bleeding from an ulcer in the nasopharynx after CIT and was treated with coil embolisation.P = 0.337).Six patients in the PT group experienced ≥ Grade 2  and 6 patients in the CIT group experienced ≥ Grade 2  late toxicities. No statistically significant difference was observed with respect to ≥ Grade 2 late toxicities between the PT and CIT groups (To our knowledge, this is the first report describing the results of both PT and CIT treatments of skull base chordoma at a single institution. Both PT and CIT treatment showed favourable LC rates and tolerable late toxicities. Tumour control and survival were significantly improved in patients who had undergone surgery before particle therapy.In this study, as the LC rate was favourable both in the PT and CIT groups with low rates of severe late toxicities, it is considered that both PT and CIT were effective and safe treatments. Currently, surgery followed by high-dose XRT is considered a standard treatment . HoweverIn this study, prognostic factor analysis showed better tumour control and survival in patients who underwent surgery before particle therapy. Other studies have also found that surgery prior to radiation therapy resulted in better outcomes , 29. HugWe found no statistically significant differences in the rates of local control and late toxicities between the PT and CIT groups, although the number of patients was small and their follow-up periods differed. As shown in Fig. Although the 5-year LC rate of 82% observed in this study was a favourable outcome, there is still room for improvement. It is considered indispensable to deliver a higher dose for local control of the skull base chordoma. Herman et al. summarized that the α/β value of the skull base chordoma was 2.0 and that local control rates were improved in proportion to the total dose . Schulz-This study has several limitations. First, it was a retrospective analysis at a single institution. Second, chordoma is a disease that takes a very long time before recurrence, and the follow-up period in this study, although slightly longer compared with other studies, was not sufficient. Third, treatment heterogeneity was noted, including total dose and fractionation. Forth, due to the small number of patients, the impact of the statistical analyses, including the comparison between the PT and CIT groups, was relatively low. However, other published studies were also retrospective, and performing a prospective study is difficult due to the rarity of this disease. Therefore, we will increase the number of patients and continue to monitor these patients to report on follow-up data.The results of proton therapy and carbon ion therapy for skull base chordoma were both favourable regarding local control and late toxicities. Both showed the potential to become a standard therapy as opposed to XRT. To increase local control, surgery before particle therapy for tumour volume reduction and separation from OARs can be considered a viable alternative.


In Cote D’Ivoire, there has been limited coverage of evidence-based sexual health services specifically supporting men who have sex with men (MSM). To date, there has been limited study of the determinants of engagement in these services including multiple intersecting stigmas and depression.1301 MSM aged 18 years and older, were recruited using respondent-driven sampling in Abidjan, Yamoussoukro, Gagnoa and Bouake, Cote d’Ivoire from January 2015 to October 2015. Inclusion criteria included anal sex with another man in the past 12 months were to complete a structured questionnaire including the Patient Health Questionnaire (PHQ)-9 to screen for depression. Chi-Square tests were used to test difference in healthcare utilization across variables, and multiple logistic regression was used to test the association between depression and health care utilization represented by HIV and sexually transmittable infection testing and treatment.Depression , being aged 25–29 years ,unemployed , being a student , being identified as male , and identifying as homosexual  were significantly associated with utilization of sexual health care services in the final multivariable model. Healthcare enacted stigma  was associated with utilizing sexual health care services, but perceived healthcare stigma, social stigma and family stigma were not.Given higher levels of depressive symptomatology among those engaging in sexual health care services, this engagement represents an opportunity for service integration which may have synergistic benefits for both sexual and mental health. Moreover, MSM in Cote D’Ivoire who had engaged in sexual health services were more likely to report having experienced health-care enacted stigma. Taken together, these results reinforce the need for stigma mitigation interventions to support sustained engagement in HIV prevention, treatment and care services as a means of reducing health disparities among MSM in Cote d’Ivoire. A leading cause of disability and global burden of disease is depression affecting more than 350 million worldwide , yet in Previous studies have consistently shown that MSM may be at higher risk for depression secondary to minority stress conceptualized as the social environmental stressors associated with being a sexual minority , 12. MinDepression can result in internalized stigma and limited self-efficacy, which may lead to impaired physical, social, and emotional health –16. In pAcross Sub-Saharan Africa, consistent data have demonstrated concentrated HIV epidemics among key populations including MSM, even in the context of more generalized epidemics , 26, 27.Same sex practices are not criminalized in Cote d’Ivoire, and the Ministry of Health defined MSM as a priority population in the National Strategic Plan for the Fight against HIV/AIDS 2011–2015 . For eviN = 1301) conducted in 5 cities in Cote d’Ivoire, including Abidjan (n = 351), Bouake (n = 351), Gagnoa (n = 150), Agboville (n = 200) and Yamoussoukro (n = 250). Individuals were eligible to participate in the study if they were aged ≥18 years and reported having anal intercourse (either receptive or insertive) in the past 12 months. Participants were required to have lived in the country for the past 3 months, and be able to give informed consent prior to participating in the study. MSM were excluded if they demonstrated mental incapacity, were under the influence of substances at the time of participation, or had any other illness preventing comprehension of the study procedures and informed consent.Data were obtained from a cross-sectional study (Participants were recruited using respondent-driven sampling (RDS) , 36, whiSeveral types of sexual behavior-related stigma were measured. MSM’s experience of each broad area of stigma  was assessed by using responses “yes” or “no” to each individual item. Each area was assessed for association with the outcome variable by using the sum of the total number of “yes” and “no” responses , 30. FamFinally, forced sex was measured by asking the participant if they have ever been coerced into having anal, vaginal or oral penetration against their will. Responses were dichotomized into “yes”/“no”.Depression and depressive symptoms were measured using the Patient Health Questionnaire (PHQ-9) , which hAlcohol use was measured using the Alcohol Use Disorders Identification Test (AUDIT), which has been shown to be valid across low and middle-income countries , 40, 47.The primary outcome was utilization of sexual health care services, measured with two indicator variables: 1) tested for sexually transmitted infections or received treatment for sexually transmitted diseases/infection (STI) in the past 12 months (yes/no), and 2) tested for HIV in the past 12 months (yes/no). These indicators of health care utilization were chosen because of the important role they play in reducing HIV infection , 49. TheData from the 5 study sites were combined and analyzed using Stata 14.0 . Chi-square tests were used to compare difference in health care use among various socio-economic indicators and other variables. Logistic regression models were used to examine the association between the outcome measure  and covariates such as depression, alcohol intake, employment, educational status and stigma, in both bivariate and multivariable models. For the HIV test variable, participants who had an HIV test more than one year prior  were not included in the final model, because one of the inclusion criteria was having anal sex in past 12 months.There were two final models: Model 1 included depression, age, education, employment status, residence , gender, sexual orientation, forceful engagement in sex, and condom use. Model 2 included variables in model 1 and stigma variables. Two separate models were used because stigma has been shown to be on the causal pathway to depression and thus depression may be acting as a mediator between stigma and utilization of sexual health care services . These vp = 0.64 and Model 2 (Hosmer-Lemeshow chi2(8) =6.19, p = 0.63)) [RDS sampling probabilities were adjusted for in the final models . The fin= 0.63)) . FinallyThe age of participants ranged from 18 to 54 years. The median age was 23 years with an interquartile range (IQR) of 21–26 years , and 19.9% (259/1301) of participants had never tested for HIV. Abidjan had the highest percentage of participants who self-reported as HIV positive  and Gagnoa the least .The prevalence of health care utilization in the last 12 months was 60.5% (787/1301). Of the participants who sought sexual health care services, 64.0% (830/1297) went to non-governmental facilities, while 13.8% (179/1297) went to either private or public facilities, but 22.2% (288/1297) had not sought sexual health care services prior. The prevalence of depression was 48.0% (624/1301), with 29.0% (377/1301) being mildly depressed, 14.5% (188/1301) moderately depressed, and 19.0% (247/1301) severely depressed.Most participants identified their gender to be male 73.9% (962/1301), 22.2% (289/1301) identifying as a woman using the two-step gender assessment and 3.8% (50/1301) identifying as a transgender woman. In terms of sexual orientation, more than half identified as bisexual 59.6% (775/1301), 38.4% (499/1301) identifying as gay or homosexual and 2.1% (27/1301) as heterosexual.Many of the participants were living with family or a spouse, 79.9% (1040/1301), and fewer were living alone , or with friends or other MSM . Participants experienced various forms of family-related stigma, such as feeling excluded from family activities , hearing discriminatory remarks from family members , and feeling rejected by friends . Stigma in health care settings was also prevalent, with 22.6% reporting being afraid to seek health care services, 15.8% (205/1301) avoiding health care services, 3.2% (42/1301) feeling not treated well in a health care center, and 9.2% (120/1301) hearing a health care worker gossip about them. For social stigma, 4.7% (61/1301) felt like police refused to protect them, 13.5% (175/1301) felt scared to walk in public, 37.0% (481/1301) reported verbal harassment, 18.8% (245/1301) reported blackmail, and 43.4% (564/1301) reported being physically hurt because they have sex with men. The prevalence of being forced to engage in sexual activity was 18.1% (236/1301).The percentage of MSM reporting alcohol consumption was 81.2% (1056/1301). More than one-third of drinkers  were classified as risky/hazardous drinkers. Of the participants who engaged in anal sex in last 30 days, 18.9% (193/1020) did not use condoms, and 81.1% (827/1020) used condoms inconsistently.Utilization of sexual health care services in the past 12 months was significantly associated with depression ; being aged 20–24 years , aged 25–29 years , and aged ≥30 years  as compared with aged < 20 years; identifying as male vs transgender , and identifying as heterosexual  , being aged 25–29 years , being unemployed , being a student , identifying as male , and identifying as homosexual , were associated with utilization of sexual healthcare services (Table This study estimated that nearly one in two MSM in Cote d’Ivoire experience depression, which is consistent with other studies conducted in countries across Sub Saharan Africa , 58. TheSimilar to many Sub-Saharan countries, the services specifically developed for MSM remain largely externally financed including stand-alone services. The challenge with this model is that there is limited coverage of education about specific sexual health services for MSM if one is not connected to these communities highlighting the importance of leveraging new technologies in promoting comprehensive sexual education. In Tanzania, a study by Dangat et al. found that the most common source of information on sexual education reported was through radio . OverallDepression and substance use including alcohol use are also common comorbidities of mental health among all adults . FurtherThis study has a number of limitations. Due to the cross-sectional nature of the data collected, temporality cannot be assessed, and therefore a causal relationship between depression and sexual healthcare service utilization cannot be established. However, depression was measured over two weeks and utilization of sexual healthcare services in a one-year period, indicating it may be likely that healthcare utilization preceded depression in these analyses. The intersectionality of mental health in general, stigma and HIV-related stigma is complex and cross-sectional data may be limited in establishing these relationships. Future studies should consider establishing this temporality through prospective data collection. Also, topics on sexuality, STIs and stigma are culturally sensitive and hence may be underreported secondary to social desirability bias. Depression has also been found to be underreported in many African settings because of social stigmas related to depression and mental health more broadly .This socDepression is prevalent among MSM in Cote d’Ivoire and associated with greater health care utilization presenting an opportunity to address the mental health needs of MSM. However, MSM with increased healthcare utilization also appear more likely to report experiences of stigma in health care settings secondary to their sexual orientation. Taken together, these data suggest the need to address stigma reduction or mitigation interventions, and better integrated mental and sexual health services are needed and may ultimately be key to reducing health disparities among MSM in Cote d’Ivoire. Integrating education about the specific needs of MSM into health care training curricula may also represent a strategy to normalize diverse sexualities and genders in Cote d’Ivoire. Ultimately, the sexual health and mental health needs are clear and so too is the intense and intersecting stigmas affecting engagement in services for MSM in Cote D’Ivoire. And the way forward includes implementing at scale evidence-based and human rights affirming strategies that will improve these services and ultimately decrease health disparities.


Nicotiana tabacum L. cv. Solaris was proposed as oil seed crop. Residues of oil extraction were successfully used in swine nutrition. The aim of this study was to explore the full potential of this innovative tobacco cultivar as multitasking feedstock non interfering with the food chain. In the triennium 2016–2018, samples from whole plant, inflorescence and stem-leaf biomass were collected in three experimental sites and analysed for chemical constituents, including fibre fractions, sugars and starch, macro-minerals and total alkaloids. The KOH soluble protein content and the amino-acid profile were also investigated as well as the biochemical methane potential. All the analyses were performed according to official methods and results were compared with values reported in literature for conventional lignocellulosic crops and agro-industry residues. The average protein content, ranging from 16.01 to 18.98 g 100 g−1 dry matter respectively for stem-leaf and whole plant samples, and their amino-acid profile are consistent with values reported for standard grass plant. These findings suggest the potential use of cv. Solaris in industrial food formulations. Moreover, considering the average content of both fibre available for fermentations  and oils and fats (7.92 g 100 g−1 dry matter), the whole plant biomass of cv. Solaris showed good attitude to anaerobic fermentation, confirmed by the biochemical methane potential of whole plant (168 Nm3 t−1 organic matter). Similarly, results allow to define the cv. Solaris biomass as a good quality forage apt to ensiling for its chemical composition. The low total alkaloids content of cv. Solaris, in average 0.3 g 100 g−1 dry matter, was previously reported not to affect growth performances and welfare traits of dairy heifers. These are the first results showing the multitasking potential use of cv. Solaris biomass, that could allow the recovery of tobacco cultivation know-how especially in marginal areas.In order to limit the smoking tobacco sector crisis, a new non-GMO  Nicotiana tabacum L. cultivation has decreased considerably in recent years because restriction of subsidies1 and smoking tobacco production is reported to account for about 219,243 tons2.The European 2. However, in Italy the crisis of the sector is particularly serious, with a reduction of both the production  and the cultivated areas, reduced from 32,000 hectares in 2007 to 14,548 hectares in 20172. Tobacco cultivation in Italy is concentrated in Campania, Umbria, Veneto and Toscana regions, where a strong cultivation know-how is available in terms of both technique and agricultural machinery.Italy is the largest tobacco producer in the European Union, with an amount of 21% of the total European tobacco productionNicotiana tabacum L. was developed for maximizing the production of seeds with high content of oil used as biofuel or biomass source1. This cultivar, named Solaris, is a non-GMO developed and patented as “energy tobacco” (PCT/IB/2007/053412)4, registered in 119 countries and granted in over 75 countries, including USA, all Africa, EuroAsia, Italy, Russia and Australia.In order to limit the crisis affecting the tobacco sector in Italy, thanks to 15 years of research, a new cultivar of The genetic improvement was addressed to a sustainable industrial “no food” culture, having no competition with plants produced as food for humans or animals.Nicotiana tabacum L. cv. Solaris compared with smoking tobacco varieties contains low levels of nicotine and maximizes the production of flowers/seeds, reducing leaf growth , and the second seed harvest carried out in the second half of October (140–150 days after transplanting) when, on the other hand, the environmental conditions can be critical.The strategy proposed by Grisan and colleagues1, the Solaris tobacco biomass has been here investigated following an alternative strategy, i.e. 50–60 days after the first collection of the plant apexes (seeds), the total biomass harvesting may occur within the same vegetative season. The large amounts of biomass harvested after the first seed collection could be used in other sectors, such as for biomethane production6, animal nutrition or food industry, consistently with a circular supply chain (circular economy) approach.Consistently with its strong vegetative capacity and its short cultivation cycle (80–90 days the first harvest)Nicotiana tabacum cv. Solaris are at the moment unknown, therefore, aiming to contribute to the advances of knowledge on this innovative culture and its potentialities, the goal of the present trial was to study the chemical characteristics, including amino acid profile and mineral content, and the potential biomethane production of cv. Solaris biomass.The aspects related to alternative uses of biomass from i.e. animal feeding and food supplement, and biomethane production.Results were then compared with literature data on conventional plant materials used as source of biomass for nutrition, Nicotiana tabacum L. cv. Solaris biomass has been carried out during the three-year period 2016–2018 in three sites located in Vicenza (45°17′25.4″N 11°30′09″E), Chieti (42°10′0″N 14°20′0″E) and Perugia (42°59′0″N 12°25′0″E) provinces, adopting conventional farming techniques of the areas1.The study on Solaris tobacco samples were made in years 2016, 2017, 2018 for Vicenza province, in 2017 for Chieti province and in 2018 for Perugia province.The scalar collections of fifteen cv. 7, HORT@ Agrometeorological Network8 and Regional Hydrographic Service9, respectively for Vicenza, Chieti and Perugia province. As depicted in Fig. Data on thermal-pluvio-metric conditions were collected by weather stations located in Vicenza, Chieti and Perugia provinces, near the study areas. Data were provided by ARPA Veneto2 for a total surface of 600 m2. Tobacco plants were transplanted after tillage carried out by ploughing (moldboard plow - depth of 50 cm) and followed by secondary harrowing in order to reduce the clod size. Nitrogen, phosphorus and potassium were distributed based on the soil analysis, in total were distributed 120 kg of N, 60 kg of P2O5, 70 kg of K2O by fertigation system. Insecticide products (z-cypermethrin) were buried on the ground to prevent attacks of terricolous cutworms in the transplantation stage. During the season, fungicide treatments  and insecticide (indoxacarb) treatments were carried out to protect the maturing capsules from fungal disease and bollworm.The field experiment was carried out in each year and site in randomized blocks with three replications. Each plot was 50 mi.e. 3.6 and 4.8 plants per m2. The plants were arranged in rows 90 cm apart and the distance between the plants on the row was 31 cm and 23 cm, respectively for the 3.6 and 4.8 plants per m2 density.In order to compare the effect of plant density, the transplant was performed mechanically on May, with two transplanting densities, The herbicide treatment was performed the day after the transplant using a clomazone based product.3 ha−1 per watering) was adopted and the total amount ranged from about 1000 m3 ha−1 (6 waterings) to about 1450 m3 ha−1 (8 waterings), according to weather conditions and practices in use by local farmers.Drip irrigation  from the end of July to the first decade of August (80–90 days after transplanting). The seeds collected were weighted and seed yield recorded.Solaris harvested by a common shredding machine for biomass collection, was weighted. Total biomass yield was recorded and sampled.The total biomass was then harvested between the months of September and October (130–150 days after transplanting), depending on altitude, rainfall and temperature. The green biomass of cv. Biomass samples were collected as fractions of whole plant (1), inflorescence , stem-leaf (3) for years 2016 and 2017, while in 2018 only the whole plant was sampled. All samples underwent natural drying and chopping (4 cm).10, plus fibrous components, including Neutral Detergent Fibre (NDF), Acid Detergent Fibre (ADF) and Acid Detergent Lignin (ADL)12, for all samples. The content of sugars 12, starch10 and macro-minerals 12 were investigated on samples from 2016. On the same samples, the nitrogenous fraction was further investigated by the analyses of KOH-soluble proteins and amino acids  after hydrolysis10. Besides that, for a more complete scenario on the analytical characteristics of the biomass, consistent with the hypothesis of the destination of cv. Solaris in anaerobic digestors or ruminant nutrition, data on fibre fractions were subjected to further evaluations, i.e. hemicellulose = NDF-ADF, cellulose = ADF-ADL, available fibre = NDF-ADL13. Moreover, the content of Non-Fibrous Carbohydrates has been calculated as the difference to 100 of the sum of the crude protein, crude oil and fat, crude ash and NDF contents, expressed on a dry matter basis13. Consistently with the hypothesis of cv. Solaris whole plant used as forage, the determination of total alkaloids was also carried out (UNI EN ISO 2881/1992) and results have been expressed as nicotine. As reported in literature, nicotine is in fact the principal tobacco alkaloid accounting for about 95% of the total alkaloid content14.Chemical analyses of all samples were carried out by an accredited laboratory  according to official methods. The analyses, performed in three replicates, were focussed on the typical constituents Solaris attitude to anaerobic fermentation, the biochemical methane potential (BMP) has been tested on whole plant samples from 2017 and 2018 by a certified laboratory , according to anaerobic digestion batch procedure (UNI EN ISO 11734/2004).As related to cv. To evaluate the effects of density, all data were processed by analysis of variance . Seed and biomass production as well as the investigated analytical characteristics were not significantly affected by the two densities that were not further considered in this paper. Results are reported as mean ± standard deviation (s.d.), on dry matter (d.m.) basis except for BMP results reported on organic matter (o.m.) basis and are compared with values reported in literature for grass and legume plants.i.e. May-October, and organic matter, crude protein, crude oil and fat, crude ash, NDF, ADF and ADL contents of whole plant cv. Solaris biomass, was examined by two tailed Pearson correlation test .Association between thermo-pluvio-metric data observed in the cultivation period, −1 per year, ranging from 65.0 to 76.6 t ha−1. The seed yield averaged per year 2.1 (±0.16) t ha−1, confirming the data observed by Grisan and colleagues1 in Northern and Centre Italy. The mean temperature values observed in the three years study during the cultivation period was significantly related to the biomass yield  but not with rainfalls, as expected effect of waterings.Table i.e. 19.32 (±3.78) g/100 g. The organic matter content of whole plant fraction was in average 82.80 ± 2.36 g/100 g dry matter (d.m.), ranging from 80.09 (±3.83) g/100 g d.m. and 85.69 (±2.37) g/100 g d.m. in stem-leaf and inflorescence fraction, respectively  and Italian sainfoin (Hedysarum coronarium L.) among the legume plants and for Italian ryegrass , cocksfoot (Dactylis glomerata L.) and sorghum (Sorghum vulgare L.) as grass plants17. Lower crude protein contents are reported for grass plants, such as triticale , barley (Hordeum vulgare L.) and corn (Zea mays L.), while higher values are found in legumes such as alfalfa (Medicago sativa L.), vetch (Vicia sativa L.), white clover (Trifolium repens L.) and red clover (Trifolium pratense L.)18. Among the raw materials with high fibre content, the same protein levels of whole plant and stem-leaf are intermediate between the protein content of alfalfa meal (17.6 g/100 g d.m)19 and dried beet pulp (9.2 g/100 g d.m.)20.The crude protein content of the inflorescence portion (24.85 ± 1.96 g/100 g d.m.) is in line with values reported in literature for the seed cakeLolium perenne L.), corn and sorghum17. According to Hagos et al.21, high levels of lipids, especially rich in unsaturated long chain fatty acids, can negatively affect the anaerobic digestion of the biomass, leading to problems in digestors such as blocking and microbial inhibition. Similarly, as far as the use of cv. Solaris as fodder is concerned, it should be noted that the lipid content of the diet intended for ruminants should not exceed 5% of dry matter consumption to prevent the depression of fibre fermentation22. In general, this constraint militates in favour of stem-leaf and whole plant fractions of cv. Solaris, as potential fodder.The content of crude oils and fats, as expected for this energy tobacco variety, is high in the inflorescence as well as in the whole plant, but it is reduced in the stem-leaf samples Table . The cruad hoc for the possible use of the plant as feedstuff. When compared to values reported for standard grass and legume plants, highest ash contents (12–15 g/100 g d.m.) are detectable in ryegrass, cocksfoot, fescue (Festuca arundinacea Scrheb.), red and white clover while the alfalfa meal has an average ash content of 11.4 g/100 g d.m. However, higher crude ash content (16–20 g/100 g d.m.) is reported for both forage and biogas biomass of cabbage (Brassica oleracea L.), sugar beet (Beta vulgaris L.) leaves and necks, and rapeseed (Brassica napus L.)17.The range of values observed for the crude ash content varies remarkably among the investigated fractions, inflorescence, whole plant and of stem-leaf Table  suggestiSolaris biomass.The thermo-pluvio- metric data observed in the cultivation period were not significantly related with the organic matter, crude protein, crude oil and fat and crude ash contents of whole plant cv. Solaris as feedstock with alternative proteins, the protein KOH soluble content in average accounts for 4.93 and 6.83 g/100 g d.m., for stem-leaf and whole plant respectively  content is consistent with values reported for both grass and legume plants  observed in the investigated fractions of cv. is Table  are consts Table  and partSolaris is given by NDF, ADF and ADL values that provide essential information on the plant cell wall composition, i.e. in terms of hemicellulose (NDF minus ADF), cellulose (ADF minus ADL) and lignin (ADL) contents whose effects on anaerobic ecosystems are very different, as well as a more precise characterization of gastrointestinal fill (NDF) of a forage, main responsible for the voluntary intake in ruminants13.The representation of fibre in cv. Solaris is close to that found in white clover and sainfoin, even though being characterized by a lower content of cell wall (NDF)17 so that legumes are considered the best forages when the ruminants’ voluntary intake is physiologically low, as it is at the onset of lactation.The amount of fibre available for the complex anaerobic ecosystem activities (NDF minus ADL), shown in Table Solaris fractions  with values consistent with the lowest levels reported in literature for grass plants (10–19.5 g/100 g d.m.)17. However, the total sugar content in both whole plant and stem-leaf cv. Solaris fractions, variable from 0.42 g/100 g d.m. of stem-leaf and 1.41 g/100 g d.m. of inflorescence, is in average consistent with values observed in fresh grass plants, whereas the starch contents (Table 17.Non-Fibrous Carbohydrates (NFC) content, including organic acids, sugars, starch, fructans and all the constituents of the cell wall that are lost during the detergent analysis of fibre21.In literature it is reported that high levels of sugars generated during anaerobic fermentations reduce methanogenesisSolaris lie at the level of a grass plant of good quality in terms of crude protein and fibrous carbohydrates contents, although presenting higher levels of lignin, close to what is observed in legume plants.From a nutritional standpoint, values reported for whole plant and stem-leaf biomass fractions appear more balanced respect to what has emerged for the inflorescences. The fractions whole plant and stem-leaf of cv. Solaris could be conserved by ensiling the biomass harvested 60 days after the seed collection. Ensiled whole plant of cv. Solaris was observed to be a good substitute for a portion of hay in dairy heifer diet26.Moreover, the content of sugars and starch suggests that cv. The comparison with compositional data reported in literature for grass and legume plants allowed us to estimate the energy content of whole plant and stem-leaf ranging between 5.33 and 5.83 MJ Net Energy of lactation (NEl)/kg d.m. and 5.48–5.94 MJ Net Energy for weight gain (NEg)/kg d.m.Solaris biomass.No significant associations were found between the thermo-pluvio-metric data recorded during the cultivation period and the fibrous components  of whole plant cv. Solaris as energy feedstock, consistently with the described compositional data, the biochemical methane potential (BMP) tested on whole plant samples by anaerobic digestion showed for the first time quite encouraging results, summarised by an average biogas production of 290 (±75) Nm3/t organic matter (or volatile solids) whose 58% is represented by biomethane, as Table Panicum virgatum L., i.e. switchgrass, is reported to produce 125 Nm3 CH4/t organic matter27 consistently with values observed for grass silage (128–140 Nm3 CH4/t organic matter), rye ensiled (140 Nm3 CH4/t organic matter) and gliricidia  leaves (165–180 Nm3 CH4/t organic matter) while higher values are reviewed for alfalfa and rapeseed crops (240 Nm3 CH4/t organic matter)27. Much lower biogas values than cv. Solaris are reported for rice (Oryza sativa L.) straw (74 Nm3 biogas/t organic matter), cotton (Gossypium L.) stalks and mixed green leaves biomass (180 Nm3 biogas/t organic matter)28.When considered cv. i.e. source of biofuel4 and protein from the seed cake for piglet feeding5.These results further extend the potential destinations for this versatile cultivar for the use in animal nutrition and in biomethane production, besides those already studied, Solaris biomass, for the first time the macro-minerals profile is reported in Table 17.Finally, as a further contribute to advanced knowledge on cv. Solaris as forage for ruminants would require attention in balancing the dietary mineral components, while its use in late pregnancy diet would be avoided in ruminants, due to the high risk of metabolic diseases in the peripartum phase.Because of the high Ca/P ratio, the eventual use of cv. 17. The average content of sodium measured in cv. Solaris samples was similar to values reported in literature for grass and legume plants17, while the potassium (K) content shows very high values. Among the factors influencing the K content of forages, also reported high for plants at early vegetative stage (1–4 g 100 g−1 d.m.), it has to be mentioned that, besides soil characteristics and environmental conditions, fertilization is one of the major cause of very high K levels of forages29. High levels of K in forages are considered etiological factors of diseases in the peripartum of the cow so that it is crucial especially in this delicate physiological phase to consider the balance between cations and anions of the diet30.Values higher than what detected in grass and legume plants are indicated also for magnesium (Mg)Nicotiana tabacum cv. Solaris multitasking potential but further investigation on qualitative aspects of analytic components are required for its use in animal nutrition.The results of the present study allow to extend the Solaris as a candidate for a competitive repositioning of innovated tobacco cultivation, especially in the marginal areas traditionally suited to tobacco and devoted to animal breeding. However, the suggested multiple potentialities of cv. Solaris raise questions about management of the crop, with reference to timing and harvesting mode, storage techniques and the development of various strategies depending on the destination of products and derivatives.Taken together, the results here presented place for the first time cv.


Background: Evidence suggests that social support impacts on participation in sport or physical activity (PA), and is associated with health benefits, although the link is complex and not well understood. The study aim was to examine whether participation in organized sports is related to body composition, physical fitness, and social correlates for PA. Methods: Cross-sectional data on 238 adolescents (90 boys and 148 girls), mean age 14.9 ± 0.8 years, who were participants in the Physical Activity and Health Longitudinal Study, were collected. The participants were divided into two groups: sport participation (SP) and non-sport participation (NSP). Height, weight, and triceps and subscapular skinfolds were assessed according to standard procedures. Weight (kg) and height (m2) were used to calculate body mass index (BMI), and skinfolds were used to calculate body fat percentage. The European Test of Physical Fitness (EUROFIT) battery of tests was used to assess physical fitness. The standardized International Physical Activity Questionnaire Short Form and Social Support for PA questionnaires were used to obtain information on PA and social correlates for PA, respectively. Participants were asked to choose between participation and non-participation in sport. Results: The SP group had lower BMI component values ; p = 0.011, d = 0.53 for sum of skinfolds (∑SKF) obtained higher physical fitness scores in selected items ; p < 0.000, d = 0.96 for maximal oxygen consumption VO2max) and received higher social support ), than the NSP group. The social support received by those participating in sport correlated positively with most fitness components ; p = 0.000, r2 = 12% for sit ups (SUP); p = 0.000, r2 = 17% for VO2max). Physical fitness components were negatively associated with most body composition components for both groups. Conclusions: The results provide a better understanding of sport participation in organized sports-related, body composition-related and physical fitness-related associations with changes in social support received by adolescents and may contribute to the development of more accurate promotive strategies to increase children’s and adolescents’ engagement in sport and PA. Children who are active through sport are more likely to be physically active in adulthood than those who do not participate in childhood sport . AlthougConcerns exist about children and adolescents who do not meet the PA guideline of 60 min (1 h) or more of PA daily ,6. One rIt is important to underline the fact that when children and adolescents participate in the recommended level of PA, multiple health benefits accrue . The WorAccording to Sheridan et al. , coachesOur previous publications on the sample under study in the Physical Activity Health Longitudinal Study (PAHLS) reported findings on body composition, physical fitness, lipid profiles, blood pressure, physical activity and the social correlates of physical activity ,36,37,38The article presents findings from cross-sectional (2011) data from PAHLS. Detailed information about the participants, method of data collection and sample size has been published elsewhere ,38. The 2 (kg/m2). Percentage body fat (%BF) was derived from the sum of triceps and subscapular skinfold (∑SKF) measurements according to an equation developed by Slaughter et al. [Anthropometric measurements of height (cm), weight (kg) and skinfolds (mm) were taken according to the standard procedures described by the International Society for the Advancement Kinanthropometry (ISAK) . BMI, asr et al. , suitabl2max) was assessed with the 20-m shuttle run test and the result was the level (minute) in which the participant stopped running. Sit and reach (SAR) is designed to measure hamstring flexibility, expressed in centimetres.The health-related physical fitness status of participants was determined by assessing their cardiorespiratory endurance, muscle strength and endurance, and flexibility, using the European Test of Physical Fitness . StandinThe standardized International Physical Activity Questionnaire Short Form version (IPAQ-SF) , and SocIn turn, the SSPA scale included nine questions, asking youth about the different types of support they receive and also give to others during a typical week. Responses were on a three-point scale ranging from 1 (never), 2 (sometimes) to 3 (every day). The questionnaire asked the following questions, “during a typical week”: (1) “How often do you encourage your friends to do physical activity or play sports?”; (2) “How often do your friends encourage you to do physical activity or sports?”; (3) “How often do your friends do physical activities or play sport with you? (4) “How often do your friends tell you that you are doing a good job at a physical activity?”; (5) “Has someone encouraged you to do physical activities or sports?”; (6) “Has someone done a physical activity or played sports with you?”; (7) “Has someone provided transportation to a place where you can do physical activities or play sports?”; (8) “Has someone watched you participate in physical activities or sports?”; and (9) “Has someone told you that you are doing well in physical activity?”. The participants were requested to choose their answer by making a mark or cross next to either never, sometimes or every day. For the purpose of this article we created a new variable named social support (SS), by adding the point scores  Cronbach’s α analyses for internal consistency gave an α score of 0.74.Before data collection for the PAHLS, permission was granted by the District Manager of the Department of Basic Education in Potchefstroom, North West Province, South Africa. The research protocol was approved by the Ethics Committee of North-West University (Ethics number: NWU-0058-01-A1) of the Potchefstroom campus. Parents or guardians gave permission for their children to participate in the study, and participants provided informed assent.t-test was used for parametric data and the Mann–Whitney U-test for non-parametric data. The effect size (ES) Cohen’s d (standardized mean differences) was calculated. Guideline values of Cohen’s d = 0.2, 0.5 and 0.8 were regarded as small, medium and large effects, respectively [2max, SAR, and SS. The correlation effect size (r) was also calculated. Guideline values of correlation effect size ρ = 0.1, 0.3 and 0.5 were regarded as small, medium and large effects, respectively [Descriptive statistics are presented as means, standard deviations and frequencies. Overall analyses were performed for sport and non-sport participation, and were also run separately for gender. The ectively  for paraectively . A simpl2max 34.25 ± 8.11 and 27.29 ± 6.42; and SAR 46.03 ± 8.62 and 45.94 ± 6.91, for the SP group and NSP group, respectively.The descriptive statistics of participants’ characteristics are presented in p < 0.001) and practically significant (ES ≥ 0.8) higher mean values for %BF and ∑SKF compared with boys. In contrast, boys had significantly (p < 0.001) and practically significant (ES ≥ 0.8) higher mean values for the majority of physical fitness items in comparison with girls. Only for SAR did girls obtain significantly (p < 0.001) and practically significant (ES ≥ 0.8) higher scores than boys. Among adolescents from the NSP group, practically significant differences were observed for the different gender groups for most physical fitness measures, except for SAR. Boys received significantly (p < 0.001) and practically significant (ES ≥ 0.8) higher results than girls for SBJ, BAH, SUP and VO2max (p < 0.002), while girls showed significantly (p < 0.001) and practically significant (ES ≥ 0.8) higher mean values for %BF and ∑SKF. Cohen’s effect size (d) values indicated large and practically significant differences for these measures.In the SP group, girls had significantly (p < 0.001), SUP (p < 0.01) and VO2max (p < 0.001). Furthermore, adolescents from SP group scored significantly and practically significantly (ES ≥ 0.5) higher for SS than their peers from NSP group (p < 0.001). In turn, adolescents from NSP group had significantly (p < 0.05) and practically significant (ES ≥ 0.5) higher body fat (%BF and ∑SKF) than those from SP group. The effect size values indicated large and practically significant differences for these measures.p < 0.02), where SP boys scored higher that peers who did not participate in sport.In boys, we found a statistically significant difference only for SUP (p < 0.01) and VO2max (p < 0.001), and also scored significantly higher for SS than girls from the NSP group (p < 0.01). The effect size values indicated large and practically significant differences for these measures.When comparing girls from the SP and NSP groups, girls who participated in sport obtained significantly and practically significantly (ES ≥ 0.5) higher results in the physical fitness items of SBJ  and social support (SS), separately for SP group and NSP group. In the SP group, BMI was negatively correlated with SBJ , BAH  and VO2max . %BF was negatively correlated with SBJ , BAH , SUP  and VO2max, . Furthermore, BMI was positively correlated with SAR . ∑SKF was negatively correlated with SBJ , BAH , SUP  and VO2max . No significant correlations were found between SS and body composition measures. SS was positively associated with BAH , SUP  and VO2max  and negatively with SAR . The correlation effect size (r) values ranged from small to large significant differences for these measures.p = 0.05) and VO2max ; and %BF and SBJ , BAH  and VO2max . Additionally, ∑SKF was negatively correlated with SBJ , BAH  and VO2max . In this group we reported positive correlation between SS and BMI  and negative correlation between SS and VO2max . The correlation effect size (r) values ranged from medium to large significant differences for these measures.Among adolescents from NSP group, negative correlations were found between BMI and BAH , %BF  and ∑SKF  and positively associated with ∑SKF . BAH was negatively associated with BMI , %BF  and ∑SKF  and positively associated with SS . SUP was negatively associated with %BF  and ∑SKF  and positively associated with SS . Also, VO2max was inversely associated with BMI , %BF  and ∑SKF  and positively associated with SS . SAR was positively associated with %BF .Simple linear regression  showed tp = 0.012, r2 = 21%) and ∑SKF . BAH was negatively associated with BMI , %BF  and SKK . VO2max was inversely associated with BMI , %BF  and ∑SKF . SAR was negatively associated with BMI  and ∑SKF .In the NSP group, we found that SBJ was negatively associated with %BF .2 = 0.016), where 50.6% of SP boys selected sometimes and 57.1% of NSP boys selected never. The SP group of boys indicated significantly often (42.0% every day and 50.6% sometimes) that their friends tell them that they are doing a good job at PA, more than boys from the NSP group (57.1% never) .In the group of boys, we found more significant differences between those who participated and those who do not participate in sport than in girls. The SP and NSP groups of boys indicated similar percentages of friends who were participating in PA or playing sport with them every day ; however, significant differences were found in the answers “sometimes” and “never” , with 27.2% of boys from the SP group indicating that they had transport ‘every day’, 45.7% of them selecting “sometimes’ and 71.4% of boys selecting the answer “never’. None of the NSP boys indicated that someone provided transportation “every day”.The SP and NSP groups of boys demonstrated significant differences in terms of support from someone who provided transportation to a place where they could participate in PA or plays sports .Finally, the SP group of boys significantly indicated that someone told them that they are doing well in PA “often” (44.4% every day and 49.4% sometimes), compared with boys from the NSP group (57.1% never)  and body composition components , both for the SP group and NSP group, were found (except for the association between SBJ and BMI in the NSP group). However, those associations were in general stronger in the SP group. In the same group of adolescents, we also reported significantly more negative associations between physical fitness and body composition  compared with the NSP group. These results highlight the important contribution of sport participation to numerous health benefits, such as improved aerobic fitness, muscular strength and endurance, increased bone mineral density and a more favourable cardiovascular risk factor profile [The results show significantly high and positive correlations for body composition components , both in SP and NSP groups. However, this finding is not surprising, as previous studies have shown that all stated variables are indicators of fatness ,48. Addi profile ,49.2max) and SS, but only for adolescents from the SP group. Similar findings to those of our study have been shown in other studies, where, among other factors, sport participation may influence cardiorespiratory fitness, which appears to have an impact on BMI and other metabolic factors [The results also show significant positive associations between selected physical fitness items  might be a key contributor to adolescents being active or participating in sport. Adolescents from the SP group reported receiving higher support (in a different context) than their peers from the NSP group. Despite far-reaching research examining the relationship between social support and participation in PA, inconsistencies have emerged. For instance, in the study of Higgins et al. , girls rBoth the girls and boys who participated in sport reported significantly higher social support than their counterparts in the NSP groups . Overall, social support was a strong predictor of physical fitness (except for SAR) only in the SP group. This finding confirms that social support is connected with physical fitness and sport participation. Girls from the SP group showed more attention while participating in PA/sport than girls from the other group. However, boys from both the SP and the NSP groups showed significant differences with regard to “support from someone who provided transportation to a place where they could do PA or plays sports”, to the disadvantage of those boys who do not participate in sport. Furthermore, we found that boys who participate in sport often got feedback from their friends that they are doing well at PA. Boys from the SP group were significantly more likely to have friends participating in PA or playing sport. Similar results were found in a study by Duncan et al. , which rLimitations need to be taken into consideration when interpreting these results. First, participation in organized sport was self-reported, which may have influenced the direction of the current findings. Second, we did not take into consideration the type of sport performed, frequency or duration. We only considered weekly practice of PA that involved training sessions organized by adults. Third, the SP and NSP group sample sizes were unequal. Interestingly, most of the examined youth participated in sports activities (208 of the 238 youth), which may be related to the PE system in South African schools, where PE is part of a multidimensional learning area known as Life Orientation  . We presOur findings indicated that participation in organized sport was associated with higher physical fitness, higher social support and lower body composition among adolescents. Sport participants were fitter, had lower body composition and obtained more social support compared with non-sport participants. The participation of adolescents in organized sports may be a way to minimize increases in body composition, and greater social support (from parents and peers) may enable more adolescents to participate in sport. Participation in organized sport conveys lasting benefits that are considered attributes of a healthy lifestyle. Therefore, the youth must be encouraged to participate in sport or PA and parents taught how to support their children. More accurate promotive strategies are needed to increase adolescent engagement in sport and general PA.


Sarcoidosis is a systemic inflammatory multi-organ disease almost always affecting the lungs. The etiology remains unknown, but the hallmark of sarcoidosis is formation of non-caseating epithelioid cells granulomas in involved organs. In Scandinavia, > 30% of sarcoidosis patients have Löfgren’s syndrome (LS), an acute disease onset mostly indicating a favorable prognosis. The impact of dysregulation of lipid mediators, which has been investigated in other inflammatory disorders, is still unknown.n = 41) and healthy controls (n = 16).Using three different liquid chromatography coupled to tandem mass spectrometry targeted platforms (LC-MS/MS), we quantified a broad suite of lipid mediators including eicosanoids, sphingolipids and endocannabinoids in bronchoalveolar lavage (BAL) fluid from pulmonary sarcoidosis patients  and its regioisomer 14,15-dihydroxyeicosa-5,8,11-trienoic acid  increased in patients with sarcoidosis. Additional shifts were observed in sphingolipid metabolism, with a significant increase in palmitic acid-derived sphingomyelin . No associations were found between these 3 lipid mediators and LS, whereas levels of SM 16:0 and 11,12-DiHETrE associated with radiological stage (p < 0.05), and levels of 14,15-DiHETrE were associated with the BAL fluid CD4/CD8 ratio.A total of 47 lipid mediators were consistently detected in BAL fluid of patients and controls. After false discovery rate adjustment, two products of the soluble epoxide hydrolase (sEH) enzyme, 11,12-dihydroxyeicosa-5,8,14-trienoic acid  contains supplementary material, which is available to authorized users. Additionally two sphingomyelins , and one hexosylceramide  , but were above the FDR threshold.One sphingolipid (SM 16:0 [There were no differences (p < 0.05) between patients with sarcoidosis and healthy controls in endocannabinoid levels  and CD4/CD8 ratio  and SM 16:0 (p = 2.5 × 10− 2) increased with radiography stage according to the Cochran-Armitage test  and SM 16:0  , associations with LS, CD4/CD8 ratio, chest radiograph stage, %FEVed Figs.  and 4. DGiven the known role of lipid mediators in inflammation , 19, andclinicaltrials.gov NCT02743468) as well as in COPD . Interestingly, the linoleic acid-derived 9(10)- and 12(13)-EpOME  were decreased in patients with sarcoidosis. Though these compounds are also metabolized via the sEH to form the corresponding vicinal diols, no differences were found between patients and controls in the BAL fluid levels of the downstream products . These data suggest that the lipid mediator dysregulations in sarcoidosis are specific for the arachidonic acid pathway, as opposed to COPD [Multiple lipid mediators were quantified in the BAL fluid, further supporting that this material is useful for monitoring biochemical process in the lung. The primary observed shifts in lipid mediators were increases in the BAL fluid concentrations of 11,12-DiHETrE and 14,15-DiHETrE with sarcoidosis  and HexCer (16:0). Sphingomyelins have been previously reported to increase in the BAL fluid of sarcoidosis patients . These sThere are some limitations in this study. First, there is an age imbalance between sarcoidosis patients and healthy controls. However, none of the dysregulated metabolites was associated with age in patients or controls. Two patients were on prednisolone treatment and four more were on local steroids at the time of sampling, but they did not exhibit different profiles in the reported lipid mediators (data not shown), which does not suggest a strong effect of steroid treatment upon lipid mediator profiles. Additionally, patients with more advanced stages of the disease would be needed in order to confirm the association of the identified markers with the sarcoidosis radiological stage. On the other hand, this study explored the lipid mediator profiles close to disease onset, minimizing any potential effects associated with ongoing treatment, and highlighting disease-specific processes. In addition, the specificity of the observed dysregulations in lipid mediator levels need to be established relative to other respiratory diseases. Lastly, an obstacle to working with BAL fluid is the lack of clear protocols to perform data normalization. Previous attempts have been made to normalize the data using for example recovery volume  or metabThis is the first study to profile lipid mediators in sarcoidosis, providing useful information on the levels of these potent biological mediators in this disease. The findings suggest that arachidonic acid-derived products of the sEH pathway as well as some sphingolipid species (particularly SM 16:0) may serve as molecular descriptors of sarcoidosis. Of particular interest are the observations that the primary altered pathways are similar to those observed in other respiratory diseases, including asthma, COPD, and cystic fibrosis, suggesting that the diseases share a similar inflammatory component. The predominant shifts were in sEH products of arachidonic acid metabolism, further stressing the potential role of this pathway in the inflammatory process in lung pathology and highlighting the targeting of this enzyme for disease treatment. It is of particular interest that these sEH products also correlated with radiological stage, further strengthening their association with more advanced disease. These findings suggest that the sEH pathway should be further investigated in sarcoidosis, both from the standpoint of potentially being involved in disease etiology and representing a novel therapeutic target for disease control.Additional file 1:Data S1. Summary of data used in the analysis. (XLSX 31 kb)Additional file 2:Table S2. Nomenclature for the compounds screened in the samples. Compounds reproducibly detected in samples and submitted to data analysis are highlighted in bold. (DOCX 196 kb)Additional file 3:Table S3. Internal standards for each of the three LC-MS/MS lipid mediator platforms. (PDF 78 kb)Additional file 4:Appendix S1. Detailed procedures employed for the extraction of eicosanoids, endocannabinoids and sphingolipids. (PDF 11 kb)Additional file 5:Appendix S2. Chromatographic and general MS conditions employed in the detection of eicosanoids, sphingolipids and endocannabinoids. (PDF 97 kb)Additional file 6:Table S4. Results for the comparisons of lipid mediator levels between healthy (n = 16) and sarcoidosis (n = 41) groups. Values have been normalized to the median value of the healthy group. Compounds with p < 0.05 are highlighted in bold. (PDF 71 kb)Additional file 7:Table S5. Spearman’s correlations between lipid mediators and age for healthy controls and patients with sarcoidosis. Compounds with p < 0.05 are highlighted in bold. (PDF 39 kb)Additional file 8:Table S6. Results of the Cochran-Armitage test with radiography stage in sphingolipids significantly differing (p < 0.05) between patients and controls. Direction of the null hypothesis is indicated for each compound. Compounds with p < 0.05 are highlighted in bold. (PDF 79 kb)


Glucocorticoid receptors (GRs) shuttle from the cytoplasm (cy) to the nucleus (nu) when bound with glucocorticoids  and alter transcriptional activity. GR activation within the fear circuit has been implicated in fear memory and post traumatic stress disorder (PTSD). However, no study to date has characterized GR internalization within the fear circuit during fear memory formation or examined how traumatic stress impacts this process. To address this, we assayed cy and nu GR levels at baseline and after auditory fear conditioning (FC) in the single prolonged stress (SPS) model of PTSD. Cy and nu GRs within the medial prefrontal cortex (mPFC), dorsal hippocampus (dHipp), ventral hippocampus (vHipp), and amygdala (AMY) were assayed using western blot. The distribution of GR in the cy and nu (at baseline and after FC) was varied across individual nodes of the fear circuit. At baseline, SPS enhanced cyGRs in the dHipp, but decreased cyGRs in the AMY. FC only enhanced GR internalization in the AMY and this effect was attenuated by SPS exposure. SPS also decreased cyGRs in the dHipp after FC. The results of this study suggests that GR internalization is varied across the fear circuit, which in turn suggests GR activation is selectively regulated within individual nodes of the fear circuit. The findings also suggest that changes in GR dynamics in the dHipp and AMY modulate the enhancing effect SPS has on fear memory persistence. Glucocorticoid receptors (GRs) are ligand-gated transcription factors. Upon binding with glucocorticoids they leave the cytoplasm (cy) and enter the nucleus (nu) as dimers  where they bind to GREs to regulate transcription –4. GRs cGlucocorticoid release during fear conditioning (FC) has been implicated in fear memory consolidation –8 and spSingle prolonged stress (SPS) refers to serial exposure to restraint, forced swim, and ether and is a validated animal model of PTSD –29. SPS These substrates were selected, because they are critical nodes of the fear circuit –45. ResuEighty-eight male Sprague-Dawley rats  obtained from Charles River Inc. were used in this study. Upon arrival, rats were housed in pairs during a five day acclimation period with ad libitum access to food and water. Following SPS and control procedures, rats were individually housed and restricted to 23g/day of standard rat chow per the manufacturer’s recommendation (LabDiet St. Louis MO) with ad libitum access to water. Experiments commenced following the animals’ acclimation period. The rats were on a 12 hour light/dark cycle and all experimental procedures were performed in the animals’ light cycle between the hours of 9:00 am and 2:00pm. All experiments were approved by the University of Delaware Institutional Animal Care and Use committee following guidelines established by the NIH.All rats were randomly assigned to the SPS or control stress group prior to SPS. SPS was conducted as previously described , 46 and SPS and control rats were randomly assigned to one of four groups: baseline, FC0, FC30, or FC60. Rats in the baseline treatment were removed from the housing colony and immediately euthanized in order to determine baseline GR levels. All other rats were removed from the housing colony and subjected to FC. FC sessions were conducted as previously described , 34 usinAll animals were euthanized via rapid decapitation and their brains were immediately extracted and flash frozen in isopentane chilled on dry ice. Brains were then stored in a -80°C freezer until further processing. To dissect brain regions, brains were thawed to -13°C cryostat (Leica CM1350) and 300 μm coronal sections through the mPFC, AMY, dHipp, and vHipp were taken and these brain regions dissected out and placed into 1.5 mL microtubes. Dissected brain regions in microtubes were then stored in a -80°C freezer.All brain sections were treated to separate cy and nu fractions using a method described by Spencer et al., . We empiThe rough pellet from the initial centrifuge treatment was used to obtain the nu fraction from dissected brain regions. The pellet was washed twice in 400 μL of buffer and then resuspended in 150 μL of buffer that had a high concentration of NaCl . Samples were placed on a fixed speed vortex mixer and the suspension was incubated in ice for 1 hour with frequent shaking. Following incubation, samples were centrifuged at 8,000 x g for 15 minutes at 4°C. The supernatants from these samples were treated as the nu fraction of brain tissue.The protein concentration of cy and nu fractions were increased using protein concentrator columns . Protein assay was then performed on each sample per manufacturer’s directions (Pierce BCA Protein Assay Kit). 0.5X Laemmli sample buffer was mixed with approximately 15 μg of protein from each sample. These samples were stored in a -80°C until western blot. Protein samples were heated at 70°C for 7 minutes before being loaded into 10% Tris-HCl polyacrylamide gels and separated by SDS polyacrylamide gel electrophoresis. Separated proteins were electrophoretically transferred from gels to nitrocellulose membranes. The membranes were subsequently left to dry for 30 minutes at 37°C followed by rehydration washes in 0.5 M Tris-buffered saline (TBS). Blots were blocked for 1 hour at room temperature in TBS containing 5% non-fat milk. Nitrocellulose membranes were probed for GR and β actin (reference protein) by incubating overnight at room temperature with a polyclonal rabbit GR antibody  and a mouse β-actin antibody  in TBS. After 18–20 hours, the membranes were subjected to several washes in 0.5 M TBS with 0.1% Tween-20 then a 3 hour incubation at room temperature with polyclonal goat anti-rabbit (800CW)  and anti-mouse IgG (680RD)  secondary antibodies in 0.5M TBS containing 0.1% Tween and 5% non-fat milk. Nitrocellulose membranes were then washed in TBS and scanned in the Li-cor Odyssey Clx scanner under the following settings: resolution– 169 μm, quality–lowest, focus offset– 0.0 mm.Freezing behavior was analyzed using ANY-maze (Stoelting Inc.) as previously described . Fear-coIn order to reduce variability in western blot data, representative rats from each independent factor (i.e. stress and condition) were always included in each protein assay and western blot. The integrated intensity (I.I.) of GR and β-actin protein bands were scored using ImageStudio. The profile curves of all bands were inspected to ensure that there was significant I.I. signal above the background within each lane in every western. For all statistical analyses cyGR or nuGR were expressed relative to cyβ-actin or nuβ-actin to yield relative GR/actin ratios. We also divided relative nuGR/actin ratios (nuGR/nuAct) into relative cyGR/actin ratios (cyGR/cyAct) to yield a single measure of GR internalization  in all brain regions.A stress x fraction (cy vs. nu) factor design was used to examine the effect of SPS on relative cyGR and nuGR levels at baseline, with fraction being a repeated measure. T-test (SPS vs. control) was used to analyze baseline nGR/cGR ratios. A stress x fraction x condition  factor design was used to examine changes in relative cyGR and nuGR levels after FC. A stress x condition factor design was used to examine changes in nGR/cGR ratios. To specifically examine how cyGR and nuGR levels changed after FC, relative cyGR, nuGR, and nGR/cGR ratios in the condition factor were expressed as a percent change from baseline . These normalized values were subjected to separate stress x condition factor designs.Main and simple effects were analyzed using analysis of variance (ANOVA), while main and simple comparisons were analyzed using independent, paired sample, or one sample t-test with a Bonferroni correction applied where appropriate. The reference value for all one sample t-tests was set to 100. Statistical significance was assumed with p < .05 for all statistical tests. = 127.615, p < .001], which suggested all rats acquired fear memory. There was also a significant stress x trial interaction . This was driven by enhanced freezing in SPS rats during FC trial 1 of FC in comparison to control rats . However, at the end of FC all animals had equivalent levels of freezing (p > .05), which suggests SPS did not alter acquisition of FC; a finding consistent with previous studies . Normalized nGR/cGR ratios were unaffected by stress (p > .05), but were significantly lower after FC in comparison to baseline . This suggests GR internalization in the vHipp was decreased after FC. These results are illustrated in Relative baseline GR/actin and nGR/cGR ratios were unaffected by stress and baseline cyGR/actin and nuGR/actin ratios were equivalent  = 57.529, p < .001] for GR/actin ratios at baseline. This reflected enhanced cyGR/actin, relative to nuGR/actin, in the dHipp. There was also a significant stress x fraction interaction , which was driven by SPS enhancement of cyGR/actin relative to nuGR/actin. This assertion was supported by significant t-test when comparing difference scores between relative cy and nu GR/actin ratios (i.e. cyGR–nuGR) for SPS vs. control rats . Independent t-test for baseline nGR/cGR ratios was not significant (ps > .05). These results are illustrated in There was a main effect of fraction  after FC. There were no significant effects of stress and/or condition (ps > .05). There were no stress and/or condition effects for nGR/cGR ratios after FC (ps > .05). These results are illustrated in  = 5.707, p = .021] that was driven by lower levels of cyGR/actin in SPS rats. This effect was pronounced at the FC60 time point. Significant one sample t-test for SPS rats , but not control rats (p > .05), also supported the assertion that cyGR/actin ratios were decreased in SPS rats after FC. There were no significant stress and/or condition effects on relative nuGR/actin or nGR/cGR ratios after FC (ps > .05). These results are illustrated in CyGR/actin ratios were higher when compared to nuGR/actin ratios  for baseline GR/actin ratios, which reflected enhanced cyGR/actin relative to nuGR/actin. This effect was attenuated in SPS rats, which was suggested by a stress x fraction interaction that approached significance . There was no effect of stress on baseline nGR/cGR ratios (p > .05). These results are illustrated in There was a significant main effect of fraction . There were no effects of stress and/or condition on GR/actin or nGR/cGR ratios after FC (ps > .05). Relative to baseline, cyGR/actin ratios increased after FC in SPS rats, but not control rats. This was suggested by a stress effect that approached statistical significance . These results are illustrated in  = 5.607, p = .024], which was driven by a failure to enhance nGR/cGR ratios after FC in SPS rats. This interpretation was supported by one sample t-test that was significant for control rats , but not SPS rats (p > .05). These findings suggests that FC enhanced GR internalization in the AMY and this effect was attenuated by SPS  = 10.147, p = .007] for baseline GR/actin ratios, which reflected enhanced cyGR/actin, relative to nuGR/actin, in the mPFC. There was no significant stress effect for baseline GR/actin ratios or nGR/cGR ratios  = 4.367, p = .047]. There were no stress and/or condition effects on GR/actin or nGR/cGR ratios after FC  changes with SPS exposure and after FC. The results suggest there is selective regulation of GR dynamics within individual neural substrates of the fear circuit at baseline and with FC. Baseline cyGR was enhanced relative to nuGR in all brain regions except for the vHipp. FC had no effect on GR dynamics in the mPFC and dHipp, but increased cy and nu GR levels in the vHipp. In spite of this there was an overall decrease in vHipp GR internalization after FC. Enhanced GR internalization after FC was only observed in the AMY. Thus, glucocorticoid release (whether at baseline or stress-induced) does not uniformly determine GR trafficking between the cy and nu within the fear circuit.SPS disrupted GR dynamics in the dHipp and AMY at baseline and after FC, with cyGRs being sensitive to SPS in both brain regions. SPS enhanced cyGRs in the dHipp, but decreased cyGRs in the AMY at baseline. These effects were inverted after FC with lower cyGRs in the dHipp of SPS rats, but enhanced cyGRs in the AMY. The enhancement in GR internalization in the AMY observed in control rats was disrupted by SPS. A previous study has observed that systematically inhibiting GR activation during FC exacerbates persistent fear memory induced by SPS exposure without having any effect on non-stressed rats . InhibitPrevious studies have also shown that GR activation in the dHipp and AMY enhance memory consolidation in non-stressed rats see , which aHow might substrate specific regulation of GR dynamics occur in the fear circuit when the ligand that activates GRs originates from a single source outside of the central nervous system ? 11β-hydroxysteroid dehydrogenase types 1 and 2  are enzymes capable of either converting inert 11-keto forms of glucocorticoids (e.g. 11 dehydrocorticosterone) into active glucocorticoid (11β-HSD1) or metabolizing glucocorticoids (11β-HSD2). Via these mechanisms substrate specific levels of glucocorticoids can be achieved within the brain , 53. BotGRS are phosphorylated at various sites, which alters GR function, including GR internalization , 59, 60.The results of this study demonstrate that GR dynamics are varied in different neural substrates that comprise the fear circuit. This suggests that basal glucocorticoid release and stress-enhanced adrenal glucocorticoid release can have varied effects on the fear circuit via local regulation of GR activation. Furthermore, the effect of traumatic stress on GR dynamics at baseline and during fear memory formation are restricted to specific nodes within the fear circuit. Previous studies have shown that glucocorticoid administration shortly after trauma , 25 and S1 Fig(DOCX)Click here for additional data file.