Evaluation of new alternative methods for the identification of estrogenic, androgenic and steroidogenic effects: a comparative in vitro/in silico study.

A suite of in vitro assays and in silico models were evaluated to identify which best detected the endocrine-disrupting (ED) potential of 10 test chemicals according to their estrogenic, androgenic and steroidogenic (EAS) potential compared to the outcomes from ToxCast. In vitro methods included receptor-binding, CALUX transactivation, H295R steroidogenesis, aromatase activity inhibition and the Yeast oestrogen (YES) and Yeast androgen screen (YAS) assays. The impact of metabolism was also evaluated. The YES/YAS assays exhibited a high sensitivity for ER effects and, despite some challenges in predicting AR effects, is a good initial screening assay. Results from receptor-binding and CALUX assays generally correlated and were in accordance with classifications based on ToxCast assays. ER agonism and AR antagonism of benzyl butyl phthalate were abolished when CALUX assays included liver S9. In silico final calls were mostly in agreement with the in vitro assays, and predicted ER and AR effects well. The efficiency of the in silico models (reflecting applicability domains or inconclusive results) was 43-100%. The percentage of correct calls for ER (50-100%), AR (57-100%) and aromatase (33-100%) effects when compared to the final ToxCast call covered a wide range from highly reliable to less reliable models. In conclusion, Danish (Q)SAR, Opera, ADMET Lab LBD and ProToxII models demonstrated the best overall performance for ER and AR effects. These can be combined with the YES/YAS assays in an initial screen of chemicals in the early tiers of an NGRA to inform on the MoA and the design of mechanistic in vitro assays used later in the assessment. Inhibition of aromatase was best predicted by the Vega, AdmetLab and ProToxII models. Other mechanisms and exposure should be considered when making a conclusion with respect to ED effects.

Antibacterial activity of free or encapsulated selected phenylpropanoids against Escherichia coli and Staphylococcus epidermidis.

AIMS: Antibacterial activities of phenylpropenes (PPs) (eugenol, isoeugenol, estragole and trans-anethole) and hydroxycinnamic acids (HCAs) (p-coumaric, caffeic and ferulic acids) were assessed against Escherichia coli and Staphylococcus epidermidis. Effect of cyclodextrin and liposome encapsulation on the PPs activity was also evaluated. METHODS AND RESULTS: All PPs inhibited the bacterial growth in the hundred micromolar range, while HCAs did not, as determined by broth macrodilution. Anethole and estragole showed a higher efficiency than eugenol and isoeugenol, and E. coli was more susceptible than S. epidermidis. Hydroxypropyl-ß-cyclodextrin/PP complexes and anethole-loaded Lipoid S100-liposomes were prepared by freeze-drying and ethanol injection respectively. Both formulations were substantially less active than free PPs. For instance, E. coli growth inhibition was about 14% for all HP-ß-CD/PP complexes evaluated at MIC50 values of free PPs (P < 0·05), and about 12% for liposomal anethole evaluated at minimal bactericidal concentration value of free anethole (P < 0·05). CONCLUSIONS: Hydrophobicity appears to be crucial for PPs antibacterial activity. Encapsulation in cyclodextrin and liposome seems to retain the PPs preventing their interaction with bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the structural features of simple phenylpropanoids related to their antibacterial activity. The limitations of conventional encapsulation systems on the activity of PPs should be considered in future applications.

Muscle activation patterns in the Nordic hamstring exercise: Impact of prior strain injury.

This study aimed to determine: (a) the spatial patterns of hamstring activation during the Nordic hamstring exercise (NHE); (b) whether previously injured hamstrings display activation deficits during the NHE; and (c) whether previously injured hamstrings exhibit altered cross-sectional area (CSA). Ten healthy, recreationally active men with a history of unilateral hamstring strain injury underwent functional magnetic resonance imaging of their thighs before and after six sets of 10 repetitions of the NHE. Transverse (T2) relaxation times of all hamstring muscles [biceps femoris long head (BFlh); biceps femoris short head (BFsh); semitendinosus (ST); semimembranosus (SM)] were measured at rest and immediately after the NHE and CSA was measured at rest. For the uninjured limb, the ST's percentage increase in T2 with exercise was 16.8%, 15.8%, and 20.2% greater than the increases exhibited by the BFlh, BFsh, and SM, respectively (P < 0.002 for all). Previously injured hamstring muscles (n = 10) displayed significantly smaller increases in T2 post-exercise than the homonymous muscles in the uninjured contralateral limb (mean difference -7.2%, P = 0.001). No muscles displayed significant between-limb differences in CSA. During the NHE, the ST is preferentially activated and previously injured hamstring muscles display chronic activation deficits compared with uninjured contralateral muscles.

Development and validation of PBPK models for genistein and daidzein for use in a next-generation risk assessment.

Introduction: All cosmetic ingredients must be evaluated for their safety to consumers. In the absence of in vivo data, systemic concentrations of ingredients can be predicted using Physiologically based Pharmacokinetic (PBPK) models. However, more examples are needed to demonstrate how they can be validated and applied in Next-Generation Risk Assessments (NGRA) of cosmetic ingredients. We used a bottom-up approach to develop human PBPK models for genistein and daidzein for a read-across NGRA, whereby genistein was the source chemical for the target chemical, daidzein. Methods: An oral rat PBPK model for genistein was built using PK-Sim® and in vitro ADME input data. This formed the basis of the daidzein oral rat PBPK model, for which chemical-specific input parameters were used. Rat PBPK models were then converted to human models using human-specific physiological parameters and human in vitro ADME data. In vitro skin metabolism and penetration data were used to build the dermal module to represent the major route of exposure to cosmetics. Results: The initial oral rat model for genistein was qualified since it predicted values within 2-fold of measured in vivo PK values. This was used to predict plasma concentrations from the in vivo NOAEL for genistein to set test concentrations in bioassays. Intrinsic hepatic clearance and unbound fractions in plasma were identified as sensitive parameters impacting the predicted Cmax values. Sensitivity and uncertainty analyses indicated the developed PBPK models had a moderate level of confidence. An important aspect of the development of the dermal module was the implementation of first-pass metabolism, which was extensive for both chemicals. The final human PBPK model for daidzein was used to convert the in vitro PoD of 33 nM (from an estrogen receptor transactivation assay) to an external dose of 0.2% in a body lotion formulation. Conclusion: PBPK models for genistein and daidzein were developed as a central component of an NGRA read-across case study. This will help to gain regulatory confidence in the use of PBPK models, especially for cosmetic ingredients.

Stereo electroencephalography in the kingdom of Saudi Arabia.

Epilepsy surgery and intracranial monitoring have a long history in the Kingdom of Saudi Arabia, spanning over 30 years. Stereo-EEG however, is a more recent offering. In this short communication, we discuss how Stereo-EEG has grown in the context of the Kingdom's healthcare model and the Vision 2030 model. We discuss the various positives of this technique and methodology as well as the various challenges that the hospitals offering Stereo-EEG have faced.

Acquisition of metastatic tissue from patients with bone metastases from breast cancer.

Biopsies of metastatic tissue are increasingly being performed. Bone is the most frequent site of metastasis in breast cancer patients, but bone remains technically challenging to biopsy. Difficulties with both tissue acquisition and techniques for analysis of hormone receptor status are well described. Bone biopsies can be carried out by either by standard posterior iliac crest bone marrow trephine/aspiration or CT-guided biopsy of a radiologically evident bone metastasis. The differential yield of these techniques is unknown. Results from three prospective studies of similar methodology were pooled. Patients underwent both an outpatient posterior iliac crest bone marrow trephine/aspiration and a CT-guided biopsy of a radiologically evident bone metastasis. Samples were assessed for the presence of malignant cells and where possible also for estrogen (ER) and progesterone receptor (PgR) expression. 40 patients were enrolled. Bone marrow aspiration/trephine biopsy was completed in 39/40 (97.5%) and CT-guided biopsy was completed in 34/40 (85%) of patients. Sufficient tumor cells for hormone receptor analysis were available in 19/39 (48.8%) and 16/34 (47%) of and bone marrow aspiration/trephine and CT-guided biopsies, respectively. Significant discordance in ER and PgR between the primary and the bone metastasis was also seen. Nine patients had tissue available from both bone marrow and CT-guided bone biopsies. ER and PgR concordance between these sites was 100 and 78%, respectively. Performing studies on human bone metastases is technically challenging, with relatively low yields regardless of technique. Given resource issues and similar success rates when comparing both techniques, bone marrow examination may be utilized first and if inadequate tissue is obtained, CT-guided biopsies can then be used.

Epidemiology of heart failure and long-term follow-up outcomes in a north-African population: Results from the NAtional TUnisian REgistry of Heart Failure (NATURE-HF).

The NATURE-HF registry was aimed to describe clinical epidemiology and 1-year outcomes of outpatients and inpatients with heart failure (HF). This is a prospective, multicenter, observational survey conducted in Tunisian Cardiology centers. A total of 2040 patients were included in the study. Of these, 1632 (80%) were outpatients with chronic HF (CHF). The mean hospital stay was 8.7 ± 8.2 days. The mortality rate during the initial hospitalization event for AHF was 7.4%. The all-cause 1-year mortality rate was 22.8% among AHF patients and 10.6% among CHF patients. Among CHF patients, the older age, diabetes, anemia, reduced EF, ischemic etiology, residual congestion and the absence of ACEI/ ARBs treatment were independent predictors of 1-year cumulative rates of rehospitalization and mortality. The female sex and the functional status were independent predictors of 1-year all-cause mortality and rehospitalization in AHF patients. This study confirmed that acute HF is still associated with a poor prognosis, while the mid-term outcomes in patients with chronic HF seems to be improved. Some differences across countries may be due to different clinical characteristics and differences in healthcare systems.

Prognostic value of absent end-diastolic flow within the first week following renal transplantation.

BACKGROUND: Doppler sonogram of the graft is used as a routine assessment in renal transplantation. When the resistance index (RI) equals 1, absent end-diastolic flow (AEDF) is observed; the prognostic value of AEDF is presently unknown. PATIENTS AND METHODS: Between 1988 and 1996, 342 patients received a first cadaveric kidney transplant in our ward. AEDF was observed in 30 patients who were compared with 60 controls who showed an RI < 0.75 within the first 7 days after transplantation. They were matched for year of transplantation (+/-1 year); recipient age (+/-2 years); recipient sex; and HLA antibodies (3 classes: 0%, 1-75%, >75%). The follow-up was 4 years. RESULTS: AEDF was observed at day 1 in 64%, at day 3 in 96%, and at day 7 in 28%. Recipient age, donor age, recipient sex, cold and warm ischemia durations, HLA A, B, and DR mismatches, and cytomegalovirus (CMV) status were not different between the 2 groups. Immediate graft function and 3- to 24-month creatinine levels were better in the control than the AEDF group. However, there was no difference in serum creatinine at 3 and 4 years or in patient and graft survivals during follow-up. CONCLUSIONS: AEDF observed within the first week following transplantation is associated with impaired renal functional recovery. However, whether AEDF is a prognostic marker of poor long-term graft function or survival remains to be proven.

Impact of a polymorphism in the IL-12p40 gene on the outcome of kidney transplantation.

A number of factors interfere with the outcome of renal transplantation. Revealing genetic factors that impact on graft outcome may have consequences for clinical practice. Interleukin-12 (IL-12), by stimulating interferon gamma (IFNgamma) production, plays a crucial role in immune responses against both graft and viral agents. An A-to-C single nucleotide polymorphism (SNP) within the 3'-untranslated region (3'UTR) of the IL-12p40 gene has been reported to be both functionally and clinically relevant. Since the impact of this SNP on kidney graft outcome has never been reported, we investigated the impact of the 3'UTR polymorphism on clinical events after transplantation among 253 kidney recipients transplanted between 1995 and 2003. The polymorphism was genotyped using the restriction fragment length polymorphism method. Our results showed that the 3'UTR polymorphism affected neither graft survival (P = .768) nor the occurrence of delayed graft function (DGF; P = .498). C allele carriers in our study displayed more acute rejections in the first year than patients with the A/A genotype, but it did not reach statistical significance (P = .108). In contrast, the C allele appeared to be a significant risk factor for cytomegalovirus infection (odds ratio = 1.77; P = .027). In conclusion, IL12B 3'UTR polymorphism did not affect graft survival, DGF, or acute rejection episodes, but had an impact on the occurrence of cytomegalovirus infection.

[Cocaine-induced acute myocarditis]. / Myocardite aiguë induite par la cocaïne.

Cocaine is a potent sympathomimetic drug usually associated with cardiotoxicity, including ventricular arrhythmia, systemic hypertension and acute myocardial infarction. It constitutes the most frequent cause of drug-related death reported by medical examiners in the US, and these events are most often related to the cardiovascular manifestations of the drug. However; to the best of our knowledge; cocaine induced acute myocarditis has very rarely been reported. We describe the case of a 19 year-old male regular user of marijuana and cocaine who was admitted for a suspicion on an acute lateral-wall myocardial infarction and in whom the final diagnosis of acute cocaine myocarditis has been made. We report diagnosis modalities and evolution.

Extracellular NK histones promote immune cell anti-tumor activity by inducing cell clusters through binding to CD138 receptor.

BACKGROUND: Natural killer (NK) cells are important anti-tumor cells of our innate immune system. Their anti-cancer activity is mediated through interaction of a wide array of activating and inhibitory receptors with their ligands on tumor cells. After activation, NK cells also secrete a variety of pro-inflammatory molecules that contribute to the final immune response by modulating other innate and adaptive immune cells. In this regard, external proteins from NK cell secretome and the mechanisms by which they mediate these responses are poorly defined. METHODS: TRANS-stable-isotope labeling of amino acids in cell culture (TRANS-SILAC) combined with proteomic was undertaken to identify early materials transferred between cord blood-derived NK cells (CB-NK) and multiple myeloma (MM) cells. Further in vitro and in vivo studies with knock-down of histones and CD138, overexpression of histones and addition of exogenous histones were undertaken to confirm TRANS-SILAC results and to determine functional roles of this material transferred. RESULTS: We describe a novel mechanism by which histones are actively released by NK cells early after contact with MM cells. We show that extracellular histones bind to the heparan sulfate proteoglycan CD138 on the surface of MM cells to promote the creation of immune-tumor cell clusters bringing immune and MM cells into close proximity, and thus facilitating not only NK but also T lymphocyte anti-MM activity. CONCLUSION: This study demonstrates a novel immunoregulatory role of NK cells against MM cells mediated by histones, and an additional role of NK cells modulating T lymphocytes activity that will open up new avenues to design future immunotherapy clinical strategies.

Early pulse pressure and low-grade proteinuria as independent long-term risk factors for new-onset diabetes mellitus after kidney transplantation.

Risk factors for new-onset diabetes after transplantation (NODAT) need to be assessed in large cohorts. We retrospectively evaluated the impact of early (3 and 6 months after transplantation) proteinuria, urinary albumin excretion (UAE) and arterial pressure on NODAT in 828 Caucasian renal transplant recipients (median follow-up: 5.3 years; 5832 patient-years). The 10- and 20-year incidence of NODAT was 15.0% and 22.0%, respectively. Low-grade (<1 g/day) (HR: 2.04 [1.25-3.33], p = 0.0042) and very low-grade (<0.3 g/day) (HR: 2.21 [1.32-3.70], p = 0.0025) proteinuria were independent risk factors for NODAT. There was a dose-dependent relationship across UAE categories (increasing risk from normoalbuminuria to macroalbuminuria) with NODAT. Tacrolimus, sirolimus and beta-blockers (HR: 1.86 [1.07-3.22], p = 0.0277) were significantly associated with NODAT even after multiple adjustments, but not diuretics, angiotensin-converting enzyme inhibitors or angiotensin-receptor blockers. Systolic arterial pressure (HR per 10 mmHg: 1.16 [1.03-1.29], p = 0.0126) and pulse pressure (HR: 1.26 [1.12-1.43], p = 0.0002) were associated with NODAT. Only pulse pressure remained significant after adjustments. Patients at highest risks had early proteinuria and pulse pressure >60 mmHg. Early low-grade proteinuria and pulse pressure (in addition to beta-blockers) constitute independent risk factors for NODAT; they may be markers of the metabolic syndrome and/or vascular damage in renal transplant recipients.

Long-term results of monoclonal anti-Il2-receptor antibody versus polyclonal antilymphocyte antibodies as induction therapy in renal transplantation.

We compared the influence of induction therapy on 5-year patient and graft survival as well as on renal function in 100 kidney graft recipients at low immunological risk treated with antilymphocyte globulin (n = 50) versus anti-IL-2R monoclonal antibody (n = 50) in a prospective multicenter study. Long-term immunosuppressive treatment included cyclosporine, mycophenolate mofetil, and a short course of steroids in all patients. Five year graft (86% vs 86%) and patient (94% vs 94%) survivals were identical in both study arms. Moreover, neither serum creatinine or proteinuria were significantly different between the two groups. Our results showed that the choice of the induction therapy seemed to not have a major impact on long-term outcomes among renal recipients at low immunological risk.

Estimation of the glomerular filtration rate through different methods in kidney transplant recipients: correlation with the creatinine clearance measurement.

The measurement of the glomerular filtration rate (GFR) is an important tool for physicians to follow kidney transplant recipients. Indeed, renal function has been shown to be predictive of graft outcome in retrospective studies. Several methods have been proposed to measure GFR. In the present study we evaluated the correlation of GFR between a reference method (calculation through the urine to plasma ratio of creatinine [UV/P] formula) and three estimation equations (Cockcroft and Gault; Nankivell; modification of diet in renal disease) in 81 kidney transplant recipients at 3 and 12 months posttransplantation. We showed a significant correlation between the three predictive formulas and UV/P, but none of the predictive equations showed an excellent correlation. The best correlation between an estimation equation and the UV/P formula was the CG formula. Further studies are required to compare the estimated GFR with better reference methods, such as the use of isotopic markers in kidney graft recipients.

Early proteinuria is a strong indicator of donor renal lesions, ischemia-reperfusion injury and immunological aggression.

BACKGROUND: Early proteinuria is associated with reduced long-term graft survival. However, the determinants and mechanisms of proteinuria early after transplantation have not been identified. METHODS: Parameters associated with proteinuria within the first 3 months following transplantation were retrospectively assessed among 484 renal transplant recipients. RESULTS: Proteinuria was more abundant in patients with a history of two or more rejection episodes (0.42 +/- 0.68 vs 0.18 +/- 0.39 g/d; P = .02). Proteinuria was greater when donor age was 60 or more (OR: 4.43; P = .003), when recipient death was due to cardiovascular causes (OR: 1.98; P = .002), or when cold (OR: 1.77; P = .006) or warm (1.21; P = .09) ischemia times were prolonged. CONCLUSIONS: Proteinuria early after transplantation was related to pretransplant renal lesions, ischemia-reperfusion, and immunologic injuries.

Minimally invasive approach for the treatment of lumbar epidural arteriovenous fistulas with intradural venous reflux.

BACKGROUND: Vascular lesions of the spinal cord are increasingly recognized. The most common types of these lesions are dural arteriovenous fistulas (AVFs) whereas, extradural AVFs are a very rare type of spinal AVF and can be associated with either extradural or intradural venous reflux. This results in neurological deficits through congestive or compressive myeloradiculopathy. These lesions must be treated to allow stabilization or improvement of neurologic status, either by endovascular therapy or microsurgical interruption. However, because some patients are not amenable to endovascular treatment, surgery is often warranted, which usually involves hemi- or bilateral laminectomy following a midline approach with bilateral muscle stripping. The main drawback of this procedure is directly related to the morbidity of the approach. Although, minimally invasive approaches are likely to overcome this drawback, there is a lack of reported experience supporting their use for treating spinal dural AVFs. CASE PRESENTATION: Two patients, aged 62 and 79 years old, presented with rapidly progressive myelopathy characterized by paraparesis and sphincter disturbance. Spinal magnetic resonance imaging showed spinal cord oedema with perimedullary flow voids in both cases. Digital subtraction angiography revealed extradural AVFs associated with perimedullary venous reflux. Endovascular therapy was not feasible. Both patients were treated with microsurgical interruption of the intradural vein through a non-expendable retractor. Complete exclusion was confirmed on postoperative angiography, resulting in resolution of spinal cord edema and improved neurological functional status at 2-year follow-up. CONCLUSION: The minimally invasive surgical treatment of spinal AVFs with epidural venous reflux is safe and effective. This approach is a valuable alternative to endovascular therapy and the standard open microsurgical approach.

Hb Doha or alpha 2 beta 2[X-N-Met-1(NA1)Val----Glu]; a new beta-chain abnormal hemoglobin observed in a Qatari female.

Structural analysis of a fast-moving hemoglobin variant, present in three members of a Qatari family, identified a Val----Glu substitution at position 1 (NA1) of the beta-chain. The introduction of this glutamic acid residue prevents the removal of the initiator methionine, thus extending the N-terminus by one residue to Met-Glu-His-Leu-Thr-. The methionine residue is blocked by an as yet not completely identified molecule. The presence of the variant in a heterozygote does not have clinical consequences.