Solfeggio-frequency music exposure reverses cognitive and endocrine deficits evoked by a 24-h light exposure in adult zebrafish.

Music therapy has long been used as a non-pharmacological intervention to improve cognitive function and mood in humans. Mounting rodent evidence also supports beneficial impact of music exposure on animal cognitive performance. The zebrafish (Danio rerio) is an important emerging aquatic animal model in translational biomedical and neuroscience research. Here, we evaluate the effects of intermittent (2-h or 6-h twice daily) and continuous (24-h) solfeggio-frequency music exposure on behavioral, cognitive and endocrine parameters in adult zebrafish whose circadian rhythm was disturbed by a 24-h light exposure. Overall, a 24-h light exposure stress evokes overt cognitive deficits in the inhibitory avoidance test and elevates zebrafish whole-body cortisol levels. However, these effects were reversed by solfeggio-frequency music exposure for 2 or 6 h twice daily, and by continuous 24-h exposure. Collectively, these findings suggest a positive modulation of cognitive and endocrine responses in adult zebrafish by environmental enrichment via the long-term exposure to music, and reinforces zebrafish as a robust, sensitive model organism for neurocognitive and neuroendocrine research.

Enantiomeric methadone quantitation on real post-mortem dried matrix spots samples: Comparison of liquid chromatography and supercritical fluid chromatography coupled to mass spectrometry.

This study describes two bioanalytical methods for the quantitation of the two methadone enantiomers in dried matrix spots using high performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) and high performance supercritical chromatography tandem mass spectrometry (HPSFC-MS/MS). Dried matrix spots were obtained by spotting 10 µL of each sample fluid on a Whatman paper. Methadone and its main metabolite, EDDP, were extracted with 100 µL methanol and subsequently injected into the LC-MS/MS and SFC-MS/MS systems. Enantiomeric separation was achieved with AGP-column for the LC conditions and with Chiralpak IH-3 in SFC. The two methods were fully validated and 93 post-mortem samples were analysed with both analytical methods. Results from validation parameters and results obtained for all post-mortem samples were compared with a significant spearman correlation of rs = 0.9978 for R-methadone and rs = 0.9981 for S-methadone. The LC method provided better results in terms of uncertainty, retention factor and resolution, whereas SFC provides better sensitivity, with lower LOD. Median R-/S-methadone ratio in peripheral blood was found equal to 1.60 (N = 32), varying from 0.79 to 4.23. The reported values were in good agreement with previously published results. Based on the results obtained here, SFC-MS/MS can be considered a reliable alternative to the widely used LC-MS/MS for the quantitation of methadone enantiomers in bioanalysis and should be evaluated for other bioanalytical methods. Both methods can be easily and quickly used in toxicological routine analysis for the methadone quantitation in human fluids matrices, even if considering that the polysaccharide coated column IH-3 used in SFC does not allow the enantiomeric EDDP separation.

Development and validation of a method for quantification of human insulin and its synthetic analogues in plasma and post-mortem sera by LC-MS/HRMS.

Analysis of human insulin and its synthetic analogues is increasingly requested for clinical monitoring, for anti-doping purposes, but also for forensic cases. Indeed, insulin analogues may be abused for suicide or homicide - whence their forensic interest. Collection and storage conditions, as well as the phenomenon of degradation make post-mortem serum samples analytically challenging and consequently, the rate of exogenous insulin administration as cause of death is undoubtedly underestimated. However, with recent technological advances and the development of new extraction techniques particularly for anti-doping analyses, detection of insulins in post-mortem samples seems to be achievable. This study describes the first validated quantitative method for analysis human insulin and its six analogues (lispro, aspart, glulisine, glargine, detemir and degludec) in plasma and post-mortem sera. Various extraction processes, namely precipitation + solid phase extraction (SPE), filtration + SPE, precipitation + SPE + immunopurification, and filtration + immunopurification, were assessed to evaluate the lowest limit of detection for all target analogues. The selected sample preparation consists of filtration step followed by immunopurification extraction with an anti-body precoated ELISA plate for plasma. For post-mortem sera, the first step of precipitation was added to remove matrix interferences. The extracts were analyzed by ultra-high-performance liquid chromatography-high resolution mass spectrometry (LC-HRMS), interfaced by electrospray (ESI). The method was validated with respect linearity, precision, accuracy, recovery, matrix effect, dilution and carryover. The limit of quantification (LOQ) in plasma was 0.5 ng/mL for human insulin and rapid-acting insulins, 1.0 ng/mL for glargine, 2.5 ng/mL for degludec and 10 ng/mL for detemir. Two types of post-mortem sera were studied based on the post-mortem interval (PMI): inferior or superior to 48 h. The obtained LOQ were the same for each analogue, independent from the PMI: 1.0 ng/mL for human insulin and rapid-acting insulins, 1.0 ng/mL for glargine, 2.5 ng/mL for degludec and 10 ng/mL for detemir. At the LOQ level, for all insulins and all samples, accuracy was between 70 and 130% and precision inferior to 30%. The validated method was applied to five subjects participating in therapeutic monitoring of insulin and to seven post-mortem cases.

Correlations Between Intratumoral Microvessel Density and Histopathological Type or Neoadjuvant Radiotherapy for Rectal Carcinoma.

PURPOSE: This study aims to evaluate intratumoral microvessel density in rectal carcinoma cases with different histopathological type (adenocarcinoma and mucinous carcinoma) and different preoperatory neoadjuvant radiotherapy status (irradiated / non-irradiated) ,thus analyzing any possible statistical correlation between these parameters. MATERIAL AND METHODS: Our prospective study consists in standard immunohistochemistry procedures using CD34, CD31 and CD105 antibodies, which were performed on 25 samples of rectal carcinoma, in order to determine intratumoral microvessel density. RESULTS: The 25 case study group was divided either by histopathological type or by prior radiotherapeutical treatment as follows: 9 cases of mucinous carcinoma versus 16 cases of adenocarcinoma and 13 cases of rectal cancer that have not received neoadjuvant radiotherapy versus 12 cases of rectal cancer with preoperatory radiotherapy. CONCLUSIONS: The number of intratumoral microvessels is higher in non-irradiated rectal tumors and in adenocarcinomas, this remark being statistical significant (with only one exception - CD34 staining in non-irradiated versus irradiated tumors) for all types of vessels (new-grown and mature). This result is due to the benefic effect of neoadjuvant radiotherapy on decreasing angiogenic activity, thus having an important prognostic value for rectal cancer.

Evaluation of an in-capillary approach for performing quantitative cytochrome P450 activity studies.

An automated in-capillary assay requiring very small quantities of reagents was developed for performing in vitro cytochrome P450 (CYP450) drug metabolism studies. The approach is based on the following: (i) hydrodynamic introduction of nanoliter volumes of substrate and enzyme solutions in the sandwich mode, within a capillary; (ii) mixing the reagents by diffusion across the interfaces between the injected solutions; (iii) collection of the capillary content at the end of the in-capillary assay; and (iv) off-line analysis of the incubation mixture by ultrahigh pressure liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). After optimizing the injection sequence of the reagents, the in-capillary approach was applied to the quantitative determination of the kinetics of drug metabolism reactions catalyzed by three CYP450 isozymes involved in human drug metabolism: CYP1A2, CYP2D6, and CYP3A4. It was demonstrated that this in-capillary method was able to provide similar kinetic parameters for CYP450 activity (e.g., Michaelis constants and turnover values) as the classical in vitro method, with a drastic reduction of reagent consumption.

Trypsin immobilization on an ethylenediamine-based monolithic minidisk for rapid on-line peptide mass fingerprinting studies.

The aim of this work was to develop a trypsin-based micro-immobilized enzyme reactor prepared on a monolithic ethylenediamine BIA Separations CIM (convective interaction media) minidisk. The micro-immobilized enzyme reactor (IMER) was integrated in a liquid chromatography system hyphenated to electrospray ionization tandem mass spectrometry to carry out on-line protein digestion and identification. The performance of this IMER was compared with that obtained using a previously developed bioreactor prepared on a conventional CIM ethylenediamine disk and with that of the commercially available Poroszyme immobilized trypsin cartridge. In this work, we showed how different proteins were identified with good recoveries using a digestion time of 10 min only.

Development of immobilized enzyme reactors based on human recombinant cytochrome P450 enzymes for phase I drug metabolism studies.

Two cytochrome P450 (CYP)-based immobilized enzyme reactors (IMERs) were developed to perform automated on-line phase I drug metabolism studies. For this purpose, biotinylated recombinant CYP2D6 or CYP3A4 reconstituted systems were anchored to the surface of two monolithic mini-columns (2 mm x 6 mm I.D.), which had been covalently grafted with NeutrAvidin. After optimization of immobilization conditions, the obtained IMERs were integrated on-line into a LC hyphenated to an electrospray ionization MS/MS system. Studies with probe substrates and a known competitive inhibitor were performed, showing the potential of CYP-based IMERs in drug metabolism. In the optimized conditions, ca. 15 experiments were carried out with each bioreactor.

Trypsin immobilization on three monolithic disks for on-line protein digestion.

The preparation and characterization of three trypsin-based monolithic immobilized enzyme reactors (IMERs) developed to perform rapid on-line protein digestion and peptide mass fingerprinting (PMF) are described. Trypsin (EC 3.4.21.4) was covalently immobilized on epoxy, carbonyldiimidazole (CDI) and ethylenediamine (EDA) Convective Interaction Media (CIM) monolithic disks. The amount of immobilized enzyme, determined by spectrophotometric measurements at 280nm, was comprised between 0.9 and 1.5mg per disk. Apparent kinetic parameters Km* and Vmax*, as well as apparent immobilized trypsin BAEE-units, were estimated in flow-through conditions using N-alpha-benzoyl-L-arginine ethyl ester (BAEE) as a low molecular mass substrate. The on-line digestion of five proteins (cytochrome c, myoglobin, alpha1-acid glycoprotein, ovalbumin and albumin) was evaluated by inserting the IMERs into a liquid chromatography system coupled to an electrospray ionization ion-trap mass spectrometer (LC-ESI-MS/MS) through a switching valve. Results were compared to the in-solution digestion in terms of obtained scores, number of matched queries and sequence coverages. The most efficient IMER was obtained by immobilizing trypsin on a CIM EDA disk previously derivatized with glutaraldehyde, as a spacer moiety. The proteins were recognized by the database with satisfactory sequence coverage using a digestion time of only 5min. The repeatability of the digestion (R.S.D. of 5.4% on consecutive injections of myoglobin 12microM) and the long-term stability of this IMER were satisfactory since no loss of activity was observed after 250 injections.

[Vaginal environment. The vaginal climate and its variations]. / La météorologie vaginale. L'atmosphère vaginale et ses variations.

Variations in the vaginal environment that occur throughout the life span are described. The nature and cybernetics of factors involved in these variations are discussed.

[Vaginal cytology in eating disorders in young girls]. / La cytologie vaginale dans les troubles de l'oralité chez la jeune fille.

Critical review of eating disorders in girls, based upon five cases:four of anorexia nervosa and one of boulimia, such problems neing accompanied from a gynecologic standpoint by amenorrhea. Interpretation of colpocytologic findings, differing between cases. Suggestion of the central nervous origin of these disorders.

[Biochemistry of Eros]. / Biochimie de l'Eros.

Analytic study of the biochemical factors in the process of pareunia: hormonal factor of embryonic genital crests, responsible for a double sexualization, both genital and encephalic, which stands for the exteriorization of a perfectly determined later behavior; attractive or repulsive exoactones (pheromones) which play a major part in inter-individual attraction. Study of those many exoactones in both sexes; hormonomediation, leading to the intervention of the hypotalamo-adeno-hypophyso-(cortico)-gonadal axis, with such effectors as sexosteroids, the hypothalamo-adenohypophyseal with the prolactin, the neuro-hypopheseal level (vasopressin, ocytocyne), and the epiphyseal level (melatonin), the terminal effector agents appearing either as libido (androgens, P-Rih), or counter-libido (oestrogens, prolactin) with the involvement of two systems of vertical and horizontal libidinization; neuromediation, with the intervention of the catecholaminergic, cholinergic, indolaminergic (serotonin); neuromodulation, with a rise in the endorphin rate when there is an orgasm; very precise enzymatic phenomena (depolymerases appearing in the vaginal content when there is an orgasm), which cannot be ignored any longer.

[Female orgasmic nodules]. / Les nodules orgastiques féminins.

During experimental female orgasm (or illation), the prominence of five stages which correspond to the orgastic raise. This raise leads to the apparition of very characteristic nodules, which are structured as they group desquamed vaginal cells together with transuded blood cells. Furthermore, the loss of pseudocrystallization of the ovulatory cervical phlegm is to be noted ("fern type crystallization"). An enzymatic explanation for these phenomena, which lead to the intervention of hyaluronidases, may be proposed.

[Vaginal ecology, climate, landscapes and populations (xenoecies and facies)]. / L'écologie vaginale, climat, paysages et peuplements (xénoecies et faciès).

Brief study of vaginal populations, the human vagina being considered as a biotopic cavity. Allusion to dynamic aspects ("vaginal climate", "landscapes") and to various bacterial populations. Introduction of the concept of xenoecies and of facies. This study is preceded by essential definitions of terms widely used in ecology.

[Bacterio-parasitic interactions. Enterobacteria and schistosomes (Salmonella - schistosome association)]. / Interactions bactério-parasitaires. Enterobactéries et schistosomidés (salmonello-schistosomo-synusies).

The description of a case imported from Chad of an association of Salmonella typhi - Schistosoma haematobium and, in this connection, a new revised general account. The association Salmonella-Schistomas is wide-spread. It corresponds to a salmonello - schistosomo - micro - association by the fixation of precise bacteria in the case of a bacteriemia on the cutaneous surface of male schistosomes in clearly defined places. These Salmonella constitute a permanent antigenic solicitation and they are capable of mobilization and rejection (bacteriuria after passage through the blood-stream) whereas bacteriosis remains asymptomatic. In fact, on the practical level, a clear and absolute distinction must be made between:--the bacteriemia with the concomitant bacteriuria, which are largely physiological in nature, very frequent and originating from various germs, connected with the emunctory function of the kidneys and which can be observed in any subject.--the schistosomal bacteriuria which are specific entities and evidence of a veritable micro-association of a symbiotic nature between the Salmonella and the schistosome, a micro-association which the bacteriuria reveals unexpectedly whereas the clinical symptoms are absent, not clearly discernible or totally misleading.