Semaglutide and health risk: Development and validation of a LC-HRMS method for testing semaglutide in whole blood and application to real cases.

The use of semaglutide, also known by its trade name Ozempic®, has been increasing worldwide in recent years due to its benefits in treating type II diabetes. Thanks to its effects on appetite regulation, in many countries it is also used to treat obesity. However, due to its promotion by social media and celebrities as a weight-loss treatment, semaglutide is misused by a non-diabetic and non-obese population and by a young public, which is the main target of these media. Following the alert by the ANSM (Agence nationale de sécurité du médicament) in France and the FDA (Food and Drug Administration) in the United States, which imposed the addition of fatal effects to the list of side effects, the misuse of semaglutide seems to be becoming a public health problem. For this reason, it seems important that a toxicology laboratory has the capacity to test for semaglutide in blood. In this study, the authors have developed and validated a method for the identification and quantification of semaglutide in whole blood using a LC-HRMS. After the addition of the internal standard (bovine insulin), the blood was subjected to protein precipitation using a mix of acetonitrile/methanol (70:30,v:v). The validation procedure demonstrated an acceptable linearity between 2 and 500 ng/mL. LOD and LOQ were 1 and 2 ng/mL, respectively. Intra and inter-day precision were below 20 % at three concentrations. The method was successfully applied to the blood samples of 3 diabetic patients under treatment of semaglutide. The samples tested positive with concentrations ranging from 31 to 70 ng/mL which fall within the limits of therapeutic blood concentrations described in the literature.

Bariatric surgery and changes in metformin distribution: About a fatal case.

A 65 year-old woman, suffering from mellitus type 2 diabetes and obesity, died at home, three days after bariatric surgery (Roux-en-Y gastric by-pass: RYGB). Her treatment, including metformin and dapagliflozin, was stopped before surgery and not postoperatively reinstalled. A forensic autopsy, toxicological and histological analyses were performed. No macroscopic or microscopic evidence of digestive perforation or peritonitis was identified, excluding an early surgery complication as the cause of death. Toxicological analysis revealed the presence of Metformin in all matrices tested, with a potentially fatal blood concentration. Death was attributed to lactic acidosis caused by a metformin overdose. With no evidence for suicide by ingestion of metformin, the authors supposed that the bariatric surgery might have caused changes in the absorption of metformin, leading to a rapid overdose and death. The only study in the literature on this subject, demonstrated a significant increase in the bioavailability of metformin following oral administration in gastric bypass patients. Thus, it can be anticipated that a therapeutic dose can become toxic when administrated to a subject who recently modified her digestive equipment. As this represents the first case of metformin overdose following bariatric surgery, further cases will be needed to confirm our initial observations.

The Effects of Probiotics, Prebiotics and Synbiotics in Non-Alcoholic Fat Liver Disease (NAFLD) and Non-Alcoholic Steatohepatitis (NASH): A Systematic Review.

Modifications in the microbiota caused by environmental and genetic reasons can unbalance the intestinal homeostasis, deregulating the host's metabolism and immune system, intensifying the risk factors for the development and aggravation of non-alcoholic fat liver disease (NAFLD). The use of probiotics, prebiotics and synbiotics have been considered a potential and promising strategy to regulate the gut microbiota and produce beneficial effects in patients with liver conditions. For this reason, this review aimed to evaluate the effectiveness of probiotics, prebiotics, and symbiotics in patients with NAFLD and NASH. Pubmed, Embase, and Cochrane databases were consulted, and PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analysis) guidelines were followed. The clinical trials used in this study demonstrated that gut microbiota interventions could improve a wide range of markers of inflammation, glycemia, insulin resistance, dyslipidemia, obesity, liver injury (decrease of hepatic enzymes and steatosis and fibrosis). Although microbiota modulators do not play a healing role, they can work as an important adjunct therapy in pathological processes involving NAFLD and its spectrums, either by improving the intestinal barrier or by preventing the formation of toxic metabolites for the liver or by acting on the immune system.

The Effects of Curcumin on Diabetes Mellitus: A Systematic Review.

Diabetes mellitus (DM) is an ensemble of metabolic conditions that have reached pandemic proportions worldwide. Pathology's multifactorial nature makes patient management, including lifelong drug therapy and lifestyle modification, extremely challenging. Currently, there is growing evidence about the effectiveness of using herbal supplements in preventing and controlling DM. Curcumin is a bioactive component found Curcuma longa, which exhibits several physiological and pharmacological properties such as antioxidant, anti-inflammatory, anticancer, neuroprotective, and anti-diabetic activities. For these reasons, our objective is to systematically review the effects of Curcuma longa or curcumin on DM. Databases such as PUBMED and EMBASE were searched, and the final selection included sixteen studies that fulfilled the inclusion criteria. The results showed that curcumin's anti-diabetic activity might be due to its capacity to suppress oxidative stress and inflammatory process. Also, it significantly reduces fasting blood glucose, glycated hemoglobin, and body mass index. Nanocurcumin is also associated with a significant reduction in triglycerides, VLDL-c, total cholesterol, LDL-c, HDL-c, serum C reactive protein, and plasma malonaldehyde. Therefore, it can be considered in the therapeutic approach of patients with DM.

First identification of a diuretic, hydrochlorothiazide, in hair: Application to a doping case and interpretation of the results.

An athlete contested an adverse analytical finding involving hydrochlorothiazide, and requested our laboratory for testing his hair. As there is no reference in the literature about identification of hydrochlorothiazide in hair, several volunteers were first enrolled (4 after a single 25 mg administration and 10 with daily therapeutic treatment). A specific method was developed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS. Hair samples were decontaminated with dichloromethane and 30 mg were incubated in buffer at pH 7.0 for 15 hours at 50°C. Then, 5 mL ethyl acetate was added for extraction. Linearity was observed for hydrochlorothiazide concentrations ranging from 5 to 2000 pg/mg. The limit of quantification was 5 pg/mg. The coefficients of variation (CVs) of repeatability and matrix effect were lower than 20%. Analysis of the 0-2-cm segment of the 4 volunteers having received a single dose, collected 1 month after administration, was negative at the limit of quantification. The hair of the 10 patients (proximal 2 cm) on daily treatment was positive with concentrations ranging from 12 to 1845 pg/mg, with no correlation between daily dose and concentration. The athlete's hair tested positive for hydrochlorothiazide at 36 pg/mg in the segment corresponding to the period of the urinary control. Since a single exposure to hydrochlorothiazide is not detectable in hair and based on the results of the patients on daily treatment, the concentration found in the athlete has been interpreted as corresponding to repeated exposures, where it was not possible to establish the dosage and the frequency.

Whiplash-Associated Disorders : Clinical and medico-legal guidelines on the methods of ascertainment.

The manuscript presents the International Guidelines developed by the Working Group on Personal Injury and Damage under the patronage of the International Academy of Legal Medicine (IALM) regarding the Methods of Ascertainment of any suspected Whiplash-Associated Disorders (WAD).The document includes a detailed description of the logical and methodological steps of the ascertainment process as well as a synoptic diagram in the form of Flow Chart.

Targeted next generation sequencing application in cardiac channelopathies: Analysis of a cohort of autopsy-negative sudden unexplained deaths.

Genetic testing for cardiac channelopathies in sudden unexplained death (SUD) has developed substantially over the last years. The Next Generation Sequencing (NGS) technology provides an unprecedented opportunity to screen for genetic variations underlying arrhythmogenic genes in a short period of time at a low cost. The present study aimed to perform genetic testing with NGS technologies on the Ion Torrent Personal Genome Machine™ (Ion PGM™) sequencer, in targeting a total of 23 genes reported to be associated with inherited cardiac channelopathies in order to identify the possible cause of death in a cohort of post-mortem cases. The molecular analyses focused on 16 cases of SUD, aged less than 35 years old. In all cases, the cause of death could not be determined after a rigorous autopsy associated with histopathological and toxicological analyses according to the guidelines of the Association for European Cardiovascular Pathology. DNA was extracted from fresh frozen tissue. An average of 200 variants was identified per case. However, after the prioritization process using a new scoring program (VaRank) and after the conjunction of clinical data and molecular findings, four "likely pathogenic" variants (including two undescribed variants), were identified in three cases (18.75%) of our cohort in the genes KCNH2, ANK2, SCN5A and RYR2. One case, who died during psychiatric hospitalization after administration of a QT prolonging drug, showed a double "likely pathogenic" variant in Long QT genes (ANK2 and SCN5A) which may have predisposed to drug-induced cardiac arrhythmias. Our study illustrates that the NGS approach based on AmpliSeq™ libraries and Ion Torrent PGM™ sequencing may be an efficient approach, integrated to post-mortem examination. Given the massive amount of information generated by NGS, a rigorous filtration strategy of variants coupled with multidisciplinary collaboration is crucial to determine the potential pathogenic role of identified variants in the cause of death.

[Whiplash and oculomotor disorders: The pet peeve of the experts ]. / Coup de lapin (whiplash) et déséquilibres oculomoteurs : la bête noire des experts .

Whiplash is a lay term combining all the complications of a cervical sprain or distortion associated with an extension. It may lead to cervical or back pain, but patients may also complain about functional visual impairment. These visual complaints: fixation disparity, heterophoria or convergence insufficiency, are frequently neglected and difficult to quantitate and treat. Establishing the relationship between the indirect cervical trauma and the visual signs may be challenging for the medical-legal expert and must include consideration of the previous oculomotor status of the patient.

3D-MSCT imaging of bullet trajectory in 3D crime scene reconstruction: two case reports.

Postmortem investigations are increasingly assisted by three-dimensional multi-slice computed tomography (3D-MSCT) and have become more available to forensic pathologists over the past 20years. In cases of ballistic wounds, 3D-MSCT can provide an accurate description of the bullet location, bone fractures and, more interestingly, a clear visual of the intracorporeal trajectory (bullet track). These forensic medical examinations can be combined with tridimensional bullet trajectory reconstructions created by forensic ballistic experts. These case reports present the implementation of tridimensional methods and the results of 3D crime scene reconstruction in two cases. The authors highlight the value of collaborations between police forensic experts and forensic medicine institutes through the incorporation of 3D-MSCT data in a crime scene reconstruction, which is of great interest in forensic science as a clear visual communication tool between experts and the court.

Prevalência e fatores de risco de leptospirose bovina no Estado do Maranhão / Prevalence and risk factors of bovine leptospirosis in the State of Maranhão, Brazil

Foram investigados a prevalência e os fatores de risco da leptospirose bovina no Estado do Maranhão. O Estado foi dividido em quatro circuitos amostrais com base em parâmetros de produção distintos que variam conforme os diferentes sistemas de produção, as práticas de manejo, a finalidade de exploração, o tamanho médio dos rebanhos e os sistemas de comercialização. Objetivou-se estudar as características epidemiológicas da leptospirose bovina no Estado do Maranhão, de modo a determinar a prevalência em bovinos e em rebanhos, detectar as sorovariedades de Leptospira spp. presentes, identificar os fatores de risco eventualmente associados à leptospirose em bovinos e diferenciar os circuitos pecuários entre si no que se refere à prevalência de leptospirose. A pesquisa foi realizada em 136 propriedades rurais pertencentes ao circuito I, no qual 841 fêmeas bovinas com idade igual ou superior a 24 meses foram analisadas; 238 do circuito II, com 2.582 fêmeas analisadas; 122 do circuito III, com 869 fêmeas analisadas; e 77 do circuito IV, com 540 fêmeas analisadas; no total, 573 propriedades e 4.832 fêmeas foram estudadas. A presença de anticorpos contra Leptospira spp. foi verificada pela técnica de soroaglutinação microscópica (SAM). Das 4.832 fêmeas bovinas analisadas, 1.904 (35,94%; IC 95% = 33,01% - 38,98%) foram reagentes. Das 573 propriedades analisadas, 380 (64,81%; IC 95% = 61,10% - 68,35%) foram consideradas positivas. As sorovariedades Hardjo e Wolffi foram as mais frequentes em todo o Estado. O circuito III foi o que apresentou menor prevalência de leptospirose em todas as comparações. As variáveis identificadas como fatores de risco de leptospirose foram: presença de equinos (p = 0,000), presença de capivaras (p = 0,034) e rebanhos bovinos com 32 ou mais fêmeas adultas (p = 0,002).

Prevalence and risk factors of bovine leptospirosis in the State of Maranhão were investigated. Based on production parameters that vary across different production systems, management practices, the purpose of exploitation, the average size of herds and market systems, the state was divided in four sampling circuits. The study aimed to investigate the epidemiological features of bovine leptospirosis in the State of Maranhão, in order to determine the prevalence of the infection in cattle and herds, to determine the occurrence of serovars of Leptospira spp., to identify risk factors associated with leptospirosis in cattle and to differentiate the livestock circuits itself regarding the prevalence of leptospirosis. The survey was conducted in 136 herds in the circuit I, in which 841 > 24 months old females were analyzed; 238 in the circuit II and 2,582 females were analyzed; 122 in the circuit III and 869 females were analyzed; 77 in the circuit IV and 540 females were analyzed; a total of 573 herds and 4,832 females were analyzed. The presence of antibodies against Leptospira spp. was verified by microscopic agglutination test (MAT). Of the 4,832 cows examined, 1,904 (35.94%, CI 95% = 33.01% - 38.98%) were positive. Of the 573 herds, 380 (64.81%, CI 95% = 61.10% - 68.35%) were positive. Serovars Hardjo and Wolffi were the most frequent in the state. The circuit III showed the lowest prevalence of leptospirosis in all comparisons. The variables presence of horses (p = 0.000), presence of capybaras (p = 0.034) and herds with up to 32 adult females (p = 0.002) were identified as risk factors for leptospirosis.

Towards child versus adult brain mechanical properties.

The characterization of brain tissue mechanical properties is of crucial importance in the development of realistic numerical models of the human head. While the mechanical behavior of the adult brain has been extensively investigated in several studies, there is a considerable paucity of data concerning the influence of age on mechanical properties of the brain. Therefore, the implementation of child and infant head models often involves restrictive assumptions like properties scaling from adult or animal data. The present study presents a step towards the investigation of the effects of age on viscoelastic properties of human brain tissue from a first set of dynamic oscillatory shear experiments. Tests were also performed on three different locations of brain (corona radiata, thalamus and brainstem) in order to investigate regional differences. Despite the limited number of child brain samples a significant increase in both storage and loss moduli occurring between the age of 5 months and the age of 22 months was found, confirmed by statistical Student's t-tests (p=0.104,0.038 and 0.054 for respectively corona radiata, thalamus and brain stem samples locations respectively). The adult brain appears to be 3-4 times stiffer than the young child one. Moreover, the brainstem was found to be approximately 2-3 times stiffer than both gray and white matter from corona radiata and thalamus. As a tentative conclusion, this study provides the first rheological data on the human brain at different ages and brain regions. This data could be implemented in numerical models of the human head, especially in models concerning pediatric population.

Destabilization and intracranial fragmentation of a full metal jacket bullet.

We report a case with an atypical entrance wound as a result of a destabilized full metal jacket bullet penetration. The destabilized bullet by an impact with the dorsal hand experiences a yawing to tumbling motion in flight. The large angle of yaw induces a larger presenting profile upon impact that contributes, associated to a rapid deceleration, to a greater mechanical force on the projectile structure and a fragmentation into core and jacket. Forensic pathologists have to be aware that the metal jacket bullet could tend to break up outside or inside the body particularly after a shooting through a target. This phenomenon induces atypical entrance wounds and atypical X-ray presentation.

Human Neck Finite Element Model Development and Validation against Original Experimental Data.

This study proposes a detailed FEM of a human volunteer's neck and proceeds to an original model validation against experimental data recorded with this human volunteer. In order to evaluate the new model against existing data a successful temporal validation of the model was obtained under frontal, lateral, oblique and rear impact. New validation parameters are based on an experimental test proceeded in the frequency domain in order to extract the volunteer's Head-Neck system's modal characteristics. In depth validation of the head neck FEM is then performed by superposing the numerical and experimental frequency response function. Model optimisation in the frequency domain permitted after significant properties modification to reproduced accurately both, the neck extension mode at 1.4 Hz and the head retraction mode at 8.8 Hz. Finally the "frequency domain optimised" FEM response was superimposed with the temporal corridors provided in the literature. It must be mentioned that the model's response in the temporal domain remains inside existing corridors after this model optimisation in the frequency domain illustrating that the temporal validation is not accurate enough. This study proposes a neck model with improved geometry description and biofidelity with special attention paid to the retraction mode, a phenomenon which is often masked in the temporal domain.

[Use of polyester urethane (Neuro-Patch) as a dural substitute. Prospective study of 70 cases]. / Utilisation du polyester urethane (Neuro-Patch) comme substitut dural. Etude prospective sur 70 cas.

Between October 1995 and March 1998, 70 patients were treated with a microporous polyester urethane dura substitute (Neuro-Patch), after brain or spinal surgery. These patients were assessed clinically and radiologicaly 10 days, 6 weeks and 1 year after surgery. Radiological evaluation used CT scan or MRI. All dura substitutes were fixed by continuous suture to the surrounding dura-mater. We studied the handling properties, the incidence of infection and of CSF leakage. Eleven patients underwent craniotomy again. This gave us the opportunity to examine the adhesion to the brain tissue and the integration of the dura substitute. Six sheets underwent histological examination. Our results show good handling properties of the material; 3 infections; 6 out of 9 radiological CSF leakage occurred from infratentorial surgery. During reoperation, no adhesion to the brain tissue or injury to the brain while detaching the dura substitute was noticed. An excellent histological integration was observed: pores of the Neuro-Patch were colonized by fibroblasts synthesizing collagen, and there was no immune or inflammatory reaction, with an actual 4 to 6 years follow-up. A Neuro-Patch can therefore be recommended as a dura substitute to repair spinal or cranial dural defects.

Determination of blood cyanide by HPLC-MS.

An original high-performance liquid chromatographic-mass spectrometric (HPLC-MS) procedure was developed for the determination of cyanide (CN) in whole blood. After the addition of K13C15N as internal standard, blood was placed in a microdiffusion device, the inner well of which was filled with a mixture of taurine (50mM in water)/naphthalene-2,3-dicarboxaldehyde (NDA, 10mM in methanol)/methanol/ concentrated (approximately 20%) ammonia solution (25:25:45:5, v/v). Concentrated H2SO4 was added to the blood sample, and the microdiffusion chamber was sealed. After 30 min of gentle agitation, 2 microL of the contents of the inner vial were pipetted and directly injected onto a NovaPak C18 HPLC column. Separation was performed by a gradient of acetonitrile in 2mM NH4COOH, pH 3.0 buffer (35-80% in 10 min). Detection was done with a Perkin-Elmer Sciex API-100 mass analyzer with an ionspray interface, operated in the negative ionization mode. MS data were collected as either TIC or SIM at m/z (299 + 191) and (301 + 193) for the derivatives formed with CN and 13C15N, respectively. Inspired by previous works dealing with the complexation of CN by NDA + taurine to form a 1-cyano [f] benzoisoindole derivative analyzed by HPLC-fluorimetry, this method appears simple, rapid, and extremely specific. Limits of detection and quantitation for blood CN are 5 and 15 ng/mL, respectively. The use of 13C15N as internal standard allows the quantitation of CN with elegance and accuracy in comparison with previously reported methods.

Fatal overdosage with sildenafil citrate (Viagra): first report and review of the literature.

A fatal case of sildenafil citrate (Viagra) overdosage is presented. The deceased was a 56-year old male found dead at home, with a past history of diabetes mellitus, hypertension, chronic alcoholism, anxio-depressive disorders, and erectile dysfunction. The main autopsy findings were cardiomegaly (650 g) with dilated cardiomyopathy, diffuse coronary atherosclerosis with no sign of acute ischaemic disease, and extensive fibrosis of the myocardium, especially affecting the cardiac conducting tissue. As measured by HPLC/MS, sildenafil concentration in postrmortem blood (6.27 microg/mL) exceeded at least four times the highest therapeutic levels previously reported. The results are discussed in the light of the literature about the cardiovascular side effects of sildenafil, with special emphasis on the recently evidenced arrhythmogenic potential of the drug. This is the first report of a fatality caused by sildenafil overdosage.