Applicability of a Chemiluminescence Immunoassay to Screen Postmortem Bile Specimens and Its Agreement with Confirmation Analysis.

Bile has emerged as an alternative matrix for toxicological investigation of drugs in suspected forensic cases of overdose in adults and intoxications in children. Toxicological investigation consists in screening and, subsequently, confirming the result with specific techniques, such as liquid chromatography with tandem mass spectrometry (LC-MS/MS). As there is no screening test on the market to test postmortem bile specimens, the novelty of this study was in investigating the applicability of a chemiluminescence immunoassay, designed for other matrices and available on the market, on bile and validate its use, testing the agreement with LC-MS/MS analysis. Bile specimens were obtained from 25 forensic cases of suspected death from overdose and intoxication. Sample preparation for bile screening consists simply in centrifugation and dilution. Confirmation analysis allows simultaneous identification of 108 drugs and was validated on bile. Kappa analysis assessed a perfect agreement (0.81-1) between the assays for benzodiazepines, methadone, opiates, cocaine, oxycodone, cannabinoids, buprenorphine and pregabalin; a substantial agreement (0.41-0.6) was reported for barbiturates. No agreement was assessed for amphetamines, due to an abundance of putrefactive amines in postmortem specimens. In conclusion, this fast and easy immunoassay could be used for initial screening of bile specimens, identifying presence of drugs, except amphetamines, with reliability.

Subcellular Effectors of Cocaine Cardiotoxicity: All Roads Lead to Mitochondria-A Systematic Review of the Literature.

Cocaine abuse is a serious public health problem as this drug exerts a plethora of functional and histopathological changes that potentially lead to death. Cocaine causes complex multiorgan toxicity, including in the heart where the blockade of the sodium channels causes increased catecholamine levels and alteration in calcium homeostasis, thus inducing an increased oxygen demand. Moreover, there is evidence to suggest that mitochondria alterations play a crucial role in the development of cocaine cardiotoxicity. We performed a systematic review according to the Preferred Reporting Items for Systemic Reviews and Meta-Analysis (PRISMA) scheme to evaluate the mitochondrial mechanisms determining cocaine cardiotoxicity. Among the initial 106 articles from the Pubmed database and the 17 articles identified through citation searching, 14 final relevant studies were extensively reviewed. Thirteen articles included animal models and reported the alteration of specific mitochondria-dependent mechanisms such as reduced energy production, imbalance of membrane potential, increased oxidative stress, and promotion of apoptosis. However, only one study evaluated human cocaine overdose samples and observed the role of cocaine in oxidative stress and the induction of apoptosis though mitochondria. Understanding the complex processes mediated by mitochondria through forensic analysis and experimental models is crucial for identifying potential therapeutic targets to mitigate or reverse cocaine cardiotoxicity in humans.

Resveratrol, a Multitasking Molecule That Improves Skeletal Muscle Health.

Resveratrol is a natural polyphenol utilized in Chinese traditional medicine and thought to be one of the determinants of the "French Paradox". More recently, some groups evidenced its properties as a calorie-restriction mimetic, suggesting that its action passes through the modulation of skeletal muscle metabolism. Accordingly, the number of studies reporting the beneficial effects of resveratrol on skeletal muscle form and function, in both experimental models and humans, is steadily increasing. Although studies on animal models confer to resveratrol a good potential to ameliorate skeletal muscle structure, function and performance, clinical trials still do not provide clear-cut information. Here, we first summarize the effects of resveratrol on the distinct components of the skeletal muscle, such as myofibers, the neuromuscular junction, tendons, connective sheaths and the capillary bed. Second, we review clinical trials focused on the analysis of skeletal muscle parameters. We suggest that the heterogeneity in the response to resveratrol in humans could depend on sample characteristics, treatment modalities and parameters analyzed; as well, this heterogeneity could possibly reside in the complexity of skeletal muscle physiology. A systematic programming of treatment protocols and analyses could be helpful to obtain consistent results in clinical trials involving resveratrol administration.

Evaluation of a Set of miRNAs in 26 Cases of Fatal Traumatic Brain Injuries.

In forensic medicine, identifying novel biomarkers for use as diagnostic tools to ascertain causes of death is challenging because of sample degradation. To that aim, a cohort (n = 26) of fatal traumatic brain injuries (TBIs) were tested for three candidate miRNAs (namely, miR-124-3p, miR-138-5p, and miR144-3p). For each case, three FFPE specimens (coup area (CA), contrecoup area (CCA), and the corpus callosum (CC)) were investigated, whereas the FFPE brain tissues of 45 subjects (deceased due to acute cardiovascular events) were used as controls. Relative quantification via the ∆∆Ct method returned significantly higher expression levels of the three candidate miRNAs (p < 0.01) in the TBI cases. No difference was detected in the expression levels of any miRNA investigated in the study among the CA, CCA, and CC. Furthermore, the analyzed miRNAs were unrelated to the TBI samples' post-mortem intervals (PMIs). On the contrary, has-miR-124-3p ahashsa-miR-144-3p were significantly correlated (p < 0.01) with the agonal time in TBI deaths. Since the RNA was highly degraded in autoptic FFPE tissues, it was impossible to analyze the mRNA targets of the miRNAs investigated in the present study, highlighting the necessity of standardizing pre-analytical processes even for autopsy tissues.

The Baron Pasquale Revoltella's Will in the Forensic Genetics Era.

In this article, we describe multiple analytical strategies that were first developed for forensic purposes, on a set of three bone samples collected in 2011. We analyzed a single bone sample (patella) collected from the artificially mummified body of the Baron Pasquale Revoltella (1795-1869), as well two femurs which allegedly belonged to the Baron's mother (Domenica Privato Revoltella, 1775-1830). Likely due to the artificial mummification procedures, the inner part of the Baron's patella allowed the extraction of high-quality DNA yields, which were successfully used for PCR-CE and PCR-MPS typing of autosomal, Y-specific, and mitochondrial markers. The samples extracted from the trabecular inner part of the two femurs yielded no typing results by using the SNP identity panel, whereas the samples extracted from the compact cortical part of the same bone samples allowed genetic typing, even by the employment of PCR-CE technology. Altogether, 10/15 STR markers, 80/90 identity SNP markers, and HVR1, HVR2, and HVR3 regions of the mtDNA were successfully typed from the Baron's mother's remains by the combined use of PCR-CE and PCR-MPS technologies. The kinship analysis showed a likelihood ratio of at least 9.1 × 106 (corresponding to a probability of maternity of 99.9999999%), and thus confirmed the identity of the skeletal remains as those of the Baron's mother. This casework represented a challenging trial for testing forensic protocols on aged bones samples. It highlighted the importance of accurately sampling from the long bones, and that DNA degradation is not blocked by freezing at -80 °C.

Liver pathology in COVID-19 related death and leading role of autopsy in the pandemic.

BACKGROUND: Information on liver involvement in patients with coronavirus disease 2019 is currently fragmented. AIM: To highlight the pathological changes found during the autopsy of severe acute respiratory syndrome coronavirus 2 positive patients. METHODS: A systematic literature search on PubMed was carried out until June 21, 2022. RESULTS: A literature review reveals that pre-existing liver disease and elevation of liver enzyme in these patients are not common; liver enzyme elevations tend to be seen in those in critical conditions. Despite the poor expression of viral receptors in the liver, it seems that the virus is able to infect this organ and therefore cause liver damage. Unfortunately, to date, the search for the virus inside the liver is not frequent (16% of the cases) and only a small number show the presence of the virus. In most of the autopsy cases, macroscopic assessment is lacking, while microscopic evaluation of livers has revealed the frequent presence of congestion (42.7%) and steatosis (41.6%). Less frequent is the finding of hepatic inflammation or necrosis (19%) and portal inflammation (18%). The presence of microthrombi, frequently found in the lungs, is infrequent in the liver, with only 12% of cases presenting thrombotic formations within the vascular tree. CONCLUSION: To date, the greatest problem in interpreting these modifications remains the association of the damage with the direct action of the virus, rather than with the inflammation or alterations induced by hypoxia and hypovolemia in patients undergoing oxygen therapy and decompensated patients.

Identification of 5F-Cumyl-PINACA, a Synthetic Cannabinoid, in the Herbal Material Used for Recreational Purposes in the Province of Trieste: Public Health Implications.

BACKGROUND: In recent years, the phenomenon of the production and trade of synthetic cannabinoids has grown, becoming a public health issue worldwide. The recent access- to the ED of the hospital of Trieste- of people who complained of episodes of hallucinations, sensation of poisoning, tachycardia, and air hunger following the inhalation of "Che Sballo platinum", have highlighted the need to perform further analysis on the contents of the packet sold as an air freshener, produced in Koper (Slovenia). OBJECTIVE: This paper wants to be an alert about the possible consequences on health due to the spreading of "Che Sballo platinum" in the province of Trieste. METHODS: The package contents were analyzed by a multi-target screening method of MRM-IDAEPI experiment. The result was then confirmed, and quantification was achieved via LC-ESI-MS/MS analysis in MRM mode using QTrap 6500 + Sinergy hydro column 100 x 2 mm 1.9 um transitions MRM1 368.3 → 250.0; MRM2 368.3 → 233.0. RESULTS: The initial screening tested negative for THC and showed positive results for 5F-Cumyl- PINACA. Quantitation result reported dose by the package of 8.5 mg of the compound. Formal notification was sent to the Italian Health Authorities (Notification No. 2021110205). CONCLUSION: Consumption of 5F-Cumyl-PINACA results in much more potent effects than marijuana. Lack of information about the actual concentration of the substance on the packaging does not allow drug users to have an adequate dosage, with possible toxic consequences on health. Further investigations must be done to discover the true extent of the phenomenon.

Recreational Cannabis Use: Regulatory Aspects, Public Opinion and Concerns for Public Health.

Cannabis represents the most used substance in the young population after alcohol and tobacco. The consumption of the substance undoubtedly represents an emerging public health issue, and the regulatory aspects are quite heterogeneous. Alternatives to drug prohibition, such as legalization or decriminalization, are a topic of growing debate among researchers, policy makers and the public. Limiting harmful use and combating illicit trafficking are the main objectives of legalization for recreational purposes, although liberalization raises critical issues that have not yet been resolved despite the experience gained in many countries. In the context of legalization, it is essential to allow the lawful production of cannabis, guarantee a supply of controlled quality cannabis, and ensure that the population understands the risks associated with the consumption of the substance. At the same time, it is essential to protect the safety and health of the population by limiting access to cannabis and preventing accidental exposure to fragile categories and children. Incomplete knowledge of law enforcement can create further complications. Incomplete knowledge of the implementation of the laws can create further complications; therefore adequate information must be favored. Scientific evidence helps to understand the potential harms and benefits of cannabis, but the impact of legalization - even in terms of reducing consumption - is not clearly documented. So, far more efforts need to be made in the future to broaden awareness of the general population on the recreational use of cannabis.

Sodium Nitrite Intoxication and Death: Summarizing Evidence to Facilitate Diagnosis.

BACKGROUND: Over the years, forensic pathology has registered the spread of new methods of suicide, such as the ingestion of sodium nitrite. Sodium nitrite causes increased methemoglobin, resulting in systemic hypoxia, metabolic acidosis, and cyanosis. Since sodium nitrite is a preservative, the ingestion of foods containing an excessive amount of this substance can also cause acute intoxication up to death. The present review is aimed at guiding health professionals in the identification and management of sodium-nitrite-related intoxications and deaths. METHODS: A systematic literature search was carried out on PubMed by following the PRISMA statement's criteria. A total of 35 studies with 132 cases were enrolled, and the data were cataloged in Microsoft Excel. To establish the causal correlation between sodium nitrite ingestion and death, the Naranjo Adverse Drug Reaction Probability Scale was used. RESULTS: In addition to the small number of cases that have currently been published, the study demonstrated that there was a general methodological discrepancy in the diagnostic process. However, some interesting results have emerged, especially in post-mortem diagnostics. CONCLUSION: Sodium-nitrite-related deaths represent a challenge for forensic pathologists; therefore, it is important to promptly recognize the essential features and perform the necessary and unrepeatable examinations for the correct diagnosis of the cause of death.

Acute Kidney Injury (AKI) in Young Synthetic Cannabinoids Abusers.

Background. Synthetic cannabinoid-related acute kidney injury represents an increasingly important public health issue due to the diagnostic challenges given by low clinical suspicion of the disease and the frequent undetectability in routine drug tests. Methods. A systematic literature search on PubMed was carried out until 31 January 2022. Case reports, case series, retrospective and prospective studies, as well as reviews on acute kidney injury related to the consumption of synthetic cannabinoid were searched. Results. The systematic review process selected 21 studies for a total of 55 subjects with synthetic cannabinoid-induced acute kidney injury. Renal damage was demonstrated by elevated serum creatinine levels in 49 patients (89%). On renal ultrasound, the most frequent finding was an increase in cortical echogenicity. Renal biopsy, performed in 33% of cases, revealed acute tubular damage, acute tubulointerstitial nephritis, and acute interstitial nephritis, in decreasing order of frequency. Conclusion. Prompt identification and treatment of synthetic cannabinoid-related acute kidney injury represent a sensitive public health goal both for the acute management of damage from synthetic cannabinoids and for the prevention of chronic kidney disease.

Spread of COVID-19 Infection in Long-Term Care Facilities of Trieste (Italy) during the Pre-Vaccination Era, Integrating Findings of 41 Forensic Autopsies with Geriatric Comorbidity Index as a Valid Option for the Assessment of Strength of Causation.

BACKGROUND: in 2020, a new form of coronavirus spread around the world starting from China. The older people were the population most affected by the virus worldwide, in particular in Italy where more than 90% of deaths were people over 65 years. In these people, the definition of the cause of death is tricky due to the presence of numerous comorbidities. OBJECTIVE: to determine whether COVID-19 was the cause of death in a series of older adults residents of nursing care homes. METHODS: 41 autopsies were performed from May to June 2020. External examination, swabs, and macroscopic and microscopic examination were performed. RESULTS: the case series consisted of nursing home guests; 15 men and 26 women, with a mean age of 87 years. The average number of comorbidities was 4. Based only on the autopsy results, the defined cause of death was acute respiratory failure due to diffuse alveolar damage (8%) or (31%) bronchopneumonia with one or more positive swabs for SARS-CoV-2. Acute cardiac failure with one or more positive swabs for SARS-CoV-2 was indicated as the cause of death in in symptomatic (37%) and asymptomatic (10%) patients. Few patients died for septic shock (three cases), malignant neoplastic diseases (two cases), and massive digestive bleeding (one case). CONCLUSIONS: Data from post-mortem investigation were integrated with previously generated Geriatric Index of Comorbidity (GIC), resulting in four different degrees of probabilities: high (12%), intermediate (10%), low (59%), and none (19%), which define the level of strength of causation and the role of COVID-19 disease in determining death.

Old and Promising Markers Related to Autophagy in Traumatic Brain Injury.

Traumatic brain injury (TBI) is one of the first causes of death and disability in the world. Because of the lack of macroscopical or histologic evidence of the damage, the forensic diagnosis of TBI could be particularly difficult. Considering that the activation of autophagy in the brain after a TBI is well documented in literature, the aim of this review is to find all autophagy immunohistological protein markers that are modified after TBI to propose a method to diagnose this eventuality in the brain of trauma victims. A systematic literature review on PubMed following PRISMA 2020 guidelines has enabled the identification of 241 articles. In all, 21 of these were enrolled to identify 24 markers that could be divided into two groups. The first consisted of well-known markers that could be considered for a first diagnosis of TBI. The second consisted of new markers recently proposed in the literature that could be used in combination with the markers of the first group to define the elapsed time between trauma and death. However, the use of these markers has to be validated in the future in human tissue by further studies, and the influence of other diseases affecting the victims before death should be explored.

"First Do No Harm". No-Fault Compensation Program for COVID-19 Vaccines as Feasibility and Wisdom of a Policy Instrument to Mitigate Vaccine Hesitancy.

Vaccines are so far proven to be safe, although related adverse events cannot be excluded. The urgency for COVID-19 vaccines determined a dilution of the general expectations of safety and efficacy of vaccination (from safe and effective to safe and effective enough). In many countries, a no-fault program was established to compensate individuals who experienced serious vaccine-related injuries. The impressive number of administrations worldwide and the legal indemnity afforded to manufacturers of approved vaccines that cannot be pursued for compensation fed the debate about the availability of a compensation model for COVID-19 vaccine-related injuries. Several European countries have long introduced a system, Vaccine Injury Compensation Programs, to compensate people who suffer physical harm because of vaccination. In Europe, COVID-19 vaccination is strongly recommended for the general population and in many states is declared mandatory for healthcare workers. In 1992, Italy edited Law no. 210 providing legal protection for individuals who reported injuries after mandatory and recommended vaccinations as a no-fault alternative to the traditional tort system. Despite its recommended nature, COVID-19 vaccination is excluded from the no-fault model in several European states, and the Italian government is called to provide clear and firm instructions for the management of the many requests for compensation. The authors provide an overview of the existing compensation models in Europe and analyse available legislative proposals.

Mors Gaudet Succurrere Vitae. The Role of Clinical Autopsy in Preventing Litigation Related to the Management of Liver and Digestive Disorders.

Over the last 50 years, the number of clinical autopsies has decreased, but their role in assessing cause of death and clinical performance is still acknowledged. Few publications have studied their role in malpractice claim prevention. The paper aims to highlight the role of clinical autopsy in preventing errors and improve healthcare quality. A retrospective study was conducted on 28 clinical autopsies performed between 2015 and 2021 on patients dead unexpectedly after procedures for the diagnosis and treatment of digestive and hepatic diseases. After an accurate analysis of medical records and consultation with healthcare professionals, all cases were subjected to autopsy and histopathology. The data obtained were analyzed and shared with the risk-management team to identify pitfalls and preventive strategies. Post-mortem evaluations confirmed the clinical diagnosis only in six cases (21.4%). Discordances were observed in 10 cases (35.7%). In the remaining 12 cases (42.9%) the clinical diagnosis was labeled as "unknown" and post-mortem examinations made it possible to document the cause of death. Post-mortem examinations can concretely enrich hospital prevention systems and improve patient safety. The methodological approach outlined certainly demonstrates that, even in the risk-management field, "mors gaudet succurrere vitae" ("death delights in helping life").

Spine surgery and fat embolism syndrome. Defining the boundaries of medical accountability by hospital autopsy.

BACKGROUND: Fat Embolism Syndrome (FES) is a clinical condition characterized by neurological, respiratory, hematological and cutaneous manifestations. Fatal FES has been described as a rare complication during or after spinal elective surgery. The investigation of the cause of death in fatalities related with spine surgery should be mandatory to exclude or confirm fat embolism; a detailed methodological approach to the body in these cases suggests to provide a cautious dissection of surgical site and collection of samples to detect embolized fat globules in vessels. METHODS: Two fatal cases of fat embolism syndrome after posterior spinal fusion are presented. CONCLUSIONS: A complete post mortem examination by means of histochemical and immunohistochemical analysis explained the cause of death and prevented medical malpractice litigation.