Search

Ethanol blood analysis is the most common request in forensic toxicology, and some studies point to positive results in approximately one-third of all unnatural deaths. However, distinguishing sober deaths from drunk deaths is not as simple as it may seem. This technical, clinical, and forensic interpretation is proposed to interpret the ethanol toxicological results, discussing several artefacts and pitfalls that must be considered, namely focusing on driving under the influence. This work is presented with a practical and objective approach, aiming to alleviate the complexities associated with clinical, physiological, pathophysiological, and toxicological aspects to enhance comprehension, practicality, and applicability of its content, especially to courts. Particularly the physical integrity of the body, the postmortem interval, putrefactive signs, anatomic place of blood collection, alternative samples such as vitreous humour and urine, the possibility of postmortem redistribution, the inclusion of preservatives in containers, and optimal temperature conditions of shipment are among some of the aspects to pay attention. Although several biomarkers related to postmortem microbial ethanol production have been proposed, their translation into forensic routine is slow to be implemented due to the uncertainties of their application and analytical difficulties. Specifically, in the interpretation of ethanol toxicological results, "not everything that can be counted counts and not everything that counts can be counted" (attributed to Albert Einstein). [ABSTRACT FROM AUTHOR]

Background: The Post-mortem interval is the time between death and the examination of a corpse. It is important to know when the crime was committed. It helps the police to start their investigations with the information available and to process cases more efficiently. Materials and Methods: The sample of vitreous homour was collected from the posterior chamber of both eyes by aspirating gradually and slowly through a puncture 5-6 mm away from the limbus using a sterile 20-gauge needle taking care to avoid tearing of any loose tissue fragments surrounding the vitreous chamber. Results: In the present study there were 80 subjects, out of these subjects' the maximum subjects were from 26-35 years and the minimum were from 66-75 years of age group. Conclusion: Our study found that an appreciable pattern of increased potassium concentration in vitreous humour was observed every 12 hours of post-mortem interval, with an approximate 2mmol/l increase. [ABSTRACT FROM AUTHOR]

Abstract Background This study aimed to explore differences in vitreous humour metabolites and metabolic pathways between patients with and without diabetic retinopathy (DR) and identify potential metabolite biomarkers. Methods Clinical data and vitreous fluid samples were collected from 125 patients (40 without diabetes, 85 with DR). The metabolite profiles of the vitreous fluid samples were analysed using ultra-high performance liquid chromatography, Q-Exactive mass spectrometry, and multivariate statistical analysis. A machine learning model based on Least Absolute Shrinkage and Selection Operator Regularized logistic regression was used to build a risk scoring model based on selected metabolite levels. Candidate metabolites were regressed to glycated haemoglobin levels by a logistic regression model. Results Twenty differential metabolites were identified between the DR and control groups and were significantly enriched in five Kyoto Encyclopedia of Genes and Genomes pathways (arginine biosynthesis; tricarboxylic acid cycle; alanine, aspartate, and glutamate metabolism; tyrosine metabolism; and D-glutamate metabolism). Ferrous ascorbate significantly contributes to poorer glycaemic control outcomes, offering insights into potential new pathogenic pathways in DR. Conclusions Disorders in the metabolic pathways of arginine biosynthesis, tricarboxylic acid cycle, alanine, aspartate, glutamate metabolism, tyrosine metabolism, and D-glutamate metabolism were associated with DR. Risk scores based on vitreous fluid metabolites can be used for the diagnosis and management of DR. Ferrous ascorbate can provide insights into potential new pathogenic pathways for DR.

Background: This study aimed to explore differences in vitreous humour metabolites and metabolic pathways between patients with and without diabetic retinopathy (DR) and identify potential metabolite biomarkers. Methods: Clinical data and vitreous fluid samples were collected from 125 patients (40 without diabetes, 85 with DR). The metabolite profiles of the vitreous fluid samples were analysed using ultra-high performance liquid chromatography, Q-Exactive mass spectrometry, and multivariate statistical analysis. A machine learning model based on Least Absolute Shrinkage and Selection Operator Regularized logistic regression was used to build a risk scoring model based on selected metabolite levels. Candidate metabolites were regressed to glycated haemoglobin levels by a logistic regression model. Results: Twenty differential metabolites were identified between the DR and control groups and were significantly enriched in five Kyoto Encyclopedia of Genes and Genomes pathways (arginine biosynthesis; tricarboxylic acid cycle; alanine, aspartate, and glutamate metabolism; tyrosine metabolism; and D-glutamate metabolism). Ferrous ascorbate significantly contributes to poorer glycaemic control outcomes, offering insights into potential new pathogenic pathways in DR. Conclusions: Disorders in the metabolic pathways of arginine biosynthesis, tricarboxylic acid cycle, alanine, aspartate, glutamate metabolism, tyrosine metabolism, and D-glutamate metabolism were associated with DR. Risk scores based on vitreous fluid metabolites can be used for the diagnosis and management of DR. Ferrous ascorbate can provide insights into potential new pathogenic pathways for DR. [ABSTRACT FROM AUTHOR]

Outbreaks of methanol poisoning have been described in the medical literature worldwide. However, the few outbreaks that have occurred in Saudi Arabia remain undocumented. This is especially noteworthy in light of the fact that Saudi Arabia is among the countries that explicitly prohibit the usage of alcoholic beverages and recreational drugs. Herein, we describe five cases of methanol poisoning in Saudi Arabia. The first three comprise patients admitted to the emergency room (ER) with signs of methanol toxicity, such as heart palpitations, vomiting, and blurred vision; otherwise, those patients were considered medically free. The remaining two cases were examined postmortem. A headspace gas chromatography-flame ionization detector was used to test blood, vitreous humor, and urine samples for methanol. Specific lethal concentrations of methanol were defined based on published case studies as 23–740 mg/dL in blood and 12–396 mg/dL in vitreous humor. In postmortem cases of our present study, samples exhibited lethal concentrations: 118 and 257 mg/dL in blood, 116.3 and 283 mg/dL in vitreous humor. In ER cases, methanol concentrations in urine samples were lower, at 7.5, 9.1, and 20.9 mg/dL; however, toxic symptoms were still observed. These case studies indicate that it is necessary to raise community awareness about the risk of methanol poisoning in order to minimize future poisoning epidemics.

Introduction: Time since death (TSD) also called, as post-mortem interval is an important element in investigation of crime and court trial. The large volume, easy accessibility, resistance to bacterial degradation and relatively inert nature makes the vitreous humour to be the most suitable for chemical analysis. Lactic acid, hypoxanthine and potassium are the common parameters studied in vitreous humour. We aimed to study the change in concentration of potassium in the vitreous humour for estimation of time since death. Material and Methods: This is a cross-sectional, hospital based study conducted in the Index medical college and hospital, Malwanchal University, Indore, from April 2019 to August 2023. All deceased cases with known time of death which were brought to mortuary were included in the current study. Body with previous ocular injury or surgery, bodies with extreme decomposition, posterior segment diseases and paediatric cases

The investigation of novel sample matrices in the forensic sciences offers several possible advantages, such as allowing for results to be obtained in cases where common sample types are absent. This review focuses on the application of gas chromatography and gas chromatography–mass spectrometry (GC-MS) for the determination of drugs in alternative sample matrices, including hair, sweat, meconium, breast milk, and vitreous humour. Less common sample types are also reported including air, cerumen, insects, and their larvae and pupae. The application of pyrolysis GC-MS (Py GC-MS) is also reviewed, showing the possibility of determining high molecular weight drugs which would commonly be unattainable by GC-MS. The application of Py GC-MS for the simulation and investigation of the underlying chemistry and the products formed in the smoking of drugs is also reported. [ABSTRACT FROM AUTHOR]

The K+ and hypoxanthine (Hx) concentrations of the vitreous humour (VH) rise gradually after death, providing a means of estimating the post-mortem interval (PMI). The correlation between these analytes and the PMI is good since the vitreous chamber is partially isolated from autolytic events occurring elsewhere; the [K +] and [Hx] recorded is thus the result of changes within the eye. The present work provides a systematic review, following PRISMA recommendations, of 36 articles (3 reviews and 33 retrospective cohort studies) discussing the many procedures and regression models that have been developed for improving PMI estimates involving VH analytes. The results of a descriptive study are also provided, highlighting the causes and distribution of mortality as registered in medico-legal autopsies performed in 2019 in Galicia (northwestern Spain), and revealing the use of these PMI estimation methods in real forensic practice. Great heterogeneity was detected in the collection of VH samples, the treatments to which they were subjected before examination, and in their conservation and analysis. A lack of reproducibility in the analytical methods employed to estimate [K +] and [Hx] was noted, as well as an absence of external validation for most of the regression formulae used to determine the PMI from analyte values. The use of methods based on high-performance liquid chromatography, focal electrophoresis, or thermogravimetric/chemometric procedures might solve the problems encountered with traditional analytical techniques, offering reliable results more quickly and effectively (even when samples are contaminated). This study recommends using flexible multiple regression models that combine physical and chemical variables, and that population databases be constructed so that models can be properly validated. [ABSTRACT FROM AUTHOR]

Introduction: The estimation of post-mortem interval (PMI) remains a major challenge in forensic science. Most of the proposed approaches lack the reliability required to meet the rigorous forensic standards. Objectives: We applied 1H NMR metabolomics to estimate PMI on ovine vitreous humour comparing the results with the actual scientific gold standard, namely vitreous potassium concentrations. Methods: Vitreous humour samples were collected in a time frame ranging from 6 to 86 h after death. Experiments were performed by using 1H NMR metabolomics and ion capillary analysis. Data were submitted to multivariate statistical data analysis. Results: A multivariate calibration model was built to estimate PMI based on 47 vitreous humour samples. The model was validated with an independent test set of 24 samples, obtaining a prediction error on the entire range of 6.9 h for PMI < 24 h, 7.4 h for PMI between 24 and 48 h, and 10.3 h for PMI > 48 h. Time-related modifications of the 1H NMR vitreous metabolomic profile could predict PMI better than potassium up to 48 h after death, whilst a combination of the two is better than the single approach for higher PMI estimation. Conclusion: The present study, although in a proof-of-concept animal model, shows that vitreous metabolomics can be a powerful tool to predict PMI providing a more accurate estimation compared to the widely studied approach based on vitreous potassium concentrations. [ABSTRACT FROM AUTHOR]

In the present study, we investigated the efficacy of vitreous humour and synovial fluid on avulsion wound healing in a rabbit’s model. The vitreous humour is a fluid that resembles gel consisting of approximately 98–99% water, little hyaluronic acid, glucose, collagen, anions, cations and ions. It is in the posterior eye chamber for the comparison with synovial fluid that consisted of hyaluronin, lubricin, proteinase, prostaglandins and collagenase. In this study, both synovial fluid and vitreous humour were collected from rabbits by aspiration of vitreous humour from the eye (postmortem) and arthrocentesis procedure was applied for collection of synovial fluid. Twelve adult rabbits were used in this study, they divided into three groups each group consisted of four animals wounded experimentally (an avulsion wound). Our results showed the influence of vitreous humour (group B) on healing of the wound is better than the synovial fluid (group C) in the clinical evaluation of shrinkage of the wound. The histo-pathologically changes also revealed that in the vitreous humour treated group (group B), the wound healing process proceeded better than other groups (control and synovial fluid groups). In conclusion, the histopathological and clinical observations demonstrated that application of vitreous humour on wound might be pivotal in improving the healing of avulsion wounds and establish a new tissue in rabbits.

Background: Time since death (TSD) is the period between the death and time of examination of a body. Numerous studies conducted on analysis of potassium in vitreous humour have shown that the vitreous potassium levels increase with rise in TSD. Studies done on postmortem biochemistry of synovial fluid have proposed a positive linear relationship between potassium and TSD. But, very few authors have compared findings of these two fluids. So, this study was undertaken to compare the accuracy of determining post-mortem interval from Sodium and Potassium levels of vitreous humour and synovial fluid. Methods: A total of 140 cases were taken with certified time of death. These were divided into 7 groups i.e. 0-6 hours, 6-12 hours, 12-18 hours, 18-24 hours, 24-30 hours, 30-36 hours and above 36 hours. Vitreous humour and Synovial fluid from knee joint were aspirated from both sides of body and were analyzed using Medica Easylyte Na/K Analyser. Results were obtained using SPSS 24 software. Conclusions: Potassium has a strong positive correlation with TSD in both fluids. Sodium has weak negative correlation. There is no difference between values of both sides. There is no statistically significant difference between values of Vitreous humour and Synovial Fluid. Hence, both can be used alternatively. [ABSTRACT FROM AUTHOR]

A main cause of vision loss in the elderly is age-related macular degeneration (AMD). Among the cellular, biochemical, and molecular changes linked to this disease, inflammation and angiogenesis appear as being crucial in AMD pathogenesis and progression. There are two forms of the disease: dry AMD, accounting for 80–90% of cases, and wet AMD. The disease usually begins as dry AMD associated with retinal pigment epithelium and photoreceptor degeneration, whereas wet AMD is associated with choroidal neovascularization resulting in severe vision impairment. The new vessels are largely malformed, leading to blood and fluid leakage within the disrupted tissue, which provokes inflammation and scar formation and results in retinal damage and detachment. MicroRNAs are dysregulated in AMD and may facilitate the early detection of the disease and monitoring disease progression. Two recent reviews of microRNAs in AMD had indicated weaknesses or limitations in four earlier investigations. Studies in the last three years have shown considerable progress in overcoming some of these concerns and identifying specific microRNAs as biomarkers for AMD. Further large-scale studies are warranted using appropriate statistical methods to take into account gender and age disparity in the study populations and confounding factors such as smoking status.

In the present study, we investigated the efficacy of vitreous humour and synovial fluid on avulsion wound healing in a rabbit’s model. The vitreous humour is a fluid that resembles gel consisting of approximately 98– 99% water, little hyaluronic acid, glucose, collagen, anions, cations and ions. It is in the posterior eye chamber for the comparison with synovial fluid that consisted of hyaluronin, lubricin, proteinase, prostaglandins and collagenase. In this study, both synovial fluid and vitreous humour were collected from rabbits by aspiration of vitreous humour from the eye (postmortem) and arthrocentesis procedure was applied for collection of synovial fluid. Twelve adult rabbits were used in this study, they divided into three groups each group consisted of four animals wounded experimentally (an avulsion wound). Our results showed the influence of vitreous humour (group B) on healing of the wound is better than the synovial fluid (group C) in the clinical evaluation of shrinkage of the wound. The histo-pathologically changes also revealed that in the vitreous humour treated group (group B), the wound healing process proceeded better than other groups (control and synovial fluid groups). In conclusion, the histopathological and clinical observations demonstrated that application of vitreous humour on wound might be pivotal in improving the healing of avulsion wounds and establish a new tissue in rabbits. [ABSTRACT FROM AUTHOR]

• PMI estimates are often based on VH [K+]. • Sample storage conditions can affect the results obtained. • Frozen VH should be thawed in a refrigerator before analysis. • VH samples should not be exposed to ambient light and temperature. Obtaining a reliable estimate of the post-mortem interval (PMI) has been a long-running challenge in forensic medicine. Several more or less successful techniques for making such estimates have been developed, but in recent years important advances have been made thanks to the detailed study of the relationship between the PMI and the analytes - in particular K+ - of the vitreous humour (VH). The extraction and pre-treatment of VH samples has been standardized, the influence of certain environmental factors on analytical results has been quantified, and some of the circumstances under which techniques become unreliable have been identified. The present work examines how the conditions to which VH samples are subject in routine practice may alter the results of their analysis. Exposure to light and ambient temperature was found to alter the values returned in determinations of VH [K+], [Na+] and [Cl-], while exposure to several freezing/thawing cycles (even with final heating) led to no significant modifications in determinations of VH [K+] and [Na+]. It is recommended that if analysis has to be delayed, VH should be frozen for storage in a refrigerator before bringing to room temperature for processing. It is also recommended that samples not be exposed to ambient light and temperature. [ABSTRACT FROM AUTHOR]

The fishing and aquaculture industries generate a huge amount of waste during processing and preservation operations, especially those of tuna. Recovering these by-products is a major economic and environmental challenge for manufacturers seeking to produce new active biomolecules of interest. A new hyaluronic acid was extracted from bluefin tuna's vitreous humour to assess its antioxidant and pharmacological activities. The characterization by infrared spectroscopy (FT-IR), nuclear magnetic resonance ((1D 1H) and 2D (1H COSY, 1H/13C HSQC)) and size exclusion chromatography (SEC/MALS/DRI/VD) revealed that the extracted polysaccharide was a hyaluronic acid with high uronic acid content (55.8 %) and a weight average molecular weight of 888 kDa. This polymer possesses significant anti-radical activity and ferrous chelating capacity. In addition, pharmacological evaluation of its anti-inflammatory and analgesic potential, using preclinical models, in comparison with reference drugs (Dexamethasone, diclofenac, and acetylsalicylate of lysine), revealed promising anti-inflammatory activity as well as interesting peripheral and central antinociceptive activity. Therefore, our new hyaluronic acid compound may therefore serve as a potential drug candidate for the treatment of pain sensation and inflammation of various pathological origins. [Display omitted] • A novel hyaluronic acid was extracted from the vitreous humour of bluefin tuna. • The physicochemical properties of the isolated hyaluronic acid were studied. • The isolated hyaluronic acid demonstrated significant pharmacological properties. • The hyaluronic acid demonstrated promising potential as an anti-inflammatory agent. [ABSTRACT FROM AUTHOR]

Outbreaks of methanol poisoning have been described in the medical literature worldwide. However, the few outbreaks that have occurred in Saudi Arabia remain undocumented. This is especially noteworthy in light of the fact that Saudi Arabia is among the countries that explicitly prohibit the usage of alcoholic beverages and recreational drugs. Herein, we describe five cases of methanol poisoning in Saudi Arabia. The first three comprise patients admitted to the emergency room (ER) with signs of methanol toxicity, such as heart palpitations, vomiting, and blurred vision; otherwise, those patients were considered medically free. The remaining two cases were examined postmortem. A headspace gas chromatography-flame ionization detector was used to test blood, vitreous humor, and urine samples for methanol. Specific lethal concentrations of methanol were defined based on published case studies as 23–740 mg/dL in blood and 12–396 mg/dL in vitreous humor. In postmortem cases of our present study, samples exhibited lethal concentrations: 118 and 257 mg/dL in blood, 116.3 and 283 mg/dL in vitreous humor. In ER cases, methanol concentrations in urine samples were lower, at 7.5, 9.1, and 20.9 mg/dL; however, toxic symptoms were still observed. These case studies indicate that it is necessary to raise community awareness about the risk of methanol poisoning in order to minimize future poisoning epidemics. [ABSTRACT FROM AUTHOR]

Eye damage during invasive aspergillosis is rarely described and biological diagnosis remains challenging. Here we report the case of a heart transplant recipient with ocular aspergillosis complicating disseminated aspergillosis. Although voriconazole was rapidly given, a decrease in visual acuity of the right eye was consistent with endophthalmitis, resulting in an emergency vitrectomy. The diagnosis was rapidly confirmed: laboratory results showed the presence of Aspergillus fumigatus in a vitreous sample. A series of systemic antifungal medications (liposomal amphotericin B, caspofungin, and voriconazole), several liposomal amphotericin B ocular injections, and pars plana vitrectomy resulted in a limited positive clinical outcome. Interestingly although standard mycological follow-up procedures were negative, Aspergillus antigen testing gave an index of 5.92 on vitreous humour, thus a new intraocular injection of liposomal amphotericin B was performed and voriconazole reinitiated. Ten other vitreous samples from patients without fungal infections were also tested, all showing indexes below 0.25. Although larger studies are needed, this case illustrates that galactomannan testing of vitreous humour could be useful for the diagnosis of fungal endophthalmitis if these data are confirmed in other patients, in particular, if standard mycology is negative and PCR is not available.

The death erases all doubts from the dead’s mind but inscribes new doubts in the living mind. The study of time since death gives the final turn in the crime to find the cause and time of death. The present study was conducted in Department of Forensic Medicine, Madras Medical College, Chennai during February 2006 to July 2006 to find out the time since death by analysing the potassium levels of vitreous humour after death. Total 75 cases were studied which is broadly classified into: Hospital cases and Non- Hospital cases irrespective of age, gender, religion, caste etc., who has died. All the data related to Non-Hospital cases were analysed to record the effect of nature and pattern of rise of vitreous humour in the deaths, whose detailed autopsy examination were also recorded subsequently for statistical analysis. From this study, it is concluded that the vitreous sodium and chloride was found to be totally unaffected and stable for 97 hours after death, irrespective of sex, age, environmental temperature, nature of death and cause of death. [ABSTRACT FROM AUTHOR]

The application of proteomic techniques to forensic science widens the range of analytical capabilities available to forensic laboratories when answering complex toxicology problems. Currently, these techniques are underutilised in post‐mortem toxicology because of the historic focus on smaller (<1,000 amu) drug molecules. Definitive confirmation of an insulin overdose by analysis of post‐mortem biological matrices is rare and challenging, however can assist coronial investigations pertaining to accidental or intentional overdoses in both diabetic and nondiabetic populations. A semiautomated micro‐solid phase extraction paired with mass spectrometry‐based insulin methodology was developed and validated for routine use in a Forensic Coronial Toxicology Laboratory. This resulting work reports the first Australian cases where synthetic insulins were confirmed by mass spectrometry in the vitreous humour of Type 1 diabetics who intentionally or accidentally overdosed on their prescription medication glargine and aspart. The detection of glargine M1 in Case 1, aspart in Case 2 and glargine M1 was indicated in Case 3. This paper highlights advancements in forensic coronial toxicology and the promising potential of proteomic analysis in a forensic context. [ABSTRACT FROM AUTHOR]

The vitreous humour (VH) is a fluid material located inside the eyeball and has been involved in postmortem studies since 1960, when Naumann first provided the concept of analysis of the biochemical changes in the VH. The special attention paid to VH by coroners is explained by its particular anatomical location, which gives it real protection against changes occurring postmortem. This fluid represents an environment less susceptible to autolytic alterations or postmortem contamination with bacteria/medications which can be found in high concentrations in other anatomical sites at the time of death. This article aims to emphasize the importance of vitreous humour in legal medicine, bringing into question anatomical and practical considerations concerning the collection and utilization of this corporeal fluid in the thanatochemical analysis of glucose. [ABSTRACT FROM AUTHOR]

Post-mortem biochemistry as a tool for the establishment of post-mortem interval (PMI) and the nature of death is very useful in Legal Medicine. Currently, several matrices are used for these investigations, although vitreous humour is the most useful for biochemical analysis due to its stability and low susceptibility to autolytic changes. The purpose of this study was to analyse to what extent the nature of death could influence in the measurement of parameters such us potassium (K+), hypoxanthine (Hx) and uric acid in the establishment of PMI. Samples of vitreous humour were collected from the eyes of 250 dead bodies (174 males, 76 females) with a mean age of 58.75 ± 20.84 years, mean PMI of 15.56 ± 6.21 hours post-mortem (hpm) and different causes of death, including natural (n = 148) and violent (n = 101) deaths. The vitreous humour was treated and the resultant supernatant was used for the biochemical analysis. Multichannel Autoanalyser was used to determine the concentrations of K+ and uric acid (mmol/L), and Hx (µmol/L) was quantified by HPLC-UV. The results showed the measurement of K+, Hx and uric acid together could be a complementary tool for establishing the PMI because these parameters provide good results. [ABSTRACT FROM AUTHOR]

Cerebrospinal fluid (CSF) and vitreous humour (VH) can be biochemically analysed post-mortem. CSF can be aspirated from the ventricular system (VA) and from lumbar puncture (LP), whereas, VH is aspirated from the orbits. This study compared the biochemical constituents between CSF and VH in 20 post-mortem subjects as well as the effect post-mortem interval and subject age have on these variables. VH samples had significantly higher sodium (Na) and chloride (Cl) and lower magnesium (Mg), potassium (K), urea (Ur) and creatinine (Cr) compared with CSF samples. No statistically significant difference was demonstrated with beta-hydroxybutyrate (BHB) and glucose levels in both VH and both CSF samples. Interestingly, some of the differences in the biochemical analysis correlated with age, but none with post-mortem interval. In addition to sampling and pre-analytical sample processing differences, there may also be ante- and post-mortem factors explaining these observations. This study suggests that post-mortem CSF and VH are biochemically different and reference ranges for VH are not recommended for use in CSF samples. [ABSTRACT FROM AUTHOR]

The estimation of post-mortem interval (PMI) procedures seem to subjectivity in assessment and less sensitivity and specificity. Biochemical determination of trace elements from closed-system samples, such as vitreous humour (VH) and synovial fluid (SF) is an alternative laboratory investigation for PMI estimation. The inductively coupled plasma-mass spectrometry (ICP-MS) provides more rapid, precise and sensitive analysis of elements samples. However, determination of electrolytes and trace elements in VH and SF by ICP-MS, and application for PMI has not been intensively investigated. In this study, VH and SF from twenty adult domesticated pig carcasses were collected at 1, 3, 6, 12, 24, 36, and 48-hour after death. Samples were analysed for total protein concentration, Ca, Cu, Fe, K and, Na using ICP-MS. The results showed that the protein concentration in VH was significantly increased in time dependent manner, but not for SF. The level of K and Fe in both VH and SF increased in time dependent manner, with a good correlation between both sample sources. The calcium concentration of both VH and SF increased in time dependent manner, but there was no correlation between both sample sources. Sodium was significantly reduced at 36-48 h, with a correlation between both sample sources. However, the copper level in VH was increased at 36-48 h but reduced in SF. In conclusion, this study showed for the first time that multiple trace elements and electrolytes could be detected, using the ICP-MS, in both of VH and SF simultaneously and additionally its correlation. [ABSTRACT FROM AUTHOR]

Aims: To compare circular (circRNA) expression levels in the vitreous humour between PDR (proliferative diabetic retinopathy) and the control groups. Methods: The present study collected vitreous humour samples of both the PDR group and the control group (composed of rhegmatogenous retinal detachment, idiopathic macular hole and idiopathic macular epiretinal membrane). All the samples were subjected to circRNA and mRNA sequencing as well as bioinformatic analysis. Results: The vitreous humour of the PDR and control groups was collected during PPV surgery. Compared to the control group, 122 upregulated and 9 downregulated circRNAs, and 818 upregulated mRNAs and 864 downregulated mRNAs were identified. We further selected 12 circRNAs to validate the RNA expression level by qPCR; results showed that with the exception of 2 downregulated circRNAs the remaining were significantly upregulated in the PDR group, which was consistent with RNA sequencing results. Bioinformatic analysis was conducted to predict possible miRNAs absorbed by circRNAs. Each circRNA could interact with at least five miRNAs. We randomly chose three miRNAs to test the expression level in the vitreous humour by qPCR and found these miRNAs were significantly downregulated in the PDR group. Conclusions: The changed profiling of circRNAs in the vitreous humour was reliable and may become a promising biomarker of DR and the circRNA–miRNA–mRNA network. It may also play an important role in the progression of DR. [ABSTRACT FROM AUTHOR]

Purpose: We determined vitreous and serum levels of high mobility group box‐1 (HMGB‐1) in patients with proliferative diabetic retinopathy (PDR) and elucidate their relationship with receptor for advanced glycation end products (RAGE), vascular endothelial growth factor (VEGF) and interleukin‐1β (IL‐1β). Methods: In this cross‐sectional study, patients with PDR who underwent vitrectomy were enrolled, and the control group included non‐diabetic eyes. Vitreous and serum samples were analysed for HMGB‐1, RAGE, VEGF and IL‐1β by ELISA. We investigated the correlation between serum and vitreous levels of each cytokine, and we analysed the influence of intravitreal anti‐VEGF treatment prior to vitrectomy on the cytokine levels in PDR. Results: Of 78 eyes of 78 patients enrolled consecutively, there were 32 PDR eyes and 46 control eyes. The serum levels were higher in diabetic than in non‐diabetic subjects for HMGB‐1, RAGE, VEGF and IL‐1β (all p < 0.001), respectively. Similarly, the vitreous levels were higher in diabetic than in non‐diabetic subjects for HMGB‐1 (p < 0.001), RAGE (p = 0.001), VEGF (p < 0.001) and IL‐1β (p < 0.001), respectively. We found a positive correlation between serum and vitreous levels of HMGB‐1 in patient with PDR (p = 0.047, R = 0.353). There was a negative correlation between serum and vitreous levels of VEGF in patient with PDR (p = 0.001, R = −0.546). For the subgroup analysis, we detected that the vitreous levels of RAGE were significantly lower in patients who underwent anti‐VEGF injection prior to vitrectomy than those who did not (p < 0.001). Conclusions: Our findings suggest that HMGB‐1 is involved in PDR disorders, and it may be a novel therapeutic target to inhibit progression of PDR. [ABSTRACT FROM AUTHOR]

Background: For the interpretation of concentrations of gamma-hydroxybutyrate (GHB) in post-mortem specimens, a possible increase due to post-mortem generation in the body and in vitro has to be considered. The influence of different storage conditions and the specimen type was investigated. Method and material: Post-mortem GHB concentrations in femoral venous blood (VB), heart blood (HB), serum (S) from VB, urine (U), cerebrospinal fluid (CSF) and vitreous humour (VH) were determined by gas chromatography–mass spectrometry after derivatisation. Various storage conditions, that is 4 °C or room temperature (RT) and the addition of sodium fluoride (NaF), were compared during storage up to 30 days. Additionally, bacterial colonisation was determined by mass spectrometry fingerprinting. Results: Twenty-six cases without involvement of exogenous GHB were examined. GHB concentrations (by specimen) at day 0 were 3.9–22.1 mg/L (VB), 6.6–33.3 mg/L (HB), < 0.5–18.1 mg/L (U), 1.1–10.4 mg/L (CSF) and 1.7–22.0 mg/L (VH). At 4 °C, concentrations increased at day 30 to 5.6–74.5 mg/L (VB), 4.6–76.5 mg/L (HB) and < 0.5–21.3 mg/L (U). At RT, concentrations rose to < 0.5–38.5 mg/L (VB), 1.2–94.6 mg/L (HB) and < 0.5–37.5 mg/L (U) at day 30. In CSF, at RT, an increase up to < 0.5–21.2 mg/L was measured, and at 4 °C, a decrease occurred (< 0.5–6.5 mg/L). GHB concentrations in VH remained stable at both temperatures (1.2–20.9 mg/L and < 0.5–26.2 mg/L). The increase of GHB in HB samples with NaF was significantly lower than that without preservation. No correlation was found between the bacterial colonisation and extent of GHB concentration changes. Conclusion: GHB concentrations can significantly increase in post-mortem HB, VB and U samples, depending on storage time, temperature and inter-individual differences. Results in CSF, VH, S and/or specimens with NaF are less affected. [ABSTRACT FROM AUTHOR]

In this study, acute carbon monoxide (CO) was used to cause the death of experimental animals through inhalation. The sample size of the study was made up of 18 rabbits divided into the control group (CG), disguised death (DD) and carbon monoxide death (CD) groups respectively. Vitreous humours were extracted using standard methodologies and biochemical parameters estimated using WHO approved methods and procedures. The findings showed a significant decrease (P < 0.05) in vitreous humour chloride ion, calcium ion, pH, uric acid and total calcium in the CDs. However, vitreous lactate dehydrogenase (LDH) of the experimental animals significantly increased (P < 0.05) only in the CD group, while creatine kinase (CK) and creatine significantly increased in both the DD and CD animal groups, although they were markedly higher in the CD. Therefore, decreases in concentrations of vitreous humour uric acid, chloride and calcium profile, and an increase in vitreous LDH could serve as possible indicators of CO poisoning, especially in scenarios of impracticable use of blood, as in rigor mortis, livor mortis or decomposition. [ABSTRACT FROM AUTHOR]

Summary: Background: Equine recurrent uveitis (ERU) has been associated with Leptospira spp. infection. No information exists concerning the prevalence of Leptospira‐associated ERU in Belgium and about the sensitivity of detection of Leptospira in different ocular media. Objectives: To establish the prevalence of intraocular Leptospira spp. in ERU‐affected and healthy eyes of horses examined at the Equine Clinic of the University of Liège by real‐time PCR and to compare the results of the aqueous and vitreous humour of the same eye. Study design: Cross‐sectional. Methods: Sixty‐six eyes from 59 client‐owned horses with a diagnosis of equine recurrent uveitis (ERU‐group) were studied from May 2015 to December 2017. Fifty healthy eyes from 28 euthanised horses for unrelated reasons examined during the same period were included in the control group. Intraocular fluids (aqueous and/or vitreous humours) from ERU‐affected eyes were sampled and analysed by real‐time PCR for Leptospira spp. Aqueous and vitreous humours from the control group were processed in the same way. Results: Both groups were comparable regarding age, sex, eye sampled (OS/OD), humours sampled (aqueous/vitreous humour) but not regarding breeds, with an over‐representation of Warmbloods and Appaloosas in the ERU‐group. The prevalence of Leptospira spp. was 30.3% (20/66 eyes) in the ERU‐group. Leptospira spp. DNA was identified in 11 aqueous and 17 vitreous humours with eight horses testing positive in both humours, nine horses testing positive for vitreous humour alone and third horses for aqueous humour alone. The phi‐correlation between aqueous and vitreous humour Leptospira‐PCR results is 0.47 suggesting a low association. All the control eyes were negative. Main limitations: The diagnostic method selected for this study (lipL32 qPCR) did not allow identification of the serovars. Conclusions: Leptospirosis is a potential cause of ERU in Belgium. Testing both intraocular media is advised whenever possible. The Summary is available in Spanish – see Supporting Information [ABSTRACT FROM AUTHOR]

Background: Although the post-mortem increase of ammonium in biological fluids is well known, ammonium analysis in vitreous humour has never been used in recent times for the determination of the post-mortem interval. The present work represents a new application of capillary electrophoresis with indirect UV detection in the field of forensic analysis. Methods: The electrophoretic separation was carried out in a running buffer made of 5 mM imidazole, 5 mM 18-crown-6 ether and 6 mM d,l-α-hydroxybutyric acid (HIBA). To overcome the lack of optical absorption of ammonium, indirect UV detection was applied. The used wavelength was 214 nm. Results: The method showed good linearity in the concentration range from 0.16 to 5.0 mM. The limit of detection, 0.039 mmol/L, was established on the basis of the linearity curve. Precision and bias studies carried out on the pure ammonium solutions and in real biological samples, revealed %RSDs well below 20%. A preliminary application to real cases where the death time was precisely known (14 bodies) was carried out plotting vitreous humour ammonium vs. post-mortem interval with a resulting good linear correlation until 100 h post-mortem. Conclusions: After validation in real cases, the present method can become a powerful tool to unravel one of the most challenging issues of forensic investigation: determination of the time of death. [ABSTRACT FROM AUTHOR]

This study reports on a smartphone-based application for the determination of time since death (TSD) from vitreous humour (VH) protein via recognition and quantification using a simple biuret method for on-the-spot analysis. In basic pH, in the presence of protein, the biuret reagent forms a protein-biuret complex resulting in a colour change that is measurable using a UV-vis spectrophotometer. The limit of detection (LOD) for Bovine serum albumin (BSA) was found to be 8 μg/ml and the limit of quantification (LOQ) was found to be 10 μg/ml. The developed method was further validated and applied for TSD determination from VH samples in the range of 3-60 h. The results showed that there is a linear correlation between the VH protein concentration and TSD, as the protein concentration decreases up to 60 h, with the equation TSD = 33.72 + (-0.27) × concentration. Further, in the present study smart approach was applied for the TSD determination. The results obtained showed that the current method proves to have a great utility up to 60 h. The linear regression equation between TSD and RGB (red, green and blue) intensities of protein concentration tup to 60 h ± 4.5 h was found to be TSD = 6.27E2+1.41*RGB intensity. The proposed method provides a smart, rapid and sensitive, as well as cost effective, TSD determination via VH protein. [ABSTRACT FROM AUTHOR]

A simple and low cost analytical device is described for the determination of ammonium in the vitreous humor suitable for inferring the post mortem interval in forensic cases. The device is based on ammonia formation from ammonium ion by means of NaOH addition to the vitreous humor sample and its detection with a pH chemical indicator in the gas phase above the vitreous humor sample. From the gas phase, ammonia diffuses through a polymeric membrane and it is trapped and detected with a droplet of pH indicator thymol blue. The color change of the droplet is measured using a smartphone camera. Under optimal conditions, the device showed a limit of detection of 0.2mM, with between days precision of ≤ 15% expressed as relative standard deviation, and an accuracy between days from 88.3% to 114.5%. This homemade gas diffusion analytical device was successfully used for the determination of ammonia in vitreous humor samples from forensic autopsies. The results obtained with the proposed method, although for a limited number, showed a close correlation with the data obtained with an instrumental analysis based on capillary electrophoresis. Moreover a significant correlation was also found between the results of the present method and the time elapsed since death by a simple evaluation of the color intensity. In conclusion, this preliminary study showed that the proposed device, after adequate validation, could be a promising tool for a presumptive estimation of the time since death directly at the crime scene. [ABSTRACT FROM AUTHOR]

Aim of the study : To produce a formula that can accurately predict postmortem interval (PMI) based on vitreous potassium levels using road traffic collision fatalities. Material and methods: Vitreous humour samples were taken from 78 individuals who had died following road traffic collisions between 2010 and 2015. Samples were obtained from both eyes and were sent for on-site analysis. Measurement of potassium was by an indirect ion-specific electrode Siemens diagnostics ADVIA 2400 chemistry system. Exact time of death was known from police reports, the time of postmortem was recorded and the postmortem interval was calculated. Linear regression was then used to analyse the relationship between the two. The impact of age was also assessed. Results: PMI was between 6 and 162 hours. As vitreous potassium increases, the PMI also increases; exhibiting a linear relationship. This is illustrated by a regression equation of PMI = 6.42[K+] – 40.94, R = 0.67 (p < 0.001). This produced a formula closely comparable with three other studies proposed in previous literature and produces estimates that may exceed one calendar day. When both age and medical intervention are accounted for there is an insignificant improvement in prediction. Conclusions : Validated methods have been used to produce a formula for prediction of PMI using vitreous potassium. Although this is specific to road traffic collisions, the methods are transferable and can be seen to be comparable with other recently published methods. Nonetheless, if greater levels of accuracy are required it is suggested that biomarkers delivering a higher level of precision should still be sought.

• The biological diagnosis of ocular fungal infection remains challenging. • Use of a galactomannan test on vitreous humour may be useful to diagnose ocular aspergillosis. • The short time-to-result allows rapid and adapted antifungal treatment. Eye damage during invasive aspergillosis is rarely described and biological diagnosis remains challenging. Here we report the case of a heart transplant recipient with ocular aspergillosis complicating disseminated aspergillosis. Although voriconazole was rapidly given, a decrease in visual acuity of the right eye was consistent with endophthalmitis, resulting in an emergency vitrectomy. The diagnosis was rapidly confirmed: laboratory results showed the presence of Aspergillus fumigatus in a vitreous sample. A series of systemic antifungal medications (liposomal amphotericin B, caspofungin, and voriconazole), several liposomal amphotericin B ocular injections, and pars plana vitrectomy resulted in a limited positive clinical outcome. Interestingly although standard mycological follow-up procedures were negative, Aspergillus antigen testing gave an index of 5.92 on vitreous humour, thus a new intraocular injection of liposomal amphotericin B was performed and voriconazole reinitiated. Ten other vitreous samples from patients without fungal infections were also tested, all showing indexes below 0.25. Although larger studies are needed, this case illustrates that galactomannan testing of vitreous humour could be useful for the diagnosis of fungal endophthalmitis if these data are confirmed in other patients, in particular, if standard mycology is negative and PCR is not available. [ABSTRACT FROM AUTHOR]

Objective and design: The purpose of this study was to investigate if diet-induced obesity (DIO) and subsequent low-level systemic inflammation would result in local increases in pro-inflammatory mediators in the vitreous humour (VH) of the eyes of rats. Methods: Sixteen male Sprague-Dawley rats were fed a high-fat/high-sucrose ( n = 9) or chow control-diet ( n = 7) for 12-weeks. RT-qPCR was conducted on RNA from VH cells and a 27-plex Luminex Assay was conducted on VH fluid and serum. Results: Increased protein levels for IL-1β, IL-6, and IL-18 in both serum and VH fluid were observed. VH protein levels for IL-13 and IL-17 were also increased. All mediators significantly increased in VH fluid were also positively correlated with percent body fat. Increased mRNA levels in VH cells for an oxidative stress molecule were accompanied by decreased mRNA levels for an antioxidant scavenger, suggesting an antioxidant/oxidant imbalance in the VH with DIO. In addition, decreased mRNA levels for TRAIL, FAS-L and TGF-β, molecules associated with immune privilege, were also significantly depressed. Conclusions: DIO-related metabolic disturbances disrupt VH homeostasis in a manner that reflects development of a pro-inflammatory environment. Prolonged exposure to such an environment may lead to overt pathologies with compromised eye function. [ABSTRACT FROM AUTHOR]