Genetic sexing of subadult skeletal remains.

When subadult skeletons need to be identified, biological sex diagnosis is one of the first steps in the identification process. Sex assessment of subadults using morphological features is unreliable, and molecular genetic methods were applied in this study. Eighty-three ancient skeletons were used as models for poorly preserved DNA. Three sex-informative markers on the Y and X chromosome were used for sex identification: a qPCR test using the PowerQuant Y target included in PowerQuant System (Promega), the amelogenin test included in ESI 17 Fast STR kit (Promega), and a Y-STR amplification test using the PowerPlex Y-23 kit (Promega). Sex was successfully determined in all but five skeletons. Successful PowerQuant Y-target, Y-amelogenin, and Y-chromosomal STR amplifications proved the presence of male DNA in 35 skeletons, and in 43 subadults female sex was established. No match was found between the genetic profiles of subadult skeletons, and the elimination database and negative control samples produced no profiles, indicating no contamination issue. Our study shows that genetic sex identification is a very successful approach for biological sexing of subadult skeletons whose sex cannot be assessed by anthropological methods. The results of this study are applicable for badly preserved subadult skeletons from routine forensic casework.

Comparison of Morphological Sex Assessment and Genetic Sex Determination on Adult and Sub-Adult 17th-19th Century Skeletal Remains.

The first step in the analysis of human skeletal remains is the establishment of the biological profile of an individual. This includes sex assessment, which depends highly on the age of the individual and on the completeness and preservation state of the remains. Macroscopic methods only provide the assessment of sex, while for sex determination, molecular methods need to be included. However, poor preservation of the remains can make molecular methods impossible and only assessment can be performed. Presented research compares DNA-determined and morphologically assessed sex of adult and non-adult individuals buried in a modern-age cemetery (17th to late 19th century) in Ljubljana, Slovenia. The aim of the study was to assess the accuracy of commonly used macroscopic methods for sex assessment on a Slovenian post-medieval population. Results demonstrate that for adults, macroscopic methods employed are highly reliable and pelvic morphology, even the sciatic notch alone, is more reliable than skull. In non-adults, macroscopic methods are not as reliable as in adults, which agrees with previous research. This study shows how morphological and molecular methods can go hand in hand when building a biological profile of an individual. On their own, each methodology presented some individuals with undetermined sex, while together, sex of all the individuals was provided. Results confirm suitability of sex assessment based on skull and especially pelvic morphology in Slovenian post-medieval adults, while in the non-adult population molecular methods are advised.

Urinary Extracellular Vesicles and Their miRNA Cargo in Patients with Fabry Nephropathy.

Current biomarkers of Fabry nephropathy lack sensitivity in detecting early kidney damage and do not predict progression of nephropathy. Urinary extracellular vesicles (uEVs) and their molecular cargo could reflect early changes in renal impairment as they are secreted by the cells lining the urinary tract. We aimed to conduct a proof-of-concept study to investigate whether analysis of uEV characteristics and expression of uEV-derived microRNAs (miRNAs) could be applicable in studies to predict the development and progression of nephropathy in Fabry disease. A total of 20 Fabry patients were divided into two groups, depending on the presence of nephropathy. Chronological urine samples collected during 10-year follow-up were used for uEVs isolation with size exclusion chromatography. Nanoparticle tracking analysis was used to determine concentration and size of uEVs. We evaluated the expression of five uEV-derived miRNAs by qPCR (miR-23a-3p, miR-29a-3p, miR-30b-5p, miR-34a-5p, miR-200a-3p). There was no difference in the concentration and size of uEVs between patients with and without nephropathy at last follow-up or longitudinally. However, we found increased expression of miR-29a-3p and miR-200a-3p in uEVs isolated from chronological samples of patients with Fabry nephropathy. This may indicate an attempt by the organism to prevent the progression of renal damage leading to end-stage renal disease as previously reported in type 1 diabetes. In addition, we found an increased expression of miR-30b-5p in the 10-year period in uEVs of patients without renal dysfunction. miR-30b-5 was reported to have a protective role in podocyte injury and may possibly be important in Fabry nephropathy. These findings indicate that uEVs and their molecular cargo could be a promising target of studies focusing on elucidation of Fabry nephropathy. Nevertheless, total concentration and size of uEVs were neither indicative of the presence nor progression of Fabry nephropathy, while the role of the analyzed miRNAs in Fabry nephropathy progression was merely indicated and needs further in-depth studies.