Osteon shape variation in the femoral diaphysis: A geometric-morphometric approach on human cortical bone microstructure in an elderly sample.

Geometric morphometrics (GMM) have been applied to understand morphological variation in biological structures. However, research studying cortical bone through geometric histomorphometrics (GHMM) is scarce. This research aims to develop a landmark-based GHMM protocol to depict osteonal shape variation in the femoral diaphysis, exploring the role of age and biomechanics in bone microstructure. Proximal, midshaft, and distal anatomical segments from the femoral diaphysis of six individuals were assessed, with 864 secondary intact osteons from eight periosteal sampling areas being manually landmarked. Observer error was tested using Procrustes ANOVA. Average osteonal shape and anatomical segment-specific variation were explored using principal component analysis. Osteon shape differences between segments were examined using canonical variate analysis (CVA). Sex differences were assessed through Procrustes ANOVA and discriminant function analysis (DFA). The impact of osteonal size on osteonal shape was investigated. High repeatability and reproducibility in osteon shape landmarking were reported. The average osteon shape captured was an elliptical structure, with PC1 reflecting more circular osteons. Significant differences in osteon shape were observed between proximal and distal segments according to CVA. Osteon shape differed between males and females, with DFA showing 52% cross-validation accuracies. No effect of size on shape was reported. Osteonal shape variation observed in this study might be explained by the elderly nature of the sample as well as biomechanical and physiological mechanisms playing different roles along the femoral diaphysis. Although a larger sample is needed to corroborate these findings, this study contributes to the best of our knowledge on human microanatomy, proposing a novel GHMM approach.

Automatic Segmentation of Osteonal Microstructure in Human Cortical Bone Using Deep Learning: A Proof of Concept.

Cortical bone microstructure assessment in biological and forensic anthropology can assist with the estimation of age-at-death and animal-human differentiation, for example. Osteonal structures within cortical bone are the key feature under analysis, with osteon frequency and metric parameters providing crucial information for the assessment. Currently, the histomorphological assessment consists of a time-consuming manual process for which specific training is required. Our work investigates the feasibility of automatic analysis of human bone microstructure images through the application of deep learning. In this paper, we use a U-Net architecture to address the semantic segmentation of such images into three classes: intact osteons, fragmentary osteons, and background. Data augmentation was used to avoid overfitting. We evaluated our fully automatic approach using a sample of 99 microphotographs. The contours of intact and fragmentary osteons were traced manually to provide ground truth. The Dice coefficients were 0.73 for intact osteons, 0.38 for fragmented osteons, and 0.81 for background, giving an average of 0.64. The Dice coefficient of the binary classification osteon-background was 0.82. Although further refinement of the initial model and tests with larger datasets are needed, this study provides, to the best of our knowledge, the first proof of concept for the use of computer vision and deep learning for differentiating both intact and fragmentary osteons in human cortical bone. This approach has the potential to widen and facilitate the use of histomorphological assessment in the biological and forensic anthropology communities.

Sex estimation in a Turkish population using Purkait's triangle: a virtual approach by 3-dimensional computed tomography (3D-CT).

Sex estimation is considered one of the first steps in the forensic identification process. Morphological and morphometrical differences between males and females have been used as means for morphoscopic and metric methods on both cranial and postcranial skeletal elements. When dry skeletal elements are not available, virtual data can be used as a substitute. The present research explores 3-dimensional (3D) scans from a Turkish population to test a sex estimation method developed by Purkait (2005). Overall, 296 individuals were used in this study (158 males and 138 females). Purkait's triangle parameters were measured on computed tomography (CT) scans obtained from both right and left femora of each patient at the Bakirkoy Dr. Sadi Konuk Training Research Hospital (Istanbul, Turkey). Intra- and inter-observer errors were assessed for all variables through technical error of measurements analysis. Bilateral asymmetry and sex differences were evaluated using parametric and non-parametric statistical approaches. Univariate and multivariate discriminant function analyses were then conducted. Observer errors demonstrated an overall agreement within and between experts, as indicated by technical error of measurement (TEM) results. No bilateral asymmetries were reported, and all parameters demonstrated a statistically significant difference between males and females. Fourteen discriminant models were generated by applying single and combined parameters, producing a total correct sex classification ranging from 78.4% to 92.6%. In addition, over 67% of the total sample was accurately classified, with 95% or greater posterior probabilities. Our study demonstrates the feasibility of 3D sex estimation using Purkait's triangle on a Turkish population, with accuracy rates comparable to those reported in other populations. This is the first attempt to apply this method on virtual data and although further validation and standardisation are recommended for its application on dry bone, this research constitutes a significant contribution to the development of population-specific standards when only virtual data are available.Key pointsCT analysis using Purkait's triangle is a suitable tool for assessment of sex in unidentified individuals.The best overall estimation rate was achieved with the F11 model, with around 92% of accuracy.The results suggested 78.4% to 92.6% correct sex identification rates.More research is needed to expand the sample set and verify the results.

Age estimation for two Mediterranean populations: rib histomorphometry applied to forensic identification and bone remodelling research.

Numerous intrinsic and extrinsic factors influence bone remodelling rates and have shown to affect the accuracy of histological aging methods. The present study investigates the rib cortex from two Mediterranean skeletal collections exploring the development of population-specific standards for histomorphometric age-at-death estimation. Eighty-eight standard ribs from two samples, Cretans and Greek-Cypriots, were processed histologically. Thirteen raw and composite histomorphometric parameters were assessed and observer error tested. The correlation between age and the parameters and the differences between sex and population subsamples were explored through group comparisons and analysis of covariance. General linear models assessed through data fit indicators and cross-validation were generated from the total dataset, and by sex and population subsamples. Most of the histological variables showed a statistically significant correlation with age with some differences observed by sex and by sample. From the twelve models generated, the optimal model for the whole sample included osteon population density (OPD), osteon perimeter, and osteon circularity producing an error of 10.71 years. When sex and samples were separated, the best model selected included OPD and osteon perimeter producing an error of 8.07 years for Greek-Cypriots. This research demonstrates the feasibility of quantitative bone histology to estimate age, obtaining errors rates in accordance with macroscopic ageing techniques. Sex and sample population differences need further investigation and inter-population variation in remodelling rates is suggested. Moreover, this study contributes to the creation of population-specific standards for Cretans and Greek-Cypriots.

Rib histomorphometry: A reliability and validation study with a critical review of histological techniques for forensic age estimation.

Fragmented human remains present a challenge for forensic experts as they attempt to identify individuals using standard forensic methods. Several histological age estimation techniques have been developed during the last fifty years to aid in this process. However, very few validation studies have been conducted in order to test their accuracy and bias, and thus, validation assessment is required as we employ them while testifying in court. Histological variables are assessed from rib thin sections from two Mediterranean samples; Cretans (N = 41) and Greek-Cypriots (N = 47). Intra and inter-observer errors are assessed through TEM analysis and Intra-class Correlation Coefficient by testing observers with different levels of experience as they collected data on osteon counts and area measurements. The relation between the variables and age is determined using correlation coefficients. Histomorphometric data are applied to four widely used age estimation formulas assessing the performance of the methods for the entire sample. Inaccuracy and bias are calculated with age estimations and known age tested for significance and proportional bias assessed. Overall, histological parameters presented acceptable intra- and inter-observer errors. All variables exhibited statistically significant correlation with age (P < 0.01). For three of the techniques, data showed a systematic underestimation of age with an increase in inaccuracy in older individuals. One of the age estimation formulas produced overestimation of young individuals yet, it more accurately estimated the age of older individuals. This validation study explores inter-population variation in bone remodeling dynamics and presents a critical evaluation on methodological issues that can affect the performance of existing histological techniques.

Bone histomorphometry of the clavicle in a forensic sample from Albania.

Forensic assessment of skeletal material includes age estimation of unknown individuals. When dealing with extremely fragmented human remains that lack macro-features used in age estimation, histological assessment of the skeletal elements can be employed. Historically, microscopic methods for age assessment used by forensic anthropologists have been available since 1965. Several skeletal elements have been used for this purpose. Among them, the clavicle has garnered very little attention. The purpose of this study is to explore the validity of clavicular histomorphometry as an age marker in a modern Balkan sample. This study examined a modern clavicular autopsy sample from Albania. The sample consisted of 33 individuals of known age and cause of death. Data were collected for micro-anatomical features including osteon population density (OPD) and cortical area. Intra- and inter-observer errors were assessed through technical error of measurement (TEM) and R coefficient. A validation study was performed in order to test the accuracy of existing histological formulae. Regression analysis was run to developed age prediction models with the best models tested through cross-validation and the comparison between OPD for the Albanian sample and a European-American sample examined. Intra- and inter-observer error TEM results demonstrated values falling within the limits of acceptance. The existing histological methods did not perform accurately on the sample under study. Regression equations for Albanians produced age estimations deviating 8 and 11 years from known age. Cross-validation on the most accurate regression formula which includes OPD as a single variable demonstrated similar mean errors. Statistically significant differences were observed between the Albanian and the European-American population when the two samples were compared. The research presented is the fifth article published and the fifth population explored on clavicular microstructure. The potential of histology to estimate age on the Albanian population is shown here; however, population effect, diet and health status might be considered. Further inclusion of individuals will corroborate our preliminary findings.

The effect of diet and sociopolitical change on physiological stress and behavior in late Roman-Early Byzantine (300-700 AD) and Islamic (902-1,235 AD) populations from Ibiza, Spain.

OBJECTIVES: This study evaluated chronological changes in physiological stress and levels of habitual loading of Ibizan populations from the Late Roman-Early Byzantine (LREB) to the Islamic period (300-1,235 AD) using measures of body size and bone cross-sectional properties to compare Urban LREB, Urban Medieval Islamic, and Rural Medieval Islamic groups. It also explored the effect of diet, modeled using stable isotopes, on physiological stress levels and behavior. MATERIALS AND METHODS: The sample comprised individuals from three archeological populations: Urban Late Roman- Early Byzantine (LREB) (300-700 AD), Medieval Urban Islamic (902-1,235 AD), and Medieval Rural Islamic. Bone lengths, femoral head dimensions, and cross-sectional properties, diaphyseal products and circumferences, were compared to assess differences in body size and habitual loading in 222 adult individuals. Ordinary least squares regression evaluated the correlations between these measures and carbon (δ13 C) and nitrogen (δ15 N) stable isotope ratios in 115 individuals for whom both isotope values and osteological measures are available. RESULTS: The Medieval Rural Islamic group had shorter stature and reduced lower limb cross-sectional properties compared to the two urban groups. Limb shape differs between Urban LREB and Urban Medieval Islamic groups. Measures of body size length were positively correlated with δ13 C values in all individuals and separately in the Urban LREB and Rural Medieval Islamic groups. δ15 N showed a positive correlation with left humerus shape in the Urban LREB sample. CONCLUSIONS: The low stature and cross-sectional properties of the Medieval Rural Islamic group may be an indicator of greater physiological stress, potentially due to poorer diet. Positive correlations between measures of body size and δ13 C values further suggest that greater access to C4 resources improved diet quality. Alternatively, this relationship could indicate greater body size among migrants from areas where individuals consumed more C4 resources.

Metric variation of the tibia in the Mediterranean: Implications in forensic identification.

Ancestry estimation from skeletal remains is a challenging task, but essential for the creation of a complete biological profile. As such, the study of human variation between populations is important for the fields of biological and forensic anthropology, as well as medicine. Cranial and dental morphological variation have traditionally been linked to geographic affinity resulting in several methods of ancestry estimation, while the postcranial skeleton has been systematically neglected. The current study explores metric variation of the tibia in six Mediterranean populations and its validity in estimating ancestry in the Mediterranean. The study sample includes 909 individuals (470 males and 439 females) from Cyprus, Greece, Italy, Spain, Portugal and Turkey. The sample was divided in two subsamples: a reference and a validation sample. Multinomial regression models were created based on the reference sample and then applied to validation sample. The first model used three variables and resulted in 57% and 56% accuracy for the two samples respectively, while the second model (6 variables) resulted in 80% and 74% respectively. Classification between groups ranged from 28% to 95% for the reference sample and from 15% to 91% for the validation sample. The highest classification accuracy was noted for the Greek sample (95% and 90% for the reference and validation sample respectively), followed by the Turkish sample (74% and 78% respectively). The Spanish, Portuguese and Italian samples presented greater morphological overlap which resulted in lower classification accuracies. The results indicate that although the tibia presents considerable variation amongst neighbour populations it is not suitable as a sole skeletal element to separate all groups successfully. A combination of different skeletal elements may be required in order to achieve the levels of reliability required for forensic applications.

Ancestry estimation of three Mediterranean populations based on cranial metrics.

The estimation of ancestry is an essential benchmark for positive identification of heavily decomposed bodies that are recovered in a variety of death and crime scenes. This is especially true when reconstructing the biological profile of the deceased as most methods for sex, age and stature estimation are population-specific. Ancestry estimation methods vary from traditional morphological assessment of cranial features and biometric quantification to computer-aided shape analysis and classification with specialised software. The current paper aims to explore population differences between three neighbouring countries (Greece, Cyprus and Turkey) that have been in constant interaction through conflicts and population movements from the ancient past to the present day, through cranial measurements. The sample consists of 160 dry crania of Greek origin, 137 dry crania of Greek-Cypriot of origin Cyprus and 380 CT scans from Turks individuals. Twelve measurements were taken in both dry and virtual skulls. Data were submitted to principal component analysis and discriminant function analysis. Intra- and inter-observer error as well as the measurement error between virtual and physical measurements were quantified using TEM, rTEM and R. Measurement error was very low in all cases. Classification accuracy for cross-validated data ranged from 74.1 to 97.9%. The highest accuracy was obtained for the Turks sample both in males and females. The results are in accordance with genetic data on the three populations. These results create great confidence in the application of the produced functions in forensic cases requiring ancestry estimation in Cyprus, specifically to unidentified individuals from the 1974 conflict. In addition, these standards can be applied in other forensic situations where ethnicity is an issue but the geographic area of origin is limited to the area encompassing Turkey, Cyprus and Greece.

Sex estimation of the tibia in modern Turkish: A computed tomography study.

The utilization of computed tomography is beneficial for the analysis of skeletal remains and it has important advantages for anthropometric studies. The present study investigated morphometry of left tibia using CT images of a contemporary Turkish population. Seven parameters were measured on 203 individuals (124 males and 79 females) within the 19-92-years age group. The first objective of this study was to provide population-specific sex estimation equations for the contemporary Turkish population based on CT images. A second objective was to test the sex estimation formulae on Southern Europeans by Kranioti and Apostol (2015). Univariate discriminant functions resulted in classification accuracy that ranged from 66 to 86%. The best single variable was found to be upper epiphyseal breadth (86%) followed by lower epiphyseal breadth (85%). Multivariate discriminant functions resulted in classification accuracy for cross-validated data ranged from 79 to 86%. Applying the multivariate sex estimation formulae on Southern Europeans (SE) by Kranioti and Apostol in our sample resulted in very high classification accuracy ranging from 81 to 88%. In addition, 35.5-47% of the total Turkish sample is correctly classified with over 95% posterior probability, which is actually higher than the one reported for the original sample (25-43%). We conclude that the tibia is a very useful bone for sex estimation in the contemporary Turkish population. Moreover, our test results support the hypothesis that the SE formulae are sufficient for the contemporary Turkish population and they can be used safely for criminal investigations when posterior probabilities are over 95%.

Accuracy and sampling error of two age estimation techniques using rib histomorphometry on a modern sample.

Most age estimation methods are proven problematic when applied in highly fragmented skeletal remains. Rib histomorphometry is advantageous in such cases; yet it is vital to test and revise existing techniques particularly when used in legal settings (Crowder and Rosella, 2007). This study tested Stout & Paine (1992) and Stout et al. (1994) histological age estimation methods on a Modern Greek sample using different sampling sites. Six left 4th ribs of known age and sex were selected from a modern skeletal collection. Each rib was cut into three equal segments. Two thin sections were acquired from each segment. A total of 36 thin sections were prepared and analysed. Four variables (cortical area, intact and fragmented osteon density and osteon population density) were calculated for each section and age was estimated according to Stout & Paine (1992) and Stout et al. (1994). The results showed that both methods produced a systemic underestimation of the individuals (to a maximum of 43 years) although a general improvement in accuracy levels was observed when applying the Stout et al. (1994) formula. There is an increase of error rates with increasing age with the oldest individual showing extreme differences between real age and estimated age. Comparison of the different sampling sites showed small differences between the estimated ages suggesting that any fragment of the rib could be used without introducing significant error. Yet, a larger sample should be used to confirm these results.