Bone protects proteins over thousands of years: extraction, analysis, and interpretation of extracellular matrix proteins in archeological skeletal remains

In a good state of preservation, bone conserves the entire protein pattern of extracellular bone matrix proteins over thousands of years. The quality of the profiles of matrix proteins isolated from ancient bones (ranging from the pre-Pottery Neolithic Phase to Early Modern Times from different archaeological sites in different geographical areas), separated by electrophoresis, is as good as those from recent bones. Molecules arising from collagenous proteins (e.g., collagen type I), from the noncollagenous group (e.g., osteonectin), and from the immune system (e.g., immunoglobulin G) were identified in Western blots by specific antibodies. A comparative study of the immunoglobulin G content of the bones of five prehistoric children showed the lowest immunoglobulin G content in a child who suffered from chronic scurvy. Ancient bone proteins were also separated by two-dimensional electrophoresis. This technique makes fractionation of the complex protein mixtures of extracellular bone matrix more reproducible. Bone retains a chemical memory of earlier metabolic stimuli in its configuration of collagenous and noncollagenous proteins. In combination with the results of the microscopic examination of ancient bone, it should be possible to obtain more reliable information on the history and the evolution of diseases, based on analysis of intact proteins.     

Probable evidence of scurvy in subadults from archeological sites in Peru

Subadult scurvy is not well documented in archeological human remains despite the existence of many biomedical references indicating that bone changes do occur in some cases and, because of this, should be observable in human burials. There are several potential reasons for this gap in our knowledge of scurvy. Not all children who suffered from scurvy died of the disease or from other causes when they had scurvy. Scurvy may not leave characteristic bone changes in every case of the disease. Some of the pathological conditions associated with scurvy have been known for many years, but these features may be rare or difficult to differentiate from other pathological conditions. Recently a lesion of the skull has been described that is probably pathognomonic for scurvy, specifically porous and sometimes hypertrophic lesions of the greater wing of the sphenoid. This lesion is bilateral and highly associated with evidence of inflammation at other anatomical sites in the skull. A survey of subadult skulls (N = 363) in the human skeletal collection from Peru at the National Museum of Natural History, Smithsonian Institution, reveals a prevalence of 10% of skulls that exhibit plausible evidence of scurvy. Some cases of scurvy also have cribra orbitalia that has been attributed to anemia. In most of the Peruvian scurvy cases, anemia is an unlikely possibility because there is no evidence of marrow hyperplasia. This highlights the need for caution in using lesions of the orbit as an indicator of anemia when there is no other evidence of this disease elsewhere in the skeleton. Anatomical evidence of scurvy offers the potential of providing new and important evidence of diet in archeological human populations.     

Effects of different kinds of cranial deformation on the incidence of wormian bones

Researchers have debated whether the presence and frequency of wormian bones (sutural bones, supernumerary bones, and ossicles) are attributable to genetic factors, environmental factors, or both. This research examines the effects of many different kinds of cranial deformation on the incidence of wormian bones. A sample of 127 deformed and undeformed crania from New World archaeological sites was examined. An undeformed cranial sample (n=35) was compared to the following cranially deformed groups: 1) occipital, 2) lambdoid, 3) annular, 4) fronto-vertico-occipital, 5) parallelo-fronto-occipital, and 6) sagittal synostosis. Three levels of degree of cultural cranial deformation were qualitatively determined. Type and number of wormian bones along each major suture were recorded for each cranium. Group means were analyzed using Kruskal-Wallis one-way ANOVA statistical tests to test the null hypothesis that cranial deformation does not have an effect on wormian bone incidence. Results indicate that all forms of cranial deformation affect the frequency of some types of wormian bones. In particular, all cranially deformed groups exhibited significantly greater frequencies of lambdoid ossicles. Apical, parieto-mastoid, and occipito-mastoid wormian bones also appeared with greater frequency in some groups of culturally deformed crania. Further, varying degrees of cultural deformation all had more lambdoid wormian bones than the undeformed group. These results suggest that wormian bone development in posteriorly placed sutures may be affected more by environmental forces than are their anteriorly placed counterparts.     

Brief communication: unusual finding at Pueblo Bonito: multiple cases of hyperostosis frontalis interna

Hyperostosis frontalis interna (HFI) is a disease characterized by excess bone growth on the internal lamina of the frontal bone and, occasionally, other cranial bones. Although the disease is fairly common in modern populations, its etiology is poorly understood. Hyperostosis frontalis interna has been identified in antiquity, primarily in the Old World, but with a much lower frequency than in modern groups. The purpose of the present study is to report multiple cases of HFI at Pueblo Bonito (Chaco Canyon, New Mexico). Twelve out of 37 adults with observable frontal bones exhibited HFI, ranging from mild to severe, including 11 females and one male. This is the first published case report of HFI in archaeological remains from the New World having a frequency comparable with modern groups. Most archaeological cases of HFI are isolated, so comparative data for multiple cases at one site are rare. The results of this study emphasize the importance of looking for HFI in archaeological remains, although it is rarely observed. Possible genetic and environmental factors for the high frequency of HFI at Chaco Canyon are considered, but additional research is needed to discover the etiology and to better understand why HFI sometimes occurs at modern frequencies in ancient populations.     

旧石器遗址出土烧骨的技术分析及其对考古学的启示

烧骨常见于各类旧石器时代遗址中,作为用火残留物的一类,烧骨对研究人类用火行为有着极为重要的意义,而目前国内针对烧骨的研究工作开展得尚不够深入。本文通过介绍目前烧骨研究领域中若干种主流的分析手段及其在考古学上的应用,以求能进一步完善国内关于旧石器时代人工用火的研究框架,同时希望能对中国古人类用火能力的相关研究提供有益的借鉴。     

Long-term effects of tissue expansion on cranial and skeletal bone development in neonatal miniature swine: clinical findings and histomorphometric correlates

Progressive tissue expansion induces significant gross, histologic, and bony changes in skulls and long bones of neonatal miniature swine. These bony changes consist of erosion underlying tissue expanders, with bony lipping and bone deposition at the periphery of the expander. Cranial suture lines underneath expanders appear effaced and convoluted. Serial CT scans reveal decreased bone thickness and volume (p less than 0.02) but identical bone density (p = 0.60) beneath expanders. Increased bone volume and thickness occur at the periphery of expanders (p less than 0.02). Bone density (CT number) is unaffected by tissue expansion in both cranial and long bones. These findings have histomorphometric correlates: Osteoclastic bone resorption occurs underneath expanders with periosteal reaction at the periphery of expanders. Cranial sutures are similarly affected, but no cranial synostosis results. No changes to the inner table of the skull or stigmata of increased intracranial pressure were observed either in CT scans or in behavioral changes in long-term animals. The pathophysiology of bony changes is a remodeling effect, not one of simple pressure deformation. Increased bone resorption and complete inhibition of bone formation occur until the pressure is removed. Cranial bone is significantly more affected than long bone. After removal of the expanders, reparative bone remodeling begins within 5 days and nearly complete healing of the cranial defects occurs within 2 months (p less than 0.02). No plagiocephaly results despite early coronal suture changes. On the basis of this study, we conclude that tissue expansion causes significant but reversible effects, readily monitored by high-resolution CT scans, on neonatal and infant cranial and long bones.     

Remodeling of the sagittal suture in osteopetrotic (op/op) mice associated with cranial flat bone growth.

It is well known that cranial flat bone experiences growth and development at the sutural interface, which is regarded as a neutral zone to control mechanical stimuli. In osteopetrotic (op/op) mice, meanwhile, cranial deformation is produced by the deficiency of osteoclasts and the subsequent defect of bone resorption. It would be a reasonable assumption that such disturbance in bone remodeling affects sutural modification and the relevant cranial flat bone development. The present study was thus conducted to examine histological features of the sagittal sutures in op/op mice, with special reference to the relevant bone remodeling. The sagittal sutures in 10-, 15-, 30-, and 60-day-old normal and op/op mice were observed microscopically. Furthermore, osteoclastic activity was evaluated on the sections stained with tartrate-resistant acid phosphatase (TRAP). The sutures of 15-day-old op/op mice showed stenosis and synostosis, and less-developed collagen fibers associated with an irregular arrangement of fibroblasts, whereas these changes were rarely found in normal mice. Osteoclasts were hardly detected in the parietal bones around the sutures of op/op mice, although the number was numerous in normal mice. These results emphasize that congenital deficiency in osteoclast produces unbalanced bone remodeling at the sutural interface and on the surfaces of the cranial bones, which is assumed to be closely related to cranial bone deformity in op/op mice.     

A quantitative study of morphological and histological changes in the skeleton of Salmo salar during its anadromous migration

A quantitative study of morphological and histological changes in the skeleton (cranial bones and vertebrae) of adult Atlantic salmon Salmo salar during its anadromous migration was performed in order to specify various aspects of its skeletal biology in relationship to this migration. At the beginning of the ascent, there was no morphological difference in the cranial bones between males and females. As the spawning season approached, males showed marked secondary sexual characters particularly allometric breeding growth of some bones of the skull. The histological analysis of the vertebral bone tissue along the vertebral axis showed that bone compacity and number of trabeculae vary depending on their localization on the vertebral axis. Moreover, bone compacity decreased significantly with the sexual maturation of the fish whereas the number of trabeculae grew in both sexes. Thus, the vertebrae (like scales) represent an important source of calcium and other elements during anadromous migration in Atlantic salmon.     

Pathological lesions attributable to vitamin deficiency in skeletal remains from Puy St. Pierre (Briançon, France)

The aim of the present study was to evaluate the presence of pathological alterations attributable to vitamin deficiency in osteological remains from the church of St. Pierre in Puy de St. Pierre, Briançon, France. We recorded alterations referable to scurvy, an acquired osteodystrophy due to vitamin C deficiency, which causes both skeletal and vascular lesions in humans. Scurvy presents a different clinical picture in children and adults: in the latter, in leaves skeletal signs on the palate and teeth. The recording of osteodystrophies attributable to vitamin deficiency allowed us to make general inferences about the nutritional profile of the group under study. The study of this pathology, as well as that of nutritional stresses, can be related to the geographical-environmental conditions of the population to which the sample belongs. This is important from the anthropological point of view, especially in order to examine alimentary habits within the context of the man-environments relationship.     

Scurvy in the Great Irish Famine: evidence of vitamin C deficiency from a mid-19th century skeletal population

Scurvy has increasingly been recognized in archaeological populations since the 1980s but this study represents the first examination of the paleopathological findings of scurvy in a known famine population. The Great Famine (1845-1852) was a watershed in Irish history and resulted in the death of one million people and the mass emigration of just as many. It was initiated by a blight which completely wiped out the potato-virtually the only source of food for the poor of Ireland. This led to mass starvation and a widespread occurrence of infectious and metabolic diseases. A recent discovery of 970 human skeletons from mass burials dating to the height of the famine in Kilkenny City (1847-1851) provided an opportunity to study the skeletal manifestations of scurvy-a disease that became widespread at this time due to the sudden lack of Vitamin C which had previously almost exclusively been provided by the potato. A three-scale diagnostic reliance approach has been employed as a statistical aid for diagnosing the disease in the population. A biocultural approach was adopted to enable the findings to be contextualized and the etiology and impact of the disease explored. The results indicate that scurvy indirectly influenced famine-induced mortality. A sex and stature bias is evident among adults in which males and taller individuals displayed statistically significantly higher levels of scorbutic lesions. The findings have also suggested that new bone formation at the foramen rotundum is a diagnostic criterion for the paleopathological identification of scurvy, particularly among juveniles.     

Ossification of the human fetal basicranium

Previous investigations of prenatal development of the human cranium have not identified the sequence of its ossification. The purpose of the present study was to elucidate the pattern of skeletal maturity of the cranial bones in the midsagittal region anterior to the foramen magnum. This study is based upon a radiographic and histochemical investigation of midsagittal tissue blocks of the cranial bases of 73 human fetuses derived from the first half of the prenatal period. A marked regularity in the ossification pattern of the bones in the midsagittal part of the human cranium was observed. Ossification starts in the frontal bone. The sequence in which the next bones ossify is occipital bone, basisphenoid bone, presphenoid bone, and ethmoid bone. The material was divided into 7 maturity stages devised for this analysis. The stages were related to general fetal size (crown-rump length) and to general fetal maturation (composite number of ossified bones in hand and foot). Skeletal development of the median part of the human cranium is not strictly correlated with the size or the stage of general maturation of the fetuses. Knowledge of normal skeletal development is necessary for understanding anomalies of development.     

The Identification of Proteoglycans and Glycosaminoglycans in Archaeological Human Bones and Teeth

Bone tissue is mineralized dense connective tissue consisting mainly of a mineral component (hydroxyapatite) and an organic matrix comprised of collagens, non-collagenous proteins and proteoglycans (PGs). Extracellular matrix proteins and PGs bind tightly to hydroxyapatite which would protect these molecules from the destructive effects of temperature and chemical agents after death. DNA and proteins have been successfully extracted from archaeological skeletons from which valuable information has been obtained; however, to date neither PGs nor glycosaminoglycan (GAG) chains have been studied in archaeological skeletons. PGs and GAGs play a major role in bone morphogenesis, homeostasis and degenerative bone disease. The ability to isolate and characterize PG and GAG content from archaeological skeletons would unveil valuable paleontological information. We therefore optimized methods for the extraction of both PGs and GAGs from archaeological human skeletons. PGs and GAGs were successfully extracted from both archaeological human bones and teeth, and characterized by their electrophoretic mobility in agarose gel, degradation by specific enzymes and HPLC. The GAG populations isolated were chondroitin sulfate (CS) and hyaluronic acid (HA). In addition, a CSPG was detected. The localization of CS, HA, three small leucine rich PGs (biglycan, decorin and fibromodulin) and glypican was analyzed in archaeological human bone slices. Staining patterns were different for juvenile and adult bones, whilst adolescent bones had a similar staining pattern to adult bones. The finding that significant quantities of PGs and GAGs persist in archaeological bones and teeth opens novel venues for the field of Paleontology.     

Lateral angle: a method for sexing using the petrous bone

We report on the results of applying the so-called lateral angle method for sex determination on skeletal remains. The lateral angle denotes the angle of the internal auditory canal in relation to the medial surface of the petrous part of the temporal bone. The method involves making a small cast of the proximal part of the internal acoustic canal and determining the angle at which the canal opens up to the surface of the petrous bone. The method has the great advantage of utilizing one of the sturdiest bone elements of the human skeleton, and may thus be especially suited for analyses of very fragmented skeletal remains or cremated bones, where the petrous bone may still be readily recognizable. The method was tested using a forensic sample of 113 petrous bones with known sex. Intra- and interobserver testing was also performed. We found a statistically significant difference in angle size between males and females (mean angle size of males, 39.3 degrees ; mean angle size of females, 48.2 degrees ; P < 0.001). There was no bilateral difference in angle size. In blind trials, 83.2% of petrous bones were assigned to the correct sex. We also tested the lateral angle method against an archaeological skeletal sample. True sex was not known for this sample; instead, sexing had been carried out by assessing pelvic and cranial morphology in independent trials. We found a higher concordance between the lateral angle and "pelvic" sex than for lateral angle and "cranial" sex. Finally, we note that subadult sexing may also be possible with this method.     

Brief Communication: A likely case of scurvy in a rural Early Classic Maya burial from Actun Uayazba Kab,Belize

A Maya burial of a late adolescent (Burial 98‐3) found in the rockshelter entrance of Actun Uayazba Kab (AUK), Belize, displays a combination of lesions that is consistent with scurvy. Signs include large, active lesions on the posterior surfaces of maxilla; relatively mild porotic hyperostosis along the midline of the skull on the parietals and occipital; cribra orbitalia; potential pinprick lesions on the greater wings of sphenoid and temporal; reactive lesions on the palate, temporal lines of frontal and parietals, and external and internal surfaces of zygomatics; small lesions on the popliteal surfaces of both femora; and periodontal disease. Identification of scurvy at AUK potentially informs the analysis of other primary burials and scattered bone found there and at other nearby sites, which often reveal evidence of nonspecific lesions that are usually attributed to anemia and infection, but that are also consistent with scurvy. The social and ecological context of this Protoclassic (0–AD 300) individual, who lived in a rural agricultural community with no evidence of complex social hierarchy, contrasts with typical discussions of disease among the Maya, which tend to focus on the degrading effects of overcrowding and resource deficiencies. While scurvy has been largely overlooked in the Maya area, this study supports earlier arguments for its presence that were based largely on clinical and ethnographic analogies and suggests the need to incorporate scurvy into broader synergistic models of ancient health. Am J Phys Anthropol 155:476–481, 2014. © 2014 Wiley Periodicals, Inc.     

Identification of semi-domesticated reindeer (Rangifer tarandus tarandus,Linnaeus 1758) and wild forest reindeer (R.t. fennicus,Lönnberg 1909) from postcranial skeletal measurements

Reindeer bones are common finds from archaeological sites from prehistoric and historic Fennoscandia. The interpretation of the reindeer bone finds, however, is often hindered by the difficulty to separate the different subspecies of reindeer using the postcranial skeletal morphology. In this paper, skeletal measurements of modern semi-domesticated reindeer and wild forest reindeer are explored with multivariate statistical methods in order to find suitable methods for subspecies identification. The results are then applied to archaeological reindeer bone finds from Northern Finland and archaeological implications of the results are discussed.     

Radiographic determination of prenatal basicranial ossification

In a previous investigation on prenatal development of the human cranial base, the sequence in which the bones develop in the midsagittal region was elucidated. The purpose of the present study was to identify fetal ossification on horizontal plane roentgenograms of the occipital and sphenoid bones in the central part of the cranial base, and establish stages in bone appearance related to general fetal developmental parameters. This study is based upon roentgenograms of the cranial base of 145 human fetuses from the first half of the prenatal period. Two different maturation patterns of the sphenoid bone were observed. The first, most common pattern is characterized by a midsagittal centre of ossification and the second by bilateral centres of ossification in the corpus of the sphenoid bone. These bilateral centres might in some cases be connected by a slight bony bridge. It appears that these different maturation patterns are maintained throughout the period investigated. The material was divided into five well-defined developmental stages for both maturation patterns and general parameters of fetal development. Mapping different aspects of ossification in normal cranial development is necessary for understanding deviations of cranial maturation and growth.     

On the trail of pulmonary tuberculosis based on rib lesions: results from the Human Identified Skeletal Collection from the Museu Bocage (Lisbon, Portugal)

In the last 20 years, studies on human identified skeletal collections have revealed a significant relationship between new bone formation on the visceral surface of ribs and pulmonary tuberculosis (TB). To improve methods of differential diagnosis of respiratory diseases in archaeological skeletons, an investigation was conducted on 197 individuals from the Human Identified Skeletal Collection of the Museu Bocage (Lisbon, Portugal). This sample included 109 males and 88 females who lived during the 19th-20th centuries, with ages at death ranging from 13-88 years. The skeletons were grouped according to cause of death: 1) pulmonary TB (N = 84); 2) pulmonary non-TB diseases (N = 49); and 3) a control group (N = 64) composed of individuals randomly selected among the extrapulmonary non-TB causes of death. The ribs, sterna, scapulae, and clavicles were macroscopically observed. New bone formation on the visceral surface of ribs was recorded in 90.5% (76/84) of individuals who died from pulmonary TB, in 36.7% (18/49) with a pulmonary non-TB disease as cause of death, and in 25.0% (16/64) of the control group. These differences were statistically significant (P < 0.001). Furthermore, in individuals with pulmonary TB, the bony lesions presented mainly as lamellar bone on the vertebral end of the upper and middle thoracic rib cage. Proliferative alterations also occurred on one sternum and in nine clavicles and eight scapulae. This work strongly supports the results of similar studies performed on other documented collections, suggesting that new bone formation on ribs, although not pathognomonic, is a useful criterion for the differential diagnosis of pulmonary TB.