Elsevier

Bone

Volume 51, Issue 3, September 2012, Pages 614-620
Bone

Review
Variation in osteocytes morphology vs bone type in turtle shell and their exceptional preservation from the Jurassic to the present

https://doi.org/10.1016/j.bone.2012.05.002Get rights and content

Abstract

Here we describe variations in osteocytes derived from each of the three bone layers that comprise the turtle shell. We examine osteocytes in bone from four extant turtle species to form a morphological ‘baseline’, and then compare these with morphologies of osteocytes preserved in Cenozoic and Mesozoic fossils. Two different morphotypes of osteocytes are recognized: flattened-oblate osteocytes (FO osteocytes), which are particularly abundant in the internal cortex and lamellae of secondary osteons in cancellous bone, and stellate osteocytes (SO osteocytes), principally present in the interstitial lamellae between secondary osteons and external cortex. We show that the morphology of osteocytes in each of the three bone layers is conserved through ontogeny. We also demonstrate that these morphological variations are phylogenetically independent, as well as independent of the bone origin (intramembranous or endochondral). Preservation of microstructures consistent with osteocytes in the morphology in Cenozoic and Mesozoic fossil turtle bones appears to be common, and occurs in diverse diagenetic environments including marine, freshwater, and terrestrial deposits. These data have potential to illuminate aspects of turtle biology and evolution previously unapproachable, such as estimates of genome size of extinct species, differences in metabolic rates among different bones from a single individual, and potential function of osteocytes as capsules for preservation of ancient biomolecules.

Graphical abstract

Highlights

► Three different morphologies of osteocytes are recognized in turtle bone. ► Osteocyte morphology is ruled by bone type in the turtle shell. ► Preservation of osteocytes in fossil bone is independent of geologic time. ► We show preservation of osteocytes in turtles from 150 Ma to Present.

Introduction

Bone is the result of phylogenetic, functional, and structural influences [1] evidenced in the hierarchical levels of bone tissue: macrostructure (cancellous and cortical bone), microstructure (Haversian systems, osteons, and lamellae), and nanostructure (mineral, collagen, and non-collagenous proteins) [2]. Of the three types of cells comprising bone, osteocytes are the most abundant, making up 95% of all cells in bone ([3] and references therein), yet little is known of osteocyte biology and function. Recent studies have begun to elucidate the role of osteocytes in bone formation, bone function, bone maintenance and bone pathology [4], [5], [6], [7], [8], [9], [10], [11], [12], but many questions regarding the fundamental biology of these cells remain. Issues that are still poorly understood include: 1) the potential variation in the morphology of osteocytes in bones with different origins (intramembranous vs endochondral) and their different roles in vertebrate body plans, (e.g., do osteocytes function differently in long bones vs bony flat plates); 2) what temporal limits exist on osteocyte preservation in the bony matrix, and whether preservation is dependent on taxon, bone type, geologic time, depositional environment or other factors; and 3) if these cells persist and can be shown to be endogenous to the organisms, can chemical/molecular analyses of these remnants shed light on the physiology, phylogeny, and/or ecology of extinct organisms across geological time. Turtles are an ideal organism to inform on these issues, because their bony shell is unique among vertebrates, in that their carapace is endochondral in origin, while the plastron has an intramembranous origin [13] (see Graphical abstract). Additionally, their shell consists of bones with three well differentiated bone types or layers [14], and a robust and continuous fossil record extending approximately up to 230 million years (Ma).

Osteocyte lacunae and associated lacunocanalicular network (LCN) have been described in many fossil specimens [15], [16], [17], including non-avian dinosaurs from the Late Cretaceous (80 Ma) of Mongolia. Two different morphs of these structures have been identified: flattened-oblate and stellate [18]. More recently, three dimensional osteocytes and blood vessel morphs have been isolated from the bone matrices of various Mesozoic (dinosaurs) and Cenozoic (mammals-birds) vertebrates [19], [20], but an in-depth examination of turtle bone for such preservation has not been previously conducted.

Here we describe osteocytes from four extant turtle taxa: sea turtle Caretta caretta (Cryptodira, Chelonidae), box turtle Terrapene carolina (Cryptodira, Testudinidae), freshwater turtle Trachemys scripta (Cryptodira, Emydidae), and freshwater side-necked turtle Podocnemis expansa (Pleurodira, Podocnemididae). We then examine the preservation of osteocytes in Cenozoic and Mesozoic fossils closely related to these extant species, and describe morphological variation in osteocytes from each of the three layers of bone that forms the shell—external cortex (EC), cancellous bone (CB), and internal cortex (IC). Finally, we compare these morphologies with osteocytes recovered from long bones (femora and humeri).

Section snippets

Sampling

Complete skeletons of each extant species were donated for research from the Amphibian and Herpetological collections from the North Carolina Museum of Natural Sciences. We studied carapace and plastron elements of C. caretta, Tr. scripta, and P. expansa, as well as the right femur of T. carolina and C. caretta. Specimens used for comparing osteocyte morphology are described in the caption of Fig. 4.

Bone histology

Eleven bone thin sections (Table 1) were taken following the procedure described in Ref. [21],

Osteocyte morphology in turtles

Two morphologically distinct types of osteocytes can be identified in ground sections of extant and extinct turtle bone or in isolated osteocytes after demineralization of each of these morphs specifically associated with variations in bone tissue (Fig. 1, Fig. 2, Fig. 3). These two morphologies are also conserved through ontogeny for each bone type, and for both shell elements (carapace and plastron), as is supported from hatchling, juvenile, and adult specimens of Mongolemys elegans from the

Discussion

Turtle osteocytes retain the typical stellate morphology mentioned by Ref. [22], also reported in non-avian dinosaurs [16] and other vertebrates [23], [24]. One notable difference, however is that FO1 osteocytes (Fig. 1a) are more abundant in the turtle shell than in any other vertebrate, possibly due to the proximity of the internal cortex to the soft body of the turtle. The very flat shape of FO1 osteocytes observed in this layer of lamellar bone may be the result of the density and

Acknowledgments

This work was supported by National Science Foundation grants OISE 0638810, EAR 0642528, and EAR 0824299, Geological Society of America Southeastern section Graduate Student Research Grant 2010 the Smithsonian Institution, the Panama Canal Authority, Mr. Mark Tupper, and SENACYT. Thanks to one anonymous reviewer for helpful comments. For access to samples of modern and fossil bones, thanks to B. Stuart and V. Schneider (North Carolina Museum of Natural Sciences, Raleigh, NC, USA). For support

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