Histological changes in the epidermis of the subdigital lamellae of Anolis carolinensis during the shedding cycle

The characteristic anoline climbing organ consists of a number of lamellar scales, on whose outer scale surface are numerous keratinized setae which contact the substrate. These setae are derived from the Oberhautchen of the epidermal generation, and as such are renewed and shed periodically along with the rest of the epidermal material. The histological development of the setae is described, and modifications of the surrounding elements are noted. The relative lengths of the setae and their congregation to form a pad unit poses certain mechanical problems during morphogenesis, simply in terms of accommodation between the functional outer epidermal generation and dermal core of each lamella. Regression of the dermal core and a distal migration of some cells permits accommodation within the lamella for the distal aspect of the Oberhautchen layer, or free margin. Additionally, changes in the gross shape of the lamella occur throughout the sloughing cycle, and a swelling of the cells of the lacunar tissue results in a gap between the stratum corneum of inner and outer epidermal generations. There is a considerable amount of variation in mitotic activity between the germinal layers of opposite sides of the lamella.     

Scanning electron microscopy of scales from different body regions of three lizard species

The Oberhautchen of scales from the dorsal, parietal, and ventral regions of Sceloporus occidentalis (Iguanidae), Gerrhonotus multicarinatus (Anguinidae), and Anniella pulchra (Anniellidae) were examined with a scanning electron microscope. At low magnification, all scales of S. occidentalis exhibit well-defined outlines of cells belonging to the Oberhautchen layer and the previously overlying clear layer. The dorsal and parietal cells of this species exhibit a minutely dentate Oberhautchen that forms tooth-like spinules 0.2 to 0.5 μ long and arranged in irregular rows. Minute pits 0.1 to 0.3 μ in diameter characterize the Oberhautchen of a ventral scale. Cell outlines are not evident on the scales of G. multicarinatus. The Oberhautchen of dorsal and parietal scales of this species is prominently laminated. Laminae are less prominent on scales of the lateral fold, and no intrinsic surface structure is evident on a ventral scale. In contrast, the fossorial anguinomorph Anniella pulchra exhibits Oberhautchen surfaces with practically no intrinsic microornamentation. However, what appear to be outlines of Oberhautchen cells are visible on the dorsal and ventral scales. These observations suggest that modifications of Oberhautchen microornamentation may have evolved to reduce friction with the substrate or other scales. The lack of pronounced microornamentation of the Oberhautchen on some body scales may indicate that a complex interdigitation between clear layer and Oberhautchen cells is not essential to the sloughing process.     

Morphogenesis of the digital pad lamellae in the embryo of the lizard Anolis lineatopus

The present study in the embryo of the lizard Anolis lineatopus describes the modality of cell proliferation responsible for the morphogenesis of the digital pad lamellae and of the epidermal stratification. After tritiated thymidine and 5-bromodeoxy-uridine administration, autoradiographic and immunocytochemical methods have been used. The lamellae originate as long, slightly slanted, undulations of the epidermis of fingers and toes. At an early stage, the epidermis consists of an outer periderm and a basal layer. Cell hypertrophy, and the prevalent cell proliferation in the longer side of the undulation with respect to the shorter side, generate the surface of the outer lamella. Under the peridermis, a shedding complex, composed by clear and oberhautchen layers, is formed and later determines the first intraepidermal shed. The first subperidermal layer derived from the basal layer is a clear layer and the first shed epidermis in the embryo is represented by periderm and clear layer. The heavily granulated clear layer in Anolis lineatopus represents the first epidermal layer produced in the embryonic epidermis, and is connected with the process of shedding. The spinulae of the underlying oberhautchen in the outer scale surface become long setae which grow toward the upper clear layer. Under the shedding complex a β-layer is produced. Autoradiographical study shows that the radioactivity stays in the basal layer for about 4 days before cells move to upper layers. At 6–8 days post-injection labelled cells are visible in the differentiated clear, oberhautchen and β-layers. Under the β-layer differentiating mesos cells are visible before the embryo hatches.     

The structure of anoline (Reptilia: Dactyloidae: Anolis) toe pads in relation to substratum conformity

Adhesive toe pads of geckos house modified components of vascular and/or connective tissues that promote conformity of the setal fields with the locomotor substratum. Similar modifications have been claimed for the digits of Anolis, but evidence for them is not compelling. Angiographic and histological investigations of Anolis failed to identify any evidence of either an intralamellar vascular reticular network or a central sinus. Instead, their vascularity more closely resembles that of lizards in general than that of pad‐bearing geckos. The loose connective tissue of the toe pads likely contributes to their general pliability and flexibility, promoting localized compliance with the substratum. Through the shedding cycle, the lamellae change shape as the replacing setae elongate. The outer epidermal generation lacunar cells on the inner lamellar faces simultaneously hypertrophy, providing for compatibility between overlapping lamellae, enabling reciprocity between them. This contributes to continuing compliance of the setal fields with the substratum. Overall, digital structure and attachment and release kinematics of the toe pads of Anolis are very similar to those of geckos exhibiting an incipient adhesive mechanism. Both lack major anatomical specializations for promoting conformity of the setae with the locomotor substratum beyond those of the seta‐bearing portions of the epidermis.     

Licht- und elektronenmikroskopische untersuchungen zur haftborstenentwicklung bei Tarentola m. mauritanica L. (Reptilia,Gekkonidae)

Zusammenfassung Kurz nach einer Hdutung wird bei Tarentola m. m. bereits die übemächste Epidermisgeneration — und somit auch die der Haftborsten —angelegt. Das geschieht vornehmlich in der Oz- (Oberhäutchenzellen-) und in der Hs-Schicht (clear layer). Zunächst entstehen die Aufspaltungen der Haftborstenenden, indem Keratinfilamentbündel nach einem bestimmten System von den Oz-Zellen aus in die Hs-Zellen einwachsen. Auf these Weise fungieren die Zellen der Hs-Schicht als Matrix der Haftborsten. Nach Abschluß dieses Prozesses werden die eigentlichen Haftborsten gebildet unter gleichzeitigem Auseinanderrücken der Hs- und Oz-Schichten. Die Hs-Schicht behdlt ihre Matrizen-Funktion bis zur anschließenden Häutung bei.

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Light and electron microscope studies of developing setae of Tarentola m. mauritanica (Rept., Gekkonidae)

Summary In Tarentola m. mauritanica the next epidermis generation but one and therefore the adhesive setae of the generation after this begin to develop shortly after a skin has been shed. This development takes place principally in the horny layer (Oz) and the clear layer. First bundles of keratin filaments radiate from the horny layer into the clear layer, thus giving rise to the split distal parts of the adhesive bristles. Thus the cells in the clear layer act as a matrix for the setae. When this stage is complete the formation of the setae proper begins, while the horny layer and the clear layer become separated from each other. The clear layer retains its function as matrix for the setae until the next time a skin is shed.

     

A whole lamella perspective on the origin of the epidermal free margin of Anolis (Reptilia: Dactyloidae) toe pads

Anoline lamellae terminate in an epidermal free margin carrying the majority of its setae. How the free margin is extruded from the body of the scale is not well understood. Two hypotheses have been advanced to account for it, one advocating distal migration of the outer epidermal layers relative to the body of the lamella, and the other proposing regression of the dermal core. Available evidence provides partial support for both. We assembled a series of specimens of Anolis grahami representing all shedding cycle stages, and prepared histological sections of the toe pads to allow measurement of appropriate lamellar components through the shedding cycle. Through its proliferative phases the lamellae increase markedly in length, with the distance between the distal tip of the dermal core and that of the lamella accounting for most of this, indicating that epidermal extrusion is responsible for production of the new free margin. The dermal core showed no evidence of regression. Concomitant with epidermal extrusion, the lacunar cells on the inner lamellar face hypertrophy and keep pace with the increasing thickness of the outer lamellar face resulting from the lengthening of the replacement setae. The integrated changes observed are consistent with continuity of functioning and alignment of the exposed setal carpet of the outer epidermal generation while ensuring that the new setal carpet is fully aligned and functional immediately after shedding. At shedding the original proportions of the lamellae are restored. Development of the new free margin results from a combination of distal displacement of Oberhäutchen cells along with arrested maturation of the epidermis in this region. Changes in length of the lamellae during the proliferative stages may impact the overall size of the adhesive toe pad, which may have consequences for assessments of the relationship between whole animal clinging ability and adhesive pad area. J. Morphol. 278:360–368, 2017. © 2017 Wiley Periodicals, Inc.     

Immunolocalization of keratin‐associated beta‐proteins in developing epidermis of lizard suggests that adhesive setae contain glycine–cysteine‐rich proteins

The localization of specific keratin‐associated beta‐proteins (formerly referred to as beta‐keratins) in the embryonic epidermis of lizards is not known. Two specific keratin‐associated beta‐proteins of the epidermis, one representing the glycine‐rich subfamily (HgG5) and the other the glycine‐cysteine medium‐rich subfamily (HgGC10), have been immunolocalized at the ultrastructural level in the lizard Anolis lineatopus. The periderm and granulated subperiderm are most immunonegative for these proteins. HgG5 is low to absent in theOberhäutchen layer while is present in the forming beta‐layer, and disappears in mesos‐ and alpha‐layers. Instead, HgGC10 is present in the Oberhäutchen, beta‐, and also in the following alpha‐layers, and specifically accumulates in the developing adhesive setae but not in the surrounding cells of the clear layer. Therefore, setae and their terminal spatulae that adhere to surfaces allowing these lizards to walk vertically contain cysteine–glycine rich proteins. The study suggests that, like in adult and regenerating epidermis, the HgGC10 protein is not only accumulated in cells of the beta‐layer but also in those forming the alpha‐layer. This small protein therefore is implicated in resistance, flexibility, and stretching of the epidermal layers. It is also hypothesized that the charges of these proteins may influence adhesion of the setae of pad lamellae. Conversely, glycine‐rich beta‐proteins like HgG5 give rise to the dense, hydrophobic, and chromophobic corneous material of the resistant beta‐layer. This result suggests that the differential accumulation of keratin‐associated beta‐proteins over the alpha‐keratin network determines differences in properties of the stratified layers of the epidermis of lizards. J. Morphol. 2013. © 2012 Wiley Periodicals, Inc.     

Immunolocalization of specific beta‐proteins in pad lamellae of the digits in the lizard Anolis carolinensis suggests that cysteine‐rich beta‐proteins provides flexibility

Knowledge of beta‐protein (beta‐keratin) sequences in Anolis carolinensis facilitates the localization of specific sites in the skin of this lizard. The epidermal distribution of two new beta‐proteins (beta‐keratins), HgGC8 and HgG13, has been analyzed by Western blotting, light and ultrastructural immunocytochemistry. HgGC8 includes 16 kDa members of the glycine‐cysteine medium‐rich subfamily and is mainly expressed in the beta‐layer of adhesive setae but not in the setae. HgGC8 is absent in other epidermal layers of the setae and is weakly expressed in the beta‐layer of other scales. HgG13 comprises members of 17‐kDa glycine‐rich proteins and is absent in the setae, diffusely distributed in the beta layer of digital scales and barely present in the beta‐layer of other scales. It appears that the specialized glycine‐cysteine medium rich beta‐proteins such as HgGC8 in the beta‐layer, and of HgGC10 and HgGC3 in both alpha‐ and beta‐layers, are key proteins in the formation of the flexible epidermal layers involved in the function of these modified scales in adaptation to contact and adhesion on surfaces. J. Morphol. 275:504–513, 2014. © 2013 Wiley Periodicals, Inc.     

Ultrastructural Features of Host-Parasite Relationship in Oral Candidiasis

In oral candidiasis, many keratinized epithelial cells and cells of Candida albicans are shed. Scales from patients with oral candidiasis were used for electron microscopic study of the epithelial-fungal relationship. Scales, scraped from the tongue and oral mucosa, were fixed for fungi. Electron microscopic observations showed cells of C. albicans outside, penetrating, or within the epithelial cells. Extracellular fungi possessed a floccular material adherent to the outer surface of the cell wall. Intracellular fungi lacked the floccular material which appeared to detach as fungi invaded the epithelial cells. Large vacuoles, which sometimes contained myelin figures, occupied the cytoplasm of fungal cells. Epithelial cells frequently contained several fungi. Discontinuous plasma membranes marked sites of fungal entry. Cytoplasmic areas devoid of fungi showed many tonofibrils, but the cytoplasm adjacent to fungi often lacked tonofibrils. Micrographs suggested that fungal cells lysed the tonofibrils. Bacteria were abundant in the scrapings, but always occupied an extracellular position.     

Histidine uptake in the epidermis of lizards and snakes in relation to the formation of the shedding complex

Mammalian epidermis utilizes histidine-rich proteins (filaggrins) to aggregate keratin filaments and form the stratum corneum. Little is known about the involvement of histidine-rich proteins during reptilian keratinization. The formation of the shedding complex in the epidermis of snakes and lizards, made of the clear and the oberhautchen layers, determines the cyclical epidermal sloughing. Differently from snakes, keratohyalin-like granules are present in the clear layer of lizards. The uptake of tritiated histidine into the epidermis of two lizards and one snake has been studied by autoradiography in sections at progressive post-injection periods. At 40 min and 1 hr post-injection keratohyalin-like granules were not or poorly labeled. At 3-22 hr post-injection most of the labeling was present over suprabasal cells destined to form the shedding complex, in keratohyalin-like granules of the clear layer, and in the forming a-layer but was low in the forming b-layer, and in superficial keratinized layers. The analysis of the shedding complex in the pad lamellae (a specialized scale used for climbing) of a gecko showed that the setae and the cytoplasm of clear cells among them are main sites of histidine uptake at 4 hr post-injection. In the snake most of the labeling at 4 hr post-injection was localized in the shedding complex along the boundary between the clear and oberhautchen layers. The present study suggests that, in the epidermis of lepidosaurian reptiles, the synthesis of a histidine-rich protein is involved in the formation of the shedding layer and, as in mammals, in a-keratinization.     

Structural organization of the toe pads in the amphibian Philautus annandalii (Boulenger, 1906)

We describe the morphology of toe pads in the Himalayan tree frog Philautus annandalii. These are expanded tips of digits and show modifications of their ventral epidermis for adhesion. The outer cells of toe pad epidermis (TPE) bear surface microstructures (0.7 × 0.2 μm), which are keratinized. Their cytoplasm contains no organelles, but pleomorphic nuclei and mucous granules (0.4–0.5 μm) that glue the keratin filaments. In the intermediate cell layer of TPE, similar keratinized microstructures as in the outer cells are present, so that when the outer layer is shed, it is ready with features for adhesion. These cells contain more keratin than the outer cells. The basal cell layer contains thin keratin bundles and usual cell organelles. The dermis contains mucous‐secreting glands, whose ducts open in the outer epidermal cell layer in channels. The dorsal epidermal cells lack surface microstructures and keratin bundles. Ultrastructural features suggest that toe pads utilize the surface microstructures for adhesion aided by mucus, in which the intermediate cell layer seems to bear the shear stress generated during locomotion. Further, TPE can expand and fit into an increased contact area of the substrate. The long, surface microstructures may also help in mechanical interlocking with rough surfaces on plants.     

Immunolocalization of large corneous beta‐proteins in the green anole lizard (Anolis carolinensis) suggests that they form filaments that associate to the smaller beta‐proteins in the beta‐layer of the epidermis

The distribution of large corneous beta‐proteins of 18–43 kDa (Ac37, 39, and 40) in the epidermis of the lizard Anolis carolinensis is unknown. This study analyses the localization of these beta‐proteins in different body scales during regeneration. Western blot analysis indicates most protein bands at 40–50 kDa suggesting they mix with alpha‐keratin of intermediate filament keratin proteins. Ac37 is present in mature alpha‐layers of most scales and in beta‐cells of the outer scale surface in some scales but is absent in the Oberhäutchen, in the setae and beta‐layer of adhesive pads and in mesos cells. In differentiating beta‐keratinocytes Ac37 is present over 3–4 nm thick filaments located around the amorphous beta‐packets and in alpha‐cells, but is scarce in precorneous and corneous layers of the claw. Ac37 forms long filaments and, therefore, resembles alpha‐keratins to which it probably associates. Ac39 is seen in the beta‐layer of tail and digital scales, in beta‐cells of regenerating scales but not in the Oberhäutchen (and adhesive setae) or in beta‐ and alpha‐layers of the other scales. Ac40 is present in the mature beta‐layer of most scales and dewlap, in differentiating beta‐cells of regenerating scales, but is absent in all the other epidermal layers. The large beta‐proteins are accumulated among forming beta‐packets of beta‐cells and are packed in the beta‐corneous material of mature beta‐layer. Together alpha‐keratins, large beta‐proteins form the denser areas of mature beta‐layer that may have a different consistence that the electron‐paler areas. J. Morphol. 276:1244–1257, 2015. © 2015 Wiley Periodicals, Inc.     

Ultrastructural and histochemical investigations of peripatus integument

The integument of Euperipatoides leukarti (Onychophora) has been studied by SEM and TEM-histochemical techniques. The thin (2-3 mu) cuticle is essentially arthropodan in design, comprising a thin, lamellate outer epicuticle, a homogeneous inner epicuticle, and an extensive fibrous procuticle. Five tanned lipoprotein lamellae constitute the outer epicuticle, the innermost showing a cross-striated pattern and the outermost also containing carbohydrates. The inner epicuticle contains untanned lipoproteins. The chitin-protein composite procuticle reveals a fibrous ultrastructure but no trace of helicoids. Distally it may be tanned, producing a sclerotin layer (exocuticle), pronounced in the sensory setae, claws and mandibles. Penetrating the epicuticle and extending into the procuticle are numerous, minute pore canals through which mobilised procuticle is resorbed prior to ecdysis. Beneath the procuticle is a monolayered epidermis traversed by tonofibrils and containing dense pigment granules. The membrane junctions are strongly convoluted and include an apical zonula adherens, septate desmosomes and a proximal lymph space. The epidermis is underlain by a thick (5-25mu) basement membrane of helicoidally organised collagen fibres. Functional and phylogetic implications of the integumental structure are discussed. Arthropod affinities are clear and the Onychophora should be included in that phylum if it is to be retained in modern taxonomic nomenclature.     

Histochemical and ultrastructural analyses of adhesive setae of lizards indicate that they contain lipids in addition to keratins

We studied the distribution of lipid material and organelles in the epidermal layers of toe pads from two species of lizards representing the two main lizard families in which adhesive scansors are found (gekkonids and polychrotids), the dull day gecko, Phelsuma dubia and the green anole, Anolis carolinensis. Although lipids are a conspicuous component of the mesos layer of squamate reptiles and function in reducing cutaneous water loss, their distribution has not been specifically studied in the highly elaborated epidermal surface of adhesive toe pads. We found that, in addition to the typical cutaneous water loss‐resistant mesos and alpha‐layer lipids, the Oberhäutchen (including the setae) on the most exterior layers of the epidermis in P. dubia and A. carolinensis also contain lipid material. We also present detailed histochemical and ultrastructural analyses of the toe pads of P. dubia, which indicate that lipid material is closely associated spatially with maturing setae as they branch during the renewal phase of epidermal regeneration. This lipid material appears associated with the packing of keratin within setae, possibly affecting permeability to water loss in the pad lamella, where the surface area is from 4–60‐fold greater compared with normal scales. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc.     

Ryocalanus spinifrons,a new species of Ryocalanidae (Copepoda: Calanoida), from the southwestern part of Sagami Bay,Japan

A new Ryocalanoid copepod, Ryocalanus spinifrons, collected by the MTD net system at a depth of 1400 m from the southwestern part of Sagami Bay, Japan, is described. The new species is morphologically very close to R. infelix Tanaka, 1956 (female unknown) from the Izu region of Sagami Bay. It is distinguished from other species by the presence of 12 long spinules on the ventral inner side of the fifth pedigerous somite, nine setae on the coxal epipodite of the maxillule and nine large robust spinules on the coxal segment of the fourth leg. The row of five robust spines on the paragnath distinguishes R. spinifrons.     

Keratinization of fish skin with special reference to the catfish Bagarius bagarius

Summary Histochemical reactions indicating keratinization have previously been demonstrated in parts of the epidermis of Bagarius bagarius. Fluorescence histochemistry and electron microscopy have now confirmed these results. Elevated areas of the epidermis are capped by a layer of dead cells with altered contents. On the outer aspect of these cells a dense layer, 18 nm thick, beneath the plasma membrane corresponds to the resistant envelope found in keratinized cells in tetrapod vertebrates. In Bagarius this layer does not extend to all faces of the keratinized cells, but a similar envelope has been detected in two other sites of piscine keratinized epidermis investigated, namely in the breeding tubercles of Phoxinus phoxinus and in the teeth of Lampetra fluviatilis. In the elevated areas of Bagarius-epidermis, the epithelial cells undergo progressive changes in cytoplasmic organization as they become more superficial. The second tier from the surface is sealed by tight junctions and is separated from the overlying keratinized cells by a sub-corneal space resembling that found in keratinized amphibian epidermis. Histochemical evidence of a high lipid content in the outer layers of the epidermis correlates with the presence of lipid inclusions and lamellated membranous profiles in the material studied by electron microscopy. Histochemical results show that the fin skin of Blennius pholis is not keratinized, but secretes a cuticle, histochemically reactive for both proteins and glycoproteins.     

Ultrastructural contributions to the identification of cell types in the lizard epidermal generation

The ultrastructure of the epidermis at different stages of the shedding cycle has been studied in Anolis carolinensis. Cells of the germinal layer are morphologically similar at all stages in the cycle. Immediately after leaving the germinal layer all daughter cells resemble one another closely. However, they later acquire specific ultrastructural features that enable them to be classified into six distinct fully differentiated types corresponding to the grouping previously set forth by light microscopy. A comparison of cytoplasmic filament size with the known X-ray diffraction data suggests that the Oberhautchen and β-layer contain a protein similar to that of avian feather; the protein in the α-layer and lacunar tissue is similar to that in mammalian hair, and the mesos layer cells probably contain a mixture of feather and hair-like proteins. The nature of the amorphous cytoplasmic material in the mature clear layer is as yet unknown.     

Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesenchymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissueengineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melanosomes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66% ± 4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type I mRNA expression in differentiated cells; radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06 pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material shortened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering.     

Fine structure of the dorsal lingual epithelium of the little tern, Sterna albifrons Pallas (Aves, Lari).

The dorsal surface of the tongue of the little tern, Sterna albifrons, has a distinctive anterior region for five-sixths of its length and a terminal posterior region. The anterior region observed by scanning electron microscopy is distinguished along its forward half by a median line from which median papillae protrude. The hind half of the anterior region has a median sulcus without papillae. The deciduous epithelium on both sides of the median line and sulcus bears scattered epithelial protrusions. The posterior lingual region has neither median papillae nor deciduous epithelium. So-called giant conical papillae are located in a transverse row between anterior and posterior regions. Delicate microridges adorn the surfaces of all outer epithelial cells in both regions. Examination of the dorsal lingual epithelium by light and electron microscopy provides histologic and cytologic criteria for distinguishing anterior and posterior regions. Basal cells are nearly alike throughout the dorsal epithelium. Intermediate layer cells of the anterior region contain numerous tonofibrils in electron-dense bundles composed of 10 nm tonofilaments. The outer layer is composed of electron-dense, well-keratinized cells, and electron-lucent epithelial protrusions are present on the exposed surface of the outermost cells. Median papillae are composed of typical keratinized cells, which are nearly filled with keratin filaments. Intermediate layer cells in the posterior region of the tongue are nearly filled with unbundled tonofilaments. There is only a very thin outer keratinized layer in this region.     

Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesenchymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissueengineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melanosomes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66% ± 4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type I mRNA expression in differentiated cells; radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06 pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material shortened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering. Supported by the Major Technology Program of Beijing Municipal Science & Technology Commission (Grant No. H060920050130) and the Major State Basic Research Development Program of China (Grant No. 2005CB522702)