Bisphosphonate affects the behavioral responses to HCl by disrupting farnesyl diphosphate synthase in mouse taste bud and tongue epithelial cells.

Little is known about the molecular mechanisms underlying drug-induced taste disorders, which can cause malnutrition and reduce quality of life. One of taste disorders is known adverse effects of bisphosphonates, which are administered as anti-osteoporotic drugs. Therefore, the present study evaluated the effects of risedronate (a bisphosphonate) on taste bud cells. Expression analyses revealed that farnesyl diphosphate synthase (FDPS, a key enzyme in the mevalonate pathway) was present in a subset of mouse taste bud and tongue epithelial cells, especially type III sour-sensitive taste cells. Other mevalonate pathway-associated molecules were also detected in mouse taste buds. Behavioral analyses revealed that mice administered risedronate exhibited a significantly enhanced aversion to HCl but not for other basic taste solutions, whereas the taste nerve responses were not affected by risedronate. Additionally, the taste buds of mice administered risedronate exhibited significantly lower mRNA expression of desmoglein-2, an integral component of desmosomes. Taken together, these findings suggest that risedronate may interact directly with FDPS to inhibit the mevalonate pathway in taste bud and tongue epithelial cells, thereby affecting the expression of desmoglein-2 related with epithelial barrier function, which may lead to alterations in behavioral responses to HCl via somatosensory nerves.

RNF43/ZNRF3 negatively regulates taste tissue homeostasis and positively regulates dorsal lingual epithelial tissue homeostasis.

Taste bud cells are renewed throughout life in a process requiring innervation. Recently, we reported that R-spondin substitutes for neuronal input for taste cell regeneration. R-spondin amplifies WNT signaling by interacting with stem-cell-expressed E3 ubiquitin ligases RNF43/ZNRF3 (negative regulators of WNT signaling) and G-protein-coupled receptors LGR4/5/6 (positive regulators of WNT signaling). Therefore, we hypothesized that RNF43/ZNRF3 may serve as a brake, controlled by gustatory neuron-produced R-spondin, for regulating taste tissue homeostasis. Here, we show that mice deficient for Rnf43/Znrf3 in KRT5-expressing epithelial stem/progenitor cells (RZ dKO) exhibited taste cell hyperplasia; in stark contrast, epithelial tissue on the tongue degenerated. WNT signaling blockade substantially reversed all these effects in RZ dKO mice. Furthermore, innervation becomes dispensable for taste cell renewal in RZ dKO mice. We thus demonstrate important but distinct functions of RNF43/ZNRF3 in regulating taste versus lingual epithelial tissue homeostasis.

Unique pattern of histogenesis of the parakeratinized epithelium on lingual prominence in the domestic goose embryos (Anser anser f. domestica).

A triangular lingual prominence (LP) is a characteristic part of the tongue in Anseriformes containing adipose tissue. The parakeratinized epithelium (PEp) covers the LP. Studies aimed to describe the histogenesis of PEp during the process of the intensive formation of the LP in domestic goose during embryonic period and to determine the structural readiness to perform a protective function. The study were conducted by using LM, SEM and TEM technique. The results revealed that on day 16th the undifferentiated epithelium of LP transformed into the typical avian multilayered epithelium. Contrary to pattern of histogenesis of parakeratinized epithelium on the lingual body, on the medial and lateral areas of the elongating and bulging LP were formed epithelial furrows. Which around 20th day, on lateral areas of LP deepened up to half of epithelium, whereas on the medial area began to fade. The ultrastructure of cells lying in furrows indicated progressive apoptosis-like degeneration. On the 25th day, shallow furrows were only present on lateral areas, where bulging of LP was continued. Whereas the epithelium on medial area started cornification by the accumulation of cytokeratin fibers. Lack of the periderm during the development of the PEp of the LP indicated its endodermal origin.

Phosphatidylinositol-3 kinase mediates the sweet suppressive effect of leptin in mouse taste cells.

Leptin is known to selectively suppress neural and taste cell responses to sweet compounds. The sweet suppressive effect of leptin is mediated by the leptin receptor Ob-Rb, and the ATP-gated K+ (KATP ) channel expressed in some sweet-sensitive, taste receptor family 1 member 3 (T1R3)-positive taste cells. However, the intracellular transduction pathway connecting Ob-Rb to KATP channel remains unknown. Here we report that phosphoinositide 3-kinase (PI3K) mediates leptin's suppression of sweet responses in T1R3-positive taste cells. In in situ taste cell recording, systemically administrated leptin suppressed taste cell responses to sucrose in T1R3-positive taste cells. Such leptin's suppression of sucrose responses was impaired by co-administration of PI3K inhibitors (wortmannin or LY294002). In contrast, co-administration of signal transducer and activator of transcription 3 inhibitor (Stattic) or Src homology region 2 domain-containing phosphatase-2 inhibitor (SHP099) had no effect on leptin's suppression of sucrose responses, although signal transducer and activator of transcription 3 and Src homology region 2 domain-containing phosphatase-2 were expressed in T1R3-positive taste cells. In peeled tongue epithelium, phosphatidylinositol (3,4,5)-trisphosphate production and phosphorylation of AKT by leptin were immunohistochemically detected in some T1R3-positive taste cells but not in glutamate decarboxylase 67-positive taste cells. Leptin-induced phosphatidylinositol (3,4,5)-trisphosphate production was suppressed by LY294002. Thus, leptin suppresses sweet responses of T1R3-positive taste cells by activation of Ob-Rb-PI3K-KATP channel pathway.

[Low expression of PTEN and high expression of STING in human tongue squamous cell carcinoma tissues are associated with poor prognosis].

Objective To investigate the expression and clinical significance of phosphatase and tension homolog deleted on chromosome ten (PTEN) and stimulator of interferon genes (STING) in human tongue squamous cell carcinoma (TSCC). Methods The expression of PTEN and STING protein in 65 pairs of TSCC and paracancerous tissues was detected by immunohistochemical EnVision method, and the relationships between PTEN, STING and clinicopathological parameters, overall survival (OS) and prognosis were analyzed by statistical methods. Results Compared with the adjacent tissues, the expression of PTEN in TSCC significantly decreased, and the expression of STING significantly increased. PTEN was negatively correlated with STING. In TSCC, the expression of PTEN and STING were correlated with pathological grade, TNM stage and lymph node metastasis. There were no significant correlations between the expression intensity of PTEN, STING and the gender and age of patients. The low expression of PTEN and the high expression of STING in TSCC tissues were significantly associated with poor prognosis and significantly shortened overall survival of patients. Conclusion TSCC patients with low expression of PTEN and high expression of STING have poor prognosis and short survival time. Combined detection of PTEN and STING expression is helpful to evaluate the degree of tumor progression and patient prognosis.

SOX2 regulates homeostasis of taste bud cells and lingual epithelial cells in posterior tongue.

Taste bud cells arise from local epithelial stem cells in the oral cavity and are continuously replaced by newborn cells throughout an animal's life. However, little is known about the molecular and cellular mechanisms of taste cell turnover. Recently, it has been demonstrated that SOX2, a transcription factor expressed in epithelial stem/progenitor cells of the oral cavity, regulates turnover of anterior tongue epithelium including gustatory and non-gustatory papillae. Yet, the role of SOX2 in regulating taste cell turnover in the posterior tongue is unclear. Prompted by the fact that there are regional differences in the cellular and molecular composition of taste buds and stem/progenitor cells in the anterior and posterior portions of tongue, which are derived from distinct embryonic origins, we set out to determine the role of SOX2 in epithelial tissue homeostasis in the posterior tongue. Here we report the differential requirement of SOX2 in the stem/progenitor cells for the normal turnover of lingual epithelial cells in the posterior tongue. Sox2 deletion in the stem/progenitor cells neither induced active caspase 3-mediated apoptotic cell death nor altered stem/progenitor cell population in the posterior tongue. Nevertheless, morphology and molecular feature of non-gustatory epithelial cells were impaired in the circumvallate papilla but not in the filiform papillae. Remarkably, taste buds became thinner, collapsed, and undetectable over time. Lineage tracing of Sox2-deleted stem/progenitor cells demonstrated an almost complete lack of newly generated basal precursor cells in the taste buds, suggesting mechanistically that Sox2 is involved in determining stem/progenitor cells to differentiate to gustatory lineage cells. Together, these results demonstrate that SOX2 plays key roles in regulating epithelial tissue homeostasis in the posterior tongue, similar but not identical to its function in the anterior tongue.

Evaluation of buccal damage associated with acute inhalation exposure to 2,4-dichlorophenoxyacetic acid (2,4-D) in mice.

BACKGROUND: The herbicide dichlorophenoxyacetic acid (2,4-D) is one of the most widely used crop spraying products in the world. Some pesticides induce the degranulation of mast cells and increase allergic responses. This is the first study to evaluate the damage to the oral mucosa after an experimental simulation of environmental inhalation exposure to the 2,4-D herbicide. The aim of this study was evaluate the possible oral damage caused by acute inhalation exposure to the herbicide 2,4-D. RESULTS: There was a difference between the exposure concentrations in relation to tissue congestion intensity (p = 0.002) and mast cell counts (p = 0.002), a difference in the evaluation of the interaction between the exposure concentrations and nebulization time in the dorsum epithelium thickness (p = 0.013), and a significant correlation between the epithelial thickness and the number of nucleoli organizing regions on the dorsum of the tongue (p = 0.048). CONCLUSIONS: Even after acute exposure, the herbicide 2,4-D had the potential to damage the oral epithelium, especially at higher doses.

Age-related taste cell generation in circumvallate papillae organoids via regulation of multiple signaling pathways.

Sense of taste is central to evaluate food before digestion. Taste stem cells undergo constant differentiation throughout the life. However, the mechanism underlying the generation of taste receptor cells is still not clear. Here, we cultured taste organoids from either Lgr5+ or Lgr5-cells, and found the preferential generation of Car4+ and Gustducin + taste receptor cells in organoids derived from Lgr5+ cells in circumvallate, foliate or fungiform papillae. Taste organoids derived from Lgr5+ cells in circumvallate papillae of neonatal mice showed stronger capacity to generate taste receptor cells compared to the organoids from Lgr5+ cells of the adult circumvallate papillae. Massive transcriptional differences were found in multiple signaling pathways including taste transduction between organoids derived from circumvallate papillae of adult and neonatal mice. Inhibiting the Notch signaling pathway by LY411575 enhanced taste receptor cell generation in organoids from circumvallate papillae and modulated multiple signaling pathways. Thus, we concluded that receptor cell generation in taste organoids was age-related and regulated via multiple signaling pathways.

Bovine tongue epithelium-derived cells: A new source of bovine mesenchymal stem cells.

Mesenchymal stem cells (MSCs) possess the ability to differentiate into multiple cell lineages, and thus, confer great potential for use in regenerative medicine and biotechnology. In the present study, we attempted to isolate and characterize bovine tongue tissue epithelium-derived MSCs (boT-MSCs) and investigate the culture conditions required for long-term culturing of boT-MSCs. boT-MSCs were successfully isolated by the collagenase digestion method and their proliferative capacity was maintained for up to 20 or more passages. We observed a significant increase in the proliferation of boT-MSCs during the 20 consecutive passages under low-glucose Dulbecco's modified Eagle's medium culture condition among the three culture conditions. These boT-MSCs presented pluripotency markers (octamer-binding transcription factor 3/4 (Oct3/4) and sex determining region Y-box2 (Sox2)) and cell surface markers, which included CD13, CD29, CD44, CD73, CD90, CD105, CD166, and major histocompatibility complex (MHC) class I (MHC-I) but not CD11b, CD14, CD31, CD34, CD45, CD80, CD86, CD106, CD117, and MHC-II at third passage. Moreover, these boT-MSCs could differentiate into mesodermal (adipocyte, osteocyte, and chondrocyte) cell lineages. Thus, the present study suggests that the tongue of bovines could be used as a source of bovine MSCs.

Muscle Progenitors Derived from Extraocular Muscles Express Higher Levels of Neurotrophins and their Receptors than other Cranial and Limb Muscles.

Extraocular muscles (EOMs) show resistance to muscle dystrophies and sarcopenia. It has been recently demonstrated that they are endowed with different types of myogenic cells, all of which present an outstanding regenerative potential. Neurotrophins are important modulators of myogenic regeneration and act promoting myoblast proliferation, enhancing myogenic fusion rates and protecting myotubes from inflammatory stimuli. Here, we adapted the pre-plate cell isolation technique to obtain myogenic progenitors from the rat EOMs, and quantified their in vitro expression of neurotrophins and their receptors by RT-qPCR and immunohistochemistry, respectively. The results were compared with the expression on progenitors isolated from buccinator, tongue and limb muscles. Our quantitative analysis of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF) and neurotrophin-3 (NT-3) transcripts showed, for the first time, that EOMs-derived cells express more of these factors and that they expressed TrkA, but not TrkB and TrkC receptors. On the contrary, the immunofluorescence analysis demonstrated high expression of p75NTR on all myogenic progenitors, with the EOMs-derived cells showing higher expression. Taken together, these results suggest that the intrinsic trophic differences between EOMs-derived myogenic progenitors and their counterparts from other muscles could explain why those cells show higher proliferative and fusion rates, as well as better regenerative properties.

Analysis of calcium signaling in live human Tongue cell 3D-Cultures upon tastant perfusion.

Bridging the gap between two-dimensional cell cultures and complex in vivo tissues, three-dimensional cell culture models are of increasing interest in the fields of cell biology and pharmacology. However, present challenges hamper live cell imaging of three-dimensional cell cultures. These include (i) the stabilization of these structures under perfusion conditions, (ii) the recording of many z-planes at high spatio-temporal resolution, (iii) and the data analysis that ranges in complexity from whole specimens to single cells. Here, we addressed these issues for the time-lapse analysis of Ca2+ signaling in spheroids composed of human tongue-derived HTC-8 cells upon perfusion of gustatory substances. Live cell imaging setups for confocal and light sheet microscopy were developed that allow simple and robust spheroid stabilization and high-resolution microscopy with perfusion. Visualization of spheroids made of HTC-8 cells expressing the G-GECO fluorescent Ca2+ sensor revealed Ca2+ transients that showed similar kinetics but different amplitudes upon perfusion of bitter compounds Salicine and Saccharin. Dose-dependent responses to Saccharin required extracellular Ca2+. From the border towards the center of spheroids, compound-induced Ca2+ signals were progressively delayed and decreased in amplitude. Stimulation with ATP led to strong Ca2+ transients that were faster than those evoked by the bitter compounds and blockade of purinergic receptors with Suramin abutted the response to Saccharin, suggesting that ATP mediates a positive autocrine and paracrine feedback. Imaging of ATP-induced Ca2+ transients with light sheet microscopy allowed acquisition over a z-depth of 100 µm without losing spatial and temporal resolution. In summary, the presented approaches permit the study of fast cellular signaling in three-dimensional cultures upon compound perfusion.

Papillary architecture in the tayra tongue.

The tayra (Eira barbara) is a mammal belonging to the Mustelidae family that occurs in all Brazilian biomes. The present work aimed to describe the morphology of the tongue of these specimens highlighting their structures and particularities that will serve as a subsidy to elucidate the anatomy of the same and for comparative studies among other species of domestic and wild animals. Five adult male specimens of E. barbara were studied, which were fixed using 10% aqueous formaldehyde solution. The tongue was removed by opening the oral cavity and separating the maxillary/mandible bone complex. Being in possession of the material, photodocumentations and collection of the fragments were made for the proper preparation of histological slides and Scanning Electron Microscopy (SEM). The lingual papillae found in tayra were mechanical: filiform and conical; and gustative: fungiform and circumvallated. Histologically, the papillae are constituted by keratinized stratified epithelium and in the innermost region, it was composed of tissue connective dense unshaped followed by a layer of muscle bundles of skeletal striated. In the region of the root of the tongue of E. barbara, there were a set of small mixed salivary glands (serous and mucous) and the punctual presence of gustatory corpuscles at the level of epithelium. The morphological description of the E. barbara tongue revealed similarity to that described in literature for other domestic and wild mammals. However, the particularity of the absence of foliate papilla and the quantitative of four papillae circumvallate in the region of the root of the tongue of this species.

Gustatory ultrastructures of an amphihaline migratory fish hilsa Tenualosa ilisha.

This study was conducted with the tongue samples of different life stages of hilsa, that is, adult Marine hilsa, adult Riverine hilsa, and Riverine juvenile hilsa, respectively. Three types of taste buds (Types I, II, and III based on their elevation from the epithelium at different levels) of the tongue, which may be to ensure full utilization of the gustatory ability of the fish were rocorded. Presence of specific taste buds indicate that the fish hilsa dwells in turbid waters with a possible preference toward diatom like planktonic food source. Enhanced expression of taste receptors (T1R1 and T1R3) and associated stimulatory G-proteins subunits on tongue also indicate occurrence of amino acid like substances that guided sensory cues for feeding by this fish. A firm regularity or stringency of the free surface of the epithelial cells may be attributed to compactly arranged microridges. These structures protect against physical abrasions potentially caused during food manoeuvring and swallowing. In our present observations, the surface architectures of the tongue of hilsa are discussed within the background of migratory adaptation of the species in the context of feeding and habitat preferences.

The expressions of some metabolic hormones (leptin, ghrelin and obestatin) in the tissues of sheep tongue.

In this study, we aimed to observe the localization and expression of peptide hormones-leptin, ghrelin and obestatin-in the sheep tongue by immunohistochemistry. For that purpose, tongues of ten adult sheep were used. Leptin expression of moderate intensity was observed in the basal and parabasal epithelial cells of the luminal epithelium, and leptin was strongly expressed in the taste buds of the circumvallate and fungiform papillae and in von Ebner's glands. Ghrelin was primarily expressed in some of the skeletal muscle cells and the smooth muscle cells of the middle layer of blood vessels. A strong expression was observed in the epithelial cells lining the base of the groove surrounding the circumvallate papillae. Obestatin expression was particularly strong in the epithelial cells of the salivary ducts. It was also stronger in the von Ebner's glands than in the seromucous glands. Leptin, ghrelin and obestatin were shown to be produced at varying levels in different cell types, including epithelial, stromal and skeletal muscle cells, as well as in ganglion neurons, neural plexuses and blood vessels in the sheep tongue. Cellular localization and expression of these peptide hormones have not been investigated in many species including sheep.

Inactivation of foot-and-mouth disease virus in epithelium samples for safe transport and processing in low-containment laboratories.

During a foot-and-mouth disease (FMD) outbreak, transport and testing of potentially infectious samples, including epithelium from suspect lesions, presents a biosafety risk, particularly in FMD-free countries. Therefore, treatment to inactivate virus prior to transport is important. Tongue epithelium from cattle infected with FMD virus (FMDV) serotype O (O ALG/3/2014 - Lineage O/ME-SA/Ind-2001d) or A (A IRN/22/2015 - Lineage A/ASIA/G-VII) was incubated in RNAlater, RNA Shield or phosphate-buffered saline (pH 7.4) at room temperature for 2, 6, 24 or 48 h. After incubation, tissues were homogenised and tested by virus titration. Viral RNA in the homogenate was quantified by RT-qPCR, used for sequencing, and transfected into LFBKαVß6 cells to recover infectious virus. RNAlater reduced A IRN/22/2015 titres by 4 log10 after 24 h, and completely after 48 h incubation. While O ALG/3/2014 was detected by VI after 2, 6 and 24 h, titration yielded no infectious virus, likely as a result of freeze-thawing. RNA Shield was cytotoxic at high concentrations but was effective at inactivating both strains after 24 h. Regardless of reagent or inactivation period, RT-qPCR, VP1 sequencing, and transfection of RNA to recover infectious virus were possible. RNA Shield appears a better choice for FMDV inactivation in tissues, however 24 h incubation is recommended.

Collection of nectar by bumblebees: how the physics of fluid demonstrates the prominent role of the tongue's morphology.

Bumblebees and some other tiny animals feed on nectar by visiting flowers in their neighborhood. Some bee species appear to be highly specialized, their tongue being adapted to specific flowers. Bombus terrestris in contrast is able to feed on a wide variety of flowers and can thus be considered as a kind of universal nectar catcher. Since plant nectars show highly variable sugar content, Bombus terrestris have developed a capture mechanism that works for almost any fluid viscosity. Their tongues are decorated with very elongated papillae forming a hairy coating surrounding a rod-like main stalk. When settled on a flower, Bombus rapidly dip their tongue into the inflorescence to catch the highly sought-after nectar. To determine the physical mechanism at the origin of this outstanding ability, the capture dynamics was followed from videos recorded during viscous fluid ingestion. Surprisingly, the volume per lap and the lapping frequency are independent of the fluid viscosity over three orders of magnitude. To explain this observation, we designed a physical model of viscous dipping with structured rods. Predictions of the model compared to observations for bees showed that the nectar is not captured with the help of viscous drag, as proposed in the Landau-Levich-Derjaguin model, but thanks to the hairy structure that traps the viscous fluid, capillary forces drastically limiting the drainage. Our approach can be transposed to others nectar foragers such as bats and hummingbirds.

Characteristic Localization of Neuronatin in Rat Testis, Hair Follicle, Tongue, and Pancreas.

Neuronatin (Nnat) is expressed in the pituitary, pancreas, and other tissues; however, the function of NNAT is still unclear. Recent studies have demonstrated that NNAT is localized in the sex-determining region Y-box 2-positive stem/progenitor cells in the developing rat pituitary primordium and is downregulated during differentiation into mature hormone-producing cells. Moreover, NNAT is widely localized in subcellular organelles, excluding the Golgi. Here, we further evaluated NNAT-positive cells and intracellular localization in embryonic and postnatal rat tissues such as the pancreas, tongue, whisker hair follicle, and testis. Immunohistochemistry revealed that NNAT was localized in undifferentiated cells (i.e., epithelial basal cells and basement cells in the papillae of the tongue and round and elongated spermatids of the testis) as well as in differentiated cells (insulin-positive cells and exocrine cells of the pancreas, taste receptor cells of the fungiform papilla, the inner root sheath of whisker hair follicles, and spermatozoa). In addition, NNAT exhibited novel intracellular localization in acrosomes in the spermatozoa. Because the endoplasmic reticulum (ER) is excluded from spermatozoa and sarco/ER Ca2+-ATPase isoform 2 (SERCA2) is absent from the inner root sheath, these findings suggested that NNAT localization in the ER and its interaction with SERCA2 are cell- or tissue-specific properties.

How to make a tongue: Cellular and molecular regulation of muscle and connective tissue formation during mammalian tongue development.

The vertebrate tongue is a complex muscular organ situated in the oral cavity and involved in multiple functions including mastication, taste sensation, articulation and the maintenance of oral health. Although the gross embryological contributions to tongue formation have been known for many years, it is only relatively recently that the molecular pathways regulating these processes have begun to be discovered. In particular, there is now evidence that the Hedgehog, TGF-Beta, Wnt and Notch signaling pathways all play an important role in mediating appropriate signaling interactions between the epithelial, cranial neural crest and mesodermal cell populations that are required to form the tongue. In humans, a number of congenital abnormalities that affect gross morphology of the tongue have also been described, occurring in isolation or as part of a developmental syndrome, which can greatly impact on the health and well-being of affected individuals. These anomalies can range from an absence of tongue formation (aglossia) through to diminutive (microglossia), enlarged (macroglossia) or bifid tongue. Here, we present an overview of the gross anatomy and embryology of mammalian tongue development, focusing on the molecular processes underlying formation of the musculature and connective tissues within this organ. We also survey the clinical presentation of tongue anomalies seen in human populations, whilst considering their developmental and genetic etiology.

Embryonic development of parakeratinized epithelium of the tongue in the domestic duck (Anas platyrhynchos f. domestica): LM, SEM, and TEM observations.

The parakeratinized epithelium is a common and widespread type of keratinized epithelium in the oral cavity in adult birds. In contrast to orthokeratinized epithelium, which mostly covers mechanical papillae and the lingual nail, parakeratinized epithelium covers almost the entire dorsal surface of the tongue in birds. The characteristic feature of parakeratinized epithelium is the presence of nuclei in the keratinized layer. The present study aimed to investigate for the first time the micro- and ultrastructural changes of parakeratinized epithelium during embryonic development and to assess the readiness of the epithelium to serve protective functions during food transport to the esophagus. Three developmental stages were distinguished: embryonic, transformation, and pre-hatching stages. The embryonic stage lasts from the 9th to the 14th day of incubation and the epithelium is composed of undifferentiated epithelial cells. The transformation stage lasts from the 15th to the 22nd day of incubation and the epithelium undergoes transformation into stratified epithelium consisting of basal, intermediate, and superficial layers. The characteristic feature of this stage is formation of the periderm with osmophilic granules. The pre-hatching stage starts on the 23rd day, and the epithelium with a fully developed keratinized layer resembles that of the epithelium in adult animals. No periderm was observed on the epithelial surface. It was confirmed that at the time of hatching the parakeratinized epithelium is fully differentiated and ready to fulfill its function during food transport. The presence of periderm is a common feature characteristic for para- and orthokeratinized epithelium in the oral cavity of birds. However, the formation of the keratinized/cornified layer is different for these two types of keratinized epithelia.

SIS-ECM Laden with GMSC-Derived Exosomes Promote Taste Bud Regeneration.

Oral cancer has a high annual incidence rate all over the world, and the tongue is the most frequently affected anatomic structure. The current standard care is ablative surgery of malignant neoplasm, followed by tongue reconstruction with free flap. However, such reconstructive modalities with postsurgery radiotherapy or chemotherapy can hardly support the functional recovery of the tongue-particularly, functional taste bud regeneration-in reconstructed areas, thus seriously affecting patients' prognosis and life quality. Using a critical-sized tongue defect model in rats, we show that combinatory transplantation of small intestinal submucosa-extracellular matrix (SIS-ECM) with gingival mesenchymal stem cells (GMSCs) or their derivative exosomes promoted tongue lingual papillae recovery and taste bud regeneration as evidenced by increased expression of CK14, CK8, and markers for type I, II, and III taste bud cells (NTPdase 2, PLC-ß2, and AADC, respectively). In addition, our results indicate that GMSCs or their derivative exosomes could increase BDNF expression, a growth factor that plays an important role in the proliferation and differentiation of epithelial basal progenitor cells into taste bud cells. Meanwhile, we showed an elevated expression level of Shh-which is essential for development, homeostasis, and maintenance of the taste bud organ-in wounded areas of the tongue among animals treated with GMSC/SIS-ECM or exosome/SIS-ECM as compared with SIS-ECM control. Moreover, our data show that GMSCs or their derivative exosomes promoted innervation of regenerated taste buds, as evidenced by elevated expressions of neurofilament and P2X3 at the injury areas. Together, our findings indicate that GMSC/SIS-ECM and exosome/SIS-ECM constructs can facilitate taste bud regeneration and reinnervation with promising potential application in postsurgery tongue reconstruction of patients with tongue cancer.