Tongue Epithelial Cells Research Articles

TNF and IL-10 in a bitter-sweet battle in taste buds

Event Abstract Back to Event TNF and IL-10 in a bitter-sweet battle in taste buds Pu Feng1, Jinghua Chai1, Liquan Huang1 and Hong Wang1* 1 Monell Chemical Senses Center, United States Taste buds are chemosensory structures widely distributed on the surface of the oral cavity and larynx. Taste cells, exposed to the oral environment, face great challenges in defense against potential pathogens. While immune cells are rarely found in taste buds, our recent studies suggest that some immune-response-associated molecules are expressed at higher levels in taste cells than in nontaste tongue epithelial cells. It remains unclear how protective responses are regulated in taste buds. In this study, we investigated the expression of the pro-inflammatory cytokine TNF and the anti-inflammatory cytokine IL-10 in the posterior taste tissues. Both TNF and IL-10 were found exclusively in subsets of type II taste cells, the receptor cells for sweet, bitter and umami tastes. Further analysis revealed that TNF was co-expressed with the sweet and umami receptor T1R3, but not with the G-protein Gustducin (mostly bitter receptor cells). In contrast, IL-10 was co-expressed with Gustducin, but not with T1R3. Meanwhile, there were few taste cells co-expressed both cytokines. These data suggest that these two cytokines are produced by different populations of taste cells in the posterior taste tissues: TNF by sweet and umami cells and IL-10 by bitter cells. Furthermore, TNF receptors were expressed in all taste cells at differential levels, while IL-10 receptors were preferentially expressed in T1R3+TNF+ cells, suggesting that TNF could act on all taste cells under inflammatory conditions, while IL-10 may preferentially target TNF-producing-taste cells to limit TNF production. This study revealed a novel regulatory mechanism of inflammation in taste buds. Acknowledgements This study was supported by NIH/NIDCD grants DC010012, DC007487 and DC011735 and NSF grant DBJ-0216310. Keywords: TNF, IL-10, cytokine, Taste Buds, Inflammation Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Innate immunity Citation: Feng P, Chai J, Huang L and Wang H (2013). TNF and IL-10 in a bitter-sweet battle in taste buds. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00081 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 08 Mar 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. Hong Wang, Monell Chemical Senses Center, Philadelphia, PA, 19104, United States, hwang@monell.org Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Pu Feng Jinghua Chai Liquan Huang Hong Wang Google Pu Feng Jinghua Chai Liquan Huang Hong Wang Google Scholar Pu Feng Jinghua Chai Liquan Huang Hong Wang PubMed Pu Feng Jinghua Chai Liquan Huang Hong Wang Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

GOLPH3 overexpression correlates with tumor progression and poor prognosis in patients with clinically N0 oral tongue cancer

BackgroundOverexpression of GOLPH3 (Golgi phosphoprotein 3, 34 kDa) is associated with the progression of many solid tumor types leading to an unfavorable clinical outcome. We aimed to investigate the clinical significance of GOLPH3 expression in the development and progression of clinically N0 (cN0) oral tongue cancer.MethodsReal-time PCR and Western blotting analyses were employed to examine GOLPH3 expression in four oral tongue cancer cell lines, primary cultured normal tongue epithelial cells (TEC), eight matched pairs of oral tongue cancer samples and adjacent noncancerous tissue samples from the same patient. Immunohistochemistry (IHC) was performed to examine GOLPH3 protein expression in paraffin-embedded tissues from 179 cN0 oral tongue cancer patients. Statistical analyses were applied to evaluate the diagnostic value and the associations of GOLPH3 expression with clinical parameters.ResultsGOLPH3 mRNA and protein was up-regulated in oral tongue cancer cell lines and cancerous tissues compared with that in primary cultured normal tongue epithelial cells (TEC) and adjacent noncancerous tissue samples. GOLPH3 protein level was positively correlated with clinical stage (P = 0.001), T classification (P = 0.001), N classification (P = 0.043) and recurrence (P = 0.009). Patients with higher GOLPH3 expression had shorter overall survival time, whereas those with lower GOLPH3 expression had longer survival time.ConclusionOur results suggest GOLPH3 overexpression is associated with poor prognosis for cN0 oral tongue cancer patients and may represent a novel and useful prognostic indicator for cN0 oral tongue cancer.

Peptidoglycan recognition protein 3 (Pglyrp3) is associated with intestinal mucosal homeostasis and protection against colitis (67.12)

Abstract Peptidoglycan recognition protein 3 (Pglyrp3) is a bactericidal protein expressed in the epithelial cells of tongue, esophagus, stomach and intestine. Pglyrp3-deficient mice compared to wild type (WT) mice have increased sensitivity to dextran sodium sulfate (DSS)-induced colitis. The increased colitis is characterized by loss of epithelial cells, severe hyperplasia of the lamina propria and ulcers. The aim of this study was to determine whether Pglyrp3 plays a role in maintaining intestinal epithelial integrity by regulating proliferation and apoptosis of epithelial cells. We report that Pglyrp3-deficient mice treated with DSS have significantly higher TUNEL positive cells, increased Annexin V staining and higher levels of cleaved caspase 3 in the epithelial cells of the colon compared to WT mice. The number of bromodeoxyuridine (BrdU) positive cells, which indicates proliferating cells, is significantly lower in Pglyrp3-deficient mice compared to WT mice. Serum levels of FITC-dextran are significantly higher in DSS-treated Pglyrp3-deficient compared to WT mice, which indicates an increased loss of epithelial lining and loss of barrier function in Pglyrp3-deficient mice. Our results indicate that Pglyrp3 is important in maintaining the epithelial lining of the colon and loss of this protein results in increased apoptosis, decreased proliferation and increased loss of barrier function, which are important contributing factors to increased colitis in Pglyrp3-deficient mice.

Down-regulation of tumor suppressor MTUS1/ATIP is associated with enhanced proliferation, poor differentiation and poor prognosis in oral tongue squamous cell carcinoma

Microtubule-associated tumor suppressor gene (MTUS1, also known as mitochondrial tumor suppressor) is a recently identified tumor suppressor gene that has been implicated in several cancer types. The expression of MTUS1 gene leads to 5 known transcript variants and codes for 5 isoforms of Angiotensin II AT2 receptor interacting protein (ATIP). In this study, we first confirmed that the down-regulation of MTUS1/ATIP was a frequent event in oral tongue squamous cell carcinoma (OTSCC) and the premalignant lesion (leukoplakia). We further demonstrated that the down-regulation of MTUS1/ATIP was correlated with poor differentiation and enhanced proliferation (Ki67 proliferation index). Statistical analysis suggests that the down-regulation of MTUS1/ATIP was associated with reduced overall survival. Isoform specific quantitative RT-PCR assays revealed that ATIP1, ATIP3a and ATIP3b were the major isoforms of the MTUS1 gene products in oral tongue epithelial cells. Significant down-regulations were observed for all 3 ATIP isoforms in OTSCC as compared to matching normal tissues. In vitro functional study showed that the restoration of ATIP1 expression led to G1 arrest, apoptosis and reduction of cell proliferation in OTSCC cell lines. These ATIP1-induced cellular changes were accompanied by reduced phosphorylation of ERK1/2 and up-regulation of p53. Taken together, these data suggest that MTUS1 plays major roles in the progression of OTSCC, and may serve as a biomarker or therapeutic target for patients with OTSCC.

A comparison of different seeding methods for constructing three dimensional urethral tissue in vitro

Objective To investigate and assess the best seeding method for constructing three dimensional urethral tissue in vitro. Methods High speed agitation decellular method was used for preparing the porcine acdlular corporous spongiosum matrix （ACSM）. Before seeding, the matrix was sterilized via soaking compound iodine solution. Rabbit tongue epithelial cells and cavernosal smooth muscle cells were isolated and cultured. Three different groups of seeding method was used in this study. Group A （sandwich seeding group） ： The smooth muscle cells and epithelial cells were seeded onto the different side of ACSM by static method. Group B （injection seeding group）.. The smooth muscle cells were injected into the scaffold. Then,epithelial cells were seeded onto the urethral surface of ACSM by static method. Group C （agitation seeding group） ： The smooth muscle cells were seeded into the scaffold by agitation method. Then, epithelial cells were seeded onto the urethral surface of ACSM by static method. After being seeded, all matrixes were cultured in vitro for 14 d. HE and immunoassay staining were used to examine the results of seeding. Results Looser matrix was obtained after using high speed agitation decellular method. Compound iodine solution could not only sterilize efficiently but also reserve the original structure of biomaterial. An intact epithelial cellularlayer onto the surface of scaffold could be observed in HE staining section after 14 d culturing in vitro. Few smooth muscle cells could be found in big space of biomaterial in group A. In group B, smooth muscle cells were restrained in some regions of the matrix. Smooth muscle cells were well distributed into the scaffold in group C. Conclusions After using high speed agitation decellular method, an ideal matrix with three dimensional structure can be obtained. Combined with agitated seeding method, three dimensional urethral tissue can be constructed. Key words: Acellular matrix; Tissue engineering; Three dimension seeding method; Ure thra

HIV Infection Induces Morphometrical Changes on the Oral (Buccal Mucosa and Tongue) Epithelial Cells

The aim of this study was to assess morphological and morphometrical alterations of oral squamous epithelial cells in type 1 HIV infected individuals. Oral smears were collected from tongue and buccal mucosa of 30 HIV infected (experimental) and 30 non-infected (control) individuals by liquid-based exfoliative cytology. The cells were morphologically analyzed and the nuclear area (NA), the cytoplasmic area (CA) and the nucleus-to-cytoplasm area ratio (NA/CA) were calculated. No morphological differences were found between the groups. The mean values of CA were decreased in tongue (P=.00006) and buccal mucosa (P=.00242) in HIV infected individual, while mean values of NA were increased (P=.00308 and .00095, respectively) in the same group. NA/CA ratio for experimental group was increased in both collected places, with P=.00001 (tongue) and P=.00000 (buccal mucosa). This study revealed that HIV infection was able to induce morphometrical changes on the oral epithelial cells.

Tongue epithelial KT-1 cell-cycle arrest by TGF-β associated with induction of p21Cip1 and p15Ink4b

Tongue epithelium continuously turns over in adults. Our previous study showed that epidermal growth factor and fibroblast growth factor-2 stimulated proliferation of KT-1 cells derived from tongue epithelium, suggesting that these signals serve as positive regulators for tongue epithelial proliferation. To investigate a negative regulation of tongue epithelial cell proliferation, we studied effects of transforming growth factor-beta (TGF-beta) on KT-1 cells. Proliferation assays showed that TGF-beta inhibited proliferation of KT-1 cells in a dose dependent manner. Cell-cycle analysis showed that TGF-beta induced G(0)/G(1) cell cycle arrest in KT-1 cells. We also examined expressions of Ink4 and Cip/Kip family mRNA by quantitative reverse transcription-polymerase chain reaction. We found that TGF-beta induced p15(Ink4b) and p21(Cip1) mRNA expressions. These results strongly suggest that G(0)/G(1) cell cycle arrest is associated with increased p15(Ink4b) and p21(Cip1) expressions. Moreover, p21(Cip1) mRNA was localized in suprabasal cells of tongue epithelium, suggesting that p21(Cip1) play a role in cell-cycle exit along with tongue epithelial differentiation. Taken together, our results suggest that TGF-beta signaling serves as negative regulator of tongue epithelial cell proliferation, and may control tongue epithelial cell differentiation through modulating expression of p21(Cip1).

Nuclear changes in tongue epithelial cells following panoramic radiography

This study aimed to investigate the effect of radiation from panoramic radiographs on the cells of the lateral border of the tongue by evaluating nuclear changes. Forty-two patients were included: 22 had one radiograph (Group I), and 20 required a repeat radiograph due to error in the first exposure (Group II). Material for the cytopathologic evaluation was collected before radiographs and 10 days later. Smears were stained with the Feulgen reaction and micronuclei, buds, broken eggs, karyorrhexis and binucleate cells were scored. The comparison of nuclear changes before and after radiation exposure in both groups revealed a statistically higher number of broken eggs, buds, karyorrhexis and binucleate cells 10 days after exposure ( P = 0.01). The number of karyorrhexis and binucleate cells was greater in group II ( P = 0.01). There was no change in the frequency of micronuclei before and after the radiographs. Radiation emitted during panoramic radiographs increased the number of nuclear anomalies (except micronuclei) in exfoliated cells of the lateral border of the tongue. This effect was more pronounced when the patients were exposed to a repeat radiograph, without however implying increased risk of irreversible tissue damage.

Various human epithelial cells express functional Toll-like receptors, NOD1 and NOD2 to produce anti-microbial peptides, but not proinflammatory cytokines

Epithelial cells may form the first barrier of defense against bacteria in human tissues. We recently revealed that oral epithelial cells generated anti-bacterial factors, such as peptidoglycan recognition proteins (PGRPs) and β-defensin 2, but not proinflammatory cytokines, such as interleukin-8 (IL-8), upon stimulation with bacterial cell-surface components. In this study, we found clear expressions of Toll-like receptor (TLR)2, TLR3, TLR4, TLR7, NOD1 and NOD2 in oral, tongue, salivary gland, pharyngeal, esophageal, intestinal, cervical, breast, lung, and kidney epithelial cells. However, tongue, salivary gland, pharyngeal, esophageal, intestinal, cervical, breast, lung, and kidney epithelial cells, as well as oral epithelial cells, did not secrete IL-6, IL-8 or monocyte chemoattractant protein-1 in response to chemically synthesized TLR and NOD agonists mimicking microbial components: TLR2 agonistic lipopeptide (Pam3CSSNA), TLR3 agonistic Poly I:C, TLR4 agonistic lipid A (LA-15-PP), TLR7 agonistic single stranded RNA (ssPoly U), NOD1 agonistic iE-DAP (γ- d-glumtamyl- meso-diaminopimelic acid), and NOD2 agonistic muramyldipeptide (MDP). Although PGRPs on oral epithelial cells were significantly up-regulated upon stimulation with these synthetic components, PGRPs on pharyngeal epithelial cells were only slightly up-regulated, and PGRPs on esophageal, intestinal and cervical epithelial cells were not up-regulated upon stimulation with the components. In contrast, stimulation with synthetic TLRs and NODs ligands induced β-defensin 2 generation in all epithelial cells examined. These findings indicate that TLR and NOD in various epithelial cells are functional receptors that induce anti-bacterial responses in general without being accompanied by inflammatory responses.

Immunohistochemical localization of aromatic l-amino acid decarboxylase in mouse taste buds and developing taste papillae

Aromatic L-amino acid decarboxylase (AADC) catalyses the decarboxylation of all aromatic L-amino acids. In mammals, AADC is expressed in many tissues besides the nervous system, and is associated with additional regulatory roles of dopamine and serotonin in a wide range of tissues. We examined the expression of AADC by using reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. RT-PCR analysis showed that mRNA of AADC was detected in the taste bud-containing epithelium of the circumvallate papilla of mice. By immunohistochemical analyses, AADC was detected in a subset of taste bud cells of fungiform, foliate, and circumvallate papillae. Double-label studies showed that AADC colocalized with serotonin, NCAM, PLCbeta2, and PGP9.5. On the other hand, AADC never colocalized with alpha-gustducin. Our results of double staining with AADC and taste cell markers indicate that only the type III cells could convert 5-hydroxytryptophan (5-HTP) to serotonin within taste buds. Taken together with previous studies, the properties of the type III cell of taste buds exactly fit into the APUD (amine and amine precursor uptake and decarboxylation) cell scheme. Furthermore, in the developing circumvallate papilla, AADC are first detected in a small number of papillary epithelial cells at E14.5. By E18.5, AADC-positive epithelial cells also express PGP9.5, which is one of marker of taste cells, and these cells have been contacted by developing nerve fibers. These results suggest that AADC expression begins at early stages of taste bud cell differentiation, and biogenic amines may act on taste bud differentiation of tongue epithelial cells, and further may regulate innervation of taste bud progenitor cells.

Epstein–Barr virus in oral diseases

Epstein-Barr virus (EBV), a B-lymphotropic gamma-herpesvirus, causes infectious mononucleosis and oral hairy leukoplakia, and is associated with various types of lymphoid and epithelial malignancies. Saliva is the main vehicle for EBV transmission from individual to individual. Recent studies have also implicated EBV in the pathogenesis of advanced types of periodontal disease. EBV DNA is detected in 60-80% of aggressive periodontitis lesions and in 15-20% of gingivitis lesions or normal periodontal sites. The periodontal presence of EBV is associated with an elevated occurrence of periodontopathic anaerobic bacteria. Moreover, EBV active infection occurs in approximately 70% of symptomatic and large-size periapical lesions. EBV and cytomegalovirus often co-exist in marginal and apical periodontitis. Periodontal therapy can markedly suppress the EBV load in periodontal pockets as well as in saliva, which has the potential to reduce the risk of viral transmission between close individuals. EBV proteins up-regulate cytokines and growth factors, which seem to play a central role in the proliferative response of tongue epithelial cells in oral hairy leukoplakia and in the cell-transformation process of EBV-associated malignancies. Further research is needed to identify the full range of EBV-related diseases in the human oral cavity.

Oregano, thyme and clove-derived flavors and skin sensitizers activate specific TRP channels

Carvacrol, eugenol and thymol are major components of plants such as oregano, savory, clove and thyme. When applied to the tongue, these flavors elicit a warm sensation. They are also known to be skin sensitizers and allergens. The transient receptor potential channel (TRPV3) is a warm-sensitive Ca2+-permeable cation channel highly expressed in the skin, tongue and nose. Here we show that TRPV3 is strongly activated and sensitized by carvacrol, thymol and eugenol. Tongue and skin epithelial cells respond to carvacrol and eugenol with an increase in intracellular Ca2+ levels. We also show that this TRPV3 activity is strongly potentiated by phospholipase C-linked, G protein-coupled receptor stimulation. In addition, carvacrol activates and rapidly desensitizes TRPA1, which may explain the pungency of oregano. Our results support a role for temperature-sensitive TRP channels in chemesthesis in oral and nasal epithelium and suggest that TRPV3 may be a molecular target of plant-derived skin sensitizers.

The Actinobacillus actinomycetemcomitans Autotransporter Adhesin Aae Exhibits Specificity for Buccal Epithelial Cells from Humans and Old World Primates

Cells of the gram-negative periodontopathogen Actinobacillus actinomycetemcomitans express a surface-exposed, outer membrane autotransporter protein, designated Aae, which has been implicated in epithelial cell binding. We constructed a mutant strain of A. actinomycetemcomitans that contained a transposon insertion in the Aae structural gene (aae) and tested the mutant to determine its ability to bind to buccal epithelial cells (BECs) isolated from healthy volunteers. Significantly fewer mutant cells than wild-type cells bound to BECs. A broad-host-range plasmid that contained an intact aae gene driven by a heterologous tac promoter restored the ability of the mutant strain to bind to BECs at wild-type levels. This plasmid also conferred upon Escherichia coli the ability to express the Aae protein on its surface and to bind to human BECs. Aae-expressing E. coli also bound to BECs isolated from six Old World primates but not to BECs isolated from four New World primates or nine other nonprimate mammals, as well as to human gingival epithelial cells but not to human pharyngeal, palatal, tongue, bronchial, or cervical epithelial cells. Our findings indicate that Aae mediates binding of A. actinomycetemcomitans to BECs from humans and Old World primates and that this process may contribute to the host range specificity and tissue tropism exhibited by this bacterium.

Epstein‐Barr Virus (EBV) Infection in Epithelial Cells In Vivo: Rare Detection of EBV Replication in Tongue Mucosa but Not in Salivary Glands

Epstein-Barr virus (EBV) is transmitted through saliva, but the cellular source is controversial. Putative reservoirs include oral epithelium and salivary glands. Tongue mucosal samples, salivary glands, and tongue carcinomas were studied, by immunohistochemistry and in situ hybridization, for evidence of EBV infection. EBV replication was seen in 1.3% of tongue mucosal samples. No latent infection was found at this site. EBV infection was detected neither in normal salivary glands nor in tongue carcinomas. Thus, EBV replication occurs infrequently in tongue epithelial cells, and salivary glands are unlikely to harbor EBV. EBV is unlikely to be involved in the pathogenesis of tongue cancer.

Epstein-Barr virus infection of polarized tongue and nasopharyngeal epithelial cells.

Epstein-Barr virus (EBV) initially enters the body through the oropharyngeal mucosa and subsequently infects B lymphocytes through their CD21 (CR2) complement receptor. Mechanisms of EBV entry into and release from epithelial cells are poorly understood. To study EBV infection in mucosal oropharyngeal epithelial cells, we established human polarized tongue and pharyngeal epithelial cells in culture. We show that EBV enters these cells through three CD21-independent pathways: (i) by direct cell-to-cell contact of apical cell membranes with EBV-infected lymphocytes; (ii) by entry of cell-free virions through basolateral membranes, mediated in part through an interaction between beta1 or alpha5beta1 integrins and the EBV BMRF-2 protein; and (iii) after initial infection, by virus spread directly across lateral membranes to adjacent epithelial cells. Release of progeny virions from polarized cells occurs from both their apical and basolateral membranes. These data indicate that multiple approaches to prevention of epithelial infection with EBV will be necessary.

Mutagenicity of the oral carcinogen 4-nitroquinoline-1-oxide in cultured BigBlue™ rat tongue epithelial cells and fibroblasts

Environmental carcinogen exposures contribute to the development of oral cancer and improved test systems for the analysis of such carcinogens are needed. We have previously isolated and characterized an epithelial cell line from the tongue of a BigBlue™ rat. Now, we have established an immortalized fibroblast cell line from the same organ. We exposed these cells to 4-nitroquinoline-1-oxide (NQO), a well-known experimental oral carcinogen in the rat and other species, and measured its cytotoxic and genotoxic ( cII transgene mutagenesis) effects. Both cell lines were very sensitive to NQO toxicity and showed dose-dependent mutant frequency responses. At the highest NQO dose tested, 70 ng/ml, the mutant frequency was elevated more than eight-fold above background for the epithelial cells and more than 25-fold for the fibroblast cells. We examined cellular parameters which could affect glutathione-dependent detoxication of mutagens. Glutathione (GSH) contents of the two cell lines were similar. Glutathione transferase (GST) activities were measured with several substrates and were generally higher in the epithelial cells. Although multiple biochemical and biological characteristics of individual cell lines are likely to determine responses to mutagens, the greater sensitivity of the fibroblast cells to NQO mutagenicity is in accord with the lower GST activity and the lower DNA content of these cells. These new cell lines are suitable for in vitro testing of chemicals as possible oral mutagens and for studies of their biochemical mechanisms of action.

Epstein-Barr virus replication in tongue epithelial cells.

Epstein-Barr virus (EBV) persistently infects B-cells in humans and can be shed into the saliva. The cellular source of infectious virus is uncertain. Hairy leukoplakia, an AIDS-associated lesion of the tongue, supports EBV replication in epithelial cells. However, the general significance of this observation has remained doubtful. Using immunohistochemistry and in situ hybridization, we demonstrate evidence of EBV replication in tongue epithelial cells in 4 of 168 samples from 84 autopsy cases. Thus, in patients who do not have AIDS, squamous epithelial cells of the tongue rarely support EBV replication. However, all individuals with evidence of EBV replication were either on immunosuppressive therapy or were terminally ill cancer patients, suggesting that an impairment of the immune system may have allowed EBV replication to occur at this site. Thus, our findings are consistent with the idea that EBV replication in oropharyngeal epithelial cells is an infrequent event.

Persistent productive Epstein-Barr virus replication in normal epithelial cells in vivo.

Productive Epstein-Barr virus (EBV) replication characterizes hairy leukoplakia, an oral epithelial lesion typically occurring in individuals infected with human immunodeficiency virus (HIV). Serial tongue biopsy specimens were obtained from HIV-infected subjects before, during, and after valacyclovir treatment. EBV replication was detected by Southern hybridization to linear terminal EBV genome fragments, reverse-transcriptase polymerase chain reaction amplification of EBV replicative gene transcripts, immunohistochemical detection of EBV replicative protein, and in situ hybridization to EBV DNA. EBV replication was detected in both hairy leukoplakia and normal tongue tissues. Valacyclovir treatment completely abrogated EBV replication in vivo, resulting in resolution of hairy leukoplakia when it was present. EBV replication returned in normal tongue epithelial cells after valacyclovir treatment. These data suggest that normal oral epithelium supports persistent EBV infection in individuals infected with HIV and that productive EBV replication is necessary but not sufficient for the pathogenesis of oral hairy leukoplakia.

Cloning and Characterization of the Gene for a New Epithelial β-Defensin: GENOMIC STRUCTURE, CHROMOSOMAL LOCALIZATION, AND EVIDENCE FOR ITS CONSTITUTIVE EXPRESSION

Mammalian beta-defensins are endogenous cysteine-rich peptide antibiotics that are produced either by epithelial cells lining the respiratory, digestive, and urogenital tracts or by granulocytes and macrophages. A growing body of evidence has implicated these peptides in host defense, particularly mucosal innate immunity. We previously reported the cloning of the full-length cDNA for a porcine beta-defensin (pBD-1), which was found to be expressed throughout the airway and oral mucosa. Here, we provide the structural organization of the pBD-1 gene, showing that the entire gene spans approximately 1.9 kilobases with two short exons separated by a 1.5-kilobase intron. Fluorescence in situ hybridization mapped the pBD-1 gene to porcine chromosome 15q14-q15. 1 within a region of conserved synteny to the chromosomal locations of human and mouse alpha- and beta-defensins. We also provide several independent lines of evidence showing that the pBD-1 gene is expressed constitutively during inflammation and infection, despite its resemblance to many inducible epithelial beta-defensins in amino acid sequence, genomic structure, and sites of expression. First, stimulation of primary porcine tongue epithelial cells with lipopolysaccharide, tumor necrosis factor-alpha, and interleukin (IL)-1beta failed to up-regulate the expression of pBD-1 mRNA. Second, pBD-1 gene expression was not enhanced in either digestive or respiratory mucosa of pigs following a 2-day infection with Salmonella typhimurium or Actinobacillus pleuropneumoniae. Last, direct transfection of the pBD-1 gene promoter into NIH/3T3 cells showed no difference in reporter gene activity in response to stimulation by lipopolysaccharide and IL-1beta. The constitutive expression of pBD-1 in airway and oral mucosa, which is consistent with a lack of consensus binding sites for nuclear factor-kappaB or NF-IL-6 in its promoter region, suggests that it may play a surveillance role in maintaining the steady state of microflora on mucosal surfaces.

Aberrant expression of carbohydrate antigens in squamous cell carcinoma of the tongue

An immunohistochemical study was performed to observe the aberrant expression of blood-related group antigens in squamous cell carcinomas of the tongue from 10 patients and 13 normal tongue tissues using seven monoclonal antibodies, A, B, H, X001, H001, Lea and Leb, and four lectins, PNA, VVA-B4, UEA-1 and LTA. In SCC, Leb antigens were expressed in all cases examined (10 cases). T and Tn antigens which were examined with PNA and VVA-B4 stains, were expressed in seven out of 10 and six out of 10 cases, respectively, while these two antigens were not found in normal tissues. A, B and H antigens were expressed in most normal tongue epithelial cells, strictly dependent on the patient blood type. But in SCC, A and B antigens were not detected in five out of seven cancer cells from blood group A, B and AB individuals, whereas H antigen was expressed in 5 out of 7 cancer cells from blood group A, B and AB individuals. These results suggest that the expression of blood group-related antigens in suppressed to that of more immature carbohydrate chains in the setting of tongue cancer.