Antimicrobial action of photodynamic therapy in root canals using LED curing light, curcumin and carbopol gel.

AIM: To evaluate the capacity of carbopol gel to maintain the intensity of a LED curing light (blueLED) along the length of prepared root canals in bovine teeth, and to assess the antimicrobial capacity of curcumin photoactivated by a LED curing light in the presence of carbopol gel. METHODOLOGY: Experiment 1: Eight straight roots of bovine incisors were standardized to a length of 15 mm, and the root canals instrumented up to a size 120 K-file. The LED curing light was irradiated inside the root canals using an aluminium collimator (1.5 mm in diameter) placed at the orifice (n = 8). Initially, the irradiation was performed in empty root canals and then repeated with the root canals filled with carbopol gel. Simple standardized photographs of the roots were taken with a digital camera in the mesial perspective during the irradiation procedure and the images analysed in OriginLab software to verify the light intensity along the length of the root. Experiment 2: Twenty dentine blocks were obtained from the cervical third of bovine incisors using a trephine bur. Biofilms were induced for 21 days on the blocks using Enterococcus faecalis (ATCC 4083) at 109 cells mL-1 . The blocks were treated according to the groups (n = 5): positive control; standard PDT (methylene blue + diode Laser); curcumin; LED curing light; and curcumin + LED curing light. After the treatment, the samples were dyed with Live/Dead BacLight Bacterial Viability solution and fluorescence images were obtained by Confocal Scanning Laser Microscopy (CSLM). Experiment 3: Thirty-two roots of bovine incisors were prepared as described in experiment 1. Their dentinal tubules were contaminated and the root canals treated according to the groups (n = 8): positive control; standard PDT; curcumin + LED curing light; curcumin + carbopol gel + LED curing light. The specimens were sectioned longitudinally and the split roots were treated with the Live/Dead dye to obtain fluorescence images by CSLM. All images were processed using BioImageL software to measure the percentage of viable bacteria and the data analysed statistically using the nonparametric Kruskal-Wallis test (α < 0.05). RESULTS: In Experiment 1, carbopol gel did not improve the intensity of LED light transmission along the root canal. In Experiment 2, a significant decrease (P < 0.05) in bacterial viability occurred in the following order: positive control < only LED curing light < only curcumin < curcumin + LED curing light = standard PDT; and in Experiment 3 positive control = curcumin + LED curing light ≤ curcumin + gel + LED curing light ≤ standard PDT. CONCLUSION: Similar disinfection effectiveness was obtained using curcumin + LED curing light and methylene blue + 660 nm LASER (standard PDT). The use of carbopol gel did not favour a greater transmission of LED light along the root canal and also resulted in less bacterial killing when used in endodontic PDT.

Mixture of alkaline tetrasodium EDTA with sodium hypochlorite promotes in vitro smear layer removal and organic matter dissolution during biomechanical preparation.

AIM: The aim of this study was to determine the following: (i) the quantity of free chlorine in mixtures of equal proportions of sodium hypochlorite (NaOCl) with trisodium ethylenediaminetetraacetic acid (EDTAHNa3 ) and alkaline tetrasodium ethylenediaminetetraacetic acid (EDTANa4 ); (ii) organic matter dissolution; and (iii) the time necessary to remove the smear layer by these irrigants alone and when mixed. METHODOLOGY: The solutions were mixed in a 1 : 1 ratio and then iodometrically titrated over time to determine the quantity of free available chlorine. The capability of organic matter dissolution by the solutions alone and the mixtures of irrigants was analysed by weighing bovine muscle tissue specimens before and after submission to the following groups (n = 10): G1 - 0.9% saline solution (control), G2 - 2.5% NaOCl, G3 - 17% EDTAHNa3 , G4 - 10% EDTANa4 , G5 - 20% EDTANa4 , G6 - 5% NaOCl + 17% EDTAHNa3 , G7 - 5% NaOCl + 10% EDTANa4 and G8 - 5% NaOCl + 20% EDTANa4 . The times necessary for smear layer removal were determinated on discs of bovine dentine with a standardized smear layer produced with SiC papers using a scanning electron microscope that did not require the samples to be sputter coated. The dentine discs were submitted to the same experimental groups previously described (n = 10) over several time periods, and the photomicrographs acquired were scored for the presence of smear layer. The parametric data of tissue dissolution were analysed using two-way anova and one-way anova with Tukey's post hoc tests (α < 0.05), whilst nonparametric data of smear layer removal were analysed by Friedman test (α < 0.05) and the Kruskal-Wallis test with Dunn's post hoc (α < 0.05). RESULTS: EDTAHNa3 caused an almost complete and immediate loss of free available chlorine from NaOCl, whilst EDTANa4 promoted a slow and concentrat-ion-dependent decline. The organic matter was not dissolved in the control group, EDTA groups or the mixture of NaOCl + 17% EDTAHNa3 group (P > 0.05). NaOCl alone and the associations of NaOCl + EDTANa4 dissolved tissue at all periods analysed (P < 0.05). The smear layer was not removed in the control and NaOCl groups (P > 0.05). The smear layer was removed at 1 min in the NaOCl + 17% EDTAHNa3 group (P < 0.05); 2 min in 17% EDTAHNa3 group (P < 0.05); and 5 min in 10% EDTANa4 , 20% EDTANa4 , 5% NaOCl + 10% EDTANa4 and 5% NaOCl + 20% EDTANa4 groups (P < 0.05). CONCLUSIONS: Alkaline EDTANa4 was slower in removing the smear layer than EDTAHNa3 , but when mixed with NaOCl during biomechanical canal preparation promoted organic matter dissolution and smear layer removal simultaneously. However, the mixing of NaOCl and EDTANa4 should be performed immediately before use to prevent the reduction of free available chlorine.

Patient-Reported Outcome Measurement in Clinical Dermatological Practice: Relevance and Feasibility of a Web-Based Portal.

BACKGROUND/AIMS: Patient-reported outcomes (PROs) are increasingly considered important. We developed a web-based application to electronically assess PROs in routine dermatological practice. We assessed (1) the relevance of PRO measurement according to health care providers and patients, (2) the feasibility of our application in routine practice according to health care providers, supporting staff and patients, and (3) barriers/facilitators for implementation according to health care providers and supporting staff. METHODS: Health care providers, supporting staff and patients completed study-specific questionnaires. Also, website statistics were analysed. RESULTS: 3/6 clinics participated, including 9 professionals and 80 patients. Both health care providers and patients rated PRO measurement as relevant. However, implementation was only moderately feasible. Time constraints and logistical problems were mentioned as barriers, and motivated patients and supportive staff as facilitators. CONCLUSION: Electronic PRO assessment in routine practice is not self-evident. Adjustments in logistics are recommended to optimize implementation, using a plan-do-study-act approach.

A Web-based, educational, quality-of-life intervention for patients with a chronic skin disease: feasibility and acceptance in routine dermatological practice.

Chronic skin diseases have a negative impact on patients' health-related quality of life (HRQoL). Patient education might contribute to HRQoL improvement. We developed a web-based, educational, HRQoL intervention for patients with a chronic skin disease. We aimed to assess 1) the feasibility of implementing the intervention in routine dermatological practice and patients' daily life, and 2) the acceptance of the intervention by health care providers and patients. Additionally, we aimed to create a patient user profile. We conducted an observational pilot study at 6 dermatological centres, including 105 outpatients. Implementation in routine practice was feasible and acceptable to health care providers. However, implementation in patients' daily life was found not to be entirely feasible. Perceived relevance by patients was low, though patients rated the intervention as convenient and attractive. No univocal user profile was found. Suggestions for improvements of the intervention, e.g. tailoring and adding blended learning components, are discussed.

Distinctive characteristics of transcriptional profiles from two epithelial cell lines upon interaction with Actinobacillus actinomycetemcomitans.

Transcriptional profiling and gene ontology analyses were performed to investigate the unique responses of two different epithelial cell lines to an Actinobacillus actinomycetemcomitans challenge. A total of 2867 genes were differentially regulated among all experimental conditions. The analysis of these 2867 genes revealed that the predominant specific response to infection in HeLa cells was associated with the regulation of enzyme activity, RNA metabolism, nucleoside and nucleic acid transport and protein modification. The predominant specific response in immortalized human gingival keratinocytes (IHGK) was associated with the regulation of angiogenesis, chemotaxis, transmembrane receptor protein tyrosine kinase signaling, cell differentiation, apoptosis and response to stress. Of particular interest, stress response genes were significantly - yet differently - affected in both cell lines. In HeLa cells, only three regulated genes impacted the response to stress, and the response to unfolded protein was the only term that passed the ontology filters. This strikingly contrasted with the profiles obtained for IHGK, in which 61 regulated genes impacted the response to stress and constituted an extensive network of cell responses to A. actinomycetemcomitans interaction (response to pathogens, oxidative stress, unfolded proteins, DNA damage, starvation and wounding). Hence, while extensive similarities were found in the transcriptional profiles of these two epithelial cell lines, significant differences were highlighted. These differences were predominantly found in pathways that are associated with host-pathogen interactions.

Orofacial rhabdomyosarcoma in neonates and young children: a review of literature and management of four cases.

Rhabdomyosarcoma (RMS) is an aggressive malignant skeletal muscle neoplasm arising from embryonal mesenchyme. It accounts for over 50% of all pediatric soft tissue sarcomas. The head and neck region is the most common site for this tumor in children. Neonatal presentation of this tumor is rare. We present the management of one neonatal case and three additional cases of orofacial RMS in children under the age of 7 years. All four patients were seen in the department of oral and maxillofacial surgery at Children's Hospital and Regional Medical Center (CHRMC) in Seattle between 1992-2000. Three of the four cases were alveolar RMS and one was botryoid sub-type of embryonal RMS. Three patients were treated with a combination of surgery, chemotherapy and radiation, while the patient with botryoid RMS was treated with surgery and chemotherapy only. The patient with congenital RMS died at 2.5 years of age due to recurrent metastatic disease. The other three patients are alive without evidence of recurrent with a mean follow up was 5.5 years (range 2.5-8.5 years). We discuss the current management, diagnosis, biological behavior, histopathology, prognosis and survival of head and neck RMS in neonates and young children.

H2O2 oxidative damage in cultured oral epithelial cells: the effect of short-term vitamin C exposure.

Smoking is the main etiology of oral cancer and generates oxygen free radicals in the oral cavity. Free radicals have been implicated in apoptosis and in DNA damage inducing alteration of the cell cycle. The antioxidant vitamin C (VC) is reported to inhibit damage induced by free radicals. We exposed cultures of normal human oral epithelial cells to hydrogen peroxide (H2O2) in the presence and absence of VC. Generation of hydroxyl radicals was measured by electron paramagnetic resonance (EPR), cell cycle alterations by flow cytometry, cell death by SYTO 11 and morphology by organotypic culture. Human primary cell culture was given four treatments - control, VC alone, H2O2 alone and VC followed by H2O2. Cell cycle analysis indicated cultures treated with H2O2 had fewer cells in G1 phase (26%) and higher number of cells in S phase (44%) compared to the control (G1 70% & S 14%). Cell cycle of 48 hour VC treatment followed by H2O2 was similar to H2O2 alone. SYTO 11 showed 22% cell death when treated with H2O2 alone compared to 9% of normal control. By organotypic culture H2O2 alone induced a two-fold cell proliferation, loss of maturation, nuclear hyperchromatism and nuclear crowding. Our results suggest that H2O2 is capable of altering the cell cycle and morphology of cultured normal human oral epithelial cells. Forty-eight hour exposure to Vitamin C does not prevent the cell cycle changes caused by hydroxyl radicals.

Phosphatidylethanolamine deficiency in membrane lipids inhibits keratinocyte intercellular networks formation.

Ethanolamine (Etn) is required for the growth of epithelial cells in culture. Without Etn, the amount of phosphatidylethanolamine (PE) in membrane lipids is reduced, and cell proliferation stops. When the membrane lipids are deficient of PE, some extracellular signaling processes become impaired. In this study, we examined the effect of Etn deprivation on the formation of intercellular networks in immortalized human oral keratinocytes. Keratinocytes proliferate with undifferentiated morphologies in a low-calcium medium, whereas they undergo differentiation to form intercellular networks in a high-calcium medium. The cells were first cultured with or without Etn supplement in a low-calcium (0.07 mM) medium, and then the calcium concentration was raised to 1.8 mM. The localization and organization of the following proteins were examined: (1) desmogleins and plakoglobin in desmosomes, (2) E-cadherin and beta-catenin in adherens junctions and (3) actin and keratin filaments in cytoskeletons. As expected, in the Etn-supplemented cells, the elevated level of calcium induced the junctional localization of the proteins associated with desmosomes and adherens junctions and also induced the formation of keratin and actin networks. On the contrary, in the Etn-deprived cells, the elevated level of calcium induced none of the above processes. The results suggest that having a sufficient amount of PE or proper phospholipid composition in the membranes is crucial for differentiation in epithelial cells.

Progression of head and neck cancer in an in vitro model.

OBJECTIVE: To identify alterations in angiogenesis and cell cycle regulation as preneoplastic cells progress to cancer in an in vitro model of head and neck tumor progression. METHODS: Immortal human gingival keratinocyte (IHGK) cells (preneoplastic) were derived from normal oral keratinocytes and were immortalized with human papillomavirus 16. Transformation of IHGK cells with a carcinogen (NNK, 4-[methylnitrosamino]-1-[3-pyridyl]-1-butanone) gave rise to IHGKN cells. We determined the growth rates, cell cycle phase, expression of cell cycle regulators, and expression of vascular endothelial growth factor along with the organotypic features of these cells and compared them with characteristics of head and neck cancer cells. RESULTS: IHGK and IHGKN cells grown in raft culture were morphologically similar to severe dysplasia and carcinoma, respectively. The proportion of cells in G(0)/G(1) was similar between IHGK and IHGKN. However, the proportion of IHGK cells was 35% greater in S phase as compared with the IHGKN cells, while a greater percentage (40%) of IHGKN cells were in G(2)/M. The expression of the other cell cycle regulators tested was unchanged. IHGK cells secreted less vascular endothelial growth factor on day 1 when compared with IHGKN (50.6 vs 245.6 pg/mL), along with a lower overall production rate (79% vs 133%). CONCLUSIONS: Transformation of IHGK cells resulted in the activation of vascular endothelial growth factor associated with angiogenesis. Inactivation of the G(1) cell cycle regulation occurred during immortalization and before transformation, and was sustained after carcinogen exposure. These alterations correspond to changes observed in patients with head and neck squamous cell carcinoma. This model can be useful in testing novel therapeutic and preventive strategies.

Response of human oral epithelial cells to oxidative damage and the effect of vitamin E.

Smoking and periodontal inflammation are various conditions with the potential to induce oxidative stress and thus DNA damage in the oral cavity. In cellular defense systems, vitamin E is considered the most powerful lipid-soluble antioxidant. To investigate whether oxygen-free radicals alter normal progression of the cell cycle and whether vitamin E prevents this damage, we exposed cultured normal human oral epithelial cells to hydrogen peroxide (H2O2) in the presence or absence of vitamin E. Two primary cell lines were analyzed for the presence of hydroxyl radical, cell cycle distribution and morphology. Each cell line received five treatments: control, ethanol only, vitamin E only, H2O2 only or vitamin E followed by H2O2. Degradation of hydroxyl radicals was detected by electron paramagnetic resonance analysis, cell cycle by flow cytometry and morphology by organotypic technique. Hydroxyl radicals were generated in H2O2-treated cells at an initial concentration, which decreased over a period of time. Cell cycle analysis showed that H2O2-treated cells differed from normal cells in that the percentage of cells in the G1 phase decreased markedly (34.3 vs. 61.2% in control) and the S phase increased (35.5 vs. 15.6% in control). Organotypic cultures treated with H2O2 demonstrated nuclear hyperchromatism, loss of maturation and prominent nucleoli, features consistent with premalignant epithelial transformation. In conclusion, our data suggest that H2O2 produced hydroxyl radicals and altered the cell cycle. Also, vitamin E may have the potential to reduce oxidative damage caused by hydroxyl radicals.

Effect of brief exposure to mitomycin C on cultured human nasal mucosa fibroblasts.

PURPOSE: To observe the effect of mitomycin C (MMC) on cultured human nasal mucosa fibroblasts. METHODS: Cultured human nasal mucosa fibroblasts were exposed to MMC (0.1-0.4 mg/ml) for 1 to 5 minutes. The viability of the fibroblasts was determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay; DNA fragmentation (apoptosis) by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL); apoptotic percentage by flow cytometry; and morphology by light microscopy. RESULTS: A portion of the fibroblasts survived the mitomycin treatment and showed evidence of regrowth within 2 to 3 days. These cells reached confluence in 5 to 7 days. The inhibition rates by MTT assay of 0.4 mg/ml MMC for 5-minute exposures was 31.3%. Dose-response effect was noted with the lower concentrations and shorter exposure times where the inhibition rates were lower (but not significantly so). DNA fragmentation was observed in fibroblasts 24 hours after MMC exposure (0.4 mg/ml) for 5 minutes compared with normal control. The apoptotic rate of fibroblasts treated by 0.4 mg/ml MMC was significantly higher than the control (p < 0.05). CONCLUSIONS: Short MMC exposure times have a variable cytotoxic effect and inhibit proliferation of cultured nasal mucosa fibroblasts. MMC also can induce apoptosis with a 5-minute exposure time. Therefore, it is possible that MMC has a complex effect in dacryocystorhinostomy.

5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

Odontogenic keratocyst: the northwestern USA experience.

Odontogenic keratocyst (OKC) is a cyst of tooth origin with an aggressive clinical behavior including a high recurrence rate. OKC demographics in the northwestern United States are presented and compared to those reported elsewhere. A total of 430 cases were obtained from 393 patients of the northwest region over a period of 15 years. Data evaluated included: site, gender, age, race, and association with bifid-rib basal cell nevus syndrome (Gorlin syndrome). Site distribution of the northwest group was similar to that of international groups. For the northwest group, the most common lesion location was the body of the mandible. Gender distribution in the northwest group appeared similar to other reports made in Denmark, England, Japan, and other regions in the United States. However, when gender distribution was compared by decade of life, the northwest group had the largest cluster of males in the fourth decade and of females in the second decade. The greatest frequency in both genders occurred in the third decade. There were 18 of 258 (6.9%) male patients with OKC under age 10 in the northwest group and nearly 80% of the patients were Caucasian. The race factor is rarely described in other reports. Gorlin's syndrome was present in 5% of the patients, with a higher distribution in the first and second decades. In conclusion, this is the first report of OKC cases from the Pacific Northwest region of the United States of America.

The effect of mitomycin C after long-term storage on human Tenon's fibroblast proliferation.

PURPOSE: To investigate the effect of mitomycin C (MMC) after long-term storage on proliferation of human Tenon's fibroblasts in vitro. METHODS: Human Tenon's fibroblasts in tissue culture were exposed for 5 minutes to MMC (0.4 mg/mL) that was either freshly prepared or had been stored for as long as 18 months at either 4 degrees C or -20 degrees C. The MTT colorimetric assay was used to determine the inhibition of proliferation as measured indirectly by mitochondrial activity. RESULTS: The inhibition rate was 88% using fresh MMC, and declined to a mean of 73% when using MMC that had been stored for as long as 18 months at 4 degrees C; this decrease was not statistically significant. The mean inhibition for MMC stored at -20 degrees C was 68%, and this was significantly less than inhibition with fresh MMC. Inhibition did not vary significantly with MMC after different storage times. CONCLUSION: Mitomycin C continues to have strong in vitro antiproliferative effects when stored for as long as 18 months at 4 degrees C or -20 degrees C. A significant decline in potency compared with fresh MMC occurs when MMC is stored at -20 degrees C.

Case challenge. Pathological condition of a newborn child.

A new born female presented at birth with a paranasal mass which appeared to be hemangiomatous. An ultrasound and aspiration biopsy of the lesion performed at the time of initial exam were not diagnostic. Due to the rapidly increasing size of the mass, the patient was referred for further evaluation to the Department of Oral and Maxillofacial Surgery at a major northwestern children's hospital. The clinical work up included a bone scan, chest and abdominal CT, and soft tissue biopsy.

Effect of vitamins A, C and E on normal and HPV-immortalized human oral epithelial cells in culture.

There is experimental and epidemiological evidence that antioxidant vitamins can inhibit carcinogenesis. Since immortalization by Human Papilloma Virus (HPV) is one possible early step towards carcinogenesis in oral epithelia, we studied the differential effect of vitamins A, C and E on HPV-immortalized oral epithelial cells (IHGK) as compared to the normal counterpart. The dose response was determined by morphology, cell cycle by flow cytometry, and growth curve by cell number. The optimum dose in terms of inhibitory effect vs. toxicity was determined for each vitamin by morphology. Optimum doses were: vitamin A--1.4 x 10(-5) M, vitamin C--10(-3) M, and vitamin E--10(-6) M for both HPV-immortalized and normal cells. Growth curve showed reduction of proliferation by all three vitamins, with vitamins A and E more effective than C for both cell types. Flow cytometry showed that vitamins A and E reduced the percentage of cells at G2 phase of cell cycle and indicated arrest in the S phase. This effect was greatest in the immortalized cells with a 50% and 35% decrease of G2 for vitamins A and E respectively, whereas the normal counterpart showed a 48% decrease for A and a 12% increase for E. By organotypic culture, the morphology was not markedly different between the vitamin-treated and the control cells, except for a slight increase in the keratinization of normal cells with vitamin A. Also noted was a reduction in number of cell layers from five layers or more for controls to only one or two for vitamin E. In conclusion, we have demonstrated that the antioxidant vitamins inhibit proliferation, and show a preferential effect on IHGK cells.

Reconstituted human oral and esophageal mucosa in culture.

We have successfully established monolayer and organotypic culture techniques for growing human oral and esophageal epithelial cells. Cells in monolayer culture were grown in serum-free medium, modified from techniques previously reported by our group. The organotypic cultures were grown in a defined medium supplemented with 10% fetal calf serum. Oral and esophageal cells were maintained in keratinocyte basal medium with pituitary extract and other supplements, and 0.05 mM calcium for 7-9 and 9-11 passages, respectively. Both cell types had similar morphology by phase contrast microscopy. When confluent, the cells were predominantly small, basaloid, and uniform and interspersed with larger, differentiated cells. By immunohistochemistry, both cell types in monolayer were positive to AE1, AE3, and 34BE12 antibodies to keratins of stratified epithelia. Oral epithelial cells in monolayer also were positive to 35BH11, representative of simple epithelial keratins, while esophageal cells were not. The esophageal cells were focally positive to K13, while the oral cells were negative. Both were negative for K19. When comparing monolayer to organotypic cultures and to in vivo specimens, there was a significant difference in the expression of keratins. Using organotypic cultures, AE1, AE3, and 34BE12 were strongly positive in both oral and esophageal cells, similar to in vivo tissues. In contrast to monolayers, both were also focally positive for K19. Esophageal cells were strongly positive for K13, while the oral cells were mildly but uniformly positive. Both were negative for keratins of simple epithelia. These two cell culture techniques offer unique opportunities to study the pathobiology, including carcinogenesis, of stable cell systems from the oral and esophageal epithelia.