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EXPRESSION OF CANCER STEM CELLS SURFACE MARKERS IN TONGUE SQUAMOUS CELL CARCINOMA CELL LINES
OOOO Volume 129, Number 1 This study aims to analyze the incidence of calcification of the stylohyoid ligament through panoramic radiographs, considering its importance in clinical routine. Five hundred digitalized panoramic radiographs taken in 2017 were analyzed considering the incidence of the calcification of the stylohyoid ligament. The material was classified according to age, sex, and the affected side. According to the analysis, it was possible to identify a high frequency of calcifications (88%). It was observed that patients aged between 21 and 30 years old were the most affected ones (32.2%). Despite its probability, no significant difference was observed regarding the presence of calcifications by sex (P = .09, Fisher exact test). As to the affected side, bilateral calcification was more frequent (60%). Regarding the affected side and the sex of the patients, it was not possible to identify a significant difference (P = .20, Fisher exact test). It is possible to conclude that the radiographic findings of the calcification of the stylohyoid ligament are frequent, calling the attention of the professional to this condition, which can bring significant symptomatology for the patient sometimes.
FIRST DETECTION OF DENGUE VIRUS IN THE SALIVA OF IMMUNOCOMPETENT MURINE MODEL. MARCOS ALEXANDRE NUNES, ARTHUR DA COSTA RASINHAS, GABRIELA CARDOSO CALDAS, FERNANDA CUNHA JACOME, ORTRUD MONIKA BARTH and, DEBORA FERREIRA BARRETO-VIEIRA Introduction: The use of saliva as an alternative detection fluid for viral diseases has been proved to be very efficient in the molecular diagnosis of viruses and in the epidemiologic study of viral diseases, a public health problems. Besides allowing the early diagnosis, viral detection in saliva is performed in a noninvasive way, which facilitates the whole process. Objective: The goal of this study is the detection viral RNA in saliva of mice experimentally infected with dengue virus type 4 (DENV-4). Study Design: Fifteen BALB/c mice were infected with 10,000 TCID50/0.1 mL of DENV-4 by intravenously. Seventytwo hours postinfection, samples of their saliva were collected. Viral RNA was extracted and subjected to real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results: The viral genome was detected in samples of 2 mice, both presenting titers of 109 RNA copies RNA/mL. While detection of DENV RNA via saliva sampling is not commonly used as a diagnosis tool due to low detection rates in human patients, the results observed in this study seem to indicate that, as in humans, saliva sampling in mice also produces low detection rates, correlating the infection in both cases. Conclusions: This study reports the first DENV detection in saliva of BALB/c immunocompetent mice experimentally infected with nonneuroadapted DENV-4.
COMPARISON OF PROGNOSTIC FACTORS OF ORAL SQUAMOUS CELL CARCINOMA BETWEEN YOUNG AND OLDER PATIENTS: A SYSTEMATIC REVIEW. ROBECI ALVES MACEDO FILHO, INGRID MORGANA FERNANDES GONCALVES, ¸ CASSIANO FRANCISCO WEEGE NONAKA and, POLLIANNA MUNIZ ALVES
ABSTRACTS
e171
Objective: Biological behavior of oral squamous cell carcinoma (OSCC) in young patients is controversial. Therefore, this study compared prognostic factors of OSCC between young and older patients, through a systemic review. Study Design: Electronic search in the databases Pubmed/Scopus/Ovid/Proquest/Cochrane Library/Web of Science/Lilacs was performed until April 2018, without filters by language or year of publication. Only cohort studies were included and evaluated by Newcastle-Ottawa scale for risk of bias. Analysis of titles and abstracts was made by 2 calibrated researchers (kappa = 0.94). Results: A total of 5370 articles were reviewed, and 10 of the most relevant articles that are suitable to the criteria were selected. All included studies had a low risk of bias. Independent of age group, male was more affected, and tongue was anatomic site more frequent. Main risk factors were smoking and alcoholism in both age groups. Most patients in both age groups were diagnosed in early clinical stages (I/II). Regarding to histologic grading, majority of OSCC was classified as moderately differentiated in both age groups. Lower survival was observed in older patients. Conclusions: Among all prognostic factors evaluated, survival showed difference in age groups, suggesting, then, that OSCC in older patients could exhibit more aggressive biologic behavior.
EFFECTS OF CAFFEIC ACID PHENETHYL ESTER ON THE EXPRESSION OF INTERLEUKINS 6 AND 8 IN ORAL SQUAMOUS CELL CARCINOMA. LEONARDO RINALDI, LEOPOLDO LUIZ ROCHA FUJII and, LUCAS NOVAES TEIXEIRA Objective: The present study evaluated the effects of caffeic acid phenethyl ester (CAPE) on cell viability and expression of interleukins (IL-) 6 and 8 in oral squamous cell carcinoma cell cultures (CAL-27). Study Design: CAL-27 cells were plated at the density of 3500 cells/well in 96-well plates and exposed to CAPE at concentrations of 0, 25, 50, 100, and 200 mM. After 24, 48, and 72 hours, the following parameters were evaluated: (1) cell viability by MTT and (2) expression of IL-6 and IL-8 by enzymelinked immunosorbent assay (ELISA). Data were submitted to the Kruskal-Wallis test, followed by Student-Newman-Keuls test (a = 5%). Results: CAL-27 cultures exposed to CAPE exhibited a reduction in cell viability compared to the nonexposed CAL-27 cultures (P < .05). The lowest cell viability was identified in cultures exposed to 200 mM CAPE in all experimental time points evaluated (P < .05). The highest IL-6 and IL-8 levels were detected in CAL-27 cultures exposed for 48 hours to 100 mM CAPE (P < .05). Conclusions: The results indicate that CAPE is able to reduce cell viability and modulate the expression of IL-6 and IL8 in CAL-27 cultures.
EXPRESSION OF CANCER STEM CELLS SURFACE MARKERS IN TONGUE SQUAMOUS CELL CARCINOMA CELL LINES. FLORENCE JUANA MARIA CUADRA, IARA AQUINO, RICARDO D. COLETTA, DEBORA CAMPANELLA BASTOS and, EDGARD GRANER
ABSTRACTS
e172
OOOO January 2020 PAES DE ALMEIDA, MARCIO AJUDARTE LOPES, RAGHU RADHAKRISHNAN, LUIZ PAULO KOWALSKI and, PABLO AGUSTIN VARGAS
Cancer stem cells (CSCs) comprise subpopulation of tumor cells associated with cancer initiation, progression, and treatment failures found in several malignancies, including oral squamous cells carcinoma (OSCC). Currently, among the methods employed for detection and isolation of CSCs the most used is flow cytometry immunophenotyping. The expression of surface markers for CSCs in SCC-9 cells, derived from tongue OSCC, has not been described in the literature. Objective: To identify and compare the expression of cell surface markers associate to CSCs in the SCC9 ZsGreen and its metastatic derivative SCC9 ZsGreen LN1 cells. Methods: The CSC subpopulations in cultured SCC9 ZsGreen and SCC9 ZsGreen LN1 were identified by flow cytometry using the following surface markers: CD44 standard (and its isoforms, CD44 v3, and CD44 v6), CD133, CD326, CD24, and CD271. Results: SCC9 ZsGreen cells were significantly more positive for CD44, CD133, and CD271 while SCC9 ZsGreen LN1 expressed more CD326. The positivity for CD24 e CD44 v3 was also higher in SCC9 ZsGreen LN1, however, without statistical significance. Conclusions: The 2 OSCC cell lines presented distinct patterns of cancer stem cells surface markers. Further experiments will be needed to better characterize the CSCs populations present in SCC9 and SCC9 ZsGreen LN1 cell lines. (FAPESP 14/ 20832-3).
Oral submucous fibrosis (OSMF) affects mainly the Indian subcontinent and its manifestations are closely linked with the habit of chewing betel. Immune system cells such as Langerhans cells (LC) and dendritic cells (DC) are linked to the immunopathogenesis of various lesions and have been studied for their ability of recognizing precursor malignant cells and to destroy them. Objective: Verify the distribution of LC and immature DC in OSMF and oral squamous cell carcinoma (OSCC). Study design: Fourteen cases of OSFM, 9 of OSCC associated to OSMF, and 40 of OSCC were retrieved. The original diagnoses were confirmed by 2 oral pathologists, who further classified OSFM in grade I, II, and III. OSCC cases were classified as well, moderately, and poorly differentiated. Immunoreactions against CD1a were performed and the number of positive LC and DC were quantified. Results: Analysis revealed a significant decrease in the number of immature LC and DC in OSCC compared to OSCCOSMF (P < .01) and OSMF (P < .01). No statistical correlation was found between OSMF and OSCC-OSMF. Conclusions: The results suggest that depletion of immature LC and DC would represent an important step for developing OSCC.
FASN INHIBITION WITH TVB-3166 IN ORAL SQUAMOUS CELL CARCINOMA CELL LINES. IARA GONCALVES ¸ DE AQUINO, FLORENCE CUADRA ZELAYA, RICARDO DELLA COLETTA, DEBORA CAMPANELLA BASTOS and, EDGARD GRANER
STUDY OF IMMUNOHISTOCHEMICAL EXPRESSION OF RANKL, OPG, IL-6, AND CTSK IN ODONTOGENIC CYSTS AND TUMORS. FERNANDA MARCELLO SCOTTI, DIOGO LENZI CAPELLA, DANIELLA SERAFIN COUTO VIEIRA and, ELENA RIET CORREA RIVERO
Introduction: Over-expression of fatty acid synthase (fasn) is associated with poor prognosis in several human cancers, including oral squamous cell carcinoma (OSCC). Objective: To evaluate the effects of the fasn inhibitor TVB-3166 on the cell viability, morphology, cell cycle and migration of SCC-9 and SCC-9 LN-1 cells derived from tongue OSCC. Study Design: Cell viability was determined using standard MTT assay after 48 hours of treatment with increasing doses of TVB-3166. IC50 was calculated with the aid of CompuSyn software. Morphology and cell migration assay were evaluated using phase contrast microscopy and cell cycle by flow cytometry. Results: TVB-3166 significantly reduced cell viability in a dose-dependent manner in both cells lines. IC50 was 30 mM and 37 mM for SCC-9 and LN-1, respectively. TVB-3166 induced morphologic changes of the cells to a fusiform pattern. In SCC-9 cells the treatment reduced cell migration, diminished the number of cells in the S phase, and increased the number of cells in G0/G1 phase. Conclusions: Our preliminary results suggest that fasn inhibition with TVB-3166 affects important processes of tumor progression in cell lines derived from OSCC.
Objective: To evaluate the receptor activator of nuclear factor-K B ligand (RANKL), osteoprotegerin (OPG), interleukin-6 (IL-6), and cathepsin K (CTSK) antigens in 34 radicular cysts (RC), 28 odontogenic keratocysts (OKC), and 25 ameloblastomas (AM). Study Design: The immunohistochemical expression of these proteins was evaluated considering the mean of percentage marked area in 10 consecutive fields (400 x), considering epithelial and connective tissue individually. The analysis of variance (ANOVA) test was used to compare the groups. A P value < .05 indicated statistically significant data. Results: RANKL was statistically higher in CR and OKC compared with AM in epithelium and connective tissue. No statistical difference was found for the OPG between the groups. Statistical difference was noted to IL-6 between all groups only in connective tissue, decreasing from CR, to AM, and to OKC. CTSK was statistically higher in AM and OKC compared with CR. Conclusions: In AM and OKC cases, the mechanism of bone resorption seems to be related to the CTSK pathway, whose expression was independent of the inflammatory component. To RC the mechanism of bone resorption is more related with RANKL and IL-6 proteins and dependent of an inflammatory component.
CD1A+ LANGERHANS CELLS ARE REDUCED IN ORAL SUBMUCOUS FIBROSIS AND ORAL SQUAMOUS CELL CARCINOMA. LUAN CESAR DA SILVA, FELIPE PAIVA FONSECA, OSLEI
HISTOLOGIC COMPARATIVE STUDY OF BONE SPECIMENS FROM BISPHOSPHONATE-RELATED OSTEONECROSIS OF THE
FIRST DETECTION OF DENGUE VIRUS IN THE SALIVA OF IMMUNOCOMPETENT MURINE MODEL. MARCOS ALEXANDRE NUNES, ARTHUR DA COSTA RASINHAS, GABRIELA CARDOSO CALDAS, FERNANDA CUNHA JACOME, ORTRUD MONIKA BARTH and, DEBORA FERREIRA BARRETO-VIEIRA Introduction: The use of saliva as an alternative detection fluid for viral diseases has been proved to be very efficient in the molecular diagnosis of viruses and in the epidemiologic study of viral diseases, a public health problems. Besides allowing the early diagnosis, viral detection in saliva is performed in a noninvasive way, which facilitates the whole process. Objective: The goal of this study is the detection viral RNA in saliva of mice experimentally infected with dengue virus type 4 (DENV-4). Study Design: Fifteen BALB/c mice were infected with 10,000 TCID50/0.1 mL of DENV-4 by intravenously. Seventytwo hours postinfection, samples of their saliva were collected. Viral RNA was extracted and subjected to real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results: The viral genome was detected in samples of 2 mice, both presenting titers of 109 RNA copies RNA/mL. While detection of DENV RNA via saliva sampling is not commonly used as a diagnosis tool due to low detection rates in human patients, the results observed in this study seem to indicate that, as in humans, saliva sampling in mice also produces low detection rates, correlating the infection in both cases. Conclusions: This study reports the first DENV detection in saliva of BALB/c immunocompetent mice experimentally infected with nonneuroadapted DENV-4.
COMPARISON OF PROGNOSTIC FACTORS OF ORAL SQUAMOUS CELL CARCINOMA BETWEEN YOUNG AND OLDER PATIENTS: A SYSTEMATIC REVIEW. ROBECI ALVES MACEDO FILHO, INGRID MORGANA FERNANDES GONCALVES, ¸ CASSIANO FRANCISCO WEEGE NONAKA and, POLLIANNA MUNIZ ALVES
ABSTRACTS
e171
Objective: Biological behavior of oral squamous cell carcinoma (OSCC) in young patients is controversial. Therefore, this study compared prognostic factors of OSCC between young and older patients, through a systemic review. Study Design: Electronic search in the databases Pubmed/Scopus/Ovid/Proquest/Cochrane Library/Web of Science/Lilacs was performed until April 2018, without filters by language or year of publication. Only cohort studies were included and evaluated by Newcastle-Ottawa scale for risk of bias. Analysis of titles and abstracts was made by 2 calibrated researchers (kappa = 0.94). Results: A total of 5370 articles were reviewed, and 10 of the most relevant articles that are suitable to the criteria were selected. All included studies had a low risk of bias. Independent of age group, male was more affected, and tongue was anatomic site more frequent. Main risk factors were smoking and alcoholism in both age groups. Most patients in both age groups were diagnosed in early clinical stages (I/II). Regarding to histologic grading, majority of OSCC was classified as moderately differentiated in both age groups. Lower survival was observed in older patients. Conclusions: Among all prognostic factors evaluated, survival showed difference in age groups, suggesting, then, that OSCC in older patients could exhibit more aggressive biologic behavior.
EFFECTS OF CAFFEIC ACID PHENETHYL ESTER ON THE EXPRESSION OF INTERLEUKINS 6 AND 8 IN ORAL SQUAMOUS CELL CARCINOMA. LEONARDO RINALDI, LEOPOLDO LUIZ ROCHA FUJII and, LUCAS NOVAES TEIXEIRA Objective: The present study evaluated the effects of caffeic acid phenethyl ester (CAPE) on cell viability and expression of interleukins (IL-) 6 and 8 in oral squamous cell carcinoma cell cultures (CAL-27). Study Design: CAL-27 cells were plated at the density of 3500 cells/well in 96-well plates and exposed to CAPE at concentrations of 0, 25, 50, 100, and 200 mM. After 24, 48, and 72 hours, the following parameters were evaluated: (1) cell viability by MTT and (2) expression of IL-6 and IL-8 by enzymelinked immunosorbent assay (ELISA). Data were submitted to the Kruskal-Wallis test, followed by Student-Newman-Keuls test (a = 5%). Results: CAL-27 cultures exposed to CAPE exhibited a reduction in cell viability compared to the nonexposed CAL-27 cultures (P < .05). The lowest cell viability was identified in cultures exposed to 200 mM CAPE in all experimental time points evaluated (P < .05). The highest IL-6 and IL-8 levels were detected in CAL-27 cultures exposed for 48 hours to 100 mM CAPE (P < .05). Conclusions: The results indicate that CAPE is able to reduce cell viability and modulate the expression of IL-6 and IL8 in CAL-27 cultures.
EXPRESSION OF CANCER STEM CELLS SURFACE MARKERS IN TONGUE SQUAMOUS CELL CARCINOMA CELL LINES. FLORENCE JUANA MARIA CUADRA, IARA AQUINO, RICARDO D. COLETTA, DEBORA CAMPANELLA BASTOS and, EDGARD GRANER
ABSTRACTS
e172
OOOO January 2020 PAES DE ALMEIDA, MARCIO AJUDARTE LOPES, RAGHU RADHAKRISHNAN, LUIZ PAULO KOWALSKI and, PABLO AGUSTIN VARGAS
Cancer stem cells (CSCs) comprise subpopulation of tumor cells associated with cancer initiation, progression, and treatment failures found in several malignancies, including oral squamous cells carcinoma (OSCC). Currently, among the methods employed for detection and isolation of CSCs the most used is flow cytometry immunophenotyping. The expression of surface markers for CSCs in SCC-9 cells, derived from tongue OSCC, has not been described in the literature. Objective: To identify and compare the expression of cell surface markers associate to CSCs in the SCC9 ZsGreen and its metastatic derivative SCC9 ZsGreen LN1 cells. Methods: The CSC subpopulations in cultured SCC9 ZsGreen and SCC9 ZsGreen LN1 were identified by flow cytometry using the following surface markers: CD44 standard (and its isoforms, CD44 v3, and CD44 v6), CD133, CD326, CD24, and CD271. Results: SCC9 ZsGreen cells were significantly more positive for CD44, CD133, and CD271 while SCC9 ZsGreen LN1 expressed more CD326. The positivity for CD24 e CD44 v3 was also higher in SCC9 ZsGreen LN1, however, without statistical significance. Conclusions: The 2 OSCC cell lines presented distinct patterns of cancer stem cells surface markers. Further experiments will be needed to better characterize the CSCs populations present in SCC9 and SCC9 ZsGreen LN1 cell lines. (FAPESP 14/ 20832-3).
Oral submucous fibrosis (OSMF) affects mainly the Indian subcontinent and its manifestations are closely linked with the habit of chewing betel. Immune system cells such as Langerhans cells (LC) and dendritic cells (DC) are linked to the immunopathogenesis of various lesions and have been studied for their ability of recognizing precursor malignant cells and to destroy them. Objective: Verify the distribution of LC and immature DC in OSMF and oral squamous cell carcinoma (OSCC). Study design: Fourteen cases of OSFM, 9 of OSCC associated to OSMF, and 40 of OSCC were retrieved. The original diagnoses were confirmed by 2 oral pathologists, who further classified OSFM in grade I, II, and III. OSCC cases were classified as well, moderately, and poorly differentiated. Immunoreactions against CD1a were performed and the number of positive LC and DC were quantified. Results: Analysis revealed a significant decrease in the number of immature LC and DC in OSCC compared to OSCCOSMF (P < .01) and OSMF (P < .01). No statistical correlation was found between OSMF and OSCC-OSMF. Conclusions: The results suggest that depletion of immature LC and DC would represent an important step for developing OSCC.
FASN INHIBITION WITH TVB-3166 IN ORAL SQUAMOUS CELL CARCINOMA CELL LINES. IARA GONCALVES ¸ DE AQUINO, FLORENCE CUADRA ZELAYA, RICARDO DELLA COLETTA, DEBORA CAMPANELLA BASTOS and, EDGARD GRANER
STUDY OF IMMUNOHISTOCHEMICAL EXPRESSION OF RANKL, OPG, IL-6, AND CTSK IN ODONTOGENIC CYSTS AND TUMORS. FERNANDA MARCELLO SCOTTI, DIOGO LENZI CAPELLA, DANIELLA SERAFIN COUTO VIEIRA and, ELENA RIET CORREA RIVERO
Introduction: Over-expression of fatty acid synthase (fasn) is associated with poor prognosis in several human cancers, including oral squamous cell carcinoma (OSCC). Objective: To evaluate the effects of the fasn inhibitor TVB-3166 on the cell viability, morphology, cell cycle and migration of SCC-9 and SCC-9 LN-1 cells derived from tongue OSCC. Study Design: Cell viability was determined using standard MTT assay after 48 hours of treatment with increasing doses of TVB-3166. IC50 was calculated with the aid of CompuSyn software. Morphology and cell migration assay were evaluated using phase contrast microscopy and cell cycle by flow cytometry. Results: TVB-3166 significantly reduced cell viability in a dose-dependent manner in both cells lines. IC50 was 30 mM and 37 mM for SCC-9 and LN-1, respectively. TVB-3166 induced morphologic changes of the cells to a fusiform pattern. In SCC-9 cells the treatment reduced cell migration, diminished the number of cells in the S phase, and increased the number of cells in G0/G1 phase. Conclusions: Our preliminary results suggest that fasn inhibition with TVB-3166 affects important processes of tumor progression in cell lines derived from OSCC.
Objective: To evaluate the receptor activator of nuclear factor-K B ligand (RANKL), osteoprotegerin (OPG), interleukin-6 (IL-6), and cathepsin K (CTSK) antigens in 34 radicular cysts (RC), 28 odontogenic keratocysts (OKC), and 25 ameloblastomas (AM). Study Design: The immunohistochemical expression of these proteins was evaluated considering the mean of percentage marked area in 10 consecutive fields (400 x), considering epithelial and connective tissue individually. The analysis of variance (ANOVA) test was used to compare the groups. A P value < .05 indicated statistically significant data. Results: RANKL was statistically higher in CR and OKC compared with AM in epithelium and connective tissue. No statistical difference was found for the OPG between the groups. Statistical difference was noted to IL-6 between all groups only in connective tissue, decreasing from CR, to AM, and to OKC. CTSK was statistically higher in AM and OKC compared with CR. Conclusions: In AM and OKC cases, the mechanism of bone resorption seems to be related to the CTSK pathway, whose expression was independent of the inflammatory component. To RC the mechanism of bone resorption is more related with RANKL and IL-6 proteins and dependent of an inflammatory component.
CD1A+ LANGERHANS CELLS ARE REDUCED IN ORAL SUBMUCOUS FIBROSIS AND ORAL SQUAMOUS CELL CARCINOMA. LUAN CESAR DA SILVA, FELIPE PAIVA FONSECA, OSLEI
HISTOLOGIC COMPARATIVE STUDY OF BONE SPECIMENS FROM BISPHOSPHONATE-RELATED OSTEONECROSIS OF THE