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Nucleus-targeted Dmp1 transgene fails to rescue dental defects in Dmp1 null mice.

Shu-Xian LIN ; Qi ZHANG ; Hua ZHANG ; Kevin YAN ; Leanne WARD ; Yong-Bo LU ; Jian-Quan FENG

International Journal of Oral Science.2014;6(3):133-141. doi:10.1038/ijos.2014.44

Dentin matrix protein 1 (DMP1) is essential to odontogenesis. Its mutations in human subjects lead to dental problems such as dental deformities, hypomineralization and periodontal impairment. Primarily, DMP1 is considered as an extracellular matrix protein that promotes hydroxyapatite formation and activates intracellular signaling pathway via interacting with αvβ3 integrin. Recent in vitro studies suggested that DMP1 might also act as a transcription factor. In this study, we examined whether full-length DMP1 could function as a transcription factor in the nucleus and regulate odontogenesis in vivo. We first demonstrated that a patient with the DMP1 M1V mutation, which presumably causes a loss of the secretory DMP1 but does not affect the nuclear translocation of DMP1, shows a typical rachitic tooth defect. Furthermore, we generated transgenic mice expressing (NLS)DMP1, in which the endoplasmic reticulum (ER) entry signal sequence of DMP1 was replaced by a nuclear localization signal (NLS) sequence, under the control of a 3.6 kb rat type I collagen promoter plus a 1.6 kb intron 1. We then crossbred the (NLS)DMP1 transgenic mice with Dmp1 null mice to express the (NLS)DMP1 in Dmp1-deficient genetic background. Although immunohistochemistry demonstrated that (NLS)DMP1 was localized in the nuclei of the preodontoblasts and odontoblasts, the histological, morphological and biochemical analyses showed that it failed to rescue the dental and periodontal defects as well as the delayed tooth eruption in Dmp1 null mice. These data suggest that the full-length DMP1 plays no apparent role in the nucleus during odontogenesis.
Animals ; Cell Nucleus ; genetics ; Codon, Initiator ; genetics ; Collagen Type I ; genetics ; Endoplasmic Reticulum ; genetics ; Extracellular Matrix Proteins ; genetics ; Familial Hypophosphatemic Rickets ; genetics ; Gene Targeting ; methods ; Genetic Vectors ; genetics ; Humans ; Introns ; genetics ; Methionine ; genetics ; Mice, Inbred C57BL ; Mice, Transgenic ; Mutation ; genetics ; Odontoblasts ; cytology ; Odontogenesis ; genetics ; Periodontal Diseases ; genetics ; Periodontal Ligament ; pathology ; Phosphoproteins ; genetics ; Promoter Regions, Genetic ; genetics ; Tooth Abnormalities ; genetics ; Tooth Eruption ; genetics ; Transcription Factors ; genetics ; Transgenes ; genetics ; Valine ; genetics ; Young Adult

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Oral microbiota and host innate immune response in bisphosphonate-related osteonecrosis of the jaw.

Smruti PUSHALKAR ; Xin LI ; Zoya KURAGO ; Lalitha V RAMANATHAPURAM ; Satoko MATSUMURA ; Kenneth E FLEISHER ; Robert GLICKMAN ; Wenbo YAN ; Yihong LI ; Deepak SAXENA

International Journal of Oral Science.2014;6(4):219-226. doi:10.1038/ijos.2014.46

Bacterial biofilms have emerged as potential critical triggers in the pathogenesis of bisphosphonate (BP)-related osteonecrosis of the jaw (ONJ) or BRONJ. BRONJ lesions have shown to be heavily colonized by oral bacteria, most of these difficult to cultivate and presents many clinical challenges. The purpose of this study was to characterize the bacterial diversity in BRONJ lesions and to determine host immune response. We examined tissue specimens from three cohorts (n=30); patients with periodontal disease without a history of BP therapy (Control, n=10), patients with periodontal disease having history of BP therapy but without ONJ (BP, n=5) and patients with BRONJ (BRONJ, n=15). Denaturing gradient gel electrophoresis of polymerase chain reaction (PCR)-amplified 16S rRNA gene fragments revealed less bacterial diversity in BRONJ than BP and Control cohorts. Sequence analysis detected six phyla with predominant affiliation to Firmicutes in BRONJ (71.6%), BP (70.3%) and Control (59.1%). Significant differences (P<0.05) in genera were observed, between Control/BP, Control/BRONJ and BP/BRONJ cohorts. Enzyme-linked immunosorbent assay (ELISA) results indicated that the levels of myeloperoxidase were significantly lower, whereas interleukin-6 and tumor necrosis factor-alpha levels were moderately elevated in BRONJ patients as compared to Controls. PCR array showed significant changes in BRONJ patients with downregulation of host genes, such as nucleotide-binding oligomerization domain containing protein 2, and cathepsin G, the key modulators for antibacterial response and upregulation of secretory leukocyte protease inhibitor, proteinase 3 and conserved helix-loop-helix ubiquitous kinase. The results suggest that colonization of unique bacterial communities coupled with deficient innate immune response is likely to impact the pathogenesis of ONJ.
Actinobacteria ; classification ; Bacteria ; classification ; Bacteroidetes ; classification ; Biofilms ; Bisphosphonate-Associated Osteonecrosis of the Jaw ; immunology ; microbiology ; Bone Density Conservation Agents ; therapeutic use ; Cathepsin G ; analysis ; Cohort Studies ; Down-Regulation ; Female ; Fusobacteria ; classification ; Gram-Negative Bacteria ; classification ; Host-Pathogen Interactions ; immunology ; Humans ; I-kappa B Kinase ; analysis ; Immunity, Innate ; immunology ; Interleukin-6 ; analysis ; Male ; Middle Aged ; Mouth ; immunology ; microbiology ; Myeloblastin ; analysis ; antagonists & inhibitors ; Nod2 Signaling Adaptor Protein ; analysis ; Periodontal Diseases ; microbiology ; Peroxidase ; analysis ; Proteobacteria ; classification ; Tumor Necrosis Factor-alpha ; analysis

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The mucosal immune system in the oral cavity-an orchestra of T cell diversity.

Rui-Qing WU ; Dun-Fang ZHANG ; Eric TU ; Qian-Ming CHEN ; WanJun CHEN

International Journal of Oral Science.2014;6(3):125-132. doi:10.1038/ijos.2014.48

The mucosal immune system defends against a vast array of pathogens, yet it exhibits limited responses to commensal microorganisms under healthy conditions. The oral-pharyngeal cavity, the gateway for both the gastrointestinal and respiratory tracts, is composed of complex anatomical structures and is constantly challenged by antigens from air and food. The mucosal immune system of the oral-pharyngeal cavity must prevent pathogen entry while maintaining immune homeostasis, which is achieved via a range of mechanisms that are similar or different to those utilized by the gastrointestinal immune system. In this review, we summarize the features of the mucosal immune system, focusing on T cell subsets and their functions. We also discuss our current understanding of the oral-pharyngeal mucosal immune system.
Epithelium ; immunology ; Humans ; Immunity, Cellular ; Immunity, Mucosal ; immunology ; Mouth Diseases ; immunology ; Mouth Mucosa ; immunology ; Pharynx ; immunology ; T-Lymphocyte Subsets ; classification ; immunology

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Metabolic activity of Streptococcus mutans biofilms and gene expression during exposure to xylitol and sucrose.

Eva-Maria DECKER ; Christian KLEIN ; Dimitri SCHWINDT ; Christiane von OHLE

International Journal of Oral Science.2014;6(4):195-204. doi:10.1038/ijos.2014.38

The objective of the study was to analyse Streptococcus mutans biofilms grown under different dietary conditions by using multifaceted methodological approaches to gain deeper insight into the cariogenic impact of carbohydrates. S. mutans biofilms were generated during a period of 24 h in the following media: Schaedler broth as a control medium containing endogenous glucose, Schaedler broth with an additional 5% sucrose, and Schaedler broth supplemented with 1% xylitol. The confocal laser scanning microscopy (CLSM)-based analyses of the microbial vitality, respiratory activity (5-cyano-2,3-ditolyl tetrazolium chloride, CTC) and production of extracellular polysaccharides (EPS) were performed separately in the inner, middle and outer biofilm layers. In addition to the microbiological sample testing, the glucose/sucrose consumption of the biofilm bacteria was quantified, and the expression of glucosyltransferases and other biofilm-associated genes was investigated. Xylitol exposure did not inhibit the viability of S. mutans biofilms, as monitored by the following experimental parameters: culture growth, vitality, CTC activity and EPS production. However, xylitol exposure caused a difference in gene expression compared to the control. GtfC was upregulated only in the presence of xylitol. Under xylitol exposure, gtfB was upregulated by a factor of 6, while under sucrose exposure, it was upregulated by a factor of three. Compared with glucose and xylitol, sucrose increased cell vitality in all biofilm layers. In all nutrient media, the intrinsic glucose was almost completely consumed by the cells of the S. mutans biofilm within 24 h. After 24 h of biofilm formation, the multiparametric measurements showed that xylitol in the presence of glucose caused predominantly genotypic differences but did not induce metabolic differences compared to the control. Thus, the availability of dietary carbohydrates in either a pure or combined form seems to affect the cariogenic potential of S. mutans biofilms.
Bacterial Load ; drug effects ; Bacteriological Techniques ; Biofilms ; drug effects ; Cariogenic Agents ; metabolism ; pharmacology ; Culture Media ; Dental Enamel ; microbiology ; Fluorescent Dyes ; Gene Expression Regulation, Bacterial ; drug effects ; Gene Expression Regulation, Enzymologic ; drug effects ; Genotype ; Glucose ; metabolism ; Glucosyltransferases ; metabolism ; Humans ; Microbial Viability ; drug effects ; Microscopy, Confocal ; Polysaccharides, Bacterial ; biosynthesis ; Streptococcus mutans ; drug effects ; enzymology ; metabolism ; Sucrose ; metabolism ; pharmacology ; Sweetening Agents ; metabolism ; pharmacology ; Tetrazolium Salts ; Time Factors ; Up-Regulation ; Xylitol ; metabolism ; pharmacology

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The clinical effectiveness of reflectance optical spectroscopy for the in vivo diagnosis of oral lesions.

Diana V MESSADI ; Fariba S YOUNAI ; Hong-Hu LIU ; Gao GUO ; Cun-Yu WANG

International Journal of Oral Science.2014;6(3):162-167. doi:10.1038/ijos.2014.39

Optical spectroscopy devices are being developed and tested for the screening and diagnosis of oral precancer and cancer lesions. This study reports a device that uses white light for detection of suspicious lesions and green-amber light at 545 nm that detect tissue vascularity on patients with several suspicious oral lesions. The clinical grading of vascularity was compared to the histological grading of the biopsied lesions using specific biomarkers. Such a device, in the hands of dentists and other health professionals, could greatly increase the number of oral cancerous lesions detected in early phase. The purpose of this study is to correlate the clinical grading of tissue vascularity in several oral suspicious lesions using the Identafi(®) system with the histological grading of the biopsied lesions using specific vascular markers. Twenty-one patients with various oral lesions were enrolled in the study. The lesions were visualized using Identafi(®) device with white light illumination, followed by visualization of tissue autofluorescence and tissue reflectance. Tissue biopsied was obtained from the all lesions and both histopathological and immunohistochemical studies using a vascular endothelial biomarker (CD34) were performed on these tissue samples. The clinical vascular grading using the green-amber light at 545 nm and the expression pattern and intensity of staining for CD34 in the different biopsies varied depending on lesions, grading ranged from 1 to 3. The increase in vascularity was observed in abnormal tissues when compared to normal mucosa, but this increase was not limited to carcinoma only as hyperkeratosis and other oral diseases, such as lichen planus, also showed increase in vascularity. Optical spectroscopy is a promising technology for the detection of oral mucosal abnormalities; however, further investigations with a larger population group is required to evaluate the usefulness of these devices in differentiating benign lesions from potentially malignant lesions.
Adult ; Aged ; Aged, 80 and over ; Antigens, CD34 ; analysis ; Biomarkers, Tumor ; analysis ; Biopsy ; methods ; Carcinoma, Squamous Cell ; blood supply ; diagnosis ; pathology ; Erythroplasia ; diagnosis ; pathology ; Female ; Humans ; Immunohistochemistry ; Leukoplakia, Oral ; blood supply ; diagnosis ; pathology ; Lichen Planus, Oral ; diagnosis ; pathology ; Male ; Middle Aged ; Mouth Neoplasms ; blood supply ; diagnosis ; pathology ; Neoplasm Grading ; Optical Imaging ; methods ; Pilot Projects ; Precancerous Conditions ; blood supply ; diagnosis ; pathology ; Spectrometry, Fluorescence ; methods ; Young Adult

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Surface roughness of zirconia for full-contour crowns after clinically simulated grinding and polishing.

Rim HMAIDOUCH ; Wolf-Dieter MÜLLER ; Hans-Christoph LAUER ; Paul WEIGL

International Journal of Oral Science.2014;6(4):241-246. doi:10.1038/ijos.2014.34

The aim of this study was to evaluate the effect of controlled intraoral grinding and polishing on the roughness of full-contour zirconia compared to classical veneered zirconia. Thirty bar-shaped zirconia specimens were fabricated and divided into two groups (n=15). Fifteen specimens (group 1) were glazed and 15 specimens (group 2) were veneered with feldspathic ceramic and then glazed. Prior to grinding, maximum roughness depth (Rmax) values were measured using a profilometer, 5 times per specimen. Simulated clinical grinding and polishing were performed on the specimens under water coolant for 15 s and 2 N pressure. For grinding, NTI diamonds burs with grain sizes of 20 µm, 10 µm, and 7.5 µm were used sequentially. The ground surfaces were polished using NTI kits with coarse, medium and fine polishers. After each step, Rmax values were determined. Differences between groups were examined using one-way analysis of variance (ANOVA). The roughness of group 1 was significantly lower than that of group 2. The roughness increased significantly after coarse grinding in both groups. The results after glazing were similar to those obtained after fine grinding for non-veneered zirconia. However, fine-ground veneered zirconia had significantly higher roughness than venerred, glazed zirconia. No significant difference was found between fine-polished and glazed zirconia, but after the fine polishing of veneered zirconia, the roughness was significantly higher than after glazing. It can be concluded that for full-contour zirconia, fewer defects and lower roughness values resulted after grinding and polishing compared to veneered zirconia. After polishing zirconia, lower roughness values were achieved compared to glazing; more interesting was that the grinding of glazed zirconia using the NTI three-step system could deliver smooth surfaces comparable to untreated glazed zirconia surfaces.
Aluminum Silicates ; chemistry ; Ceramics ; chemistry ; Crowns ; Dental Materials ; chemistry ; Dental Polishing ; instrumentation ; methods ; Dental Prosthesis Design ; Dental Veneers ; Diamond ; chemistry ; Humans ; Materials Testing ; Microscopy, Electron, Scanning ; Particle Size ; Potassium Compounds ; chemistry ; Pressure ; Surface Properties ; Time Factors ; Water ; chemistry ; Yttrium ; chemistry ; Zirconium ; chemistry

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Comparison of the rheological properties of four root canal sealers.

Seok Woo CHANG ; Young-Kyu LEE ; Qiang ZHU ; Won Jun SHON ; Woo Cheol LEE ; Kee Yeon KUM ; Seung Ho BAEK ; In Bog LEE ; Bum-Soon LIM ; Kwang Shik BAE

International Journal of Oral Science.2015;7(1):56-61. doi:10.1038/ijos.2014.33

The flowability of a root canal sealer is clinically important because it improves the penetration of the sealer into the complex root canal system. The purpose of this study was to compare the flowabilities of four root canal sealers, measured using the simple press method (ISO 6876), and their viscosities, measured using a strain-controlled rheometer. A newly developed, calcium phosphate-based root canal sealer (Capseal) and three commercial root canal sealers (AH Plus, Sealapex and Pulp Canal Sealer EWT) were used in this study. The flowabilities of the four root canal sealers were measured using the simple press method (n=5) and their viscosities were measured using a strain-controlled rheometer (n=5). The correlation between these two values was statistically analysed using Spearman's correlation test. The flow diameters and the viscosities of the root canal sealers were strongly negatively correlated (ρ=-0.8618). The viscosity of Pulp Canal Sealer EWT was the lowest and increased in the following order: AH Plus Materials Testing ; Rheology ; Root Canal Filling Materials ; Temperature ; Viscosity

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Expression of cyclin-dependent kinase inhibitor 2A 16, tumour protein 53 and epidermal growth factor receptor in salivary gland carcinomas is not associated with oncogenic virus infection.

Ellen SENFT ; Juliana LEMOUND ; Angelika STUCKI-KOCH ; Nils-Claudius GELLRICH ; Hans KREIPE ; Kais HUSSEIN

International Journal of Oral Science.2015;7(1):18-22. doi:10.1038/ijos.2014.28

It is known that human papillomavirus (HPV) infection can cause squamous cell neoplasms at several sites, such as cervix uteri carcinoma and oral squamous carcinoma. There is little information on the expression of HPV and its predictive markers in tumours of the major and minor salivary glands of the head and neck. We therefore assessed oral salivary gland neoplasms to identify associations between HPV and infection-related epidermal growth factor receptor (EGFR), cyclin-dependent kinase inhibitor 2A (CDKN2A/p16) and tumour protein p53 (TP53). Formalin-fixed, paraffin-embedded tissue samples from oral salivary gland carcinomas (n=51) and benign tumours (n=26) were analysed by polymerase chain reaction (PCR) analysis for several HPV species, including high-risk types 16 and 18. Evaluation of EGFR, CDKN2A, TP53 and cytomegalovirus (CMV) was performed by immunohistochemistry. Epstein-Barr virus (EBV) was evaluated by EBV-encoded RNA in situ hybridisation. We demonstrated that salivary gland tumours are not associated with HPV infection. The expression of EGFR, CDKN2A and TP53 may be associated with tumour pathology but is not induced by HPV. CMV and EBV were not detectable. In contrast to oral squamous cell carcinomas, HPV, CMV and EBV infections are not associated with malignant or benign neoplastic lesions of the salivary glands.
Alphapapillomavirus ; isolation & purification ; Cohort Studies ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; Female ; Humans ; Male ; Middle Aged ; Polymerase Chain Reaction ; Receptor, Epidermal Growth Factor ; metabolism ; Salivary Gland Neoplasms ; metabolism ; virology ; Tumor Suppressor Protein p53 ; metabolism ; Tumor Virus Infections ; metabolism ; virology

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Bacterial entombment by intratubular mineralization following orthograde mineral trioxide aggregate obturation: a scanning electron microscopy study.

Jun Sang YOO ; Seok-Woo CHANG ; So Ram OH ; Hiran PERINPANAYAGAM ; Sang-Min LIM ; Yeon-Jee YOO ; Yeo-Rok OH ; Sang-Bin WOO ; Seung-Hyun HAN ; Qiang ZHU ; Kee-Yeon KUM

International Journal of Oral Science.2014;6(4):227-232. doi:10.1038/ijos.2014.30

The time domain entombment of bacteria by intratubular mineralization following orthograde canal obturation with mineral trioxide aggregate (MTA) was studied by scanning electron microscopy (SEM). Single-rooted human premolars (n=60) were instrumented to an apical size #50/0.06 using ProFile and treated as follows: Group 1 (n=10) was filled with phosphate buffered saline (PBS); Group 2 (n=10) was incubated with Enterococcus faecalis for 3 weeks, and then filled with PBS; Group 3 (n=20) was obturated orthograde with a paste of OrthoMTA (BioMTA, Seoul, Korea) and PBS; and Group 4 (n=20) was incubated with E. faecalis for 3 weeks and then obturated with OrthoMTA-PBS paste. Following their treatments, the coronal openings were sealed with PBS-soaked cotton and intermediate restorative material (IRM), and the roots were then stored in PBS for 1, 2, 4, 8 or 16 weeks. After each incubation period, the roots were split and their dentin/MTA interfaces examined in both longitudinal and horizontal directions by SEM. There appeared to be an increase in intratubular mineralization over time in the OrthoMTA-filled roots (Groups 3 and 4). Furthermore, there was a gradual entombment of bacteria within the dentinal tubules in the E. faecalis inoculated MTA-filled roots (Group 4). Therefore, the orthograde obturation of root canals with OrthoMTA mixed with PBS may create a favorable environment for bacterial entombment by intratubular mineralization.
Aluminum Compounds ; therapeutic use ; Calcification, Physiologic ; physiology ; Calcium Compounds ; therapeutic use ; Crystallization ; Dental Pulp Cavity ; microbiology ; Dentin ; microbiology ; Drug Combinations ; Enterococcus faecalis ; ultrastructure ; Humans ; Methylmethacrylates ; therapeutic use ; Microscopy, Electron, Scanning ; Oxides ; therapeutic use ; Root Canal Filling Materials ; therapeutic use ; Root Canal Obturation ; methods ; Root Canal Preparation ; instrumentation ; Silicates ; therapeutic use ; Time Factors ; Zinc Oxide-Eugenol Cement ; therapeutic use

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Sphere-forming-like cells (squamospheres) with cancer stem-like cell traits from VX2 rabbit buccal squamous cell carcinoma.

Yuk-Kwan CHEN ; Anderson Hsien-Cheng HUANG ; Li-Min LIN

International Journal of Oral Science.2014;6(4):212-218. doi:10.1038/ijos.2014.32

Previous studies have demonstrated that spheroid type cells grown under suspension culture conditions have cancer stem cell (CSC) traits in a number of cancers, but this phenomenon has not yet been reported in the VX2 rabbit oral cancer model. Hence, this study aimed to study the spheroid cells from VX2 rabbit buccal squamous cell carcinomas (SCCs) and assess their CSC characteristics. Five adult male New Zealand white outbred rabbits were used to generate VX2 rabbit buccal SCC. Sphere-forming cell culture was performed for the VX2 rabbit buccal SCC specimens. The self-renewal capability; cluster of designation (CD) 44, CD133, acetaldehyde dehydrogenase 1 (ALDH1), B cell-specific Moloney murine leukemia virus integration site 1 (Bmi-1), Nestin, octamer-binding transcription factor 4 (Oct4) and reduced expression protein-1 (Rex-1) expression with reverse transcription-polymerase chain reaction (RT-PCR); chemoresistance to cisplatin and 5-fluorouracil; and in vivo tumorigenicity of spheroid cell transplantation in nude mice were evaluated to determine the CSC characteristics of the resulting spheroid cells. We successfully obtained spheroid cells from the VX2 rabbit OSCC tissues. The spheroid cells exhibited CSC traits, including the expression of CSC and stem cell markers (CD44, Bmi-1, Nestin, Oct4 and Rex-1), capacity to generate new spheroid colonies within 1 week of reseeding from single-dissociated spheroid cells, chemoresistance capacity and generation of tumour xenografts (with histological features resembling those of the original VX2 rabbit buccal SCC) from the transplantation of 10(3) undifferentiated spheroid cells into nude mice. In summary, we demonstrated that spheroid cells with CSC cell traits can be derived from VX2 rabbit buccal SCCs, indicating that this animal cancer model is applicable for studying CSCs in human oral cancers.
AC133 Antigen ; Animals ; Antigens, CD ; analysis ; Antineoplastic Agents ; pharmacology ; Carcinoma, Squamous Cell ; pathology ; Cell Culture Techniques ; Cisplatin ; pharmacology ; DNA-Binding Proteins ; analysis ; Disease Models, Animal ; Drug Resistance, Neoplasm ; Fluorouracil ; pharmacology ; Glycoproteins ; analysis ; Heterografts ; transplantation ; Hyaluronan Receptors ; analysis ; Isoenzymes ; analysis ; Male ; Mice ; Mice, Nude ; Mouth Neoplasms ; pathology ; Neoplasm Transplantation ; Neoplastic Stem Cells ; classification ; Nestin ; analysis ; Octamer Transcription Factor-3 ; analysis ; Peptides ; analysis ; Polycomb Repressive Complex 1 ; analysis ; Proto-Oncogene Proteins ; analysis ; Rabbits ; Retinal Dehydrogenase ; analysis ; Spheroids, Cellular ; classification

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