Ghost cells unveiled: A comprehensive review.

BACKGROUND: Ghost cells (GCs) are cells with distinct intracytoplasmic keratinization, which leads to the preservation of the cellular outline with a clear area corresponding to the previous nucleus location. GCs may show various patterns, such as degeneration, tissue granulation, and calcification. Their true nature and the mechanism regulating the conversion of odontogenic epithelial cells into GCs remain unclear. GC keratinization is different from normal keratinization as they are larger than keratotic squames, are frequently vacuolated, and have prominent nuclear membrane remnants. Few cystic lesions, odontogenic tumors, and non-odontogenic tumors, such as calcifying odontogenic cyst, craniopharyngioma, pilomatrixoma, odontoma, dentinogenic ghost cell tumor, and ghost cell odontogenic carcinoma, exhibit GCs as a typical feature. The Wnt and Notch signaling pathways play a role in the histogenesis of the neoplasms. HIGHLIGHT: The review clarifies the various proposed hypotheses of the histogenesis of GCs, including molecular pathogenesis. Diagnostic workup for the identification of GCs, including special staining and immunohistochemistry, has been extensively discussed. A stepwise algorithm for identifying odontogenic and non-odontogenic lesions containing GCs has been proposed. Additionally, the prognostic role of GCs in the lesions has been elucidated. CONCLUSION: Among the various hypotheses of the origin of GCs, we suggest that aberrant keratinization is the most accepted based on various immunohistochemical studies and special staining characteristics. GCs are a distinct characteristic entity of many odontogenic and non-odontogenic lesions; however, it remains controversial whether their presence has any pathognomonic role in the biological nature of these lesions.

Role of hypoxia-related proteins in invasion of ameloblastoma cells: crosstalk between NOTCH1, hypoxia-inducible factor 1α, a disintegrin and metalloproteinase 12, and heparin-binding epidermal growth factor.

AIMS: Ameloblastoma AME is a benign tumour characterized by local invasiveness, high recurrence rates, and diverse histological patterns. The oxygen concentration is reduced in specific areas of the tumour microenvironment, which leads to intratumoral hypoxia. Crosstalk between NOTCH1, a disintegrin and metalloproteinase 12 (ADAM-12), hypoxia-inducible factor 1α (HIF-1α) and heparin-binding epidermal growth factor (HB-EGF) under hypoxic conditions has been implicated in invadopodia formation, tumour invasiveness, and metastasis development. The aim of this study was to analyse the expression of these proteins, in order to further elucidate the mechanisms underlying AME invasiveness. METHODS AND RESULTS: Twenty cases of AME, eight calcifying cystic odontogenic tumours CCOTs and 10 samples of dental follicle were used to investigate the expression of these proteins by immunohistochemistry with the primary antibodies anti-NOTCH1, anti-ADAM-12, anti-HIF-1α, and anti-HB-EGF. Immunostaining results were expressed as the percentage of stained area in images acquired in an AxioScope microscope equipped with an AxioCamHRc camera and a × 40 objective. The results showed that immunoexpression of all proteins was higher in the AME samples than in the CCOT and dental follicle samples (P < 0.05). CONCLUSIONS: AME showed an increased presence of proteins associated with tumour invasiveness, which indicates a possible role of these proteins in the biological behaviour of this tumour.

Ghost cells in pilomatrixoma, craniopharyngioma, and calcifying cystic odontogenic tumor: histological, immunohistochemical, and ultrastructural study.

BACKGROUND: Pilomatrixoma, craniopharyngioma, and calcifying cystic odontogenic tumor are the main entities presenting ghost cells as an important histological feature, in spite their quite different clinical presentation; it seems that they share a common pathway in the formation of these cells. The aim of this study is to examine and compare the characteristics of ghost and other cells that form these lesions. METHODS: Forty-three cases including 21 pilomatrixomas, 14 craniopharyngiomas, and eight calcifying cystic odontogenic tumors were evaluated by immunohistochemistry for cytokeratins, CD138, ß-catenin, D2-40, Glut-1, FAS, CD10 and also by scanning electron microscopy. RESULTS: The CKs, CD138, ß-catenin, Glut-1, FAS, and CD10 were more often expressed by transitional cells of craniopharyngioma and calcifying cystic odontogenic tumor, compared with pilomatrixoma. Basaloid cells of pilomatrixoma showed strong positivity for CD138 and CD10. Differences on expression pattern were identified in transitional and basal cells, as ghost cells were negative for most antibodies used, except by low expression for cytokeratins. By scanning electron microscopy, the morphology of ghost cells were similar in their fibrillar cytoplasm, but their pattern varied from sheets in pilomatrixoma to small clusters in craniopharyngioma and calcifying cystic odontogenic tumor. CONCLUSIONS: Mechanisms involved in formation of ghost cells are unknown, but probably they follow different pathways as protein expression in the basal/transitional cells was not uniform in the three tumors studied.

Clinicopathologic analysis and syndecan-1 and Ki-67 expression in calcifying cystic odontogenic tumors, dentinogenic ghost cell tumor, and ghost cell odontogenic carcinoma.

OBJECTIVE: Benign and malignant tumor cells can express altered adhesion properties, and these features can be associated with their proliferative and invasive characteristics. This study aimed to evaluate syndecan-1 and Ki-67 expression in ghost cell-containing odontogenic tumors. STUDY DESIGN: Clinical data were retrieved from laboratory records, and hematoxylin-eosin-stained slides and sections, labeled with monoclonal antibodies anti-syndecan-1 and anti-Ki-67 using the immunoperoxidase technique, were evaluated. RESULTS: Included were 21 central calcifying cystic odontogenic tumors (CCOTs) (4 associated with odontoma), 2 peripheral CCOTs, 1 dentinogenic ghost cell tumor, and 1 ghost cell odontogenic carcinoma (GCOC). Syndecan-1 was mainly expressed in cells resembling stellate reticulum and in stromal cells from the fibrous capsule. The mean Ki-67 labeling index was 4.1% (49.3% for GCOC), but it was not associated with syndecan-1 expression. CONCLUSIONS: Syndecan-1 is variably expressed in cells resembling the stellate reticulum, stromal cells, and basal cells and might be associated with the biology of these tumors.

Detection of cytokeratins in ghost cells of calcifying cystic odontogenic tumor indicates an altered keratinization and hair follicle differentiation for their development.

Calcifying cystic odontogenic tumors (CCOTs) are benign cystic lesions of odontogenic origin characterized by an ameloblastoma-like epithelium and the presence of a group of cells named ghost cells. The pattern of cytokeratin (Ck) expression on these lesions remains unclear and needs to be clarified. To this end, the expression of Ck6, Ck13, Ck14, Ck18, and Ck19 in the epithelium lining of 7 cases of CCOTs was evaluated by immunohistochemistry. For this, the epithelium lining was divided into 3 distinct regions: basal layer, suprabasal layer, and the compartment composed of ghost cells. In this study, 6 cases (85.7%) were classified as type 1 and 1 (14.3%) as type 4. All cases were negative for Ck13 and Ck18, despite the epithelial layer, as well as in the ghost cells. Ck6 was only positive in the ghost cells. Positivity for Ck14 and Ck19 was found in the basal and suprabasal layers, including the ghost cells. The results showing positivity for Ck14 and Ck19 in all of the analyzed cases reinforce CCOT as being of odontogenic origin, and the restricted expression of Ck6 in the ghost cells may be indicative that these cells suffer an altered differentiation into hair follicles in CCOTs.

Matrix metalloproteinases, tissue inhibitors of metalloproteinases, and growth factors regulate the aggressiveness and proliferative activity of keratocystic odontogenic tumors.

OBJECTIVE: The objective of this preliminary study was to evaluate the expression of matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs) and growth factors in keratocystic odontogenic tumors (KOTs). STUDY DESIGN: The expression of MMPs, TIMPs, growth factors, and the extracellular signal-regulated kinase (ERK) 1/2 signaling pathway were assessed by immunohistochemistry in 15 cases of KOT and 4 cases of calcifying cystic odontogenic tumor (CCOT). RESULTS: KOT samples expressed significantly higher amounts of MMPs, TIMPs, growth factors, epidermal growth factor receptor (EGFR), and ERK compared with CCOT samples, with the exception of MMP-2 and TIMP-1. CONCLUSIONS: MMP-9, TIMP-2, EGF and transforming growth factor α act together and likely regulate the proliferation and aggressiveness of KOT. ERK-1/2 serves as the transducer of signals generated by these proteins, which signal through the common receptor, EGFR. This process may be related to the increased proliferation and aggressiveness observed in KOT.

Evaluation of osteopontin and CD44v6 expression in odontogenic cystic lesions by immunohistochemistry.

Odontogenic cysts are common lesions with different biological behavior. Odontogenic keratocysts (OKCs) and calcifying odontogenic cysts (COCs) with ameloblastoma-like epithelium are more aggressive than dentigerous cysts (DCs) and radicular cysts (RCs). Therefore, they were included in the list of odontogenic tumors by WHO. Osteopontin (OPN) is a calcium-binding glycoprotein present in many normal tissues. It plays a role in the migration and invasion of transformed epithelial cells. Binding of OPN to its receptor CD44v6 can enhance cell motility and migration. The purpose of this study was to compare the expression of these markers between odontogenic cysts of varying biological behavior. We examined OPN and CD44v6 expression in tissue sections of 14OKCs, 14COCs, 14RCs and 14DCs by immunohistochemistry. OPN and CD44v6 immunostaining was observed in all lining epithelial cells of the studied lesions with different degrees. The highest level of OPN and CD44v6 expression was found in OKCs, followed by COCs, RCs and DCs. Comparison of both markers among four groups revealed significant differences (P<0.001). Our findings suggest that higher level of OPN and CD44v6 expression in epithelial cells of some lesions such as OKC and COC can explain the local aggressive behavior of them.

Melan-A/Mart-1- or HMB-45-positive melanocytes are not present in calcifying cystic odontogenic tumors (calcifying odontogenic cysts): a study in 13 Caucasian patients.

Melanin pigment and melanocytes may be found in odontogenic cysts and tumors, particularly calcifying cystic odontogenic tumor (CCOT). In the present study we investigated the immunohistochemical expression of the Melan-A/Mart-1 and HMB-45 antigens in 13 Caucasians patients with CCOT. Melan-A/Mart-1- and HMB-45-positive melanocytes were not seen in any of the cases. Our findings are in agreement with the assumption that pigmentation in odontogenic lesions may be a racial phenomenon.

Notch signaling and ghost cell fate in the calcifying cystic odontogenic tumor.

Notch signaling is an evolutionarily conserved mechanism that enables adjacent cells to adopt different fates. Ghost cells (GCs) are anucleate cells with homogeneous pale eosinophilic cytoplasm and very pale to clear central areas (previous nucleus sites). Although GCs are present in a variety of odontogenic lesions notably the calcifying cystic odontogenic tumor (CCOT), their nature and process of formation remains elusive. The aim of this study was to investigate the role of Notch signaling in the cell fate specification of GCs in CCOT. Immunohistochemical staining for four Notch receptors (Notch1, Notch2, Notch3 and Notch4) and three ligands (Jagged1, Jagged2 and Delta1) was performed on archival tissues of five CCOT cases. Level of positivity was quantified as negative (0), mild (+), moderate (2+) and strong (3+). Results revealed that GCs demonstrated overexpression for Notch1 and Jagged1 suggesting that Notch1-Jagged1 signaling might serve as the main transduction mechanism in cell fate decision for GCs in CCOT. Protein localizations were largely membranous and/or cytoplasmic. Mineralized GCs also stained positive implicating that the calcification process might be associated with upregulation of these molecules. The other Notch receptors and ligands were weak to absent in GCs and tumoral epithelium. Stromal endothelium and fibroblasts were stained variably positive.

[Expression of p-p38MARK, uPA and Ki-67 in epithelial odontogenic tumour].

OBJECTIVE: to examine the expression of phosphorylated p38 mitogen-activated protein kinase (p-p38MAPK), urokinase plasminogen activator (uPA) and Ki-67. METHODS: the specimens of ameloblastoma (AB), keratocystic odontogenic tumour (KCOT), calcifying epithelial odontogenic tumor (CEOT), adenomatoid odontogenic tumour (AOT), calcifying cystic odontogenic tumour (CCOT) and 5 tooth germs were examined immunohistochemically for the expression of p-p38MAPK, uPA and Ki-67. RESULTS: p-p38MAPK was detected both in the cytoplasm and the nucleus of the epithelial odontogenic tumour cells and uPA in the cytoplasm of the epithelial odontogenic tumour cells. Among the 65 cases, there were 17 (26%), 51 (78%) and 62 cases (95%) of positive expression of p-p38MAPK, uPA and Ki-67 protein respectively. Furthermore, there was a statistically significant difference in the expression of p-p38MAPK, uPA and Ki-67 between epithelial odontogenic tumor group and tooth germ group (P < 0.05). There were significant correlations among the expression of p-p38MAPK, uPA and Ki-67 (P < 0.05). CONCLUSIONS: the p38MAPK-signaling pathway could promote tumour growth and invasion in epithelial odontogenic tumour by up-regulating uPA protein expression and may play a role in oncogenesis, invasion and proliferation of epithelial odontogenic tumour.

Matrix metalloproteinases, TIMPs and growth factors regulating ameloblastoma behaviour.

AIMS: Ameloblastoma is an odontogenic neoplasm with local invasiveness and recurrence. We have previously suggested that growth factors and matrix metalloproteinases (MMPs) influence ameloblastoma invasiveness. The aim was to study expression of MMPs, tissue inhibitor of metalloproteinases (TIMPs) and growth factors in ameloblastoma. METHODS AND RESULTS: Thirteen cases of solid/multicystic ameloblastoma were examined. As a control, calcifying cystic odontogenic tumour (CCOT), a non-invasive odontogenic neoplasm with ameloblastomatous epithelium was also studied. Immunohistochemistry detected MMPs, TIMPs and growth factors in ameloblastoma and CCOT. The labelling index (LI) of MMP-9 and TIMP-2 was significantly higher in ameloblastoma compared with CCOT. The LI of epidermal growth factor (EGF), transforming growth factor (TGF)-alpha and epidermal growth factor receptor (EGFR) was also increased in ameloblastoma. This neoplasm showed greater expression of MMPs, TIMPs and growth factors compared with CCOT. We then analysed these molecules in ameloblastoma cells and stroma. Ameloblastoma cells exhibited increased LI of MMP-1, -2 and EGFR. We found a positive correlation between EGF and TIMP-1, and between TGF-alpha and TIMP-2. It is known that signals generated by growth factors are transduced by the ERK pathway. Ameloblastoma stroma exhibited the phosphorylated (activated) form of ERK. CONCLUSIONS: These results suggest an interplay involving growth factors MMPs and TIMPs that may contribute to ameloblastoma behaviour. Signals generated by this molecular network would be transduced by ERK 1/2 pathway.

Immunohistochemical analysis of p53 and PCNA expression in calcifying odontogenic cyst.

Calcifying odontogenic cyst (COC) is a developmental odontogenic cyst in the jaw. Because of its diverse histopathologic features and biological behavior, there has long been confusion with regard to its nature as a cyst or neoplasm. This study evaluated the proliferative activity of 57 COC samples, including simple cyst (10 cases), cystic neoplasm (34 cases), solid neoplasm (6 cases) and combined lesion (7 cases) by p53 and PCNA immunohistochemical staining. For assessment of p53 and PCNA positivity, the number of positively stained cells with brown-stained nuclei was counted in 1000 cells from each sample. p53 and PCNA expression in the solid neoplasm subtype were significantly higher when compared to cystic neoplasm and simple cyst (P < 0.05). The lowest p53 and PCNA expression was found in the simple cyst subtype. p53 and PCNA expression in the basal and suprabasal layers was significantly higher in the solid subtype when compared to others, and the difference between COC groups was significant. The results demonstrated that within benign types of COC, the amount of p53 and PCNA in proliferative epithelium is significantly higher when compared to non-proliferative epithelium. p53 and PCNA markers are possible parameters for differentiation of COC subtypes.

Methylation frequencies of cell-cycle associated genes in epithelial odontogenic tumours.

OBJECTIVE: The benign epithelial odontogenic tumours constitute a group of lesions derived from epithelial elements of the tooth-forming apparatus. This group includes lesions of different biological behaviour, such as ameloblastoma, calcifying cystic odontogenic tumour (CCOT) and adenomatoid odontogenic tumour (AOT). The pathogenesis of these neoplasms remains uncertain and the occurrence of methylation in cell-cycle related genes may be involved in their development. The aim of this study was to investigate the methylation status of P16, P21, P27, P53 and RB1 genes in epithelial odontogenic tumours. DESIGN: Methylation-specific polymerase chain reaction (MSP) was used to evaluate the presence of methylation in 13 samples of ameloblastoma, six samples of CCOT, three samples of AOT and 14 samples of dental follicles, included as control. RESULTS: Our results showed a distinct methylation profile in each group. In ameloblastoma, the highest methylated genes were P16 and P21, while in CCOT the P21 and RB1 genes were the most commonly methylated genes. Only the P16 and P21 genes were methylated in the AOT samples. In the dental follicle samples, P16, P27 and RB1 genes were commonly methylated. A high percentage of the odontogenic tumours analysed showed methylation of the P21 gene, in contrast to dental follicles. CONCLUSIONS: Epithelial odontogenic tumours show a distinct methylation profile in cell-cycle associated genes. In addition to this, the current findings show that epigenetic alterations are common events in epithelial odontogenic tumours.

Immunoexpression of keratins in the calcifying cystic odontogenic tumor epithelium.

Ameloblastomatous epithelium containing clusters of ghost cells is the typical histopathology of calcifying cystic odontogenic tumor (CCOT). This paper aimed to assess keratins AE1-AE3, K7, K10/13, K14, K18, K19, vimentin, laminin, and collagen IV in 08 CCOTs to discuss their histopathogenesis. Similarity to the immunoprofile of the stratified squamous epithelium was seen in the with the basal layer expressing K14 and the upper cells expressing K10/13. When compared to the immunoprofile of the normal odontogenic epithelium, of odontogenic tumor epithelia and of the ghost cells described in the literature, it was possible to suggest that the CCOT epithelium differentiates towards squamous type.

The expression of NF-kappaB, Ki-67 and MMP-9 in CCOT, DGCT and GCOC.

Calcifying odontogenic cysts (COCs) represent a group of rare odontogenic lesions with a diversity of clinicopathological and behavioral features. According to the WHO classification of head and neck tumors in 2005, COC has been divided into calcifying cystic odontogenic tumor (CCOT), dentinogenic ghost cell tumor (DGCT) and ghost cell odontogenic carcinoma (GCOC). With few reports available on its immunohistochemical profile, this study investigated the histopathological features and the expression of nuclear factor kappaB (NF-kappaB), Ki-67 and matrix metalloproteinase-9 (MMP-9) in CCOT, DGCT and GCOC. According to the WHO classification of head and neck tumors in 2005, 26 cases of the so-called COC were diagnosed as CCOT (n=14), DGCT (n=7) and GCOC (n=5), respectively. The specimens of 26 COCs and 10 classic ameloblastomas (as control group) were examined by immunohistochemistry using anti-NF-kappaB p65, anti-Ki-67 and anti-MMP-9 antibodies and by in situ hybridization(ISH)using anti-MMP-9 mRNA probes. Immunohistochemical reactivity for NF-kappaB was mainly detected in the cytoplasm of tumor cells, and nuclear reactivity was only seen in few tumor cells in COC and classic ameloblastomas. Rate of nuclear staining was less than 1%. The expression of Ki-67 in GCOC was significantly higher than those in CCOT (p<0.001), DGCT and ameloblastoma (p<0.005). In COCs and ameloblastomas, expression of MMP-9 mRNA and protein was detected in tumor cells as well as in stromal cells. The positive staining for MMP-9 protein was detected in stromal cells of all GCOC cases and was significantly stronger than those in CCOT and DGCT groups (p<0.05). NF-kappaB may minimally affect the progression and local invasiveness of CCOT, DGCT and GCOC. GCOC show significantly higher proliferative activity than CCOT and DGCT. MMP-9 in stroma is associated with invasive ability of the CCOT, DGCT and GCOC.

Beta-catenin gene alterations in a variety of so-called calcifying odontogenic cysts.

Calcifying odontogenic cysts (COCs) evidence a variety of histopathological features, in addition to biological behaviors. Several systems have been proposed for the classification of COCs. However, the molecular changes underlying their development remain largely unknown. In an effort to elucidate the participation of beta-catenin gene mutation in COC development, we evaluated the genetic alterations and expression of beta-catenin in different COC subtypes with archival paraffin blocks. beta-catenin gene mutations were detected in all subtypes. Immunohistochemically, all cases of COCs evidenced cytoplasmic and nuclear beta-catenin expression. Beta-catenin gene mutations and beta-catenin overexpression have been identified as characteristics of COCs. Therefore, the constitutive activation of Tcf/Lef-dependent transcription appears to be intimately involved in their development. These findings indicate that aberrations of the Wnt signaling pathway may perform a crucial role in the development and differentiation status of the odontogenic epithelium in COCs via the deregulation of cell proliferation.

Calcifying odontogenic cyst associated with an orthokeratinized odontogenic cyst.

Odontogenic tumors composed of two or more distinct types of lesions are unusual. In this paper, a case of an odontogenic lesion characterized by simultaneous occurrence of areas of calcifying odontogenic cyst (COC) and orthokeratinized odontogenic cyst (OOC) is described. The lesion was asymptomatic and presented at the radiographic examination as a unilocular well-delimited radiolucency extending from left incisor to right premolar area in the mandible. To date, this is the first report of COC associated with an OOC.

Calcifying epithelial odontogenic tumor: an immunohistochemical case study.

Calcifying epithelial odontogenic tumor (CEOT) is a rare benign odontogenic tumor. A case of CEOT in a 25-year-old female is presented here. Histologically, the case showed sheets of polyhedral epithelial cells with deep eosinophilic cytoplasm and prominent nuclei. Nuclear pleomorphism and hyperchromatism were evident. Globules of amyloid-like material among the tumor cells were prominent. Also found was a small area demonstrating a cribriform pattern. Immunohistochemical studies with antibodies against basement membrane proteins (laminins 1 and 5, collagen type IV and fibronectin), pan-cytokeratins AE1/AE3, vimentin, S-100 protein and CD 1a were performed. Tumor cells expressed laminins 1 and 5, fibronectin, cytokeratins and vimentin. The amyloid-like material was not reactive to all antibodies examined. A number of dendritic cells among sheets of tumor cells were revealed with strong staining for S-100 protein and CD 1a. These dendritic cells are likely to be Langerhans cells. Hence, immunohistochemistry is a useful method to study the variant of CEOT.

[Expression of HOX C13 in odontogenic tumors].

PURPOSE: To study the expression of HOXC13 mRNA in odontogenic tumors. METHODS: HOXC13 mRNA was detected in 47 cases of ameloblastoma (AB), 3 cases of calcifying cystic odontogenic tumor (CCOT), 3 cases of ameloblastic fibroma (AF), 10 cases of keratocystic odontogenic tumor (KCOT) and 2 cases of calcifying epithelial odontogenic tumor (CEOT) by in situ hybridization, and 7 cases of normal oral mucosa were selected as control. SPSS10.0 software package was used for chi(2) test. RESULTS: HOXC13 mRNA was positively expressed in all odontogenic epithelium except AF. The positive ratios were 97.9% in AB, 100% in CCOT, 100% in CEOT, 70.0% in KCOT epithelium and 42.9% in the normal oral mucosa. There was significant difference among AB, KCOT and normal mucosa (chi(2)=21.665, P<0.001). CONCLUSIONS: The genesis and development of odontogenic tumors are related to the high expression of HOXC13. The expression of HOXC13 mRNA in the odontogenic lesions has heterogeneity.

[Expression of nuclear factor-kappaB, Ki-67 and matrix metalloproteinase-9 in calcifying odontogenic cyst].

OBJECTIVE: To evaluate the expression of nuclear factor-kappaB (NF-kappaB), Ki-67 and matrix metalloproteinase-9 (MMP-9) in calcifying odontogenic cyst (COC), in order to investigate the proliferation and invasion of COC. METHODS: Twenty-six cases of COC were classified into calcifying cystic odontogenic tumor (CCOT), dentinogenic ghost cell tumor (DGCT) and ghost cell odontogenic carcinoma (GCOC) based on the WHO classification of odontogenic tumors in 2005. The specimens of COC and 10 classic ameloblastoma (AB) were examined immunohistochemically to determine the expression of NF-kappaB p65, Ki-67 and MMP-9. RESULTS: NF-kappaB was mainly detected in the cytoplasm of most tumor cells, but was only detected in the nucleus of few tumor cells (rate of nuclear staining < 1%). The expression of Ki-67 was significantly higher in GCOC than in CCOT (P < 0.001), DGCT (P < 0.05) and AB (P < 0.005). MMP-9 was detected both in tumor cells and stromal cells. GCOC showed significantly higher percentage of MMP-9 positive cases in stromal cells than CCOT, DGCT and AB (P < 0.05). CONCLUSIONS: NF-kappaB may minimally affect the progression and invasion of COC. GCOC shows significantly higher proliferative activity and aggressiveness than CCOT and DGCT. MMP-9 in stroma may play a key role in the invasion of GCOC.