Immunohistochemical analysis of pulmonary and pleural neoplasms with monoclonal antibodies B72.3 and CSLEX-1

Sequential paraffin sections of 222 epithelial lung tumors comprising all common histologic types, and 31 pleural mesotheliomas of all variants were immunostained with monoclonal antibodies (Mabs) B72.3 and CSLEX-1. Reactivity with Mabs B72.3 and CSLEX-1 respectively was noted in 7/57 and 4/57 squamous carcinomas, in 44/70 and 60/70 adenocarcinomas, 9/16 and 11/16 bronchioloalveolar carcinomas, 8/25 and 14/25 large cell undifferentiated carcinomas, 3/3 and 3/3 adenosquamous carcinomas, 0/11 and 0/11 carcinoids, 0/10 and 2/10 well differentiated neuroendocrine (NE) carcinomas, 4/13 and 5/13 intermediate cell NE carcinomas, 0/17 and 0/17 small cell NE carcinomas, and 0/31 and 1/31 mesotheliomas. In most instances, both Mabs stained the same tumors; however, reactivity with CSLEX-1 was more intense and extensive, and involved more cases. Therefore, regardless of conventional histologic type, staining with Mabs B72.3 and CSLEX-1 defines 4 subsets of lung tumors: one expressing both antigens, two expressing one but not the other, and one expressing neither. The possible biological and/or clinical significance of these subsets remains undetermined. When correlated with conventional histologic tumor types, our findings indicate: 1). both of these Mabs recognize most but not all adenocarcinomas and bronchioloalveolar carcinomas, and since CSLEX-1 stained more cases than B72.3, it may be argued that the former is a broader exocrine phenotype marker than the latter; 2). both of these Mabs select exocrine subsets of large cell undifferentiated carcinomas; 3). both of these Mabs stain exocrine cell subpopulations in well differentiated and intermediate cell NE carcinomas but not in carcinoids or small cell NE carcinomas, and 4). except for rare cases, neither B72.3 nor CSLEX-1 reacts with mesotheliomas regardless of variant.(ABSTRACT TRUNCATED AT 250 WORDS)     

The establishment of two rat colonic carcinomas in tissue culture. A basic prerequisite for standardized experiments on the biology of colonic carcinomas in vivo

Two rat colonic carcinomas (DMH-Co-1 and DMH-Co-2) derived from dimethyl-hydrazine-induced metastasizing adenocarcinomas were established as permanent cell lines. By means of electron microscopy, immunofluorescence microscopy and biochemical analysis of cytoskeletal components, it has been shown that both tumor cell lines retain in vitro the phenotypic characteristics of the primary tumors. The in vitro growth properties revealed only minor differences between the two cell lines. After retransplantation in vivo, DMH-Co-2 gave rise to moderately differentiated adenocarcinomas, whereas the tumors arising from DMH-Co-1 exhibited a continuum of differentiation encompassing adenocarcinomas, undifferentiated carcinomas and squamous cell carcinomas. These permanent cell lines offer the opportunity for isolating divergent subpopulations by in vitro cloning and facilitate standardized experiments on their biological behaviour in vivo.     

Fine needle aspiration cytology of atypical carcinoid of the lung

The cytologic features of eight atypical carcinoid tumors of the lung, as observed in fine needle aspiration (FNA) specimens, are described in detail. They were compared with 21 pulmonary squamous-cell carcinomas, 16 adenocarcinomas, 5 small-cell undifferentiated carcinomas, 3 large-cell undifferentiated carcinomas and 1 typical carcinoid tumor. Atypical carcinoid tumor was easily distinguished from the other pulmonary neoplasms in most instances. Only two poorly differentiated squamous-cell carcinomas (one of which had atypical carcinoid as a component) and one small-cell undifferentiated carcinoma had similar cytologic features. One atypical carcinoid also had cytologic features similar to small-cell undifferentiated carcinoma. Because atypical carcinoid and small-cell undifferentiated carcinoma, at times, may be difficult to separate in FNA specimens, surgical resection of all stage I neoplasms with cytologic features evocative of either neoplasm is recommended.     

Analysis of differentiation antigens on normal and carcinogen-altered rat epithelial cells in vivo and in culture

Two murine monoclonal antibodies, 3BG8 and 9BG8, which were raised against a rat tracheal squamous-cell-carcinoma cell line, recognize cell-surface antigens on normal rat squamous epithelium (skin, esophagus, vagina, and cornea) as well as on carcinogen-exposed, immortalized, rat tracheal epithelial cells. Monoclonal antibody 3BG8 binds to a 115-kilodalton cell-surface protein on undifferentiated basal cells of the epithelium, while the binding of the other antibody, 9BG8, occurs in both differentiated and undifferentiated populations of normal squamous epithelium and squamous cell carcinomas. Undifferentiated tracheal carcinomas bound only the 3BG8 antibody. No binding of either antibody was detected on normal tracheal mucociliary epithelium. Only under conditions that induce squamous differentiation of rat tracheal epithelium was binding of 3BG8 and 9BG8 detected. For reasons which are not clear at present, 9BG8 dramatically inhibits the growth of normal tracheal and esophageal cells in primary culture, whereas only 3BG8 affects the growth of carcinogen-altered tracheal cell lines. Based on antigen characterization and distribution, it is concluded that the 3BG8 and 9BG8 epitopes are localized on differentiation antigens which differ from others that have been previously described.     

Genetic and epigenetic alterations of tumor suppressor and tumor-related genes in gastric cancer

Both genetic and epigenetic alterations of tumor suppressor and tumor-related genes involved in the pathogenesis of gastric cancer are reviewed here, and molecular pathways of gastric carcinogenesis are proposed. Gastric carcinomas are believed to evolve from native gastric mucosa or intestinal metaplastic mucosa that undergoes genetic and epigenetic alterations involving either the suppressor pathway (defects in tumor suppressor genes) or mutator pathway (defects in DNA mismatch repair genes). Methylation of E-cadherin in native gastric mucosa results in undifferentiated carcinomas (suppressor pathway), while methylation of hMLHI results in differentiated foveolar-type carcinomas (mutator pathway). The majority of differentiated gastric carcinomas however, arise from intestinal metaplastic mucosa and exhibit structural alterations of tumor suppressor genes, especially p53. They appear to be related to chronic injury, perhaps due to Helicobacter pylori infection. Approximately 20% of differentiated carcinomas (ordinary-type) have evidence of mutator pathway tumorigenesis. Mutations of E-cadherin are mainly involved in the progression of differentiated carcinomas to undifferentiated tumors. The molecular pathways of gastric carcinogenesis depend on the histological background, and gastric carcinomas show distinct biological behaviors as a result of discernible cellular genetic and epigenetic alterations.     

Comparison of the intratumor DNA ploidy distribution pattern between differentiated and undifferentiated gastric carcinoma

OBJECTIVE: To compare the differences in intratumor DNA ploidy distribution pattern between differentiated and undifferentiated carcinoma of the stomach. STUDY DESIGN: Two hundred five cases of surgically resected gastric carcinoma were studied, comprising 125 differentiated and 80 undifferentiated carcinomas. DNA ploidy was determined by means of flow cytometry in stepwise sections of the entire tumor. The intratumor DNA ploidy distribution pattern was compared between these two histologic types. RESULTS: Differentiated carcinomas comprised 35 cases of predominantly diploid (28%) and 90 cases of predominantly aneuploid (72%) tumors, while undifferentiated carcinomas comprised 46 cases of predominantly diploid (57.5%) and 34 cases of predominantly aneuploid (42.5%) (P < .01). The frequency of cases showing predominantly aneuploidy among the differentiated carcinomas was significantly higher than among the undifferentiated carcinomas at stages IA and IIIA. The rate of cases showing heterogeneity was lower among the early-stage cases than among the advanced-stage cases in both histologic types. Among the differentiated carcinomas, there were 22 cases that had more than six DNA stemlines in each tumor, whereas among undifferentiated carcinomas, there were six such cases (P < .05). CONCLUSION: There were more cases with high malignant potential among the differentiated carcinoma cases than among the undifferentiated carcinoma cases in the present series.     

Casein phosphopeptides modulate proliferation and apoptosis in HT-29 cell line through their interaction with voltage-operated L-type calcium channels

At the intestinal level, proliferation and apoptosis are modulated by the extracellular calcium concentration; thus, dietary calcium may exert a chemoprotective role on normal differentiated intestinal cells, while it may behave as a carcinogenesis promoter in transformed cells. Calcium in milk is associated with casein and casein phosphopeptides (CPPs), hence is preserved from precipitation. CPPs were demonstrated to induce uptake of extracellular calcium ions by in vitro intestinal tumor HT-29 cells but only upon differentiation. Here, the hypothesis that CPPs could differently affect proliferation and apoptosis in undifferentiated and differentiated HT-29 cells through their binding with calcium ions was investigated. Results showed that CPPs protect differentiated intestinal cells from calcium overload toxicity and prevent their apoptosis favoring proliferation while inducing apoptosis in undifferentiated tumor cells. The CPP effect on undifferentiated HT-29 cells, similar to that exerted by ethyleneglycol-O, O'-bis(2-aminoethyl)-N, N, N', N'-tetraacetic acid (EGTA), is presumably due to the ability in binding the extracellular calcium. The effect on differentiated HT-29 cells is coupled to the interaction of CPPs with the voltage-operated L-type calcium channels, known to activate calcium entry into the cells under depolarization and to exert a mitogenic effect: the use of an agonist potentiates the cell response to CPPs, while the antagonists abolish the response to CPPs (36% of examined cells) or reduce both the percentage of responsive cells and the increase of intracellular calcium concentration. Taken together, these results confirm the potentialities of CPPs as nutraceuticals/functional food and also as modulators of cellular processes connected to the expression of a cancer phenotype.     

Terminally differentiated postmitotic tumor cells in a rat rhabdomyosarcoma cell line

A permanent rat rhabdomyosarcoma cell line (BA-HAN-1C) has been established, the phenotype of which is characterized by the coexistence of undifferentiated mononuclear cells and differentiated multinuclear myotube-like giant cells. The failure of attempts to separate these two cell types by repeated recloning procedures indicates their close histogenetic relationship and suggests that differentiation in this tumor proceeds in a similar manner to that in normal striated muscle where postmitotic myotubes arise from mononuclear myoblasts by fusion. The morphologically undifferentiated mononuclear tumor cells were shown to be actively proliferating and to incorporate thymidine methyl-3H(3H-TdR). The myotube-like giant cells neither incorporated 3H-TdR nor underwent mitosis or exhibited any clonogenic potential. After retransplantation into syngenic rats, tumor growth was markedly retarded when the tumor cell inoculum contained a high percentage of myotube-like giant cells. These data show that proliferative activity in this rhabdomyosarcoma cell line is confined to the mononuclear tumor cell compartment, the multinuclear myotube-like giant cells having withdrawn from the cell cycle and represent terminally differentiated postmitotic cells. This cell line should provide a valuable tool for further investigation of coherent aspects of proliferation and differentiation using various differentiation inducers.     

Breast cancer, a stem cell disease

Breast cancer is a first magnitude problem of public health worldwide. There is increasing evidence that this cancer is originated in and maintained by a small population of undifferentiated cells with self-renewal properties. This small population generates a more differentiated pool of cells which represents the main mass of the tumor, resembling the hierarchical tissue organization of the normal breast. These cancer stem cells seem to share a similar phenotype with their normal counterparts but they display dysfunctional patterns of proliferation and differentiation, and they no longer respond to normal physiological controls that ensure a balanced cellular turnover. The origin of these cancer stem cells is controversial; it is not well known if they are originated from normal stem cells or from more differentiated progenitors where a de novo stem cell program is activated by the oncogenic insult. Here we review the origin of breast cancer stem cells and their role in the pathogenesis of cancer development, together with their implications in breast cancer progression, treatment and prognosis.     

Oncogenic and anti-apoptotic activity of NF-kappa B in human thyroid carcinomas

Thyroid cancer includes three types of carcinomas classified as differentiated thyroid carcinomas (DTC), medullary thyroid carcinomas, and undifferentiated carcinomas (UTC). DTC and medullary thyroid carcinomas generally have a good prognosis, but UTC are usually fatal. Consequently, there is a need for new effective therapeutic modalities to improve the survival of UTC patients. Here we show that NF-kappa B is activated in human thyroid neoplasms, particularly in undifferentiated carcinomas. Thyroid cell lines, reproducing in vitro the different thyroid neoplasias, also show basal NF-kappa B activity and resistance to drug-induced apoptosis, which correlates with the level of NF-kappa B activation. Activation of NF-kappa B in the DTC cell line NPA renders these cells resistant to drug-induced apoptosis. Stable expression of a super-repressor form of I kappa B alpha (I kappa B alpha M) in the UTC cell line FRO results in enhanced sensitivity to drug-induced apoptosis, to the loss of the ability of these cells to form colonies in soft agar, and to induce tumor growth in nude mice. In addition, we show that FRO cells display a very low JNK activity that is restored in FRO-I kappa B alpha M clones. Moreover, inhibition of JNK activity renders FRO-I kappa B alpha M clones resistant to apoptosis induced by chemotherapeutic agents. Our results indicate that NF-kappa B plays a pivotal role in thyroid carcinogenesis, being required for tumor growth and for resistance to drug-induced apoptosis, the latter function very likely through the inhibition of JNK activity. Furthermore, the strong constitutive NF-kappa B activity in human anaplastic thyroid carcinomas, besides representing a novel diagnostic tool, makes NF-kappa B a target for the development of novel therapeutic strategies.     

Resveratrol affects undifferentiated and differentiated PC12 cells differently,particularly with respect to possible differences in mitochondrial and autophagic functions

Since resveratrol is considered to exert a unique dual effect, protective for normal cells but toxic to tumor cells, its action on undifferentiated (original) and differentiated PC12 cells was analyzed, because undifferentiated cells are tumorigenic and differentiated ones are neuronal in nature. Compared to resveratrol-untreated cells in both undifferentiated and differentiated cell groups, cells treated with different doses of resveratrol, at dosages of 1, 10 and 100 μM, showed the following alterations. Dying/dead cells were significantly increased in a dose-dependent manner in undifferentiated cells, but they were unchanged at doses of up to 10 μM resveratrol in differentiated cells. In living cells, neurites were short in undifferentiated cells, but drastically elongated with an increased number in differentiated cells. The expression of SIRT1 was drastically reduced in undifferentiated cells, but stable in differentiated cells. SIRT3 was significantly enhanced in a dose-dependent manner at resveratrol doses of up to 10 μM in both cells, with reduction and more enhanced at a dosage of 100 μM in undifferentiated and differentiated cells, respectively. Mitochondrial number and ATP synthase β subunit expression was unaltered at doses of up to 10 μM and were significantly reduced at doses of 100 μM in undifferentiated cells, but they were significantly increased in a dose-dependent manner, with a slight reduction in the ATP synthase at doses of 100 μM, in differentiated cells. In a dose-dependent manner, the number of autophagosomes and the LC3-II/LC3-I ratio were significantly less in undifferentiated cells and greater in differentiated cells. Also, in a dose-dependent manner, the expression of phosphorylated AMP-activated kinase (AMPK) was significantly less in undifferentiated cells and greater in differentiated cells. Resveratrol-induced AMPK suppression and activation, possibly through the modulation of SIRT protein activity, may thus be related to the inhibition and promotion of mitochondrial and autophagic functions, leading to cell death and survival in undifferentiated and differentiated cells, respectively.     

Diffusion-Weighted Imaging Using a Readout-Segmented,Multishot EPI Sequence at 3 T Distinguishes between Morphologically Differentiated and Undifferentiated Subtypes of Thyroid Carcinoma—A Preliminary Study

BACKGROUND: Thyroid carcinomas represent the most frequent endocrine malignancies. Recent studies were able to distinguish malignant from benign nodules of the thyroid gland with diffusion-weighted imaging (DWI). Although this differentiation is undoubtedly helpful, presurgical discrimination between well-differentiated and undifferentiated carcinomas would be crucial to define the optimal treatment algorithm. Therefore, the aim of this study was to investigate if readout-segmented multishot echo planar DWI is able to differentiate between differentiated and undifferentiated subtypes of thyroid carcinomas. PATIENTS AND METHODS: Fourteen patients with different types of thyroid carcinomas who received preoperative DWI were included in our study. In all lesions, apparent diffusion coefficient (ADC)min, ADCmean, ADCmax, and D were estimated on the basis of region of interest measurements after coregistration with T1-weighted, postcontrast images. All tumors were resected and analyzed histopathologically. Ki-67 index, p53 synthesis, cellularity, and total and average nucleic areas were estimated using ImageJ version 1.48. RESULTS: Analysis of variance revealed a statistically significant difference in ADCmean values between differentiated and undifferentiated thyroid carcinomas (P = .022). Spearman Rho calculation identified significant correlations between ADCmax and cell count (r = 0.541, P = .046) as well as between ADCmax and total nuclei area (r = 0.605, P = .022). CONCLUSION: DWI can distinguish between differentiated and undifferentiated thyroid carcinomas.     

Four Jointed Box 1 Promotes Angiogenesis and Is Associated with Poor Patient Survival in Colorectal Carcinoma

Angiogenesis, the recruitment and re-configuration of pre-existing vasculature, is essential for tumor growth and metastasis. Increased tumor vascularization often correlates with poor patient outcomes in a broad spectrum of carcinomas. We identified four jointed box 1 (FJX1) as a candidate regulator of tumor angiogenesis in colorectal cancer. FJX1 mRNA and protein are upregulated in human colorectal tumor epithelium as compared with normal epithelium and colorectal adenomas, and high expression of FJX1 is associated with poor patient prognosis. FJX1 mRNA expression in colorectal cancer tissues is significantly correlated with changes in known angiogenesis genes. Augmented expression of FJX1 in colon cancer cells promotes growth of xenografts in athymic mice and is associated with increased tumor cell proliferation and vascularization. Furthermore, FJX1 null mice develop significantly fewer colonic polyps than wild-type littermates after combined dextran sodium sulfate (DSS) and azoxymethane (AOM) treatment. In vitro, conditioned media from FJX1 expressing cells promoted endothelial cell capillary tube formation in a HIF1-α dependent manner. Taken together our results support the conclusion that FJX1 is a novel regulator of tumor progression, due in part, to its effect on tumor vascularization.     

Notch1 Is a 5-Fluorouracil Resistant and Poor Survival Marker in Human Esophagus Squamous Cell Carcinomas

Notch signaling involves the processes that govern cell proliferation, cell fate decision, cell differentiation and stem cell maintenance. Due to its fundamental role in stem cells, it has been speculated during the recent years that Notch family may have critical functions in cancer stem cells or cancer cells with a stem cell phenotype, therefore playing an important role in the process of oncogenesis. In this study, expression of Notch family in KYSE70, KYSE140 and KYSE450 squamous esophageal cancer cell lines and virus transformed squamous esophageal epithelial cell line Het-1A was examined by quantitative RT-PCR. Compared to the Het-1A cells, higher levels of Nocth1 and Notch3 expression in the cancer cell lines were identified. Due to the finding that NOTCH3 mainly mediates squamous cell differentiation, NOTCH1 expression was further studied in these cell lines. By Western blot analyses, the KYSE70 cell line which derived from a poorly differentiated tumor highly expressed Notch1, and the Notch1 expression in this cell line was hypoxia inducible, while the KYSE450 cell line which derived from a well differentiated tumor was always negative for Notch1, even in hypoxia. Additional studies demonstrated that the KYSE70 cell line was more 5-FU resistant than the KYSE450 cell line and such 5-FU resistance is correlated to Notch1 expression verified by Notch1 knockdown experiments. In clinical samples, Notch1 protein expression was detected in the basal cells of human esophagus epithelia, and its expression in squamous cell carcinomas was significantly associated with higher pathological grade and shorter overall survival. We conclude that Notch1 expression is associated with cell aggressiveness and 5-FU drug resistance in human esophageal squamous cell carcinoma cell lines in vitro and is significantly associated with a poor survival in human esophageal squamous cell carcinomas.     

HMGA1-pseudogene overexpression contributes to cancer progression

Two pseudogenes for HMGA1, whose overexpression has a critical role in cancer progression, have been identified. They act as decoy for miRNAs that are able to target the HMGA1 gene then enhancing cell proliferation and migration. Moreover, these pseudogenes contain sequences that are potential target sites for cancer-related miRNAs. Interestingly, HMGA1 pseudogenes are highly expressed in human anaplastic thyroid carcinomas, that is one of the most aggressive tumor in mankind, but almost undetectable in well differentiated thyroid carcinomas.     

Value of demonstration of cytokeratin and leukocyte common antigen in poorly differentiated malignant tumors in fine needle aspirates

Immunoperoxidase staining for cytokeratin and leukocyte common antigen (LCA) was applied to aspirated material from 21 well-differentiated malignant tumors ("control cases," which could be cytologically categorized) and 32 poorly differentiated malignant tumors ("study cases," whose tumor type was not clearly evident from study of the aspirate). Fifteen (48%) of the 32 poorly differentiated malignant tumors were classified as carcinomas (12 cases) or lymphomas (3 cases) by positive staining for cytokeratin or LCA, respectively. Good cytohistopathologic correlation was observed in all 7 control cases and in 11 cases of the study cases for which histologic specimens were available. The addition of staining for desmin helped in categorizing one undifferentiated tumor as a rhabdomyosarcoma. Immunostaining for vimentin was not found to be valuable in the categorization of these tumors.     

The localization of Fos B, a member of transcription factor AP-1 family, in rat odontoblasts and pulpal undifferentiated ectomesenchymal cells

It has been proposed that cellular proliferation and differentiation are accomplished by AP-1 components but different components can be responsible for different functions. The aim of this study was to compare the localization of Fos B, which is a component of AP-1, in postmitotic differentiated and undifferentiated cells via Fos B immunoreactivity. For this purpose, maxillary incisor teeth from 10 Wistar rats were obtained and Fos-B was investigated immunohistochemically in formalin-fixed, paraffin-embedded tooth sections containing odontoblasts, which are postmitotic differentiated cells, and pulpal undifferentiated ectomesenchymal cells. No significant differences in percentage of Fos B-positive cells were observed between the two cell types (p>0.05). These findings suggest that Fos B, a component of AP-1 family, seems to have a negligible effect on differentiation and proliferation in odontoblasts and pulpal undifferentiated ectomesenchymal cells.