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Summary: A 13‐year‐old Thoroughbred/Oldenburg gelding presented as a tertiary referral for a 6‐year history of immune‐mediated keratitis (IMMK) after the development of corneal and limbal nodules in the presence of medical therapy. A keratectomy extending to the limbus was performed to remove the affected cornea and limbal nodules. Two corneal‐limbal samples were submitted for histopathology and initially diagnosed as marked lympho‐histiocytic stromal keratitis with epithelial hyperplasia, keratinisation and keratinocyte apoptosis consistent with IMMK. Additional testing with immunohistochemical staining for CD3, CD20, PAX5 and IBA1 revealed large, atypical B‐lymphocytes on a background of histiocytes and mature T‐lymphocytes, with concern for T‐cell‐rich large B‐cell lymphoma. PCR antigen receptor rearrangement clonality testing was also performed and was diagnostic for large B‐cell lymphoma. Polymerase chain reaction (PCR) for equine herpesvirus 5 (EHV‐5) was positive. Following keratectomy, a short course of neomycin/polymyxin B/dexamethasone ophthalmic ointment was prescribed; no other treatment nor adjunct therapy was performed. Eighteen months post‐operation, the case remains visual and comfortable on no medications. [ABSTRACT FROM AUTHOR]

Background: Although intestinal smooth muscle cells (ISMCs) play an important role in the remodeling of the intestinal structure, considerably less is known about the molecular mechanisms that mediate the development and growth of ISMCs. A possible reason for this lag is the lack of cell lines that recapitulate ISMCs in vivo. Methods: In this study, we separated the primary ISMCs from the rat intestinal tract and integrated the Simian Vacuolating virus 40 Large T antigen (SV40 LT) gene into the genome of the primary ISMC to construct an immortalized cell line named ISMC‐Hc. Key Results: ISMC‐Hc proliferated persistently without any signs of senescence up to 50 passages and without neoplasticity. Analysis of the genome isolated from ISMC‐Hc confirmed that the SV40LT gene recombined in the genome, and mRNA reverse transcription PCR suggested that SV40LT could be expressed normally. In addition, ISMC‐Hc had few morphological differences compared with the primary ISMC. Furthermore, ISMC‐Hc showed the expression of the specific protein markers (alpha‐smooth muscle actin and desmin) through immunofluorescence analysis. Further studies showed that ISMC‐Hc had enhanced contractility and expressed the glial‐derived neurotrophic factor, nerve growth factor, ciliary neurotrophic factor, and leukemia inhibitory factor after co‐stimulation with IL‐1 beta and TNF‐alpha. Conclusions and Inferences: ISMC‐Hc showed characteristics similar to that of primary ISMC and can be used as an in vitro model to explore the cellular and molecular mechanisms of ISMC. Additionally, the immortalized ISMCs could help investigate the basic functional mechanisms of intestinal diseases. [ABSTRACT FROM AUTHOR]

Pioglitazone (PIO), an oral hypoglycemic agent, is used in the treatment of type 2 diabetes. Some studies have suggested that an increased risk of bladder cancer with PIO exposure, while the others reported there is no such relationship. Therefore, it is doubtful whether PIO can increase the risk of bladder cancer. The effects of PIO on DNA damage and/or transformation of human bladder cells are not fully known. We investigated the effects of PIO on cytotoxicity, DNA single and double strand breaks and repair and neoplastic transformation in human bladder cells (hTU1) treated with 10, 20, and 40 μM PIO for 24, 48 and 72 hr. PIO decreased cell viability in a concentration‐dependent manner. Increased levels of comet parameters showed that PIO and its metabolites can significantly induce DNA double strand breaks at all concentrations tested. PIO also significantly induced the formation of phosphorylated H2AX and p53 binding protein 1 foci. DNA damage was not repaired in a 24 hr recovery period. PIO can also induce malignant transformation of human bladder cells exhibiting loss of contact inhibition and anchorage independent growth. This is the first study to indicate that PIO can induce DNA damage and malignant transformation, reduce or alter the DNA repair capacity in human bladder cells. From these results, we suggest that patients with diabetes treated with PIO may have an increased risk of bladder cancer. [ABSTRACT FROM AUTHOR]

Neoplastic transformation is a rare but serious complication of Paget's disease of bone (PDB), occurring in fewer than 1% of individuals with polyostotic disease. Their prognosis is poor, with less than 50% surviving 5 years. In 2016, the genetic alteration of giant cell tumor (GCT) complicating PDB was identified as a founder germline mutation (P937R) in the ZNF687 gene. However, the study population was exclusively of Italian descent, and patients of different ethnic origins were not studied. To fill this gap, herein we performed mutation analysis of ZNF687 in a GCT in the pelvis of a 45‐year‐old black American woman with polyostotic PDB. The P937R mutation in ZNF687 was found in her tumor but, as expected, the ancestral haplotype that characterizes the Italian GCT/PDB patients was not found. Furthermore, we identified two additional Italian GCT/PDB patients with this ZNF687 mutation, now constituting a cohort of 18 GCT/PDB cases, all harboring the identical mutation. We also searched for ZNF687 mutations in a unique collection of tumor tissues derived from Italian PDB patients, including 28 osteosarcomas (OS/PDB), 8 undifferentiated sarcomas (SRC/PDB), 1 fibrosarcoma (FS/PDB), and 1 chondrosarcoma (CS/PDB). We identified the P937R mutation in one SRC/PDB and a different ZNF687 mutation (R331W) in 1 of 28 pagetic osteosarcomas. Thus, whereas GCT/PDB pathogenesis globally seems to involve the P937R mutation in ZNF687, other neoplasms associated with PDB seem to be less related to mutations in this gene. Finally, we identified the G34W mutation in the H3F3A gene in the maxillary tumor masses of two PDB patients, defining them as conventional GCT rather than GCT/PDB. Thus, combined molecular analysis of H3F3A and ZNF687 is essential to clarify the origin and diagnosis of tumors in PDB. © 2020 American Society for Bone and Mineral Research. [ABSTRACT FROM AUTHOR]

Neoplastic transformation of Spirocerca lupi induced esophageal nodules carries a poor prognosis. Clinical, clinicopathological, endoscopic, and radiographic characteristics may be indicative of neoplastic transformation but variable sensitivity and specificity of these parameters makes their use questionable. We hypothesized that CT would be a better diagnostic modality to discriminate between non-neoplastic and neoplastic nodules. In this prospective study of 38 dogs, the appearance and perfusion characteristics of confirmed spirocercosis-induced neoplastic and non-neoplastic esophageal nodules were described using survey CT and triple phase dynamic CT angiography (CTA). Pre- and post-contrast early arterial, late arterial, and venous CTA images were evaluated. Non-neoplastic nodules were smooth and nonmineralized with a higher proportion of hypoattenuating necropurulent cavities compared to neoplastic nodules that had a more irregular surface, with 93% having mineralized foci and rarely any hypoattenuating pockets. Non-neoplastic nodules were significantly more perfused than neoplastic nodules with the difference being up to 23 Hounsfield units. The difference was most marked in the early and late arterial phases ( P = 0.0005 and 0.00005, respectively). Ratios of the normal esophagus adjacent to the neoplastic and non-neoplastic nodules did not differ significantly from each other. Perfusion findings demonstrated relative hypoperfusion of the esophageal sarcomas. Findings from the current study indicated that CT characteristics of relative postcontrast hypoperfusion, combined with nodule irregularity and mineralization warrant a high level of concern for neoplastic transformation in canine spirocercosis-induced esophageal nodules. [ABSTRACT FROM AUTHOR]

Cytochrome P450 2A13 (CYP2A13), mainly expressed in human respiratory tract, is highly efficient in the metabolic activation of aflatoxin (AF) B1 (AFB1) and is assumed to play a role in human lung tumorigenesis in airborne AFB1 exposure. To validate the assumption, we exposed human bronchial epithelial (BEAS-2B) cells stably expressing CYP2A13 (B-2A13), CYP1A2 (B-1A2) and CYP2A6 (B-2A6) to 0.1-10 nM AFB1 for 30-50 passages. B-2A13 cells showed increased sensitivity to 0.1 nM AFB1-induced neoplastic transformation and the formation of tumors in nude mice were observed at passage 30 (P30) while it occurred at P50 B-1A2 cells. B-2A6, similar to vector control, showed no neoplastic transformation in this condition. Additionally, AFB1-DNA adducts and 8-OHdG significantly increased in transformed P40 B-2A13, in parallel with the upregulation of p-ATR, p-BRCA1, Mre11, Rad50 and Rad51. However, the apoptosis of P40 cells was near normal, while the expression of Bax, C-Caspase 3 and C-PARP increased passage-dependently. Inhibition of ATR (ATR siRNA or NU6027) reversely increased the apoptosis of P40 B-2A13 cells in parallel with the upregulation of Bax, C-Caspase 3 and C-PARP, suggesting that ATR plays an important role in maintaining cell survival via antiapoptosis. Additionally, activation of ATR was necessary to neoplastic transformation since blockage of ATR in P40 cells inhibited DNA damage repair response and anchorage-independent growth. Our data demonstrated that CYP2A13 played a critical role in AFB1-induced neoplastic transformation. ATR-mediated the dysfunction of apoptosis and DNA damage repair might be involved. These results help establish a linkage between airborne AFB1 and human respiratory carcinoma. [ABSTRACT FROM AUTHOR]

Epileptic seizures are frequent manifestations of brain tumors. However, biopsy specimens of patients who undergo neurosurgical removal of circumscribed foci to control chronic recurrent pharmacoresistant seizures often reveal tumor entities that are rare in general brain tumor series. The spectrum of these 'long-term epilepsy-associated neoplasms' comprises highly differentiated glial and glioneuronal tumors that show a benign biologic behavior and clinical course, and that rarely relapse. Several entities are well recognizable on the basis of histopathologic and immunohistochemical characteristics. An intriguing functional aspect of these tumors, sometimes collectively referred to as 'epileptomas,' is their prominent epileptogenicity, which may represent a clinical feature indicating rather than causing the generally benign biologic behavior of these tumors. A frequent feature of respective neoplasms is their coincidence with dysplastic lesions in the vicinity of the tumor itself. The recent advent of new molecular markers, including genomic alterations leading to activation of the protooncogene BRAF and impaired function of isocitrate dehydrogenase ( IDH1), provides excellent new tools in the differential diagnosis of low grade brain tumors, and provides intriguing implications to further develop the pathogenetic concepts of these neoplasms. Despite this progress, a number of tumors from patients with chronic epilepsy show combinations of cytologic, histologic, and immunohistochemical characteristics that challenge the current neuropathologic classification schemes. Attempts are currently ongoing to develop further classification schemes. [ABSTRACT FROM AUTHOR]

Oxidative stress, caused by reactive oxygen species ( ROS), is a major contributor to inflammatory bowel disease ( IBD)-associated neoplasia. We mimicked ROS exposure of the epithelium in IBD using non-tumour human colonic epithelial cells ( HCEC) and hydrogen peroxide ( H2O2). A population of HCEC survived H2O2-induced oxidative stress via JNK-dependent cell cycle arrests. Caspases, p21WAF1 and γ-H2 AX were identified as JNK-regulated proteins. Up-regulation of caspases was linked to cell survival and not, as expected, to apoptosis. Inhibition using the pan-caspase inhibitor Z- VAD- FMK caused up-regulation of γ-H2 AX, a DNA-damage sensor, indicating its negative regulation via caspases. Cell cycle analysis revealed an accumulation of HCEC in the G1-phase as first response to oxidative stress and increased S-phase population and then apoptosis as second response following caspase inhibition. Thus, caspases execute a non-apoptotic function by promoting cells through G1- and S-phase by overriding the G1/S- and intra-S checkpoints despite DNA-damage. This led to the accumulation of cells in the G2/M-phase and decreased apoptosis. Caspases mediate survival of oxidatively damaged HCEC via γ-H2 AX suppression, although its direct proteolytic inactivation was excluded. Conversely, we found that oxidative stress led to caspase-dependent proteolytic degradation of the DNA-damage checkpoint protein ATM that is upstream of γ-H2 AX. As a consequence, undetected DNA-damage and increased proliferation were found in repeatedly H2O2-exposed HCEC. Such features have been associated with neoplastic transformation and appear here to be mediated by a non-apoptotic function of caspases. Overexpression of upstream p- JNK in active ulcerative colitis also suggests a potential importance of this pathway in vivo. [ABSTRACT FROM AUTHOR]

Fundamental to eukaryotic cell function, nucleocytoplasmic transport can be regulated at many levels, including through modulation of the importin/exportin (Imp/Exp) nuclear transport machinery itself. Although Imps/Exps are overexpressed in a number of transformed cell lines and patient tumor tissues, the efficiency of nucleocytoplasmic transport in transformed cell types compared with nontransformed cells has not been investigated. Here we use quantitative live cell imaging of 3 isogenic nontransformed/transformed cell pairs to show that nuclear accumulation of nuclear localization signal (NLS)-containing proteins, but not their NLS-mutated derivatives, is increased up to 7-fold in MCF10CA1h human epithelial breast carcinoma cells and in simian virus 40 (SV40)-transformed fibroblasts of human and monkey origin, compared with their nontransformed counterparts. The basis for this appears to be a significantly faster rate of nuclear import in transformed cell types, as revealed by analysis using fluorescence recovery after photobleaching for the human MCF10A/MCF10CA1h cell pair. Nuclear accumulation of NLS/nuclear export signal-containing (shuttling) proteins was also enhanced in transformed cell types, experiments using the nuclear export inhibitor leptomycin B demonstrating that efficient Exp-1-mediated nuclear export was not impaired in transformed compared with nontransformed cells. Enhanced nuclear import and export efficiencies were found to correlate with 2- to 4-fold higher expression of specific Imps/Exps in transformed cells, as indicated by quantitative Western blot analysis, with ectopic expression of Imps able to enhance NLS nuclear accumulation levels up to 5-fold in nontransformed MCF10A cells. The findings indicate that transformed cells possess altered nuclear transport properties, most likely due to the overexpression of Imps/Exps. The findings have important implications for the development of tumor-specific drug nanocarriers in anticancer therapy. [ABSTRACT FROM AUTHOR]

Abstract: Human chorionic gonadotropin (hCG), a hormone produced during pregnancy, can elicit life-long refractoriness to carcinogenesis by differentiation of the breast epithelium. Human breast epithelial cells MCF-10F form tubules in collagen, mimicking the normal ductules. We have shown that 17 β-estradiol (E2) alter the ductulogenic pattern of these cells. The effect of the recombinant hCG (rhCG) in vitro was evaluated on the transformation of MCF-10F induced by E2. MCF-10F cells were treated with 70nM E2 alone or in combination with 50IU/ml rhCG during 2 weeks, while the controls were treated with DMSO (the solvent in which E2 was dissolved) or rhCG alone. At the end of treatment, the cells were plated in type I collagen matrix (3D-cultures) for detecting 2 main phenotypes of cell transformation, namely the loss of ductulogenic capacity and the formation of solid masses. Although E2 significantly increased solid mass formation, this effect was prevented when MCF-10F cells were treated with E2 in combination with rhCG. Furthermore, E2 increased the main duct width (p <0.001), and caused a disruption of the luminal architecture, whereas rhCG increased the length of the tubules (p <0.001) and produced tertiary branching. In conclusion, rhCG was able to abrogate the transforming abilities of estradiol, and had the differentiating property by increasing the branching of the tubules formed by breast epithelial cells in collagen. These results further support our hypothesis, known as the terminal differentiation hypothesis of breast cancer prevention, that predicts that hCG treatment results in protection from tumorigenic changes by the loss of susceptible stem cells 1 through a differentiation to refractory stem cells 2 and increase differentiation of the mammary gland. [Copyright &y& Elsevier]

While bile acids are a risk factor for tumorigenesis induced by reflux disease, the mechanisms by which they contribute to neoplasia remain undefined. Here, we reveal that in gastroesophageal junction (GEJ) cells bile acids activate a tissue-specific developmental program defining the intestinal epithelial cell phenotype characterizing GEJ metaplasia. Deoxycholic acid (DCA) inhibited phosphorylation of EGF receptors (EGFRs) suppressing the proto-oncogene AKT. Suppression of EGFRs and AKT by DCA actuated an intestine-specific cascade in which NF-κB transactivated the tissue-specifi c transcription factor CDX2. In turn, CDX2 orchestrated a lineage-specific differentiation program encompassing genes characterizing intestinal epithelial cells. Conversely, progression from metaplasia to invasive carcinoma in patients, universally associated with autonomous activation of EGFRs and/or AKT, was coupled with loss of this intestinal program. Thus, bile acids induce intestinal metaplasia at the GEJ by activating the lineage-specifi c differentiation program involving suppression of EGFR and AKT, activating the NF-κB-CDX2 axis. Induction of this axis provides the context for lineage-addicted tumorigenesis, in which autonomous activation of AKT corrupts adaptive intestinal NF-κB signaling, amplifying tumorigenic programs. [ABSTRACT FROM AUTHOR]

Carcinogenesis is a multi-step process involving genetic alterations and non-genotoxic mechanisms. The in vitro cell transformation assay allows the monitoring of the neoplastic phenotype by foci formation in suitable cells (e.g. C3H10T1/2 mouse embryo fibroblasts) showing aberrant morphology of massive build-up, polar and multi-layered densely stained cells. The classification of transformed foci in C3H cells relies on light microscopy scoring by a trained human expert based on standard rules. This procedure is time-consuming and prone, in some cases, to subjectivity, thereby leading to possible over- or under-estimation of the carcinogenic potential of tested compounds. Herewith we describe the in vitro neoplastic transformation induced by B[a]P and CdCl2, and the development of a foci classifier based on image analysis and statistical classification. The image analysis system, which relies on ‘spectrum enhancement’, is quantitative and extracts descriptors of foci texture and structure. The statistical classification method is based on the Random Forest algorithm. We obtained a classifier trained by using expert's supervision with a 20% classification error. The proposed method could serve as a basis to automate the in vitro cell transformation assay. [ABSTRACT FROM AUTHOR]

The expression of the high mobility group I (HMGI)-C chromatin component was shown previously to be essential for the establishment of the neoplastic phenotype in retrovirally transformed thyroid cell lines. To identify possible targets of the HMGI-C gene product, we have analyzed the AP-1 complex in normal, fully transformed and antisense HMGI-C-expressing rat thyroid cells. We show that neoplastic transformation is associated with a drastic increase in AP-1 activity, which reflects multiple compositional changes. The strongest effect is represented by the dramatic junB and fra-1 gene induction, which is prevented in cell lines expressing the antisense HMGI-C. These results indicate that the HMGI-C gene product is essential for the junB and fra-1 transcriptional induction associated with neoplastic transformation. The inhibition of Fra-1 protein synthesis by stable transfection with a fra-1 antisense RNA vector significantly reduces the malignant phenotype of the transformed thyroid cells, indicating a pivotal role for the fra-1 gene product in the process of cellular transformation. [ABSTRACT FROM AUTHOR]