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We have recently shown that IFNγ, produced during cancer therapy, induces expression of the Bcl3 proto‐oncogene in ovarian cancer (OC) cells, resulting in their increased proliferation, migration, and invasion, but the mechanisms are unknown. Here, we demonstrate that the IFNγ‐induced Bcl3 expression is dependent on JAK1 and STAT1 signaling, and on p65 NFκB. Furthermore, the IFNγ‐induced Bcl3 expression is associated with an increased occupancy of Ser‐727 phosphorylated STAT1 and acetylated histone H3 at the Bcl3 promoter. Our data indicate that Bcl3 promotes expression of the pro‐inflammatory chemokine interleukin‐8 (IL‐8) in OC cells. These findings identify Bcl3 as a novel target of IFNγ/JAK1/STAT1 signaling and suggest that targeting the JAK1/STAT1 pathway may suppress IFNγ‐induced Bcl3 expression in OC. [ABSTRACT FROM AUTHOR]

The proto-oncogene Bcl3 induces survival and proliferation in cancer cells; however, its function and regulation in ovarian cancer (OC) remain unknown. Here, we show that Bcl3 expression is increased in human OC tissues. Surprisingly, however, we found that in addition to promoting survival, proliferation, and migration of OC cells, Bcl3 promotes both constitutive and interferon-γ (IFN)-induced expression of the immune checkpoint molecule PD-L1. The Bcl3 expression in OC cells is further increased by IFN, resulting in increased PD-L1 transcription. The mechanism consists of an IFN-induced, Bcl3- and p300-dependent PD-L1 promoter occupancy by Lys-314/315 acetylated p65 NF-kB. Blocking PD-L1 by neutralizing antibody reduces proliferation of OC cells overexpressing Bcl3, suggesting that the pro-proliferative effect of Bcl3 in OC cells is partly mediated by PD-L1. Together, this work identifies PD-L1 as a novel target of Bcl3, and links Bcl3 to IFNγ signaling and PD-L1-mediated immune escape. [ABSTRACT FROM AUTHOR]

Objective To describe a Hokkaido dog, one of the traditional Japanese breeds that was affected by Collie eye anomaly (CEA), and to report the genotype of this dog and the Hokkaido dog allelic frequency of the CEA-associated mutation. Case A nine-month-old intact female Hokkaido dog without any obvious visual disturbance was diagnosed ophthalmoscopically with CEA. Severe choroidal hypoplasia was observed in the bilateral temporal area adjacent to the optic nerve head, appearing as whitish areas. Therefore, the dog was suspected of possessing the CEA-associated mutation that was previously reported as an intronic 7.8-kilo base deletion in the canine NHEJ1 gene. Procedures SYBR Green-based real-time PCR with a melting curve analysis, conventional PCR with agarose gel electrophoresis, and direct DNA sequencing were carried out to determine the genotype of the dog. Furthermore, a preliminary genotyping survey was carried out in 17 Hokkaido dogs from three kennels using the real-time PCR method, and the pedigree relationships were analyzed using their pedigree papers. Results The Hokkaido dog affected by CEA was proven to possess the CEA-associated mutation. Of these 17 Hokkaido dogs, 12 dogs were heterozygous carriers and five dogs were affected by this mutation. The preliminary genotyping survey and pedigree analysis demonstrated that the allelic frequency of the CEA-associated mutation is very high in Hokkaido dogs. Conclusion These data suggest that the Hokkaido breed is highly susceptible to CEA because of the known CEA-associated mutation much like the Collie-related breeds. [ABSTRACT FROM AUTHOR]

The article looks at the role of rapid genotyping assays in detecting deletion mutation and assessing mutant allele frequency in Border Collie dogs. The study was performed using microchip electrophoresis for evaluating allele frequency and deletion mutation of surveyed Border Collie dogs in Japan. Results of the study showed that carrier and mutant allele frequencies in Border Collie dogs were at 0.49 and 0.25 percent, respectively.

The article discusses a study on the use of SYBR Green-based real-time polymerase chain reaction (PCR) in order to develop a novel rapid genotyping technique for Collie eye anomaly (CEA). It notes the advantages of SYBR Green-based real-time PCR when combined with Flinders Technology Associates filter paper (FTA) cards. Continuous selective breeding is identified as an effective method of decreasing high CEA incidences. Such technique is recommended for large-scale screening in breeding populations of Collie-related breeds.

The pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8) induces proliferation and invasion of solid tumor cells. In many types of solid cancer, including triple-negative breast cancer (TNBC), the IL-8 expression is induced by proteasome inhibition. In this chapter, we describe a protocol for the analysis of TNBC cell invasion induced by IL-8 in response to proteasome inhibition by bortezomib (BZ). Using this approach, we show that BZ increases the invasion ability of TNBC cells, and that the BZ-increased TNBC cell invasion is suppressed by IκB kinase (IKK) inhibition, which also decreases the IL-8 expression. The experimental protocol includes the cell invasion assay, microscopic evaluation of the invading cells, and quantitative analysis of the obtained images. This protocol should be applicable also for measurement of chemokine-induced tumor cell invasion in other types of cancer cells.

Interferon-γ (IFNγ) is a pleiotropic cytokine that signals to many different cell types. IFNγ has both antitumor functions and pro-tumorigenic effects and regulates different aspects of cell physiology, including metabolism. Cancer cells undergo a complex rearrangement of metabolic pathways that allows them to satisfy the needs of increased proliferation, and many cancer cells redirect glucose metabolism from oxidative phosphorylation to aerobic glycolysis. In this chapter, we describe a protocol that utilizes the Agilent Seahorse XFp Analyzer to assess mitochondrial respiration and glycolysis in ovarian cancer cells treated with IFNγ.

IFNγ is a pleiotropic cytokine that has both antitumor functions and pro-tumorigenic effects. Recent studies have shown that IFNγ induces expression of the immune checkpoint PD-L1 in ovarian cancer (OC) cells, resulting in their increased proliferation and tumor growth. Here, we tested the hypothesis that IFNγ induces migration of OC cells. Using the scratch wound healing assay, our results demonstrate that IFNγ promotes OC cell migration, thus adding to the complexities of IFNγ pro-tumorigenic mechanisms. This chapter describes analysis of the IFNγ-induced migration of OC cells by the wound healing assay followed by quantification of the obtained images using ImageJ software.

Ovarian cancer (OC) is the most common cause of cancer deaths among gynecological malignancies. OC ascites contain multicellular spheroid aggregates, which exhibit increased pro-survival signaling, invasive behavior, and chemotherapeutic resistance. OC cells are characterized by an increased expression of the pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8), which increases their survival and migration, thus contributing to OC metastasis and angiogenesis. While previous studies have shown that IL-8 increases proliferation of OC cells grown in monolayer cultures, the effect of IL-8 on proliferation of OC cells grown in 3D spheroids has not been investigated. The spheroid 3D culture assays have been particularly useful in translational research since they allow cell-to-cell interactions that resemble tumor growth in vivo, while allowing easy cell manipulations and visualization. Here, we used the 3D spheroid culture assay to investigate the effect of IL-8 on OC cell proliferation. Using this assay, our results show that IL-8 significantly increases proliferation of OC cells grown in 3D spheroids.