Spatial Localization of Eubacterial 16S rRNA in Early Pregnancy Placenta and Decidua.

Bacteria derived from the maternal circulation have been suggested to seed the human placenta during pregnancy leading to development of an intrinsic placental microbiome; however, other data indicates these bacteria are artifactual contaminants. Limited research on the localization of bacteria in human placental tissue is available, which may help differentiate resident placental bacteria from contaminants. This study spatially localizes bacteria in situ in normal late first to early second trimester human placenta by 16S rRNA chromogenic in situ hybridization and demonstrates patterns consistent with both contaminants and intraparenchymal signals. These results suggest that placental microbiome studies may benefit from spatial strategies that can exclude surface contamination.

High-Risk HPV CISH Detection in Cervical Biopsies with Weak and/or Focal p16 Immunohistochemical Positivity.

In cervical biopsies, for diagnosis of Human Papilloma Virus (HPV) related conditions, the immunohistochemical staining for p16 has a diagnostic value only if diffusely and strongly positive, pattern named "block-like". "Weak and/or focal (w/f) p16 expression" is commonly considered nonspecific. In our previous study, we demonstrated the presence of high-risk HPV (hrHPV) DNA by LiPa method in biopsies showing w/f p16 positivity. The aim of the present study was to investigate the presence of hrHPV-DNA by CISH in the areas showing w/f p16 expression. We assessed the presence of hrHPV16, 18, 31, 33, 51 by CISH in a group of 20 cervical biopsies showing w/f p16 expression, some with increased Ki67, and in 10 cases of block-like expression, employed as control. The immunohistochemical p16 expression was also assessed by digital pathology. hrHPV-CISH nuclear positivity was encountered in 12/20 cases of w/f p16 expression (60%). Different patterns of nuclear positivity were identified, classified as punctate, diffuse and mixed, with different epithelial distributions. Our results, albeit in a limited casuistry, show the presence of HPV in an integrated status highlighted by CISH in w/f p16 positive cases. This could suggest the necessity of a careful follow-up of the patients with "weak" and/or "focal" immunohistochemical patterns of p16, mainly in cases of increased Ki67 cell proliferation index, supplemented with molecular biology examinations.

Hepatic cytokine-inducible SH2-containing protein (CISH) regulates gluconeogenesis via cAMP-responsive element binding protein (CREB).

Impairment of gluconeogenesis is a key factor responsible for hyperglycemia in patients with type 2 diabetes. As an important member of the suppressors of cytokine signaling (SOCS) protein family, many physiological functions of cytokine-inducible SH2-containing protein (CISH) have been described; however, the role of hepatic CISH in gluconeogenesis is poorly understood. In the present study, we observed that hepatic CISH expression was reduced in fasted wild-type (WT) mice. Overexpression of CISH decreased glucose production in mouse primary hepatocytes, while silencing of CISH had the opposite effects. In addition, adenovirus-mediated hepatic CISH overexpression resulted in improved glucose tolerance and decreased gluconeogenesis in WT and leptin receptor-deficient diabetic (db/db) mice. In contrast, adenovirus-mediated hepatic CISH knockdown impaired glucose tolerance and increased gluconeogenesis in WT mice. We also generated liver-specific CISH knockout (LV-CISH KO) mice and discovered that these mice had a similar phenotype in glucose tolerance and gluconeogenesis as mice injected with adenoviruses that knockdown CISH expression. Mechanistically, we found that CISH overexpression decreased and CISH knockdown increased the mRNA and protein levels of glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase 1 (PEPCK), two key enzymes involved in gluconeogenesis, in vitro, and in vivo. Moreover, we discovered that the phosphorylation of cAMP-responsive element binding protein 1 (CREB), a transcription factor of G6pase and Pepck, was required for regulating gluconeogenesis by CISH. Taken together, this study identifies hepatic CISH as an important regulator of gluconeogenesis. Our results also provide important insights into the metabolic functions of the SOCS protein family and the potential targets for the treatment of diabetes.

Cytokine-inducible SH2 domain containing protein contributes to regulation of adiposity, food intake, and glucose metabolism.

The cytokine-inducible SH2 domain containing protein (CISH) is the founding member of the suppressor of cytokine signaling (SOCS) family of negative feedback regulators and has been shown to be a physiological regulator of signaling in immune cells. This study sought to investigate novel functions for CISH outside of the immune system. Mice deficient in CISH were generated and analyzed using a range of metabolic and other parameters, including in response to a high fat diet and leptin administration. CISH knockout mice possessed decreased body fat and showed resistance to diet-induced obesity. This was associated with reduced food intake, but unaltered energy expenditure and microbiota composition. CISH ablation resulted in reduced basal expression of the orexigenic Agrp gene in the arcuate nucleus (ARC) region of the brain. Cish was basally expressed in the ARC, with evidence of co-expression with the leptin receptor (Lepr) gene in Agrp-positive neurons. CISH-deficient mice also showed enhanced leptin responsiveness, although Cish expression was not itself modulated by leptin. CISH-deficient mice additionally exhibited improved insulin sensitivity on a high-fat diet, but not glucose tolerance despite reduced body weight. These data identify CISH as an important regulator of homeostasis through impacts on appetite control, mediated at least in part by negative regulation of the anorexigenic effects of leptin, and impacts on glucose metabolism.

The Role of Cytokine-Inducible SH2 Domain-Containing Protein (CISH) in the Regulation of Basal and Cytokine-Mediated Myelopoiesis.

Cytokine-inducible SH2 domain-containing protein (CISH) is a member of the suppressor of cytokine signaling (SOCS) family of negative feedback regulators shown to play crucial roles in lymphoid cell development and function as well as appetite regulation. It has also been implicated in the control of signaling downstream of the receptors for the cytokines granulocyte/macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) in myeloid cells. To investigate the physiological role of CISH in myelopoiesis, mice deficient in CISH were analyzed basally and in response to administration of these cytokines. CISH knockout (KO) mice possessed basally elevated neutrophils in the blood, bone marrow, and spleen compared to wild-type (WT) mice. During GM-CSF-induced myelopoiesis, the frequency of neutrophils, myeloid dendritic cells (DCs), and CFU-M in the bone marrow was higher in the KO, as were the neutrophils and CFU-G in the spleen. In contrast, no differences were observed between KO and WT mice during G-CSF-induced myelopoiesis apart from an elevated frequency of CFU-G and CFU-M in the spleen. This work has identified a role for CISH in the negative regulation of granulopoiesis, including that mediated by GM-CSF.

[Enhanced Natural Killers with CISH and B2M Gene Knockouts Reveal Increased Cytotoxicity in Glioblastoma Primary Cultures].

In an experimental study using the CRISPR/Cas9 system, "enhanced" NK cell lines with knockout of CISH, the gene for the CIS protein (a negative regulator of NK cytotoxicity), as well as two lines with a knocked-out ß2-microglobulin gene, which provides membrane exposure of MHC class I, were obtained from two parental lines of human natural killers (YT wild type and YT-VAV1^(+) overexpressing the VAV1 cytotoxicity enhancing protein). The knockout efficiency was determined by real-time PCR as well as by flow cytometry with specific antibodies. The resulting CISH^(-/-) or B2M^(-/-) knockout lines were tested for cytotoxicity in primary monolayer cultures of human glioblastoma multiforme. The cytotoxicity of the lines was assessed using a cell analyzer that records the cell index based on cell impedance. YT-CISH^(-/-) has been shown to be significantly more effective than wild-type YT in eliminating primary glioblastoma cells in an in vitro cell monolayer experiment. The cytotoxicity of the YT-VAV1^(+)-CISH^(-/-) and YT-VAV1^(+)B2M^(-/-) lines against glioblastoma cells was the highest, but overall, it did not significantly differ from the initially increased cytotoxicity of the YT-VAV1^(+) line. The lines of NK-like cells obtained may serve as a prototype for the creation of "enhanced" allogeneic and autologous NK- and CAR-NK cells for the immunotherapy of glioblastoma multiforme.

MYC status in HIV-associated plasmablastic lymphoma: dual-colour CISH, FISH and immunohistochemistry.

AIMS: We utilised chromogenic and fluorescence in-situ hybridisation (CISH and FISH) to evaluate MYC gene copy numbers and rearrangements within HIV-associated plasmablastic lymphomas (PBLs). Thereafter, clinicopathological features were explored retrospectively. METHODS AND RESULTS: Sixty-seven (n = 67) patients were included and the HIV seropositive status was confirmed in 98% (63 of 64) with a median viral load of 55 587 (IQR 273 582) copies/ml and median CD4 count of 170 (IQR 249) cells/µl. The mean age was 41 ± 10.1 years and females comprised 54%. PBL was documented predominantly at extra-oronasal topographic regions. Starry-sky (SS) appearance was evident in 33% in association with monomorphic morphology (P-value 0.02). c-MYC protein was expressed in 81% and latent EBV infection was detected in 90%. EBER ISH-positive status and MYC rearrangement occurred in 67% of HIV PBL. MYC aberrations included MYC rearrangement (70%), low-level increase in MYC gene copy numbers (43%), concurrent MYC rearrangement and increased MYC gene copy numbers (49%) as well as low-level chromosome 8 polysomy (6%). MYC aberrations in HIV PBLs were significantly associated with SS appearance (P -0.01), monomorphic morphology (P - 0.03), c-MYC protein expression ≥40% (P - 0.03) and mortality (P - 0.03). There was advanced stage (Ann Arbor III/IV) at presentation (77%) and the median overall survival for HIV PBL was 75 days (95% CI 14-136). CONCLUSION: Majority of the HIV-associated PBL tumours harbour MYC aberrations. Due to the persistently inferior survival outcome of HIV-associated PBL in the era of antiviral treatment, targeted and/or intensified therapy of oncogenic MYC may need to be explored in future.

Genetic Predisposition to the Mortality in Septic Shock Patients: From GWAS to the Identification of a Regulatory Variant Modulating the Activity of a CISH Enhancer.

The high mortality rate in septic shock patients is likely due to environmental and genetic factors, which influence the host response to infection. Two genome-wide association studies (GWAS) on 832 septic shock patients were performed. We used integrative bioinformatic approaches to annotate and prioritize the sepsis-associated single nucleotide polymorphisms (SNPs). An association of 139 SNPs with death based on a false discovery rate of 5% was detected. The most significant SNPs were within the CISH gene involved in cytokine regulation. Among the 139 SNPs associated with death and the 1311 SNPs in strong linkage disequilibrium with them, we investigated 1439 SNPs within non-coding regions to identify regulatory variants. The highest integrative weighted score (IW-score) was obtained for rs143356980, indicating that this SNP is a robust regulatory candidate. The rs143356980 region is located in a non-coding region close to the CISH gene. A CRISPR-Cas9-mediated deletion of this region and specific luciferase assays in K562 cells showed that rs143356980 modulates the enhancer activity in K562 cells. These analyses allowed us to identify several genes associated with death in patients with septic shock. They suggest that genetic variations in key genes, such as CISH, perturb relevant pathways, increasing the risk of death in sepsis patients.

TRPM1 (melastatin) expression is an independent predictor of overall survival in clinical AJCC stage I and II melanoma patients.

BACKGROUND: The expression of TRPM1 (melastatin) mRNA is an independent marker, as measured by radioactive in situ hybridization (RISH), of disease-free survival in primary cutaneous melanoma (PM). The aim of the study was to determine if chromogenic in situ hybridization (CISH) can reproduce results examining diagnostic and prognostic utility of TRPM1 mRNA expression in melanocytic proliferations as measured by RISH. METHODS: The expression of TRPM1 mRNA was detected by CISH in melanocytic nevi (MN, n = 61), PM (n = 145) and metastatic melanomas (MMs, n = 15). RESULTS: A progressive loss of TRPM1 was found moving from MN to PM to MM. The histologic stepwise model of melanoma progression revealed that loss of TRPM1 occurred at the transition of RGP PM to VGP PM. As a diagnostic marker, TRPM1 gradient loss showed 93.8% sensitivity and 52.4% specificity for PM. Loss of TRPM1 mRNA correlated with melanoma aggressiveness markers and was independent predictor of disease-free and overall survival. The corresponding survival curves for degree of melanoma pigmentation matched those for degree of loss of TPRM1 mRNA. CONCLUSION: Loss of TRPM1 mRNA expression appears to be a crucial event in the progression of melanoma to a more malignant, metastatic phenotype.

Characterization of porcine cytokine inducible SH2-containing protein gene and its association with piglet diarrhea traits.

OBJECTIVE: The cytokine inducible SH2-containing protein (CISH), which might play a role in porcine intestine immune responses, was one of the promising candidate genes for piglet anti-disease traits. An experiment was conducted to characterize the porcine CISH (pCISH) gene and to evaluate its genetic effects on pig anti-disease breeding. METHODS: Both reverse transcription polymerase chain reaction (RT-PCR) and PCR were performed to obtain the sequence of pCISH gene. A pEGFP-C1-CISH vector was constructed and transfected into PK-15 cells to analysis the distribution of pCISH. The sequences of individuals were compared with each other to find the polymorphisms in pCISH gene. The association analysis was performed in Min pigs and Landrace pigs to evaluate the genetic effects on piglet diarrhea traits. RESULTS: In the present research, the coding sequence and genomic sequence of pCISH gene was obtained. Porcine CISH was mainly localized in cytoplasm. TaqI and HaeIII PCR restriction fragment length polymorphism (RFLP) assays were established to detect single nucleotide polymorphisms (SNPs); A-1575G in promoter region and A2497C in Intron1, respectively. Association studies indicated that SNP A-1575G was significantly associated with diarrhea index of Min piglets (p<0.05) and SNP A2497C was significantly associated with the diarrhea trait of both Min pig and Landrace piglets (p<0.05). CONCLUSION: This study suggested that the pCISH gene might be a novel candidate gene for pig anti-disease traits, and further studies are needed to confirm the results of this preliminary research.

A Chromogenic In Situ Hybridization (CISH) Assay for Detection of HBV RNA, DNA, and cccDNA in Liver Tissue.

Hepatitis B virus (HBV) developed highly intricates mechanisms exploiting host resources for its multiplication within a constrained genetic coding capacity. With the aid of a series of classical analytical methods such as ultrafiltration, and Southern and Northern blots, a general framework of HBV life cycle has been established. However, this picture still lacks many key histological contexts which involves pathophysiological changes of hepatocytes, non-parenchymal cells, infiltrated leukocytes, and associated extracellular matrix. Here, we describe a CISH protocol modified from the ViewRNA assay that allows direct visualization of HBV RNA, DNA, and cccDNA in liver tissue of chronic hepatitis B patients. By coupling it with immunohistochemistry and other histological stains, much richer information regarding the HBV-induced pathological changes can be harvested.

Chromogenic LMO2 mRNA ISH Expression Correlates with LMO2 Protein and Gene Expression and Captures Their Survival Impact in Diffuse Large B-Cell Lymphoma, NOS.

BACKGROUND: LMO2 is a relevant gene involved in B-cell ontogeny and a survival predictor of aggressive large B-cell lymphomas (aLBCL). Most studies assessing LMO2 mRNA expression have relied on microarray platforms or qRT-PCR methods, overlooking tissue morphology. In this study, we evaluate LMO2 RNA expression by chromogenic in situ hybridization (CISH) in normal tissue and in a series of 82 aLBCL. METHODS: LMO2 CISH was performed in formalin-fixed paraffin-embedded tissues, scored by three different methods, and correlated with a transcriptome panel. RESULTS: We obtained statistically significant results correlating the methods of evaluation with LMO2 protein expression and gene expression results. Normal tonsil tissue showed high levels of LMO2, particularly within the light zone of the germinal center. Conversely, in aLBCL, a notable reduction in LMO2 expression was noted, remarkably in cases carrying MYC rearrangements. Furthermore, significant results were obtained through overall survival and Cox regression survival analysis, incorporating International Prognostic Index data alongside LMO2 expression levels. CONCLUSIONS: We show a reliable method to identify LMO2 mRNA expression by CISH, effectively capturing many of the reported biologic features of LMO2.

BMSCs-derived Exosome CISH Alleviates Myocardial Infarction by Inactivating the NF-κB Pathway to Stimulate Macrophage M2 Polarization.

Current myocardial infarction (MI) treatments are suboptimal, necessitating deeper pathogenesis understanding of MI. This research explored how exosomes (Exo) derived from bone marrow mesenchymal stem cells (BMSCs) contribute to MI mitigation and their therapeutic potential. Isolated BMSCs was identified by microscope, flow cytometry, alizarin red and oil red O staining. Exo were identified by TEM, NTA and western blot. HE staining, masson staining, and cardiac function parameters were used to assess the cardiac function in MI mice. TUNEL staining, western blot and qRT-PCR were used to detect apoptosis, inflammatory factors and M1/M2 markers. The NF-κB pathway activation was detected through western blot assays. Immunofluorescence, qRT-PCR, western blot, and flow cytometry were employed to evaluate macrophage polarization. MI mice showed cardiac injury, increased apoptosis and inflammation, while BMSCs-Exo treatment alleviated these effects. In MI mice, the macrophage M1 polarization was increased and the NF-κB pathway was activated, whereas BMSCs-Exo treatment reversed these changes. Furthermore, CISH expression was reduced in MI mice, but was elevated with BMSCs-Exo treatment. In vitro, LPS shifted RAW264.7 cells to M1 phenotype and activated the NF-κB pathway, yet BMSCs-Exo shifted them to M2 phenotype and inhibited the NF-κB pathway. Mechanistically, BMSCs-Exo induced macrophage M2 polarization by transmitting CISH to inhibit NF-κB activation. BMSCs-Exo mitigates MI by transmitting CISH to inhibit the NF-κB pathway, promoting macrophages to M2 type. This implies BMSCs-Exo could be a useful treatment for MI, and CISH could be a potential therapy target.

Enhancing the utility of chromosome 6 and 8 testing in uveal melanoma biopsies.

BACKGROUND: The aim of this study was to evaluate the significance of testing the gain of chromosome 8 and the gain of chromosome 6 as prognostic markers in histopathological samples of enucleated eyes in with uveal melanoma. METHODS: This is a retrospective study of 54 enucleated eyes. The status of chromosomes 3, 8 and 6 was tested by CISH, and FISH was used in a few samples. A follow-up for the detection of metastases was conducted in all patients. The statistical significance of chromosomal abnormalities as a prognostic factor for the development of metastases was determined. RESULTS: The study group consists of 54 patients (average age 63 years), 28 men (51.9%) Monosomy 3 together with gain of chromosome 8 was found in 10 samples (18.5%). Both chromosomal abnormalities were detected in 6 (11%) patients. No chromosomal abnormality in 3 or 8 was detected in 21 (38.9%) patients. Abnormalities of chromosome 6 were present in 6 (11%) patients. Progression free survival after 5 years was 33.3% (95% CI 0.0; 83.3) in these patients. CONCLUSIONS: Our findings indicate a correlation between progression-free survival and the presence of changes in chromosome 3 and e 8 in uveal melanomas. The results underline the necessity of testing for both chromosomal aberrations.

CSF1 expression in xanthogranulomatous epithelial tumor/keratin-positive giant cell-rich tumor.

"Xanthogranulomatous epithelial tumor" (XGET) and "keratin-positive giant cell-rich soft tissue tumor" (KPGCT), two recently described mesenchymal neoplasms, likely represent different aspects of a single entity. Both tumors are composed of only a small minority of tumor cells surrounded by large numbers of non-neoplastic inflammatory cells and histiocytes, suggesting production of a paracrine factor with resulting "landscape effect," as seen in tenosynovial giant cell tumor. Recent evidence suggests that the paracrine factor in XGET/KPGCT may be CSF1, as in tenosynovial giant cell tumor. We hypothesized that CSF1 is overexpressed in XGET/KPGCT. To test our hypothesis, we performed quantitative real time PCR (qPCR) for CSF1 expression and CSF1 RNAscope chromogenic in situ hybridization (CISH) on 6 cases of XGET/KPGCT. All cases were positive with CSF1 CISH and showed increased expression of CSF1 by qPCR. Our findings provide additional evidence that the CSF1/CSF1R pathway is involved in the pathogenesis of XGET/KPGCT. These findings suggest a possible role for CSF1R inhibition in the treatment of unresectable or metastatic XGET/KPGCT.

Methylation profiling of SOCS1, SOCS2, SOCS3, CISH and SHP1 in Philadelphia-negative myeloproliferative neoplasm.

Janus kinase-signal transducer and activator of transcription (JAK/STAT) signalling, pivotal in Philadelphia-negative (Ph-ve) myeloproliferative neoplasm (MPN), is negatively regulated by molecules including SOCSs, CISH and SHP1. SOCS1, SOCS2 and SOCS3 methylation have been studied in MPN with discordant results. Herein, we studied the methylation status of SOCS1, SOCS2 and SOCS3, CISH and SHP1 by methylation-specific polymerase chain reaction (MSP) in cell lines and 45 diagnostic marrow samples of Ph-ve MPN. Moreover, we attempted to explain the discordance of methylation frequency by mapping the studied MSP primers to the respective genes. Methylation was detected in normal controls using SOCS2 MSP primers in the 3'translated exonic sequence, but not primers around the transcription start site in the 5' untranslated regions (5'UTR). SOCS1, SOCS2, SOCS3 and CISH were completely unmethylated in primary MPN samples and cell lines. In contrast, methylation of SHP1 was detected in 8.9% primary marrow samples. Moreover, SHP1 was completely methylated in K562 cell line, leading to reversible SHP1 silencing. A review of methylation studies of SOCS1 and SOCS3 showed that spuriously high rates of SOCS methylation had been reported using MSP primers targeting CpG sites in the 3'translated exonic sequence, which is also methylated in normal controls. However, using MSP primers localized to the 5'UTR, methylation of SOCS1, SOCS2 and SOCS3 is infrequent across all studies. In summary, methylation of SOCS1, SOCS2, SOCS3 and CISH is infrequent in Ph-ve MPN. Appropriate MSP primers are important for accurate estimation of the methylation frequency. The role of SHP1 methylation in the pathogenesis of MPN warrants further investigation.

Role of Cytokine-Inducible SH2 Domain-Containing (CISH) Protein in the Regulation of Erythropoiesis.

The cytokine-inducible SH2 domain-containing (CISH) protein was the first member of the suppressor of cytokine signaling (SOCS) family of negative feedback regulators discovered, being identified in vitro as an inducible inhibitor of erythropoietin (EPO) signaling. However, understanding of the physiological role played by CISH in erythropoiesis has remained limited. To directly assess the function of CISH in this context, mice deficient in CISH were characterized with respect to developmental, steady-state, and EPO-induced erythropoiesis. CISH was strongly expressed in the fetal liver, but CISH knockout (KO) mice showed only minor disruption of primitive erythropoiesis. However, adults exhibited mild macrocytic anemia coincident with subtle perturbation particularly of bone marrow erythropoiesis, with EPO-induced erythropoiesis blunted in the bone marrow of KO mice but enhanced in the spleen. Cish was expressed basally in the bone marrow with induction following EPO stimulation in bone marrow and spleen. Overall, this study indicates that CISH participates in the control of both basal and EPO-induced erythropoiesis in vivo.

Immunohistochemical Detection of the Expressed BRCA1 and BRCA2 Proteins in Microenvironment of Malignant Breast Cancerous Tissues Infected with Human Mammary Tumor Virus.

OBJECTIVE: The objective of the study was to examine the prevalence of HMTV infection, its associations with breast malignant tissues, and the expression of BRCA1 and BRCA2 proteins. METHODS: One hundred archival breast tissues, 40 biopsies from female patients with breast cancer (BC), and 20 healthy breast tissues from the control group were used in the study. Immunohistochemical analysis was conducted to detect the expressed BRCA1 and BRCA2 proteins. Digoxigenin-labeled HMTV probes were used in chromogenic in situ hybridization for the identification of HMTV in breast tumor tissues. The complementary sequence sites of the HMTV probe sequences were stained by NBT/BCIP as blue signals. RESULTS: There were 12 out of 40 (30%) benign breast tumorous tissues and 14 out of 40 (35%) BC tissues, while healthy control breast tissues were 10% (2 out of 20 tissues). Positive immunohistochemical (IHC) reactions for BRCA2 protein were observed in 12 out of 40 BC tissues (30.0%), 25% of benign breast tumorous tissues, and 5% of the control group. A significant (p < 0.05) statistical difference in the percentages of HMTV in the studied groups was found. CONCLUSION: HMTV might contribute to the development of subsets of benign and malignant breast tumors. The observed rates of defective or mutated BRCA1 and BRCA2 genes in healthy tissues indicate a role in the development of breast tumors.

Expression of CISH, an Inhibitor of NK Cell Function, Increases in Association with Ovarian Cancer Development and Progression.

Epithelial ovarian cancer (OVCA), a fatal malignancy of women, disseminates locally. Although NK cells mount immune responses against OVCA, tumors inhibit NK cells, and the mechanism is not well understood. Cytokines stimulate NK cells; however, chronic stimulation exhausts them and induces expression of cytokine-inducible SH2-containing protein (CISH). Tumors produce anti-inflammatory cytokine interleukin (IL)-10 which may induce NK cell exhaustion. The goal of this study was to examine if CISH expression in NK cells increases during OVCA development and to determine the mechanism(s) of OVCA-induced CISH expression in NK cells. Normal ovaries (n = 7) were used for CISH, IL-10 and GRP78 expression. In tumor ovaries, CISH was examined in early and late stages (n = 14 each, all subtypes) while IL-10 and GRP78 expression were examined in early and late stage HGSC (n = 5 each). Compared to normal, the population of CISH-expressing NK cells increased and the intensity of IL-10 and GRP78 expression was significantly higher in OVCA (p < 0.05). CISH expression was positively correlated with IL-10 expression (r = 0.52, r = 0.65, p < 0.05 at early and late stages, respectively) while IL-10 expression was positively correlated with GRP78 expression (r = 0.43, r = 0.52, p < 0.05, respectively). These results suggest that OVCA development and progression are associated with increased CISH expression by NK cells which is correlated with tumor-induced persistent cellular stress.