Probes for INS

In the present study, we evaluated expression of IFN-γ, IL-17, TNF-α, IL-10 and TGF-β by mucosal cells, including WC1+ γδ T cells, in ileal tissues taken from non-infected cattle and cattle naturally infected with Mycobacterium avium subsp paratuberculosis (MAP). Infected cattle were either in the subclinical or clinical stage of infection. We hypothesized that the cytokine profile of the WC1+ γδ T cell subset would be different between subclinical and clinical cattle. Our data indicate a significant increase in the numbers of WC1+ γδ T cells expressing IL-10 in clinical cattle compared to subclinical and non-infected cattle. We observed a significant increase in TGF-β expression by non-WC1+ cells in clinically infected cattle. Expression of IFN-γ, IL-17 and TNF-α in mucosal cells, including the WC1+ γδ T cell subset, was identified in all examined groups. However, our data indicate that the stage of infection did not significantly influence expression of these proinflammatory cytokines. This study demonstrates changes in the cytokine mRNA expression profile of mucosal cells in the ileum, and specifically WC1+ γδ T cells, as cattle progress to the clinical disease. The change is characterized by an increase in expression of anti-inflammatory cytokines.

Inflammatory cytokines, like tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), are elevated in ovarian cancer. Differences in cytokine expression by histologic subytpe or ovarian cancer risk factors can provide useful insight into ovarian cancer risk and etiology. We used ribonucleic acid (RNA) in-situ hybridization to assess TNF-α and IL-6 expression on tissue microarray slides from 78 epithelial ovarian carcinomas (51 serous, 12 endometrioid, 7 clear cell, 2 mucinous, 6 other) from a population-based case control study. Cytokine expression was scored semi-quantitatively and odds ratios (OR) and 95% confidence intervals (CI) were calculated using polytomous logistic regression. TNF-α was expressed in 46% of the tumors while sparse IL-6 expression was seen only 18% of the tumors. For both markers, expression was most common in high grade serous carcinomas followed by endometrioid carcinomas. Parity was associated with a reduced risk of TNF-α positive (OR = 0.3, 95% CI: 0.1-0.7 for 3 or more children versus none) but not TNF-α negative tumors (p-heterogeneity = 0.02). In contrast, current smoking was associated with a nearly three fold increase in risk of TNF-α negative (OR = 2.8, 95% CI: 1.2, 6.6) but not TNF-α positive tumors (p-heterogeneity = 0.06). Our data suggests that TNF-α expression in ovarian carcinoma varies by histologic subtype and provides some support for the role of inflammation in ovarian carcinogenesis. The novel associations detected in our study need to be validated in a larger cohort of patients in future studies.

MicroRNA-21 (miR-21) is upregulated in many cancers including colon cancers and is a prognostic indicator of recurrence and poor prognosis. In colon cancers, miR-21 is highly expressed in stromal fibroblastic cells and more weakly in a subset of cancer cells, particularly budding cancer cells. Exploration of the expression of inflammatory markers in colon cancers revealed tumor necrosis factor alpha (TNF-α) mRNA expression at the invasive front of colon cancers. Surprisingly, a majority of the TNF-α mRNA expressing cells were found to be cancer cells and not inflammatory cells. Because miR-21 is positively involved in cell survival and TNF-α promotes necrosis, we found it interesting to analyze the presence of miR-21 in areas of TNF-α mRNA expression at the invasive front of colon cancers. For this purpose, we developed an automated procedure for the co-staining of miR-21, TNF-α mRNA and the cancer cell marker cytokeratin based on analysis of frozen colon cancer tissue samples (n = 4) with evident cancer cell budding. In all four cases, TNF-α mRNA was seen in a small subset of cancer cells at the invasive front. Evaluation of miR-21 and TNF-α mRNA expression was performed on digital slides obtained by confocal slide scanning microscopy. Both co-expression and lack of co-expression with miR-21 in the budding cancer cells was noted, suggesting non-correlated expression. miR-21 was more often seen in cancer cells than TNF-α mRNA. In conclusion, we report that miR-21 is not linked to expression of the pro-inflammatory cytokine TNF-α mRNA, but that miR-21 and TNF-α both take part in the cancer expansion at the invasive front of colon cancers. We hypothesize that miR-21 may protect both fibroblasts and cancer cells from cell death directed by TNF-α paracrine and autocrine activity.

The cytokine interleukin-1β (IL-1β) is known to stimulate proinflammatory immune responses and impair β-cell function and viability, all critical events in the pathogenesis of type 1 diabetes (T1D). Here we evaluate the effect of SER140, a small peptide IL-1β receptor antagonist, on diabetes progression and cellular pancreatic changes in female nonobese diabetic (NOD) mice. Eight weeks of treatment with SER140 reduced the incidence of diabetes by more than 50% compared with vehicle, decreased blood glucose, and increased plasma insulin. Additionally, SER140 changed the endocrine and immune cells dynamics in the NOD mouse pancreas. Together, the data suggest that SER140 treatment postpones the onset of diabetes in female NOD mice by interfering with IL-1β activated pathways.

Human papillomavirus (HPV) infection is associated with several genetic alterations including oncogene amplification, leading to increased aggression of tumors. Recently, a relationship between HPV infection and oncogene amplification has been reported, but this finding remains controversial. This study therefore investigated relationships between HPV infection and amplification of genes in the epidermal growth factor receptor (EGFR) signaling cascade in oral squamous cell carcinoma (OSCC). Extracted DNA from 142 formalin-fixed paraffin-embedded (FFPE) OSCC tissues was performed to investigate the copy number of EGFR, KRAS, c-myc and cyclin D1 genes using real-time polymerase chain reaction (RT-PCR) and compared with calibrators. A tissue microarray of OSCC tissues was used for detection of c-Myc expression and HPV infection by immunohistochemistry and HPV E6/E7 RNA in situ hybridization, respectively. HPV infection was also investigated using PCR and RT-PCR. Of the 142 OSCC samples, 81 (57%) were HPV-infected cases. The most frequently amplified gene was c-myc (55.6%), followed by cyclin D1 (26.1%), EGFR (23.9%) and KRAS (19.7%). Amplification of c-myc was significantly associated with levels of its protein product. EGFR amplification was also significantly associated with amplification of genes in the signaling cascade: KRAS (50.0%), c-myc (34.2%) and cyclin D1 (46.0%). Interestingly, HPV infection was significantly associated with amplification of both EGFR (76.5%) and cyclin D1 (73.0%). Only cyclin D1 amplification was significantly associated with severity of OSCC histopathology. HPV infection may play an important synergistic role in amplification of genes in the EGFR signaling cascade, leading to increased aggression in oral malignancies.