Infiammazione e cancro: l'interazione COX-2/CA-IX promuove il potenziale invasivo e la sopravvivenza in ipossia delle cellule del cancro colon-rettale

Cyclooxygenase-2/Carbonic anhydrase-IX up-regulation promotes invasive potential and hypoxia survival in colorectal cancer cells Purpose: Cyclooxygenase-2 (COX-2) is a major mediator of inflammation, playing a pivotal role in colorectal carcinogenesis. Hypoxia is an universal hallmark of solid tumour in vivo. This investigation was prompted by the observation that in colorectal cancer cells the expression of COX-2 protein is positively correlated with that of the hypoxia survival gene Carbonic Anhydrase-IX (CA-IX). Experimental Design: Since COX-2 gene expression and activity is increased in hypoxia, and that CA-IX is expressed also in normoxia in colorectal cancer cells, we tested the hypothesis that COX-2 activity in normoxia, as well as in hypoxia may be functionally linked to that of CA-IX gene. We investigated the role of COX-2 and CA-IX in colorectal cancer cell lines. In this regard, we performed RNA interference to knockdown COX-2 gene in vitro and immunohistochemistry to evaluate the protein expression of COX-2 and CA-IX in human colon cancer tissue specimens ex vivo. Results: We found that COX-2, by PGE2 production, controls CA-IX gene expression in an ERK dependent manner. In line with this finding, we also showed that the COX-2 inhibition by a specific short harpin COX-2 RNA (shCOX-2) or by a specific drug (SC-236), down-regulated CA-IX expression in colon cancer cells. We then exposed colon cancer cells to hypoxia stimuli and found that COX-2/CA-IX interplay promoted hypoxia survival. Moreover, we also report that COX-2/CA-IX interplay triggers Matrix Metalloproteinase 2/9 (MMP-2/9) activation and enhances the invasiveness of colorectal cancer cells. Thus given our above observations, we found that CA-IX and COX-2 protein expressions correlate with more aggressive stage colorectal cancer tissues ex vivo. Conclusions: Taken together these data indicate that COX-2/CA-IX interplay promotes an aggressive phenotype (hypoxia survival and invasiveness) which can be modulated in vitro by COX-2 selective inhibition and which may play a role in determining the biological aggressiveness of colorectal tumours. Moreover, in vitro and ex vivo data also suggest that the signatures of inflammation (COX-2) and hypoxia (CA-IX) may be difficult to be disentangled in colon cancer, being both responsible for the up-regulation of the same pathways.

Diabete ed ischemia critica degli arti inferiori. Studio del danno della parete arteriosa e del ruolo dell'endotelio circolante nella riparazione tessutale e nella neoangiogenesi

Objectives In diabetic and non diabetic patients with peripheral artery obstructive disease (PAOD), we sought to establish whether the vascular wall damage, the mature circulating endothelium and the "in situ" neoangiogenesis are related with each other. Design In the peripheral blood of diabetic patients suffering critical ischaemia associated with peripheral artery disease, low levels and poor function of circulating endothelial progenitor cells (EPCs) were observed. Moreover, circulating endothelial cells (CECs) have been described in different conditions of vascular injury. In this type of disorders, which are all characterized by endothelial damage, neoangiogenesis plays a key role. Materials In the study we recruited 22 diabetic and 16 non diabetic patients, all of them suffering PAOD and critical ischaemia; healthy subjects and multiorgan donors have also been considered like controls. Methods Histopathologic characterization was performed on arterial tissue samples under a light microscope. Flow cytofluorimetric analysis was used to quantify CECs in peripheral blood samples. "In situ" expression of the Vascular Endothelial Growth Factor (VEGF) and Metalloproteinase 9 (MMP-9) transcripts was quantified in a Real Time-PCR analysis. Circulating VEGF concentration was determined by an ELISA assay. Results Arterial wall from diabetic patients, compared with non diabetic subjects, revealed a higher incidence of serious lesions (60% vs 47%) and a lower number of capillaries (65% vs 87%). Mean number of CECs/ml was significantly increased in all patients, compared to healthy controls (p=0.001). Compared to healthy subjects, VEGF transcripts expression resulted significantly higher in diabetic patients and in all patients (p<0.05) and a similar result was obtained in the MMP-9 transcripts expression. Serum VEGF concentration was significantly increased in PAOD patients correlated with controls (p=0.0431). Conclusions Our study demonstrates that in all patients considered, probably, regressive phenomenons prevail on reparative ones, causing an inesorable and progressive degeneration of the vascular wall, worse by diabetes. The vascular damage can be monitored by determining CECs number and its severity and development are emphasized by the MMP-9 transcripts expression. The "in situ" VEGF increased expression seems to be the evidence of a parietal cells bid to induce local angiogenesis. This reparing mechanism could induce the EPCs mobilitation by means the release of VEGF from the arterial wall. The mechanism, however, is ineffective like demonstrated by the EPCs reduced number and activities observed in patients suffering PAOD and critical ischaemia.

Developing a radiomic based machine learning model to identify patients with resected non-small-cell lung cancer at high risk of relapse

Background There is a wide variation of recurrence risk of Non-small-cell lung cancer (NSCLC) within the same Tumor Node Metastasis (TNM) stage, suggesting that other parameters are involved in determining this probability. Radiomics allows extraction of quantitative information from images that can be used for clinical purposes. The primary objective of this study is to develop a radiomic prognostic model that predicts a 3 year disease free-survival (DFS) of resected Early Stage (ES) NSCLC patients. Material and Methods 56 pre-surgery non contrast Computed Tomography (CT) scans were retrieved from the PACS of our institution and anonymized. Then they were automatically segmented with an open access deep learning pipeline and reviewed by an experienced radiologist to obtain 3D masks of the NSCLC. Images and masks underwent to resampling normalization and discretization. From the masks hundreds Radiomic Features (RF) were extracted using Py-Radiomics. Hence, RF were reduced to select the most representative features. The remaining RF were used in combination with Clinical parameters to build a DFS prediction model using Leave-one-out cross-validation (LOOCV) with Random Forest. Results and Conclusion A poor agreement between the radiologist and the automatic segmentation algorithm (DICE score of 0.37) was found. Therefore, another experienced radiologist manually segmented the lesions and only stable and reproducible RF were kept. 50 RF demonstrated a high correlation with the DFS but only one was confirmed when clinicopathological covariates were added: Busyness a Neighbouring Gray Tone Difference Matrix (HR 9.610). 16 clinical variables (which comprised TNM) were used to build the LOOCV model demonstrating a higher Area Under the Curve (AUC) when RF were included in the analysis (0.67 vs 0.60) but the difference was not statistically significant (p=0,5147).

Valutazione dell'attività delle metalloproteinasi 2 e 9 nelle urine e nel tessuto renale di cani normali e affetti da nefropatia

Matrix metalloproteinases (MMP) are a large family of proteinases that remodel extracellular matrix (ECM) component. Recent data suggest a role for MMPs in a number of renal pathophysiologies, associated with an imbalance of ECM syntesis and degradation, which may result in an accumulation of ECM molecules and renal fibrosis. The aim of this study is to elucidate the role of pro and activated MMP-2 and 9 in urine and renal tissue of healty and nephropatic dogs. Renal tissue of 8 healty dogs and either renal tissue and urine of 9 nephropatic dogs was collected and analize using zimographic method, which is been validated in this study. Either MMPs zimographic bands were present in almost all samples. In particular, pro and activated MMP-9 zimographic bands were poorly represent in renal tissue of healty dogs, whereas were very represent in nephropatic dogs. Pro and activated MMP-2 was present in either tissue of healty and nephropatic dogs. In urine of nephropatic dogs, pro and activated MMP-9 was more evident than MMP-2, but there was not correlaction with renal tissue levels, therefore urine levels of MMPs have poorly usefulness in diagnostic pratice. The values of Pro and activated MMP-9 in nephropatic dogs were significantly higher compared with normal dogs (p < 0,05), whereas there was not statistically meaningful for Pro and activated MMP-2. In conclusion, in this study we have validated a zimographic method for renal tissue of dogs and we have illustrated the changes in nephropatic dogs, which may be useful for further study.