A comparative Proteomics Analysis Identified Differentially Expressed Proteins in Pancreatic Cancer–Associated Stellate Cell Small Extracellular Vesicles

Human pancreatic stellate cells (HPSCs) are an essential stromal component and mediators of pancreatic ductal adenocarcinoma (PDAC) progression. Small extracellular vesicles (sEVs) are membrane-enclosed nanoparticles involved in cell-to-cell communications and are released from stromal cells within PDAC. A detailed comparison of sEVs from normal pancreatic stellate cells (HPaStec) and from PDAC-associated stellate cells (HPSCs) remains a gap in our current knowledge regarding stellate cells and PDAC. We hypothesized there would be differences in sEVs secretion and protein expression that might contribute to PDAC biology. To test this hypothesis, we isolated sEVs using ultracentrifugation followed by characterization by electron microscopy and Nanoparticle Tracking Analysis. We report here our initial observations. First, HPSC cells derived from PDAC tumors secrete a higher volume of sEVs when compared to normal pancreatic stellate cells (HPaStec). Although our data revealed that both normal and tumor-derived sEVs demonstrated no significant biological effect on cancer cells, we observed efficient uptake of sEVs by both normal and cancer epithelial cells. Additionally, intact membrane-associated proteins on sEVs were essential for efficient uptake. We then compared sEV proteins isolated from HPSCs and HPaStecs cells using liquid chromatography–tandem mass spectrometry. Most of the 1481 protein groups identified were shared with the exosome database, ExoCarta. Eighty-seven protein groups were differentially expressed (selected by 2-fold difference and adjusted p value ≤0.05) between HPSC and HPaStec sEVs. Of note, HPSC sEVs contained dramatically more CSE1L (chromosome segregation 1–like protein), a described marker of poor prognosis in patients with pancreatic cancer. Based on our results, we have demonstrated unique populations of sEVs originating from stromal cells with PDAC and suggest that these are significant to cancer biology. Further studies should be undertaken to gain a deeper understanding that could drive novel therapy.     

LncRNA TP73-AS1 enhances the malignant properties of pancreatic ductal adenocarcinoma by increasing MMP14 expression through miRNA -200a sponging

Pancreatic ductal adenocarcinoma (PDAC) is an invasive and aggressive cancer that remains a major threat to human health across the globe. Despite advances in cancer treatments and diagnosis, the prognosis of PDAC patients remains poor. New and more effective PDAC therapies are therefore urgently required. In this study, we identified a novel host factor, namely the LncRNA TP73-AS1, as overexpressed in PDAC tissues compared to adjacent healthy tissue samples. The overexpression of TP-73-AS1 was found to correlate with both PDAC stage and lymph node metastasis. To reveal its role in PDCA, we targeted TP73-AS1 using LnRNA inhibitors in a range of pancreatic cancer (PC) cell lines. We found that the inhibition of TP73-AS1 led to a loss of MMP14 expression in PC cells and significantly inhibited their migratory and invasive capacity. No effects of TP73-AS1 on cell survival or proliferation were observed. Mechanistically, we found that TP73-AS1 suppressed the expression of the known oncogenic miR-200a. Taken together, these data highlight the prognostic potential of TP73-AS1 for PC patients and highlight it as a potential anti-PDAC therapeutic target.     

Establishment and characterization of a new,spontaneously immortalized,pancreatic ductal cell line from the Syrian golden hamster

Spontaneously immortal pancreatic cell lines are not available. By use of a defined culture medium, such a line (TAKA-1) was established from the Syrian golden hamster. Cytological, cytogenetic, molecular biological, enzymatic and receptor patterns as well as antigenicity were studied and were compared with those of the normal hamster pancreatic ductal cells in vivo. TAKA-1 cells grew exponentially in a monolayer on collagen gel in a defined medium but did not proliferate in soft agar. Ultrastructurally, the cells closely resembled the normal hamster pancreatic ductal cells. Similarities and dissimilarities were found between the normal ductal cells and TAKA-1 cells. Similarities included the presence of cytokeratin, carbonic anhydrase and some tumor-associated antigens. However, unlike the normal ductal cells, TAKA-1 cells expressed blood group A angigen and anti-vimentin, showed affinity to selected lectins, and an abnormality of chromosome 3, which is suggested to be associated with immortality. Moreover, unlike the hamster pancreatic ductal cancer cells but like the normal hamster pancreatic ductal cells, TAKA-1 cells did not have a c-Ki-ras mutation. EGF, TGF- and secretin, but not CCK or GRP, bound to the TAKA-1 cells. TAKA-1 cells produced TGF-, and their growth was stimulated by exogenous EGF in serum-free medium. This cell line presents a suitable model for biologic and pathologic study of the hamster pancreatic ductal cells in vitro.     

FXR在胃粘膜肠化生及胃癌中的表达及其意义研究

目的:研究FXR在胃炎,胃粘膜肠化生及胃癌组织中的表达,分析其在胃癌发生中的意义。方法:采用免疫组化方法检测FXR在55例胃炎组织,61例胃黏膜肠化生组织及61例胃癌组织中的表达,利用统计学方法 SPSS17.0软件分析其在三种组织中的表达变化,结合文献回顾,分析FXR在胃癌发生中的意义。结果:FXR在胃黏膜肠化生中的表达明显高于胃炎组织(P0.05),而在胃癌组织中,FXR的表达显著低于胃粘膜肠化生组织(P0.05)。结论:FXR是一个潜在的胃癌发生生物标记物,其具体机制有待于进一步探索。     

The intact parasympathetic nerve promotes submandibular gland regeneration through ductal cell proliferation

ObjectivesSalivary gland regeneration is closely related to the parasympathetic nerve; however, the mechanism behind this relationship is still unclear. The aim of this study was to evaluate the relationship between the parasympathetic nerve and morphological differences during salivary gland regeneration.Materials and MethodsWe used a duct ligation/deligation‐induced submandibular gland regeneration model of Sprague‐Dawley (SD) rats. The regenerated submandibular gland with or without chorda lingual (CL) innervation was detected by haematoxylin–eosin staining, real‐time PCR (RT‐PCR), immunohistochemistry and Western blotting. We counted the number of Ki67‐positive cells to reveal the proliferation process that occurs during gland regeneration. Finally, we examined the expression of the following markers: aquaporin 5, cytokeratin 7, neural cell adhesion molecule (NCAM) and polysialyltransferases.ResultsIntact parasympathetic innervation promoted submandibular gland regeneration. The process of gland regeneration was significantly repressed by cutting off the CL nerve. During gland regeneration, Ki67‐positive cells were mainly found in the ductal structures. Moreover, the expression of NCAM and polysialyltransferases‐1 (PST) expression in the innervation group was significantly increased during early regeneration and decreased in the late stages. In the denervated submandibular glands, the expression of NCAM decreased during regeneration.ConclusionsOur findings revealed that the regeneration of submandibular glands with intact parasympathetic innervation was associated with duct cell proliferation and the increased expression of PST and NCAM.     

Development and validation of glycolysis-related prognostic score for prediction of prognosis and chemosensitivity of pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy with aggressive biological behaviour. Its rapid proliferation and tumour growth require reprogramming of glucose metabolism or the Warburg effect. However, the association between glycolysis-related genes with clinical features and prognosis of PDAC is still unknown. Here, we used the meta-analysis to correlate the hazard ratios (HR) of 106 glycolysis genes from MSigDB by the cox proportional hazards regression analysis in 6 clinical data sets of PDAC patients to form a training cohort, and a single group of PDAC patients from the TCGA, ICGC, Arrayexpress and GEO databases to form the validation cohort. Then, a glycolysis-related prognosis (GRP) score based on 29 glycolysis prognostic genes was established in 757 PDAC patients from the training composite cohort and validated in 267 ICGC-CA validation cohort (all P < .05). In addition, including PADC, the prognostic value was also confirmed in other 7 out of 30 pan-cancer cohorts. The GRP score was significantly related to specific metabolism pathways, immune genes and immune cells in the patients with PADC (all P < .05). Finally, by combining with immune cells, the GRP score also well-predicted the chemosensitivity of patients with PADC in the TCGA cohort (AUC = 0.709). In conclusion, this study developed a GRP score for patients with PDAC in predicting prognosis and chemosensitivity for PDAC.     

Cell-to-cell interaction analysis of prognostic ligand-receptor pairs in human pancreatic ductal adenocarcinoma

Cell-to-cell interactions (CCIs) through ligand-receptor (LR) pairs in the tumor microenvironment underlie the poor prognosis of pancreatic ductal adenocarcinoma (PDAC). However, there is scant knowledge of the association of CCIs with PDAC prognosis, which is critical to the identification of potential therapeutic candidates. Here, we sought to identify the LR pairs associated with PDAC patient prognosis by integrating survival analysis and single-cell CCI prediction. Via survival analysis using gene expression from cancer cohorts, we found 199 prognostic LR pairs. CCI prediction based on single-cell RNA-seq data revealed the enriched LR pairs associated with poor prognosis. Notably, the CCIs involved epithelial tumor cells, cancer-associated fibroblasts, and tumor-associated macrophages through integrin-related and ANXA1–FPR pairs. Finally, we determined that CCIs involving 33 poor-prognostic LR pairs were associated with tumor grade. Although the clinical implication of the set of LR pairs must be determined, our results may provide potential therapeutic targets in PDAC.     

Intratumor heterogeneity of biomarker expression in breast carcinomas

Small biopsy samples are used increasingly to assess the biomarker expression for prognostic information and for monitoring therapeutic responses prior to and during neoadjuvant therapy. The issue of intratumor heterogeneity of expression of biomarkers, however, has raised questions about the validity of the assessment of biomarker expression based on limited tissue samples. We examined immunohistochemically the expression of HER-2neu (p185^erbB-2), epidermal growth factor receptor (EGFR), Bcl-2, p53, and proliferating cell nuclear antigen (PCNA) in 30 breast carcinomas using archived, paraffin embedded tissue and determined the extent of intratumor heterogeneity. Each section was divided into four randomly oriented discrete regions, each containing a portion of the infiltrating carcinoma. For each tumor, the entire lesion and four regions were analyzed for the expression of these markers. Scores of both membrane and cytoplasmic staining of HER-2neu and EGFR, scores of cytoplasmic staining of Bcl-2, and scores of nuclear staining of both p53 and PCNA were recorded. The intensity of staining and the proportion of immunostained cells were determined. A semiquantitative immunoscore was calculated by determining the sum of the products of the intensity and corresponding proportion of stained tumor cells. We analyzed both invasive (IDC) and in situ (DCIS) carcinomas. The Wilcoxon signed-rank test was used for paired comparisons between overall and regional immunoscores and between overall and regional percentages of stained cells. Spearman's correlation coefficients were used to assess the level of agreement of overall biomarker expression with each of the regions. Generalized linear models were used to assess overall and pair-wise differences in the absolute values of percent changes between overall and regional expression of biomarkers. For IDCs, there were no statistically significant differences in the expression of the biomarkers in terms of either the percentage of cells staining or the immunoscores when comparing the entire tumor with each region except for the lower EGFR expression of arbitrarily selected region 1 and lower p53 expression of region 1 compared to that of the entire tumor section. For DCIS, there were no statistically significant differences in the expression of the biomarkers between the entire tumor and each region except in PCNA of region 2 compared to that of entire tumor section. Positive correlation of immunoscores was observed between the entire tumor and each region as well as across all four regions for IDC. Similar observations were noted with DCIS except for HER-2neu and PCNA. No statistically significant differences were observed in the absolute values of percent changes of biomarker expression between overall and the four regions for both DCIS and IDC. Therefore, no significant intratumor heterogeneity in the expression of HER-2neu, Bcl-2, and PCNA was observed in IDC. Minor regional variations were observed for EGFR and p53 in IDC. Similarly, no significant regional variation in the expression of markers was observed in DCIS except for PCNA.