Chronic hyperglycemia induces trans-differentiation of human pancreatic stellate cells and enhances the malignant molecular communication with human pancreatic cancer cells

BACKGROUND: Diabetes mellitus is linked to pancreatic cancer. We hypothesized a role for pancreatic stellate cells (PSC) in the hyperglycemia induced deterioration of pancreatic cancer and therefore studied two human cell lines (RLT-PSC, T3M4) in hyperglycemic environment. METHODOLOGY/PRINCIPAL FINDINGS: The effect of chronic hyperglycemia (CHG) on PSCs was studied using mRNA expression array with real-time PCR validation and bioinformatic pathway analysis, and confirmatory protein studies. The stress fiber formation (IC: αSMA) indicated that PSCs tend to transdifferentiate to a myofibroblast-like state after exposure to CHG. The phosphorylation of p38 and ERK1/2 was increased with a consecutive upregulation of CDC25, SP1, cFOS and p21, and with downregulation of PPARγ after PSCs were exposed to chronic hyperglycemia. CXCL12 levels increased significantly in PSC supernatant after CHG exposure independently from TGF-β1 treatment (3.09-fold with a 2.73-fold without TGF-β1, p<0.05). The upregualtion of the SP1 transcription factor in PSCs after CHG exposure may be implicated in the increased CXCL12 and IGFBP2 production. In cancer cells, hyperglycemia induced an increased expression of CXCR4, a CXCL12 receptor that was also induced by PSC's conditioned medium. The receptor-ligand interaction increased the phosphorylation of ERK1/2 and p38 resulting in activation of MAP kinase pathway, one of the most powerful stimuli for cell proliferation. Certainly, conditioned medium of PSC increased pancreatic cancer cell proliferation and this effect could be partially inhibited by a CXCR4 inhibitor. As the PSC conditioned medium (normal glucose concentration) increased the ERK1/2 and p38 phosphorylation, we concluded that PSCs produce other factor(s) that influence(s) pancreatic cancer behaviour. CONCLUSIONS: Hyperglycemia induces increased CXCL12 production by the PSCs, and its receptor, CXCR4 on cancer cells. The ligand-receptor interaction activates MAP kinase signaling that causes increased cancer cell proliferation and migration

Outcome analysis and risk factors for postoperative colonic ischaemia after aortic surgery

Purpose!#!Colonic ischaemia (CI) represents a serious complication after aortic surgery. This study aimed to analyse risk factors and outcome of patients suffering from postoperative CI.!##!Methods!#!Data of 1404 patients who underwent aortic surgery were retrospectively analysed regarding CI occurrence. Co-morbidities, procedural parameters, colon blood supply, procedure-related morbidity and mortality as well as survival during follow-up (FU) were compared with patients without CI using matched-pair analysis (1:3).!##!Results!#!Thirty-five patients (2.4%) with CI were identified. Cardiovascular, pulmonary and renal comorbidity were more common in CI patients. Operation time was longer (283 ± 22 vs. 188 ± 7 min, p &amp;lt; 0.0001) and blood loss was higher (2174 ± 396 vs. 1319 ± 108 ml, p = 0.0049) in the CI group. Patients with ruptured abdominal aortic aneurysm (AAA) showed a higher rate of CI compared to patients with intact AAA (5.4 vs. 1.9%, p = 0.0177). CI was predominantly diagnosed by endoscopy (26/35), generally within the first 4 postoperative days (20/35). Twenty-eight patients underwent surgery, all finalised with stoma creation. Postoperative bilateral occlusion and/or relevant stenosis of hypogastric arteries were more frequent in CI patients (57.8 vs. 20.8%, p = 0.0273). In-hospital mortality was increased in the CI group (26.7 vs. 2.9%, p &amp;lt; 0.0001). Survival was significantly reduced in CI patients (median: 28.2 months vs. 104.1 months, p &amp;lt; 0.0001).!##!Conclusion!#!CI after aortic surgery is associated with considerable perioperative sequelae and reduced survival. Especially in patients at risk, such as those with rAAA, complicated intraoperative course, severe cardiovascular morbidity and/or perioperative deterioration of the hypogastric perfusion, vigilant postoperative multimodal monitoring is required in order to initiate diagnosis and treatment

Impact of the histone deacetylase inhibitor 4-phenylbutyrate on the clearance of apoptotic pancreatic carcinoma cells by human macrophages

Histone deacetylase inhibitors have been found to have potent anticancer activities, partly induced by tumour cell apoptosis. The clearance of apoptotic tumour cells is an important mechanism of antitumour immune surveillance. The aim of this study was to assess the impact of 4-phenylbutyrate.(4-PB) and its immunological effects on the macrophage clearance of apoptotic pancreatic ductal adenocarcinoma (PDAC) cells. To this end, a co-culture system of human macrophages from donors and PDAC patients, and PDAC cell lines (T3M4, PANC-1 and AsPC-1) was established to study the effect of 4-PB. Apoptosis and phagocytic activity were analysed using flow cytometry, and phagocytosis was confirmed by confocal microscopy. Further, p21 expression was quantified by immunoblot analysis. 4-PB treatment (0-10 mM) resulted in a dose-dependent induction of tumour cell apoptosis in two of the cell lines (T3M4 and PANC-1), but it also induced human macrophage apoptosis. The apoptotic effect of gemcitabine on PDAC cells was further enhanced by 4-PB. Moreover, 4-PB led to a dose-dependent overexpression of the cell cycle regulator p21 in tumour cells. In co-culture, apoptotic PDAC cells were phagocytosed by donor macrophages and phagocytosis was increased through tumour cell exposure to 4-PB and/or gemcitabine, whereas phagocytosis of PANC-1 cells was reduced using macrophages of PDAC patients treated with 4-PB. The 4-PB treatment induced human macrophage expression of the pro-angiogenic IL-8 and simultaneously inhibited inflammatory cytokine release through modulation of IL-10 and TNF alpha after phagocytosis of apoptotic PDAC cells. In conclusion, the 4-PB treatment activated tumour cell death in PDAC cells, resulting in tumour cell phagocytosis by macrophages. The latter were characterized by an anti-inflammatory and pro-angiogenic cytokine response demonstrating adverse, tumour-promoting effects of macrophages on tumour cells. Thus, the potential of 4-PB as an anticancer agent against PDAC cannot be reliably assessed without taking into account the complex tumour microenvironment

Experimental <it>in vivo</it> and <it>in vitro</it> treatment with a new histone deacetylase inhibitor belinostat inhibits the growth of pancreatic cancer

Abstract Background Treatment options for pancreatic ductal adenocarcinoma (PDAC) are limited. Histone deacetylase inhibitors are a new and promising drug family with strong anticancer activity. The aim of this study was to examine the efficacy of in vitro and in vivo treatment with the novel pan-HDAC inhibitor belinostat on the growth of human PDAC cells. Methods The proliferation of tumour cell lines (T3M4, AsPC-1 and Panc-1) was determined using an MTT assay. Apoptosis was analysed using flow cytometry. Furthermore, p21Cip1/Waf1 and acetylated histone H4 (acH4) expression were confirmed by immunoblot analysis. The in vivo effect of belinostat was studied in a chimeric mouse model. Antitumoural activity was assessed by immunohistochemistry for Ki-67. Results Treatment with belinostat resulted in significant in vitro and in vivo growth inhibition of PDAC cells. This was associated with a dose-dependent induction of tumour cell apoptosis. The apoptotic effect of gemcitabine was further enhanced by belinostat. Moreover, treatment with belinostat increased expression of the cell cycle regulator p21Cip1/Waf1 in Panc-1, and of acH4 in all cell lines tested. The reductions in xenograft tumour volumes were associated with inhibition of cell proliferation. Conclusion Experimental treatment of human PDAC cells with belinostat is effective in vitro and in vivo and may enhance the efficacy of gemcitabine. A consecutive study of belinostat in pancreatic cancer patients alone, and in combination with gemcitabine, could further clarify these effects in the clinical setting.</p

Growth inhibition of pancreatic cancer by experimental treatment with 4-phenylbutyrate is associated with increased expression of connexin 43

Histone deacetylase inhibitors are a new and promising drug family with a strong anticancer activity and potent modulation of connexin expression. The restoration of connexins in cancer cells has been suggested as a possible mechanism to control tumor progression. The aim of this study was to investigate the effects of 4-phenylbutyrate (4-PB) on the growth of human pancreatic cell lines in vitro and in vivo with a focus on connexin modulation. The proliferation of tumor cells was determined using an MTT assay, and the effect of 4-PB in vivo was studied in a chimeric mouse model. The expression and localization of connexin 43 (Cx43) were assessed by Western blot, immunofluorescence microscopy, and immunohistochemistry. Antitumoral activity was assessed by immunohistochemistry for Ki-67 and histone H4. Treatment with 4-PB resulted in the significant in vitro and in vivo growth inhibition of pancreatic tumor cells. The reduction of the xenograft tumor volume was associated with the inhibition of histone deacetylation and decrease in cell proliferation. Treatment with 4-PB caused a significant increase in the Cx43 expression in T3M4 cells (up to 2.8-fold). The newly synthesized Cx43 was localized in the cytoplasm but not on the cell membrane. Treatment with 4-PB inhibited the proliferation of human pancreatic tumor cells in vitro and in vivo and increased the expression of Cx43. Therefore, 4-PB might be useful in the therapy of pancreatic cancer