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Aim Pathological complete response (pCR) and clinical outcomes [overall survival (OS), disease-free survival (DFS), locoregional control (LC)] were evaluated in a single-institution experience of different schedules of neoadjuvant chemoradiotherapy (CRT) for patients with locally advanced rectal cancer (LARC). Patients and methods Data for 322 patients with LARC were retrospectively analyzed. pCR was evaluated according to Mandard tumor regression grade (TRG). The Kaplan-Meier method was used to estimate OS, DFS and LC. Results Three hundred and three (94.1%) patients underwent surgery. pCR was observed in 81 patients (26.7%), with TRG1-2 rate of 41.8%. The 5- and 10-year OS, DFS and LC rates were 82.5%±2.5% and 65.5%±3.8%, 81.2%±2.4% and 79.3%±2.9%, 93.1%±1.7% and 90.5%±2.1%, respectively. Conclusion Neoadjuvant CRT in LARC patients resulted in favorable long-term oncological outcomes, with a high pCR rate and acceptable toxicity.

To compare tumor detectability and conspicuity of standard b = 1000 s/mm2 (b1000) versus ultrahigh b = 2000 s/mm2 (b2000) diffusion-weighted imaging (DWI) in rectal cancer. Fifty-five patients for a total of 81 3T DWI-MR scans were retrospectively evaluated by two differently experienced readers. A comparison between b1000 and b2000 for tumor detectability and conspicuity was performed. The conspicuity was qualitatively and quantitatively assessed by using three-point scale and whole tumor volume manual delineation, respectively. Receiver-operating characteristic curve (ROC) with area under the curve (AUC) analysis provided diagnostic accuracy in tumor detectability of restaging MR scans. Qualitative scores and quantitative features including mean signal intensity, variance, 10th percentile and 90th percentile, were compared using the Wilcoxon test. Interobserver agreement (IOA) for qualitative and quantitative data was calculated using Cohen’s Kappa and intraclass correlation coefficient (ICC) respectively. Diagnostic accuracy was comparable between b1000 and b2000 for both readers (p > 0.05). Overall quality scores were significantly better for b2000 than b1000 (2.29 vs 1.65 Reader 1, p = 0.01; 2.18 vs 1.69 Reader 2, p = 0.04). IOA was equally good for both b values (k = 0.86 b1000, k = 0.86 b2000). Quantitative analysis revealed more uniform signal (measured in variance) of b2000 in both healthy surrounding tissue (p

Background Pancreatic Ductal Adenocarcinoma (PDAC) is an aggressive and lethal disease, lacking effective therapeutic approaches. Available therapies only marginally prolong patient survival and are frequently coupled with severe adverse events. It is therefore pivotal to investigate novel and safe pharmacological approaches. We have recently identified the ABC transporter, ABCC3, whose expression is dependent on mutation of TP53, as a novel target in PDAC. ABCC3-mediated regulation of PDAC cell proliferation and tumour growth in vivo was demonstrated and was shown to be conferred by upregulation of STAT3 signalling and regulation of apoptosis. Methods To verify the potential of ABCC3 as a pharmacological target, a small molecule inhibitor of ABCC3, referred to here as MCI-715, was designed. In vitro assays were performed to assess the effects of ABCC3 inhibition on anchorage-dependent and anchorage-independent PDAC cell growth. The impact of ABCC3 inhibition on specific signalling pathways was verified by Western blotting. The potential of targeting ABCC3 with MCI-715 to counteract PDAC progression was additionally tested in several animal models of PDAC, including xenograft mouse models and transgenic mouse model of PDAC. Results Using both mouse models and human cell lines of PDAC, we show that the pharmacological inhibition of ABCC3 significantly decreased PDAC cell proliferation and clonal expansion in vitro and in vivo, remarkably slowing tumour growth in mice xenografts and patient-derived xenografts and increasing the survival rate in a transgenic mouse model. Furthermore, we show that stromal cells in pancreatic tumours, which actively participate in PDAC progression, are enriched for ABCC3, and that its inhibition may contribute to stroma reprogramming. Conclusions Our results indicate that ABCC3 inhibition with MCI-715 demonstrated strong antitumor activity and is well tolerated, which leads us to conclude that ABCC3 inhibition is a novel and promising therapeutic strategy for a considerable cohort of patients with pancreatic cancer. Electronic supplementary material The online version of this article (10.1186/s13046-019-1308-7) contains supplementary material, which is available to authorized users.

BACKGROUND Although laparoscopic cholecystectomy (LC) is the gold standard for symptomatic gallbladder disease, a single-incision approach may be a new challenge in order to achieve minimization of surgical trauma. Single-site robotic cholecystectomy (SSRC) is able to offset the ergonomic limitation of laparoscopic single-site cholecystectomy and improves cosmesis. METHODS We present a single-institution initial experience of SSRC for cholecystolithiasis. Intra-operative and post-operative data of patients were reviewed to assess the technical feasibility and cosmetic outcome. RESULTS We evaluated a series of 27 consecutive patients retrospectively analyzed and prospectively collected who underwent SSRC. One patient was excluded from the final analysis because they converted to open procedure. The female/male ratio was 17/9, with mean age of 48 ± 12 years. The body mass index mean value was 26.0 ± 4.2. The mean operative time was 99.6 ± 21.5 minutes. No intra- or post-operative complications and readmissions were recorded. At 12 months follow up, every patient received the Body Image Questionnaire (BIQ) and a Photo Series Questionnaire. We recorded three patients (11.5%) with post-operative incisional hernia. Scores of the BIQ subscale for body image perception were 6 ± 1.2, while the scores of scar cosmesis were 21.1 ± 3.0. A statistically significant improvement in scar self-rating from T0 to T1 (P

Neoadjuvant chemo-radiotherapy (CRT) followed by total mesorectal excision (TME) represents the standard treatment for patients with locally advanced (≥ T3 or N+) rectal cancer (LARC). Approximately 15% of patients with LARC shows a complete response after CRT. The use of pre-treatment MRI as predictive biomarker could help to increase the chance of organ preservation by tailoring the neoadjuvant treatment. We present a novel machine learning model combining pre-treatment MRI-based clinical and radiomic features for the early prediction of treatment response in LARC patients. MRI scans (3.0 T, T2-weighted) of 72 patients with LARC were included. Two readers independently segmented each tumor. Radiomic features were extracted from both the “tumor core” (TC) and the “tumor border” (TB). Partial least square (PLS) regression was used as the multivariate, machine learning, algorithm of choice and leave-one-out nested cross-validation was used to optimize hyperparameters of the PLS. The MRI-Based “clinical-radiomic” machine learning model properly predicted the treatment response (AUC = 0.793, p = 5.6 × 10–5). Importantly, the prediction improved when combining MRI-based clinical features and radiomic features, the latter extracted from both TC and TB. Prospective validation studies in randomized clinical trials are warranted to better define the role of radiomics in the development of rectal cancer precision medicine.

Splenic flexure (SF) cancer is not a common condition and its treatment is still under discussion. Although laparoscopic surgery is well accepted for the treatment of colon cancer at any stage, complete mesocolon excision (CME) with selective vascular ligation using the laparoscopic approach for SF cancer remains technically demanding and represents a real challenge for surgeons. We present a single-institution experience of laparoscopic CME for SF cancer. Intra-operative, pathologic, and post-operative data of patients who underwent laparoscopic SF resection were reviewed to assess the technical feasibility and oncologic safety. Technical features, histopathology, morbidity, and mortality were evaluated. From February 2015 to October 2017, a minimally invasive approach was proposed to 17 patients (M/F 14/3) affected by splenic flexure cancer. In all patients, the procedure was completed by laparoscopy. The anastomosis was completed intra-corporeally in 89% of cases. The distal margin was 3.1 ± 2.6 cm and the proximal margin was 6.5 ± 3.3 cm from the tumor site. The number of mean harvested nodes was 13.9 ± 7. The mean operative time was 215.5 ± 65 min, and blood loss was 80 ± 27. In one case, a laparoscopic partial gastrectomy was associated due to tumor invasion. The mean post-operative stay was 6.7 ± 3.3 days. Readmission was necessary for two patients. No major morbidity was recorded. Despite the wide spread and increasing confidence in laparoscopic colectomy, SF resection remains one of the most challenging procedures in colorectal surgery with a complex learning curve. SF resection with CME and CVL is feasible and safe for the treatment of early-stage and locally advanced SF cancer.

Introduction. Despite the widespread use of the robotic technology, only a few studies with small sample sizes report its application to pancreatic diseases treatment. Our aim is to present the results of a multicenter study on the safety and feasibility of robot-assisted distal pancreatectomy (RDP). Materials and Methods. All RDPs for benign, borderline, and malignant diseases performed in 5 referral centers from 2008 to 2016 were included. Perioperative outcomes were evaluated. Results. Two hundred thirty-six patients were included. Spleen preservation was performed in 114 cases (48.3%). Operative time was 277.8 ± 93.6 minutes. Progressive improvement in operative time was observed over the study period. Conversion rate was 6.3%. Morbidity occurred in 102 cases (43.2%), mainly due to grade A fistulas. Reoperation was required in 10 patients. Postoperatively, 2 patients died of sepsis due to a grade C fistula. Hospital readmission was necessary in 11 cases. A R0 resection was always achieved, with a mean number of 16.2 ± 15 harvested lymph nodes. Conclusion. To our knowledge, this is one of the largest RDP series. Safety and feasibility including the low conversion rate, the high spleen preservation rate, the adequate operative time, and the acceptable morbidity and mortality rates confirm the validity of this technique. Appropriate oncological outcomes have been also obtained.

Background Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies and is not a clinically homogeneous disease, but subsets of patients with distinct prognosis and response to therapy can be identified by genome-wide analyses. Mutations in major PDAC driver genes were associated with poor survival. By bioinformatics analysis, we identified protocadherins among the most frequently mutated genes in PDAC suggesting an important role of these genes in the biology of this tumor. Promoter methylation of protocadherins has been suggested as a prognostic marker in different tumors, but in PDAC this epigenetic modification has not been extensively studied. Thus, we evaluated whether promoter methylation of three frequently mutated protocadherins, PCDHAC2, PCDHGC5 and PCDH10 could be used as survival predictors in PDAC patients. Methods DNA extracted from 23 PDACs and adjacent non-neoplastic pancreatic tissues were bisulfite treated. Combined Bisulfite Restriction Analysis (COBRA) coupled to denaturing high-performance liquid chromatography (dHPLC) detection and bisulfite genomic sequencing (BGS) were used to determine the presence of methylated CpG dinucleotides in the promoter amplicons analyzed. Results In an exploratory analysis, two protocadherins showed the same pattern of CpG methylation in PDAC and adjacent non-neoplastic pancreatic tissues: lack of methylation for PCDHAC2, complete methylation for PCDHGC5. Conversely, the third protocadherin analyzed, PCDH10, showed a variable degree of CpG methylation in PDAC and absence of methylation in adjacent non-neoplastic pancreatic tissues. At Kaplan–Meier analysis, high levels of PCDH10 methylation defined according to the receiver operating characteristic (ROC) curve analysis were significantly associated with worse progression-free survival (PFS) rates (P = 0.008), but not with overall survival (OS). High levels of PCDH10 methylation were a prognostic factor influencing PFS (HR = 4.0: 95% CI, 1.3–12.3; P = 0.016), but not the OS. Conclusions In this study, we show for the first time that the methylation status of PCDH10 can predict prognosis in PDAC patients with a significant impact on the outcome in terms of progression-free survival. High levels of PCDH10 promoter methylation could be useful to identify patients at high risk of disease progression, contributing to a more accurate stratification of PDAC patients for personalized clinical management.

Pancreatic cancer (PC), the fourth leading cause of cancer-related deaths, is characterized by high aggressiveness and resistance to chemotherapy. Pancreatic carcinogenesis is kept going by derangement of essential cell processes, such as proliferation, apoptosis, metabolism and autophagy, characterized by rhythmic variations with 24-h periodicity driven by the biological clock. We assessed the expression of the circadian genes ARNLT, ARNLT2, CLOCK, PER1, PER2, PER3, CRY1, CRY2 and the starvation-activated histone/protein deacetylase SIRT1 in 34 matched tumor and non-tumor tissue specimens of PC patients, and evaluated in PC derived cell lines if the modulation of SIRT1 expression through starvation could influence the temporal pattern of expression of the circadian genes. We found a significant down-regulation of ARNLT (p = 0.015), CRY1 (p = 0.013), CRY2 (p = 0.001), PER1 (p 0.0001), PER2 (p 0.001), PER3 (p = 0.001) and SIRT1 (p = 0.017) in PC specimens. PER3 and CRY2 expression levels were lower in patients with jaundice at diagnosis ( 0.05). Having adjusted for age, adjuvant therapy and tumor stage, we evidenced that patients with higher PER2 and lower SIRT1 expression levels showed lower mortality (p = 0.028). Levels and temporal patterns of expression of many circadian genes and SIRT1 significantly changed upon serum starvation in vitro, with differences among four different PC cell lines examined (BXPC3, CFPAC, MIA-PaCa-2 and PANC-1). Serum deprivation induced changes of the overall mean level of the wave and amplitude, lengthened or shortened the cycle time and phase-advanced or phase-delayed the rhythmic oscillation depending on the gene and the PC cell line examined. In conclusion, a severe deregulation of expression of SIRT1 and circadian genes was evidenced in the cancer specimens of PC patients, and starvation influenced gene expression in PC cell lines, suggesting that the altered interplay between SIRT1 and the core circadian proteins could represent a crucial player in the process of pancreatic carcinogenesis.

Summary Background There is substantial uncertainty regarding the optimal surgical treatment for chronic pancreatitis. Short-term outcomes have been found to be better after duodenum-preserving pancreatic head resection (DPPHR) than after partial pancreatoduodenectomy. Therefore, we designed the multicentre ChroPac trial to investigate the long-term outcomes of patients with chronic pancreatitis within 24 months after surgery. Methods This randomised, controlled, double-blind, parallel-group, superiority trial was done in 18 hospitals across Europe. Patients with chronic pancreatitis who were planned for elective surgical treatment were randomly assigned to DPPHR or partial pancreatoduodenectomy with a central web-based randomisation tool. The primary endpoint was mean quality of life within 24 months after surgery, measured with the physical functioning scale of the European Organisation for Research and Treatment of Cancer QLQ-C30 questionnaire. Primary analysis included all patients who underwent one of the assigned procedures; safety analysis included all patients who underwent surgical intervention (categorised into groups as treated). Patients and outcome assessors were masked to group assignment. The trial was registered, ISRCTN38973832. Recruitment was completed on Sept 3, 2013. Findings Between Sept 10, 2009, and Sept 3, 2013, 250 patients were randomly assigned to DPPHR (n=125) or partial pancreatoduodenectomy (n=125), of whom 226 patients (115 in the DPPHR group and 111 in the partial pancreatoduodenectomy group) were analysed. No difference in quality of life was seen between the groups within 24 months after surgery (75·3 [SD 16·4] for partial pancreatoduodenectomy vs 73·0 [16·4] for DPPHR; mean difference −2·3, 95% CI −6·6 to 2·0; p=0·284). The incidence and severity of serious adverse events did not differ between the groups. 70 (64%) of 109 patients in the DPPHR group and 61 (52%) of 117 patients in the partial pancreatoduodenectomy group had at least one serious adverse event, with the most common being reoperations (for reasons other than chronic pancreatitis), gastrointestinal problems, and other surgical morbidity. Interpretation No differences in quality of life after surgery for chronic pancreatitis were seen between the interventions. Results from single-centre trials showing superiority for DPPHR were not confirmed in the multicentre setting. Funding German Research Foundation (DFG).

Adenocarcinomas of Vater's papilla (PVAC) may originate from either the pancreatic duct or the intestinal epithelium. Conflicting data have been reported about the frequency of the 2 anatomical entities and their influence on patients' prognosis. To ascertain the anatomical origin of PVAC in a family member of a Lynch syndrome kindred, we searched for microRNA (miRNA) expression profiles on resected tumor specimens. The support vector machine was trained on our previous miRNAs expression data sets of pancreatic and colorectal tissue samples and used to classify the site of origin of the tumor in our patient. The support vector machine worked by contrasting the profiles of miRNAs in patients with pancreatic ductal and colorectal cancers to that of our patient, which was finally classified as pancreatic ductal adenocarcinoma accordingly to alterations of 55 miRNAs. The PVAC might be originated from ductal epithelium rather than from the intestinal mucosa of the papilla in the case at issue. Alteration of miR-548b-3p, miR-551a, miR-21, miR-92a, miR-let-7i, and miR-181a* emerged as potentially associated with cancer genetic susceptibility in PVAC.

Despite advances in clinical management of pancreatic cancer (PC), there is still room for improvement in early detection, diagnosis, and treatment strategies. The role of microRNAs (miRNAs) in tumor biology might pinpoint an alteration in expression of miRNAs as new diagnostic/prognostic biomarkers.Expression levels of miR-143 and miR-21 and correlations with clinicopathological features were analyzed in 26 matched pairs of tumor and adjacent noncancerous tissue samples collected from patients with PCs, including 18 pancreatic ductal adenocarcinomas (PDACs) and 8 adenocarcinomas of Vater's papilla (PVACs).Compared to normal tissues, miR-143 was up-regulated in both PDAC and PVAC tumor samples (P = 0.0028 and P = 0.039, respectively). Conversely, alterations in miR-21 expression were significantly different in PDAC versus PVAC samples (P = 0.0049). Tumor levels of miR-21 were associated with preoperative serum levels of CA 19-9 (r = 0.63, P = 0.0022), whereas miR-143 expression was negatively correlated to lymph node spreading (r = -0.64; P = 0.0004). Correlation between miR-143 and miR-21 expression levels in patients with PDAC was observed (r = 0.53, P = 0.023).Deregulation of miR-143 and miR-21 may reflect histological features and biological behavior of different PCs. Association data with clinical parameters might indicate a prognostic significance for miR-143 and miR-21 in PCs.

Background and aim The neuropeptide Substance P, plays a key role in modulating neuroimmune interactions in patients with inflammatory bowel diseases. We analyzed Substance P serum levels in patients with ulcerative colitis and Crohn's disease, to detail the involvement of the neuropeptide in the pathophysiology of these disorders. Methods Serum samples were collected from 61 patients with ulcerative colitis (24 with active and 37 with inactive disease), 66 patients with Crohn's disease (29 with active and 37 with inactive disease) and 45 healthy subjects, enrolled into the study. Neuropetide serum levels were measured by means of an ELISA/EIA. Associations with disease activity and patients clinical features were also taken into account. Results Compared to controls, Substance P serum levels were significantly increased in both patients with ulcerative colitis and Crohn's disease, ( p ). In patients with ulcerative colitis, levels paralleled disease activity ( p = 0.014 ), and the amount of the neuropeptide was considerably decreased during clinical and endoscopic remission of the disease, ( p = 0.025 ). Conversely, median Substance P levels did not differ between patients with active and inactive Crohn's disease. However, levels of the neuropeptide were more often elevated in patients with inactive and stricturing/fistulizing Crohn's disease, ( p = 0.002 ). Conclusions Data underline that Substance P might exerts important immunomodulatory functions in inflammatory bowel disease. This study suggests a potential role for Substance P serum levels in monitoring intestinal inflammation in patients with inflammatory bowel disease.

Germline genetic variability might contribute, at least partially, to the survival of pancreatic ductal adenocarcinoma (PDAC) patients. Two recently performed genome-wide association studies (GWAS) on PDAC overall survival (OS) suggested (P < 10(-5)) the association between 30 genomic regions and PDAC OS. With the aim to highlight the true associations within these regions, we analyzed 44 single-nucleotide polymorphisms (SNPs) in the 30 candidate regions in 1722 PDAC patients within the PANcreatic Disease ReseArch (PANDoRA) consortium. We observed statistically significant associations for five of the selected regions. One association in the CTNNA2 gene on chromosome 2p12 [rs1567532, hazard ratio (HR) = 1.75, 95% confidence interval (CI) 1.19-2.58, P = 0.005 for homozygotes for the minor allele] and one in the last intron of the RUNX2 gene on chromosome 6p21 (rs12209785, HR = 0.88, 95% CI 0.80-0.98, P = 0.014 for heterozygotes) are of particular relevance. These loci do not coincide with those that showed the strongest associations in the previous GWAS. In silico analysis strongly suggested a possible mechanistic link between these two SNPs and pancreatic cancer survival. Functional studies are warranted to confirm the link between these genes (or other genes mapping in those regions) and PDAC prognosis in order to understand whether these variants may have the potential to impact treatment decisions and design of clinical trials.

OBJECTIVE: Generation and maintenance of pain in chronic pancreatitis (CP) have been shown to be partially attributable to neuroimmune interactions, which involve neuropeptides such as substance P (SP). So far, expression of SP receptors NK-2R, NK-3R, the SP-encoding gene preprotachykinin A (PPT-A), and the SP degradation enzyme neutral endopeptidase (NEP) and their relation to pain in CP have not been determined. METHODS: Tissue samples from patients with CP (n = 25) and from healthy donors (n = 20) were analyzed for PPT-A, NK-2R, NK-3R, and NEP expression using quantitative RT-PCR. NEP protein levels were examined by immunoblot analysis and its localization was determined using immunohistochemistry. A scoring system was used to grade the extent of fibrosis on hematoxylin and eosin- and Masson-Trichrome-stained sections. Messenger RNA levels and the extent of pain were analyzed for correlations. RESULTS: In CP tissues, NK-2R and PPT-A expression was increased, whereas NK-3R and NEP mRNA levels were comparable with normal pancreas. Overexpression of NK-2R was related to the intensity, frequency, and duration of pain in CP patients. NK-1R and NEP expression was significantly related to the extent of fibrosis. CONCLUSIONS: Expression of NK-2R and PPT-A is increased in CP and is associated with pain. Failure to up-regulate NEP may contribute to the disruption of the neuropeptides loop balance in CP and thus may exacerbate the severe pain syndrome.

Background Neuropeptides, such as substance P (SP), are mediators of neurogenic inflammation and play an important role in inflammatory disorders. To further investigate the role of the SP pathway in inflammatory bowel disease (IBD), we analyzed the following in normal intestinal tissue specimens and in tissue specimens from patients with Crohn's disease (CD) and ulcerative colitis (UC): neurokinin receptor-1 (NK-1R); its isoforms (NK-1R-L and NK-1R-S); its ligand SP, encoded by preprotachykinin-A (PPT-A); and the SP-degradation enzyme, neutral endopeptidase (NEP). Methods Real-time quantitative reverse transcription–polymerase chain reaction was used to simultaneously determine the expression of NK-1R-L, NK-1R-S, and PPT-A. Protein levels of NK-1R and NEP were determined by immunoblot analysis. Results In noninflamed small-bowel tissue samples of CD patients, PPT-A mRNA expression was significantly increased, whereas there was no difference between inflamed or noninflamed UC and normal intestinal tissue samples. Examining subgroups of diverse intestinal segments from CD and UC samples with various levels of inflammation revealed no differences in NK-1R-L and NK-1R-S mRNA expression, whereas there was a tendency toward overall lower NK-1R-S mRNA copy numbers. Immunoblot analysis showed upregulation of NK-1R protein levels in cases of IBD, with more pronounced enhancement in cases of CD than in UC. For NEP, there were no differences in protein levels in normal, CD, and UC intestinal tissues. Comments These observations suggest a contribution of SP and its receptor, NK-1R, in the local inflammatory reaction in IBD and particularly in ileal CD. Moreover, significant upregulation of PPT-A mRNA in the noninflamed ileum of these patients suggests an influence of inflamed intestines on their healthy counterparts.

The incidence and death rate of pancreatic ductal adenocarcinoma (PDAC) have increased in recent years, therefore the identification of novel targets for treatment is extremely important. Interactions between cancer and stromal cells are critically involved in tumour formation and development of metastasis. Here we report that PDAC cells secrete BAG3, which binds and activates macrophages, inducing their activation and the secretion of PDAC supporting factors. We also identify IFITM-2 as a BAG3 receptor and show that it signals through PI3K and the p38 MAPK pathways. Finally, we show that the use of an anti-BAG3 antibody results in reduced tumour growth and prevents metastasis formation in three different mouse models. In conclusion, we identify a paracrine loop involved in PDAC growth and metastatic spreading, and show that an anti-BAG3 antibody has therapeutic potential.

Clearance of apoptotic or necrotic cells by phagocytes protects the surrounding tissue from toxic intracellular factors and reduces probable tissue damage following an inappropriate inflammatory reaction.1 Phosphatidylserine (PS) is one of the key factors in induction of phagocytosis of apoptotic cells. It is the most abundant anionic phospholipid of the plasma membrane, which is located at the inner leaflet of the cell surface.2 The loss of asymmetry and the appearance of this molecule on the outer leaflet of the cell membrane is regarded as one of the essential steps in apoptosis and subsequently in phagocytosis of apoptotic cells.3–7 If apoptotic cells are unable to express PS on their surface, it is almost impossible for them to be phagocytosed.6 However, in addition to the presentation of PS, apoptotic cells display numerous receptors on their surfaces, such as CD 36, αvβ3, αvβ5 integrins, CD14 and CD68. These are necessary in order for phagocytes to approach apoptotic cells, but only the interplay with the PS receptor enables their internalization.8 Binding of PS on the apoptotic cell surface to the PS-receptor (PSR) on phagocytes is an active noninflammatory process which is associated with the down-regulation of proinflammatory mediators such as interleukin (IL)-1β, IL-8, IL-10, granulocyte macrophage colony-stimulating factor, tumor necrosis factor-α, leukotriene C4 and thromboxane B2, while the production of anti-inflammatory mediators (transforming growth factor [TGF]-β1, prostaglandin E2, and platelet-activating factor) is increased in human monocyte-derived macrophages.9,10 Therefore, abnormalities of apoptotic cell clearance may contribute to the pathogenesis of chronic inflammatory diseases.5,9 Tissue destruction in chronic pancreatitis is histopathologically associated with a large amount of infiltrating macrophages.11 In addition, infiltrates of cytotoxic cells are localized at the border of parenchyma and fibrosis, suggesting that cell-mediated cytotoxicity is involved in cell damage in this disease.12,13 Morphologically, cell damage and apoptosis in chronic pancreatitis are associated with up-regulation of pro-apoptotic genes,14 the FAS/FAS-l system,15 caspase-116 and p75NTR17 in atrophic acinar cells, proliferating cells of ductal origin and in acinar cells redifferentiating to form tubular structures. Apoptotic mechanisms in the margin regions of the parenchyma close to stromal fibroblasts may be responsible for increased acinar cell redifferentiation, death, and phagocytosis with replacement by stromal fibroblasts.18–20 Since apoptosis is active in chronic pancreatitis and large numbers of macrophages are present in CP tissue samples, we investigated whether the presence of macrophages is PS/PSR-mediated and whether PS/PSR interaction is a major component in apoptotic cell phagocytosis in CP.

The etiopathogenesis of cholecystolithiasis is not well defined. Primary dysmotility of the organ, due to fibrosis of the gallbladder wall or muscular dysfunction, is suggested as a crucial factor. Transforming growth factor-beta (TGF-beta) and connective tissue growth factor (CTGF) are involved in several fibrotic disorders and play a critical role in fibrogenesis, thereby changing the physiological function of the organs. In the present study we analyzed the role of TGF-beta and its downstream target CTGF in patients with cholecystolithiasis.Gallbladders were obtained from 16 individuals undergoing surgery for symptomatic cholecystolithiasis. Normal human gallbladder tissue samples from five individuals without any history of gallbladder disease were obtained through an organ donor transplantation program. Northern blot analysis, in situ hybridization, and immunohistochemistry were used to analyze the expression of TGF-beta1 and CTGF in the gallbladder tissue samples.By northern blot analysis there was an enhanced TGF-beta1 mRNA expression (eightfold increase; P0.04) in the cholecystolithiasis tissue samples in comparison with normal controls. There was also a concomitant increase in CTGF (41-fold increase; P0.01). By in situ hybridization and immunohistochemistry, CTGF mRNA was localized mainly in the mucosa layer, while intensive staining of the smooth muscle cells with TGF-beta1 and CTGF was observed. In addition, TGF-beta1 immunoreactivity was also localized in the fibroblasts and inflammatory cells. TGF-beta1 m-RNA levels showed a significant relationship with the degree of fibrosis in the tissue samples (P0.04, r = 0.5).Our data indicate that TGF-beta and CTGF are involved in ultrastructural tissue changes in patients with cholecystolithiasis. Activation of the "TGF-beta pathway," predominantly in the remaining mucosa and submucosal layer, indicates that extracellular matrix (ECM) synthesis with subsequent gallbladder wall fibrosis is an important step in gallbladder dysfunction in this disorder.

Connective tissue growth factor (CTGF), which is regulated by transforming growth factor-ss (TGFss), has recently been implicated in the pathogenesis of fibrotic diseases and tumor stroma. Inasmuch as generation of desmoplastic tissue is characteristic for pancreatic cancer, it is not known whether it gives pancreatic cancer cells a growth advantage or is a reaction of the body to inhibit cancer cell progression. In the present study we analyzed the expression and localization of CTGF and evaluated whether it influences the prognosis of pancreas cancer. Tissue samples were obtained from 25 individuals (6 women, 19 men) undergoing pancreatic resection for pancreatic cancer. Tissue samples from 13 previously healthy organ donors (5 women, 8 men) served as controls. Expression of CTGF was studied by Northern blot analysis. In situ hybridization and immunohistochemistry localized the respective mRNA moieties and proteins in the tissue samples. Northern blot analysis revealed that pancreatic cancer tissue samples exhibited a 46-fold increase in CTGF mRNA expression ( p0.001) over that of normal controls. In vitro studies confirmed that pancreatic stellate cells are the major source of CTGF mRNA expression and revealed a large variance in basal and TGFss-induced CTGF expression in cultured pancreatic cancer cells. This could also be confirmed by in situ hybridization, indicating that CTGF mRNA signals were located principally in fibroblasts, with only weak signals in the cancer cells. High CTGF mRNA levels in the tissue samples correlated with better tumor differentiation ( p0.03). In addition, patients whose tumors exhibited high CTGF mRNA levels (onefold increase above normal controls) lived significantly longer than those whose tumors expressed low CTGF mRNA levels (none to onefold) ( p0.04 multivariate analysis). Our present data indicate that CTGF, as a downstream mediator of TGFss, is overexpressed in connective tissue cells and to a lesser extent in pancreatic cancer cells. Because patients with high CTGF mRNA expression levels have a better prognosis, our findings indicate that the desmoplastic reaction provides a growth disadvantage for pancreatic cancer cells.

Background Pancreatic ductal adenocarcinoma (PDAC) is an extremely aggressive malignancy, characterized by largely unsatisfactory responses to the currently available therapeutic strategies. In this study we evaluated the expression of genes involved in gemcitabine uptake in a selected cohort of patients with PDAC, with well-defined clinical-pathological features. Methods mRNA levels of hENT1, CHOP, MRP1 and DCK were evaluated by means of qRT-PCR in matched pairs of tumor and adjacent normal tissue samples collected from PDAC patients treated with gemcitabine after surgical tumor resection. To detect possible interaction between gene expression levels and to identify subgroups of patients at different mortality/progression risk, the RECursive Partitioning and Amalgamation (RECPAM) method was used. Results RECPAM analysis showed that DCK and CHOP were most relevant variables for the identification of patients with different mortality risk, while hENT1 and CHOP were able to identify subgroups of patients with different disease progression risk. Conclusion: hENT1, CHOP, MRP1 and DCK appear correlated to PDAC, and this interaction might influence disease behavior.

15-25% of appendices removed from patients with suspected appendicitis appear normal on histological examination. The cause of pain in such patients is unknown. Since the content of neuropeptides seems to be altered in chronic inflammation, we investigated possible changes in peptidergic innervation for substance P (SP), vasoactive intestinal peptide (VIP), and growth-associated protein-43 (GAP-43).Appendices classified as showing acute appendicitis, non-acute appendicitis (clinical signs of acute appendicitis, but histologically not inflamed), or normal were processed for SP, VIP, and GAP-43 immunocytochemistry. The density of SP immunostaining was assessed by digitised morphometry.31 appendix specimens were studied (16 acute, 15 non-acute). 16 specimens were used as controls. Expression of GAP-43 was increased in the non-acute appendices. We observed larger amounts of SP-immunoreactive and VIP-immunoreactive nerves in the mucosal layer of the appendix in patients with non-acute appendicitis than in controls and patients with acute appendicitis (mean % area SP-immunoreactive 0.0496 [SD 0.0113] non-acute, 0.0221 [0.0049] acute, 0.0229 [0.0068] controls). In addition, a close spatial relation between SP-immunoreactive and VIP-immunoreactive nerve fibres and lymphoid cells was detected in the outer zone of lymph follicles.Neuroproliferation in the appendix, in association with an increase in neurotransmitters SP and VIP, may be involved in the pathophysiology of acute right abdominal pain in the absence of an acute inflammation of the appendix. Our data, together with increasing knowledge about the way in which the nervous system and immune cells interact, suggest that neuroimmune appendicitis is a distinct pathological entity.

A tumour score for venous invasion in patients with pancreatic adenocarcinoma was evaluated by means of computed tomography (CT), in order to improve the assessment of medical treatment and clinical outcome with special attention to borderline resectable disease.Fifty-six consecutive patients who underwent curative surgical resection for pancreatic cancer were analysed. On the basis of CT criteria, tumour involvement of the portal vein (PV) and superior mesenteric vein (SMV) was graded according to an adapted 4-point scale: score 1, definite absence of invasion; score 2, probable absence of invasion; score 3, probable presence of invasion; score 4, definite presence of invasion. Correlations between the venous infiltration scores and the patients' clinical features were also evaluated.After radiological evaluation of PV and SMV grades of infiltration, 21/56 (37 %) and 37/56 (66 %) patients, respectively, were found to have borderline resectable disease. The 4-point scale achieved a sensitivity of 80 %, a specificity of 96 % and an accuracy of 93 % in the evaluation of the PV, and a sensitivity of 100 %, a specificity of 94 % and an accuracy of 95 % in the evaluation of the SMV. Analysis of the distribution of clinical characteristics by PV and SMV infiltration showed that both scores correlated with the presence of distal metastasis (p = 0.016 and p = 0.028, respectively), and resection margins status (p = 0.015 and p = 0.006, respectively).This adapted tumour score is reliable for assessing venous invasion and might improve preoperative staging in patients with borderline resectable pancreatic cancer.

Controversy prevails on the impact of preoperative biliary drainage (PBD) on postoperative complications and clinical outcome of pancreatic cancer. We determined whether PBD is associated with increased morbidity and mortality rates after pancreaticoduodenectomy.A total of 131 consecutive patients who underwent pancreaticoduodenectomy (93 jaundiced, 38 with no jaundice) were included in this study. Overall, 57 % of jaundiced patients underwent PBD, while 43 % were not drained. The impact of PBD on postoperative morbidity and mortality was evaluated by means of logistic regression analysis. The Kaplan-Meier method was applied to determine the effect of PBD on survival of patients with malignant lesions.Mortality and morbidity rate was 3 % and 54.6 %, respectively. PBD was demonstrated to be the unique predictor of complications (odds ration [OR] = 10.18; 95 % confidence interval [CI], 3.65-28.39, p 0.001). The jaundiced patients who were drained exhibited high frequencies of wound infection (p 0.001), post-pancreatectomy haemorrhage (p = 0.0185) and hyperglycaemia (p 0.001). In addition, an increased frequency of pancreatic fistula emerged among drained patients compared to those who were not drained (p = 0.036). PBD did not affect survival of patient with malignant lesions.With the exception of the classical indications, PBD should be carefully evaluated in patients with resectable pancreatic cancer.

Colorectal carcinogenesis relies on loss of homeostasic mechanisms regulating cell proliferation, differentiation and survival. These cell processes have been reported to be influenced independently by transcription factors activated downstream of the Wnt pathway, such as SOX9 and β-catenin, and by the nuclear receptor PPARγ. The purpose of this study was to explore the expression levels and functional link between SOX9, β-catenin and PPARγ in the pathogenesis of colorectal cancer (CRC). We evaluated SOX9, β-catenin and PPARγ expression levels on human CRC specimens by qPCR and immunoblot detection. We tested the hypothesis that PPARγ activation might affect SOX9 and β-catenin expression using four colon cancer cell lines (CaCo2, SW480, HCT116, and HT29 cells). In CRC tissues SOX9 resulted up-regulated at both mRNA and protein levels when compared to matched normal mucosa, β-catenin resulted up-regulated at protein levels, while PPARG mRNA and PPARγ protein levels were down-regulated. A significant relationship was observed between high PPARG and SOX9 expression levels in the tumor tissue and female gender (p=0.005 and p=0.04, respectively), and between high SOX9 expression in the tumor tissue and age (p=0.04) and microsatellite instability (MSI), in particular with MSI-H (p=0.0002). Moreover, treatment with the synthetic PPARγ ligand rosiglitazone induced different changes of SOX9 and β-catenin expression and subcellular localization in the colon cancer cell lines examined. In conclusion, SOX9, β-catenin and PPARγ expression levels are deregulated in the CRC tissue, and in colon cancer cell lines ligand-dependent PPARγ activation unevenly influences SOX9 and β-catenin expression and subcellular localization, suggesting a variable mechanistic role in colon carcinogenesis.

The objective of this study is to evaluate the impact of a fast-track protocol in a high-volume center for patients with pancreatic disorders.The concept of fast-track surgery allowing accelerated postoperative recovery is accepted in colorectal surgery, but efficacy data are only preliminary for patients undergoing major pancreatic surgery. We aimed to evaluate the impact of a modified fast-track protocol in a high-volume center for patients with pancreatic disorders.Between February 2005 and January 2010, 145 subjects had resective pancreatic surgery and were enrolled in the program. Essential features of the program were no preanaesthetic medication, upper and lower air-warming device, avoidance of excessive i.v. fluids perioperatively, effective control of pain, early reinstitution of oral feeding, and immediate mobilization and restoration of bowel function following surgery. Outcome measures were postoperative complications such as pancreatic fistula, delayed gastric emptying, biliary leak, intra-abdominal abscess, post-pancreatectomy hemorrhage, acute pancreatitis, wound infection, 30-day mortality, postoperative hospital stay, and readmission rates.On average, patients were discharged on postoperative day 10 (range 6-69), with a 30-day readmission rate of 6.2%. Percentage of patients with at least one complication was 38.6%. Pancreatic anastomotic leakage occurred in seven of 101 pancreatico-jejunostomies, and biliary leak in three of 109 biliary jejunostomies. Postoperative hemorrhage occurred in ten (6.9%) patients and wound infection in nine (6.2%) cases. In-hospital mortality was 2.7%. Fast-track parameters, such as normal food and first stool, correlated significantly with early discharge (0.05). At multivariate analysis, lack of jaundice, and resumption of normal diet by the 5th postoperative day were independent factors of early discharge.Fast-track programs are feasible, easy, and also applicable for patients undergoing a major surgery such as pancreatic resection.

Hepato-pancreatico-biliary (HPB) surgery encompasses major hepatic resection and pancreatic surgery, both procedures of high complexity with a potentially high complication rate. The establishment of centres of excellence with a high patient volume has lowered the complication and increased the resection rate. Besides this, increased life expectancy and improved general health status have increased the number of elderly patients eligible for major surgery. Because elderly patients have more co-morbidities and decreased life expectancy, the benefit of these procedures must be critically evaluated in such patients. Analysis of the literature on this subject demonstrated that pancreatico-duodenectomy can be performed safely in selected elderly patients (80 years of age or older), with morbidity and mortality rates approaching those observed in younger patients. This aspect was also confirmed by cost analysis studies that reported similar data in both groups. Similar findings are also reported for major hepatic resection in elderly patients with either hepatocellular carcinoma (HCC), Klatskin tumour or gallbladder carcinoma. Nevertheless, those elderly patients who will benefit from surgery must be critically selected.

Inflammatory bowel disease consists of Crohn’s disease (CD) and ulcerative colitis (UC). A major clinical problem in some patients is to differentiate clearly between these entities, which is important when planning appropriate medical and surgical treatment. Connective tissue growth factor (CTGF), a novel peptide involved in fibrotic disorders, was analyzed in the present study in CD and UC patients to evaluate its possible role in these two disorders. Twenty-five normal human intestinal tissue samples were obtained through an organ donor program. CD tissues were obtained from 28 individuals undergoing partial intestinal resection (17 small bowel; 11 large bowel) due to complications of the disease. UC tissue samples were obtained from 16 patients undergoing colectomy due to complications of the disease. Expression of CTGF was studied by Northern blot analysis. In situ hybridization was used to localize mRNA moieties in the tissue samples. Northern blot analysis revealed an average 5-fold increase in CTGF mRNA expression in 89% (25/28) of CD tissue samples by comparison with normal controls (p < 0.0001). In contrast, in UC samples CTGF mRNA levels were comparable to those of normal controls. However, UC tissue samples exhibited enhanced TGF-β1 mRNA levels (4-fold; p < 0.05). In situ hybridization in CD samples showed CTGF mRNA localized especially in fibroblasts within the submucosal layer, around lymph follicles and in some areas of intense damage in the proximity of the luminal surface, whereas inflammatory cells were devoid of any CTGF mRNA signal. The present data indicate that CTGF plays a different role in IBD and might be useful, especially in those cases with unusual disease presentation, to better differentiate UC and CD. In addition, our data indicate a crucial role for CTGF in CD, where fibrosis and stenosis are frequent complications that require surgery.

To evaluate the parameters that mediate fibrogenesis in chronic pancreatitis (CP).Connective tissue growth factor (CTGF), which is regulated by transforming growth factor beta (TGF-beta), has recently been implicated in skin fibrosis and atherosclerosis. In the present study, the authors analyzed the concomitant presence of TGF-beta1 and its signaling receptors-TGF-beta receptor I, subtype ALK5 (TbetaR-I(ALK5)), and TGF-beta receptor II (TbetaR-II)-as well as CTGF and collagen type I in the pancreatic tissue of patients undergoing surgery for chronic pancreatitis.CP tissue samples were obtained from 40 patients (8 women, 32 men) undergoing pancreatic resection. Tissue samples of 25 previously healthy organ donors (12 women, 13 men) served as controls. The expression of TGF-beta1, TbetaR-I(ALK5), TbetaR-II, CTGF, and collagen type I was studied by Northern blot analysis. By in situ hybridization and immunohistochemistry, the respective mRNA moieties and proteins were localized in the tissue samples.Northern blot analysis showed that CP tissue samples exhibited concomitant enhanced mRNA expression of TGF-beta1 (38-fold), TbetaR-II (5-fold), CTGF (25-fold), and collagen type I (24-fold) compared with normal controls. In addition, TbetaR-I(ALK5) mRNA was increased in 50% of CP tissue samples (1.8-fold). By in situ hybridization, TGF-beta1, TbetaR-I(ALK5), and TbetaR-II mRNA were often seen to be colocalized, especially in the ductal cells and in metaplastic areas where atrophic acinar cells appeared to dedifferentiate into ductal structures. In contrast, CTGF was located in degenerating acinar cells and principally in fibroblasts surrounding these areas. Moreover, CTGF mRNA expression levels correlated positively with the degree of fibrosis in CP tissues.The concomitant overexpression of CTGF, collagen type I, TGF-beta1, and its signaling receptors in CP suggests that these proteins contribute to enhanced extracellular matrix synthesis and accumulation, resulting finally in the fibrogenesis observed in CP.