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The integrin αvβ6 is up‐regulated in numerous carcinomas, where expression commonly correlates with poor prognosis. αvβ6 promotes tumour invasion, partly through regulation of proteases and cell migration, and is also the principal mechanism by which epithelial cells activate TGF‐β1; this latter function complicates therapeutic targeting of αvβ6, since TGF‐β1 has both tumour‐promoting and ‐suppressive effects. It is unclear how these different αvβ6 functions are linked; both require actin cytoskeletal reorganization, and it is suggested that tractive forces generated during cell migration activate TGF‐β1 by exerting mechanical tension on the ECM‐bound latent complex. We examined the functional relationship between cell invasion and TGF‐β1 activation in pancreatic ductal adenocarcinoma (PDAC) cells, and confirmed that both processes are αvβ6‐dependent. Surprisingly, we found that cellular functions could be biased towards either motility or TGF‐β1 activation depending on the presence or absence of epidermal growth factor receptor pathway substrate 8 (Eps8), a regulator of actin remodelling, endocytosis, and GTPase activation. Similar to αvβ6, we found that Eps8 was up‐regulated in >70% of PDACs. In complex with Abi1/Sos1, Eps8 regulated αvβ6‐dependent cell migration through activation of Rac1. Down‐regulation of Eps8, Sos1 or Rac1 suppressed cell movement, while simultaneously increasing αvβ6‐dependent TGF‐β1 activation. This latter effect was modulated through increased cell tension, regulated by Rho activation. Thus, the Eps8/Abi1/Sos1 tricomplex acts as a key molecular switch altering the balance between Rac1 and Rho activation; its presence or absence in PDAC cells modulates αvβ6‐dependent functions, resulting in a pro‐migratory (Rac1‐dependent) or a pro‐TGF‐β1 activation (Rho‐dependent) functional phenotype, respectively. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

BACKGROUND and AIMS: Crohn's disease (CD) is associated with nutritional deficiencies, altered plasma concentrations of polyunsaturated fatty acids (PUFA) and an anti-inflammatory response to fish oil that contains n-3 PUFA. This suggests that, in CD, immune cells may have altered n-3 PUFA composition with functional consequences. The aim of this study is to investigate n-3 and n-6 PUFA composition and synthetic function of peripheral blood mononuclear cells (PBMC) in the basal state. METHODS: A case control study of 52 adult CD patients and healthy, age- and sex-matched controls. Composition of PBMC and plasma phospholipids were measured by gas chromatography and production of tumour necrosis factor-alpha, prostaglandin E2 (PGE2) and interferon-gamma (IFN-gamma) by PBMC were measured by ELISA. RESULTS: CD was associated with higher concentrations of eicosapentaenoic acid and other n-3 PUFA, and lower arachidonic acid (AA) (n-6 PUFA) in PBMC. This was not explained by differences in dietary fat intake. Lower rates of production of PGE2 and IFN-gamma by PBMC were noted in quiescent and active CD, respectively, compared to controls. CONCLUSIONS: CD is associated with a greater availability, and not a deficiency, of n-3 PUFA in PBMC, but lower concentrations of AA, and lower rates of production of PGE2 and IFN-gamma, compared to healthy controls.

The follow-up of inflammatory bowel disease (IBD) patients is challenging due to the relapsing remitting nature of the diseases, the wide spectrum of severity and complexity as well as the need for monitoring of long-term complications and drug treatments. Conventional outpatient follow-up lacks flexibility for patients and there are competing pressures for clinic time. Alternative follow-up pathways include telephone clinics, self-management programmes or discharging patients. The IBD virtual clinic (VC) is a further option. Patients with an established diagnosis for >2 years, who have been stable for >1 year, do not have primary sclerosing cholangitis and who give their consent, are entered into the VC system. Two months before their annual follow-up is due patients are sent blood test forms and a simple questionnaire with an information sheet. If they meet any of the criteria on the questionnaire, they are asked to contact the IBD specialist nursing team to discuss their situation. The blood test results and the patient's database entry are reviewed to ensure that they are not due surveillance investigations. The patients and their GPs then receive a letter informing them of their management plan. We currently follow-up 20% of the Southampton IBD cohort using the VC. The VC system is an innovative, efficient and patient-responsive method for following up mild to moderate IBD. It is well liked by patients but is dependent on a well-maintained database with good integration of IT systems and requires both clerical and IBD nurse specialist support.

An integrated pancreatic disease unit needs to deliver high-quality care both to patients with malignant and non-malignant pancreatic disease. The regionalisation of pancreatic cancer services which followed the publication of policy frameworks by the Department of Health and NHS executive led to the development of disease-site-specialised high-volume multidisciplinary teams. As the majority of patients with pancreatic cancer are not suitable for surgery, partner hospitals within a region need to provide access to a wide range of non-surgical treatment. The implementation of such working may require pooling of local resources to create networks of equivalence to tertiary centres. The provision of care to non-malignant pancreatic disease can benefit from this type of working and services can be modelled on, and integrate with, cancer services. One way of achieving this is to establish working groups based upon diseases rather than traditional departments, which can deliver standardised and optimal care with a patient-centred approach. However, this poses a number of potential problems. This review examines how an integrated pancreatic unit may be developed in district general and larger hospitals, and also describes our experience in developing such a unit.

Leukotrienes derived from the 5-lipoxygenase pathway are proinflammatory lipid mediators that possibly play a role in inflammatory bowel diseases. The expression of 5-lipoxygenase pathway proteins has not previously been examined in colonic mucosa in inflammatory bowel disease.Quantitative immunohistochemical analyses showed that, compared to those of the control subjects (n = 9), colonic biopsies from patients with active inflammatory bowel disease (n = 17) had 3- to 7-fold higher mean counts of cells expressing 5-lipoxygenase (P = 0.03), 5-lipoxygenase-activating protein (P = 0.005), and the leukotriene A(4) hydrolase (P = 0.004), which make up the biosynthetic pathway of the potent neutrophil chemotaxin leukotriene B(4). Immunoexpression of the leukotriene C(4) synthase was unaltered (P0.2). The increased representation of leukotriene B(4)-pathway enzymes was associated with higher counts of neutrophils (P = 0.0001), macrophages (P = 0.03), eosinophils (P = 0.0004), CD8(+) T cells (P0.001), activated T cells (P0.05), and B cells (P0.05) but not of mast cells (P0.9). These eicosanoid and cellular changes were most marked in the subgroup of patients with ulcerative colitis (n = 9), and were absent in patients with quiescent disease (n = 6). The anomalies in the 5-lipoxygenase pathway were accompanied as expected by more cells immunostaining for cytokine-inducible COX-2 (P = 0.004, n = 17), but this study also revealed a greater number of cells expressing COX-1 in the samples from the patients in the ulcerative colitis subgroup (P = 0.03, n = 9).The 5-lipoxygenase data provide a cellular basis for increased tissue synthesis of the leukotriene B(4), as reflected in the colonic mucosa and rectal dialysates of patients with active inflammatory bowel disease, which contributes to neutrophil influx and colonic injury. The COX-1/COX-2 data highlight the ambiguous functional role of prostanoid pathways in inflammatory bowel diseases.

Crohn's disease is associated with altered bone turnover that may be influenced by nutritional status, the systemic inflammatory response, cytokine production by circulating (peripheral blood) mononuclear cells (PBMC) and antioxidant micronutrient intake. High-dose fish oil is associated with reductions in disease relapse and inflammatory markers, and modulates PBMC function. The effect of fish oil plus antioxidants on bone turnover and PBMC function (the production of interferon-γ and prostaglandin E2) in Crohn's disease was investigated in a randomised-controlled trial. Patients with currently or recently raised biochemical markers of inflammation (C-reactive protein ≧6·9 mg/l or erythrocyte sedimentation rate ≧18 mm/h) received fish oil (providing 2·7 g/d EPA and DHA) and antioxidants (vitamins A, C and E, and Se) (n 31) or placebo (n 30) for 24 weeks. Bone turnover was assessed by measuring the concentrations of urinary deoxypyridinoline (bone resorption) and serum osteocalcin (bone formation). Fish oil plus antioxidants were associated with increases in EPA, DHA Se in plasma (all Pr −0·33, P=0·009). There were no differences between the groups at 24 weeks in the response of deoxypyridinoline or osteocalcin or their ratio, or in nutritional status. Dietary supplementation in Crohn's disease with high intakes of EPA and DHA, as fish oil, plus antioxidants was associated with a modulated production of interferon-γ by PBMC but not altered indices of bone turnover.

Purpose: The purpose of this study was to determine the role of functional interactions between pancreatic cancer cells and pancreatic stellate cells (PSCs) in the formation of the desmoplastic reaction (DR) in pancreatic cancer and to characterize the effect of type I collagen (the predominant component of the DR) on pancreatic cancer cell phenotype. Experimental Design: PSCs and type I collagen were identified in sections of pancreatic cancer using immunohistochemistry, and their anatomic relationship was studied. Interactions among pancreatic cancer cell lines (MIA PaCa-2, Panc-1, and AsPC-1), primary cultures of human PSCs, and type I collagen were investigated in a series of tissue culture models. Results: In vivo, the DR causes gross distortion of normal pancreas, bringing cancer cells into close contact with numerous PSCs and abundant type I collagen. In tissue culture models of pancreatic cancer, conditioned media from each cell line increased PSC [3H]thymidine incorporation up to 6.3-fold that of controls, and AsPC-1 cells also increased PSC collagen synthesis 1.3-fold. Type I collagen was observed to increase long-term survival of pancreatic cancer cells treated with 5-fluorouracil, by up to 62% in clonogenic assays. This was because type I collagen increased the proliferation of cancer cells ([3H]thymidine incorporation was up to 2.8-fold that of cells cultured on tissue culture plastic) and reduced apoptosis of AsPC-1 cells in response to 5-fluorouracil (by regulating mcl-1). Conclusions: These experiments elucidate a mechanism by which the DR in pancreatic cancer may form and, via the collagen within it, promote the malignant phenotype of pancreatic cancer cells, suggesting significant detriment to the host.

Background: Crohn's disease is associated with reduced bone density. The power of simple markers of systemic inflammation to identify higher rates of bone loss, in Crohn's disease, is uncertain. This relationship and the role of circulating (peripheral blood) mononuclear cells were investigated in a case-control study. Methods: Urinary deoxypyridinoline/creatinine and serum osteocalcin concentrations were compared in male and premenopausal females with "active" Crohn's disease (C-reactive protein > or = 10 and/or erythrocyte sedimentation rate > or = 20) (n = 22) and controls with "quiescent" Crohn's disease (C-reactive protein Results Active Crohn's disease was associated with a higher deoxypyridinoline/creatinine (P = 0.02) and deoxypyridinoline/creatinine:osteocalcin ratio (P =0.01) compared with quiescent Crohn's disease, but similar osteocalcin (P = 0.24). These were not explained by vitamin D status, dietary intake or nutritional status. However, production of interferon-gamma by concanavalin A-stimulated peripheral blood mononuclear cells was lower in active Crohn's disease (P = 0.02) and correlated negatively with the deoxypyridinoline/creatinine:osteocalcin ratio (r = -0.40, P = 0.004). Conclusion: In Crohn's disease, raised C-reactive protein and erythrocyte sedimentation rate may indicate higher rates of bone loss and, if persistent, the need to assess bone mass even where disease symptoms are mild. This may be partly explained by altered production of interferon-gamma by peripheral blood mononuclear cells

Background: Chronic pancreatitis is characterised clinically by early exocrine insufficiency, with diabetes mellitus occurring as a late phenomenon. This is mirrored pathologically by extensive acinar cell destruction and islet preservation. The mechanisms underlying this differential rate of cellular destruction are unknown. Aims: To test the hypothesis that acinar loss and islet preservation in chronic pancreatitis occurs due to differential epithelial kinetics and investigate the role of inflammatory cells and cell cycle associated molecules. Methods: Archival tissue from six chronic pancreatitis cases was compared with six normal controls using TUNEL and immunohistochemistry for CD3, CD20, CD68, MIB-1, Bcl-2, Bax, Fas, Fas ligand, retinoblastoma protein (Rb), and tissue inhibitor of metalloproteinases 1 (TIMP-1) and 2 (TIMP-2). Results: The acinar cell apoptotic index (AI) and proliferation index were higher in chronic pancreatitis than controls. T lymphocytes diffusely infiltrated fibrous bands and acini but rarely islets. Acinar Bcl-2 expression exceeded islet expression in chronic pancreatitis and controls while Bax was strongly expressed by a subset of islet cells and weakly by centroacinar cells. Islet Fas and Fas ligand expression exceeded acinar expression in chronic pancreatitis and controls. Acinar Rb expression was higher in chronic pancreatitis than in controls. Islets in chronic pancreatitis and controls showed intense TIMP-1 and TIMP-2 expression. Conclusion: Apoptosis plays a significant role in acinar loss in chronic pancreatitis. Acinar Bcl-2 and islet Bax expression indicates complex AI control. Increased acinar Rb expression in chronic pancreatitis may differentially promote acinar loss. Fas ligand expression may be restricted to islet cell membranes through TIMP-1 expression and inhibit islet damage by promoting apoptosis of cytotoxic T lymphocytes.

Background: Exocrine (acinar) destruction and endocrine (islet) preservation in chronic pancreatitis (CP) is related to differential apoptotic indices within these cellular compartments. Aim: To define the role of cell cycle regulatory proteins in the control of pancreatic epithelial cell apoptosis. Methods: Formalin-fixed, paraffin-embedded tissue from six cases of CP and eight normal controls (N) were studied using immunohistochemistry for BCl-2, Bax, retinoblastoma protein (Rb) and Fas ligand (Fas-L). Labelling patterns were assessed using semi-quantitative (intensity-proportion [I-P]) or fully quantitative (labelling index [LI]) analysis. Results: Rb protein was expressed at a higher level by acinar cells in CP than N (mean acinar cell Rb LI 1.45% in CP and 0.437% in N; p

This is an excerpt from the content: Pancreatic fecal elastase-1 (FE-1) has become the first-line test of pancreatic function. Our experience as a referral center for pancreatic disease suggests that in the minds of most gastroenterologists, a low FE-1 is diagnostic of pancreatic insufficiency. The paper by Evans and colleagues in the current issue of this journal highlights some weaknesses of that assumption in patients with Celiac disease. FE-1, first described by Balo and Banga in 1950 [1], is one of a family of protease enzymes secreted by acinar cells. FE-1 is well documented in quantitative studies to be stable during intestinal transit and studies demonstrate significant correlation between fecal elastase and levels of other pancreatic enzymes such as duodenal lipase, amylase, trypsin, and bicarbonate concentration [2–6]. Fecal elastase is concentrated in human feces compared with pancreatic juice and is a simple, non-invasive and inexpensive test [2, 7, 8]. As a result, FE-1 has replaced “tube tests” and fecal fat ...

The author gives an anecdotal account of his life with developmental prosopagnosia (DP). He was not formally diagnosed until the age of 53 and has evolved a complicated strategy for recognizing people based on non-facial physical features and context. He describes his experiences through infancy, school, university life and courtship, work and family life. He believes that he has lived a full and successful life despite DP but that some aspects of his social and work life were impaired by face-blindness. In his experience people react positively and helpfully if the consequences of DP are explained to them, and this improves social interactions and communications.

Pancreatic adenocarcinoma has one of the worse prognoses of any cancer with a 5-year survival of only 3%. Pancreatic cancer displays one of the most prominent stromal reactions of all tumors and it is evident that this is a key contributing factor to disease outcome. The tumor microenvironment of pancreatic cancer harbors a wide spectrum of cell types and a complex network of mechanisms which all serve to promote tumor progression. It is clear that the symbiotic relationship between pancreatic cancer cells and stellate cells is the chief factor creating this unique tumor milieu. Pancreatic stellate cells play critical roles in evasion of cancer cell apoptosis, invasion and metastases, angiogenesis, and promotion of an immunosuppressive environment, all key hallmarks of malignancy. Existing treatments for pancreatic cancer focus on targeting the cancer cells rather than the whole tumor, of which cancer cells represent a small proportion. It is now increasingly evident that research targeted towards the interactions between these cell types, ideally at an early stage of tumor development, is imperative in order to propel the way forward to more effective treatments.

He is a distinguished colleague and I am proud we are on first name terms: had I met him at the conference rather than the airport I would not have walked past. I was embarrassed, so my wife explained my problem, and his reply was unusual: “You must write a review for the BMJ so that others can understand your problem and benefit from your experience. Accounts of disability and how people cope are uplifting and help even those not afflicted.” I have severe inherited (developmental) prosopagnosia, or face blindness. The term prosopagnosia was coined in 1947, but it has been widely recognised outside the context of brain injury only in the past decade. Its purest form is limited to facial recognition, but I also have problems with inanimate and animate objects and in interpreting facial signs of emotion and sex. I often fail to recognise my children or even my wife. Prosopagnosia has shaped my life. At every stage I have failed to acknowledge friends and, more distressingly, those …

Epidemiological studies have been published worldwide in recent decades describing the incidence, mortality, aetiology and trends of chronic pancreatitis. Accumulated evidence suggests that chronic pancreatitis is increasing in incidence and hospital admission rates are rising accordingly. Alcoholic chronic pancreatitis was previously more common in the developed world than elsewhere, but is now increasing worldwide due to growing per capita alcohol consumption in each nation. Supporting alcohol and smoking cessation in individual patients is essential to slow disease progression and improve overall health, as most patients will die of cirrhosis, cardiovascular disease or smoking related cancers rather than chronic pancreatitis. The socioeconomic impact of chronic pancreatitis is difficult to quantify as little data exists, however given the rising incidence the costs to health care and society are likely to increase. This chapter will describe the epidemiology and aetiology of chronic pancreatitis worldwide and discusses the factors that influence its socioeconomic impact.

Aims: To test the hypothesis that single nucleotide polymorphisms (SNPs) within genes (or their promoter regions) encoding cytokines, growth factors, and intercellular adhesion molecules modulate the risk of development of chronic pancreatitis (CP). Methods: DNA was extracted from peripheral blood leucocytes or formalin fixed, paraffin wax embedded tissue from 53 patients with CP and 266 healthy controls. SNPs within the interleukin 1β (IL-1β), IL-6, IL-8, tumour necrosis factor α (TNFα) and vascular endothelial growth factor (VEGF) gene promoter regions and the transforming growth factor β1 (TGFβ1) and intercellular cell adhesion molecule 1 (ICAM-1) genes were genotyped by the amplification refractory mutation system polymerase chain reaction or 5′ nuclease (Taqman®) techniques. Patient–control comparisons were made using 2 × 2 contingency tables and χ2 analyses. Results: A non-significant decrease in the frequency of the IL-8 −251 AA genotype and a non-significant increase in the frequency of the ICAM-1 +469 GA genotype was seen in patients compared with controls. No associations were identified between SNPs in the promoter regions of the IL-1β, IL-6, or TNFα proinflammatory cytokines genes or the TGFβ1 and VEGF genes and susceptibility to CP. Conclusions: This preliminary study suggests that genetic polymorphism within several cytokine genes is unlikely to influence susceptibility to CP, but the possible role of IL-8 and ICAM-1 polymorphisms in the development of this disease requires further investigation.

BACKGROUND: Crohn disease (CD) is associated with osteoporosis and other extraintestinal manifestations that might be mediated by cytokines from circulating (peripheral blood) mononuclear cells (PBMCs). Fish oil rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) reduces disease activity in patients with CD with raised laboratory markers of inflammation and in healthy subjects alters PBMC function. OBJECTIVE: We investigated the effect of fish oil plus antioxidants on cytokine production by PBMCs from patients with CD with raised C-reactive protein concentrations (>/=6.9 mg/L) or erythrocyte sedimentation rates (>/=18 mm/h). DESIGN: A randomized placebo-controlled trial of fish oil (2.7 g EPA and DHA/d; n = 31) or placebo (olive oil; n = 31) for 24 wk was conducted in patients with CD. The fish-oil group additionally received an antioxidant preparation (vitamins A, C, and E and selenium). Exclusion criteria included corticosteroid use. Fatty acid composition was measured by gas chromatography. Production of tumor necrosis factor alpha, interferon gamma (IFN-gamma), and prostaglandin E(2) (PGE(2)) was measured by enzyme-linked immunosorbent assays after stimulation with mitogen and endotoxin (lipopolysaccharide). RESULTS: Fish-oil plus antioxidant dietary supplementation was associated with higher EPA and DHA incorporation into PBMCs (P < 0.001) and lower arachidonic acid (P = 0.006) and lower production of IFN-gamma by mitogen-stimulated PBMCs (P = 0.012) and of PGE(2) by lipopolysaccharide-stimulated PBMCs (P = 0.047). CONCLUSION: Dietary supplementation with fish oil plus antioxidants is associated with modified PBMC composition and lower production of PGE(2) and IFN-gamma by circulating monocytes or macrophages. The response of extraintestinal manifestations of CD should be investigated in a randomized controlled trial.

To the clinician, the fibrosis of chronic pancreatic pancreatitis appears relentless and unregulated. To an extent this is a feature of the natural history of the disease. Although the presentations of our patients vary, some with episodes of acute pancreatitis; some with chronic pain and others with exocrine and endocrine failure; however, all present with established fibrosis. Whilst we may witness the progression of this disease over many years we do not see the early stages. The reasons for this are both anatomical and physiological; by comparison with the liver or kidney, the pancreas is relatively difficult to image and biopsy, and there are no routine blood tests to mark the early stages of dysfunction. As a result our understanding of the processes initiating and driving fibrosis in the pancreas is relatively poor. However, there are some clues.

Pancreatic stellate cells mediate fibrosis in chronic pancreatitis. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs)-1 and -2 are crucial modulators of fibrosis. Transforming growth factor-beta (TGF-beta) is a key regulator of extracellular matrix production and myofibroblast proliferation. We have examined MMP and TIMP synthesis by transformed cultured pancreatic stellate cells and their regulation by TGF-beta 1. By Northern analysis they expressed mRNAs for procollagen 1, TIMP-1, TIMP-2, and MMP-2. Expression of membrane type-1 MMP was confirmed by Western blotting. By immunohistochemistry these enzymes localized to fibrotic areas in human chronic pancreatitis. Active TGF-beta 1 constitutes 2 to 5% of total TGF-beta 1 secreted by pancreatic stellate cells; they express TGF-beta receptors I and II. Exogenous TGF-beta 1 (10 ng/ml) significantly increased procollagen-1 mRNA by 69% and collagen protein synthesis by 34%. Similarly TGF-beta 1 at 0.1, 1, and 10 ng/ml significantly reduced cellular proliferation rate by 37%, 44%, and 44%, respectively, whereas pan-TGF-beta-neutralizing antibody increased proliferation by 40%. TGF-beta1 (10 ng/ml) down-regulated MMP-9 by 54% and MMP-3 by 34% whereas TGF-beta 1-neutralizing antibody increased MMP-9 expression by 39%. Pancreatic stellate cells express both mediators of matrix remodeling and the regulatory cytokine TGF-beta 1 that, by autocrine inhibition of MMP-3 and MMP-9, may enhance fibrogenesis by reducing collagen degradation.

s / Pancreatology 14 (2014) S1eS129 S72 MDSCs expansion and imbalance is in part Smad4-dependent and requires the co-presence of exosomes and soluble factors.

Background Tumour cell motility requires reorganisation of the actin cytoskeleton. The epidermal growth factor receptor pathway substrate 8 (EPS8) regulates actin remodelling, through interactions with numerous intracellular proteins, including integrins. The integrin αvβ6 is overexpressed in many carcinomas, and promotes two key features of pancreatic cancer—aggressive local invasion, and activation of transforming growth factor β1 (TGFβ1) leading to desmoplasia. We aimed to investigate the role of EPS8 in αvβ6-dependent functions in pancreatic cancer. Methods We used immunohistochemistry to examine the expression of αvβ6, EPS8, and the EPS8-binding partners SOS1 and ABI1 in human pancreatic cancer specimens using tissue microarrays. Cell motility was investigated using transwell assays and organotypic cultures after RNA interference. Primary pancreatic stellate cells were isolated from fresh tissue to optimise these organotypic cultures. TGFβ1 activation was measured using mink lung epithelial cells expressing a TGFβ1-responsive luciferase construct. Findings αvβ6, EPS8, SOS1, and ABI1 were upregulated in more than 70% of human pancreatic cancer tissues compared with neighbouring non-cancerous pancreatic tissue. αvβ6-dependent cell motility in panceatic cancer was dependent on EPS8, SOS1, and the small GTPase Rac1. Knockdown of any of these proteins inhibited cell motility. Conversely, EPS8 knockdown promoted αvβ6-dependent TGFβ1 activation, in a Rho-dependent manner. Interpretation EPS8 promotes motility in pancreatic cancer, modulated through αvβ6, yet inhibits αvβ6-dependent TGFβ1 activation. EPS8 might therefore function as a master regulator of cell invasion in pancreatic cancer by acting as a molecular switch, differentially regulating small GTPases (Rac1 and Rho) and thereby altering cell function between cell motility and TGFβ1 activation. Funding UK Medical Research Council.

Patients on long-term oral corticosteroids have an increased risk of low bone mass and fragility fractures. Fracture risk rises soon after commencement of corticosteroid therapy and it is possible that these agents adversely influence bone architecture disproportionately to their effect on bone mass. The best means of assessing bone status in patients using corticosteroids remains uncertain, but quantitative ultrasound of the calcaneus may provide evidence of microarchitectural changes not detected by dual-energy X-ray absorptiometry (DXA). Patients with Crohn's disease have an increased risk of low bone mineral density (BMD), the etiology of which is multifactorial but includes corticosteroid use. We studied 118 consecutive patients with Crohn's disease, 21 of whom used continuous oral corticosteroids, 70 of whom were intermittent users, and 27 who had never used the drug. All patients received DXA of the lumbar spine, hip and calcaneus and quantitative ultrasound (QUS) of the calcaneus. The different techniques were compared using a femoral neck T-score < or = -1.5 as the threshold of corticosteroid-induced osteoporosis. When compared with the femoral neck T-score, there were no significant differences between the predictive values of lumbar spine DXA, calcaneal DXA or calcaneal QUS to identify low femoral neck BMD. However, the absolute T-score required to give similar discriminatory capacity to femoral neck T-score varied substantially (T= -0.81 to -1.5) between the different measurement techniques and sites.

We have examined the expression of p75, a member of the TNF receptor superfamily in hepatic stellate cells (HSC) and pancreatic stellate cells (PSC). Activated HSC and PSC were demonstrated by Western blot analysis to express p75. p75 was immunolocalized to cells with a myofibroblast-like morphology in the fibrotic bands of six fibrotic and cirrhotic liver biopsies and three biopsies of fibrotic human pancreas. Immunostaining of parallel sections indicated that these cells were α-smooth muscle actin-positive, identifying them as activated HSC and PSC, respectively. HSC apoptosis in tissue culture in the presence of serum was quantified after addition of 0.1 to 100 ng/ml of nerve growth factor (NGF) a ligand for p75, by in situ counting of apoptotic bodies after addition of acridine orange. HSC demonstrated a significant increase in apoptosis in response to 100 ng/ml NGF (0.05 > P by Wilcoxon's rank; n = 7) after 24 hours. NGF 100 ng/ml had no effect on HSC proliferation, but reduced total HSC DNA by 19% relative to control after 24 hours ( n = 3). These data demonstrate that activated HSC express p75 and respond to NGF stimulation by undergoing apoptosis. We therefore report p75 as a novel marker of activated HSC and suggest that signaling via ligand binding to p75 may provide a mechanism for selective apoptosis of HSC.

Introduction EGFR pathway substrate 8 (Eps8) is an adapter protein that sits at the heart of a complex system regulating re-organisation of the actin cytoskeleton. Eps8 has been shown to be involved in cell motility and EGFR internalisation, via interactions with multiple intracellular proteins, including integrins. αvβ6 is an epithelial specific integrin, not present in healthy tissue but over-expressed in many carcinomas. αvβ6 promotes tumour invasion and is a key activator of TGFβ, which modulates tumour cell EMT and the desmoplastic response characteristically present in PC. The aim of this study was to investigate whether there is a link between Eps8 and αvβ6 in pancreatic cancer invasion. Methods We used immunochemistry to examine the expression of Eps8 and αvβ6 in normal pancreas and PC in vivo using tissue microarrays. PC cell motility was investigated using Transwell® assays. siRNA interference was used to down-regulate genes of interest. The ability of PC cells to activate TGFβ was measured using TGFβ-responsive mink lung epithelial cells. Cell surface αvβ6 levels were assessed with FACS analysis. Endocytosis of αvβ6 was suppressed using Clathrin siRNA to examine the role of endocytosis in αvβ6 functions. Pancreatic stellate cells (PSCs) were isolated from primary pancreatic resection tissue and used to optimise 3D pancreatic organotypic cultures. Results Eps8 and αvβ6 were up-regulated in > 70% PC in vivo but were not generally detected in normal pancreas. An organotypic model of pancreatic cancer was developed using primary PSCs to study αvβ6 functions. αvβ6 promoted PC cell invasion and TGFβ activation. Interestingly, Eps8 knockdown suppressed αvβ6-dependent motility but conversely, promoted αvβ6-dependent TGFβ activation. Although inhibition of αvβ6 endocytosis also increased TGFβ activation, this was not Eps8 dependent, nor did Eps8 knockdown affect total cell surface levels of the integrin. Conclusion αvβ6 and Eps8 are over-expressed in PC. αvβ6 promotes both invasion and TGFβ activation, and Eps8 appears to act as a molecular switch between these functions. How this relates to disease progression is unclear; in late stage disease, where canonical TGFβ signalling is dysregulated (e.g. SMAD4 mutations which are seen in > 50% of PCs) both αvβ6 functions are likely to be tumour promoting. However, in premalignancy when canonical TGFβ signalling acts as a tumour suppressor, we speculate that up-regulation of Eps8 may result in a pro-invasive phenotype and facilitate tumour development. Eps8 may therefore act as a master regulator of PC invasion by switching the cell function between motility and TGFβ activation at critical time points in PC progression. Disclosure of Interest None Declared

Introduction It is suggested that T-lymphocytes play a role in the pathogenesis of chronic pancreatitis (CP), but little is known about the composition of T-cell subsets in this disease and previous studies have been discordant. 1–4 We therefore aimed to characterise T-helper (Th) and T-regulatory (Treg) lymphocytes in CP tissue and peripheral blood. Methods Peripheral blood mononuclear cells were isolated from 15 patients with CP (all male, median age 48.2 years) and 14 controls (10 male, four female, median age 56.1 years). Mononuclear cells were also isolated from the pancreatic tissue of four CP patients (all male, median age 41 years) using enzymatic and mechanical digestion, followed by density gradient centrifugation. The mononuclear cells were stimulated with PMA and ionomycin (not Treg cells), and analysed using a FACSAria flow cytometer. Live CD3 + CD4 + Th1, Th2, Th17 and Treg cells were identified as IFN-γ + , IL-13 + , IL-17 + (IFN-γ ± ), and CD25 + FoxP3 + CD127 lo/− respectively. Statistical analysis was performed using a Mann–Whitney U Test. Results The peripheral blood of CP patients comprised a significantly higher percentage of Th1 cells (15.2% vs 8.11%; p=0.03), Th2 cells (2.00 % vs 1.17%; p=0.03), Th17 cells (1.23% vs 0.41%; p=0.003), dual secreting IFN-γ + IL-17 + Th17 cells (0.11% vs 0.03%; p=0.003) and Treg cells (6.30% vs 4.40%; p=0.05) compared to controls. CP patients who consume excess alcohol have significantly more Th1 cells than non-drinkers (23.7% vs 9.81%; p=0.01). The T-helper cell infiltrate in CP tissue was mainly composed of Th1 cells (26.1%–57.2%, median 41.5%). Th17 cells were also seen (0.95%–3.80%, median 2.1%), including IFN-γ + IL-17 + Th17 cells (0.22%–2.39%, median 0.37%). No discernible Th2 cells were identified and few or no Treg cells were seen in CP tissue. Conclusion This work is the first to demonstrate a significant increase in the number of Th17 cells in the peripheral blood of CP patients, and to clearly demonstrate that Th1 cells are the principal T-helper cell found in CP tissue along with appreciable numbers of Th17 cells. Interestingly there is no polarisation of the peripheral blood T-helper cell response in CP towards either a Th1 or Th2 phenotype. It appears therefore that the blood of CP patients is primed to respond non-specifically to inflammatory stimuli. Intriguingly CP patients who consume excess alcohol have more peripheral blood Th1 cells. Alcohol increases gut permeability causing high circulating levels of lipopolysaccharide which is known to generate Th1 cell responses. 5 These combined features may contribute to the pathogenesis of CP. Competing interests None declared. References 1. Hunger RE . Gastroenterology 1997. 2. Ebert MP . Am J Gastro 1998. 3. Miyoshi H . Pancreas 2008. 4. Schmitz-Winnenthal H . Gastroenterology 2010. 5. Bode C . J Hepatol 1987.

Introduction Mucosal-Associated Invariant T cells (MAIT) are recently described human T lymphocytes with possible functions in mucosal antibacterial host defence.1 MAIT express a highly conserved T cell receptor (TCR) alpha chain consisting of an invariant Vα7.2-Jα33 rearrangement.2 Recent evidence suggests a role for T lymphocytes in the pathogenesis of chronic pancreatitis (CP), but little is known about the composition of T cell subsets in this disease.34 Intestinal bacterial overgrowth is common in chronic pancreatitis patients.5 We hypothesised that this antigenic load may promote MAIT infiltration of the pancreas and used flow cytometry to study T lymphocytes in resected pancreatic tissue and blood of patients with chronic pancreatitis. Methods Fresh resected pancreatic tissue from five patients with CP (5 males, aged 35–62, median age 43) was diced finely, enzymatically digested and passed through a nylon mesh. Lymphocytes were then isolated using density gradient centrifugation. Peripheral blood mononuclear cells (PBMCs) were isolated from the blood of the CP patients and four control patients (2 male, 2 female, aged 31–75, median age 41) using widely established techniques. Isolated tissue lymphocytes and PBMCs were analysed with fluorochrome conjugated antibodies (anti- CD3; CD4; CD8; CD161; TCR Vα7.22 and a live/dead stain) according to standard techniques using a FACSAria. Results The ratio of CD4:CD8 T cells found in pancreatic tissue compared to blood in patients with CP trended towards a higher ratio in blood, however the trend was not statistically significant. Double negative (DN) and CD8 MAIT cells (TCR Vα7.2+ CD161) were identified in pancreatic tissue and blood in all patients. The proportion of DN MAIT cells was significantly higher in pancreatic tissue compared to blood in patients with CP: 19.4% (5.7–39.6%) versus 5.2% (1.9–11.1%); p Conclusion These preliminary data show that while the distribution of CD4 and CD8 lymphocytes in CP tissue and blood is similar, DN MAIT cells are over-represented in tissue. This suggests a specific role for DN MAIT cells in local immune responses in CP. Further analysis using a larger patient cohort and functional studies to assess the possible role of these cells in the regulation of pancreatic inflammation are required.

Introduction Pancreatic stellate cells (PSC) are exposed to supranormal concentrations of insulin via the islet–acinar portal system. We initially hypothesised that this might influence PSC numbers by effects on proliferation or apoptosis. This effect could be mediated via insulin receptors or via receptors for insulin-like growth factor (IGF-1). In this context, we examined the role of IGF-1 and insulin in the prevention of pancreatic stellate cell apoptosis. Following initial experiments, it became apparent that PSC can synthesise IGF-1, we therefore examined whether IGF-1 might be an autocrine growth factor in cultured stellate cells. The aims were to determine (1) the expression of IGF-1 and insulin receptor by PSC; (2) the secretion of IGF-1 by pancreatic stellate cells; (3) the effects of IGF-1 on PSC proliferation and apoptosis. Methods Culture-activated rat PSCs were used between passages 1 and 4 and serum deprived prior to all experiments. PSC proliferation was determined by [3H]-thymidine incorporation. Apoptosis was determined with acridine orange counting of apoptotic bodies and caspase 3 activity assays. Western blotting and immunocytochemistry were used to demonstrate the presence of IGF-1 and insulin receptors. Real-time quantitative PCR was used to detect receptor mRNA. A sandwich ELISA was used to determine the secretion of IGF-1, and real-time quantitative PCR use to confirm mRNA expression. Results Activated PSC express protein and mRNA for IGF-1 and insulin receptors. IGF-1 secretion is upregulated 2.40-fold following serum withdrawal compared with 16% FCS (see Abstract 001). This upregulation is also seen at the level of mRNA in response to serum withdrawal. Exogenous IGF-1 (100 ng/ml) and insulin (10 000 ng/ml) prevented cycloheximide-induced PSC apoptosis in a concentration-dependent manner, 63% at 24 h, and 71% at 24 h, respectively. There was moderate effect on PSC proliferation following exposure to IGF-1 but not with insulin. Conclusion These data suggest that IGF-1 has an autocrine role in reducing PSC apoptosis on serum withdrawal. Inhibiting IGF-1 expression or blocking the IGF-1 receptor may increase susceptibility to apoptosis. This represents a potential therapeutic target for prevention or treatment of pancreatic fibrosis.

BACKGROUND—Hepatitis E (HEV) is the primary cause of enterically transmitted non-A non-B hepatitis worldwide. Case reports of HEV in individuals in the United Kingdom relate to travel to endemic areas or contact with individuals who have visited these areas. CASE REPORTS—Four individuals presented with acute hepatitis to a hepatology clinic in a teaching hospital. Serology confirmed acute hepatitis E in all four. Investigation by the Communicable Disease Control Department established no links between the cases, no travel to an endemic area, and no contacts. CONCLUSION—Contrary to current belief, community acquired hepatitis E virus infection occurs sporadically in the United Kingdom and should be considered as a cause of seronegative hepatitis. Keywords: hepatitis E; viral hepatitis; epidemiology

This article describes changes in divorce laws in the United States, England and Wales, France, and Sweden in the areas of obtaining a divorce, spousal support, child support and child custody. Also, the article discusses the relation between individuals' values and their evaluations of the effects of changes in divorce laws on family members, as well as the relation between legal change and cultural values. In addition to providing valuable descriptive information about divorce laws from an international perspective, comparative analyses provide unique insights into the relation between culture and laws pertaining to divorce beyond that provided by an analysis of any single country. This analysis not only provides information about the laws of several Western countries, but also highlights the complex relation between cultural values and the legal aspects of divorce. To further the understanding of this complex relation, the authors' recommend that future researchers must incorporate non-Western countries into their analyses.

This article provides an update of the status of laws affecting stepfamilies. Four legal areas are reviewed: (a) stepparents supporting their stepchildren during the marriage and following divorce, (b) stepchildren inheriting from a deceased stepparent, (c) stepparents receiving custody of stepchildren following termination of the remarriage, and (d) stepparents visiting stepchildren following divorce. The review demonstrates that, in some areas more than others, legislatures and courts have become more sensitive to the possibly enduring nature of the stepparent-stepchild relationship. Suggestions for legal reform and implications for family professionals are discussed.