Christopher Marshall

Hepatitis C Virus Core and Nonstructural Protein 3 Proteins Induce Pro- and Anti-inflammatory Cytokines and Inhibit Dendritic Cell Differentiation

Antiviral immunity requires recognition of viral pathogens and activation of cytotoxic and Th cells by innate immune cells. In this study, we demonstrate that hepatitis C virus (HCV) core and nonstructural protein 3 (NS3), but not envelope 2 proteins (E2), activate monocytes and myeloid dendritic cells (DCs) and partially reproduce abnormalities found in chronic HCV infection. HCV core or NS3 (not E2) triggered inflammatory cytokine mRNA and TNF-α production in monocytes. Degradation of I-κBα suggested involvement of NF-κB activation. HCV core and NS3 induced production of the anti-inflammatory cytokine, IL-10. Both monocyte TNF-α and IL-10 levels were higher upon HCV core and NS3 protein stimulation in HCV-infected patients than in normals. HCV core and NS3 (not E2) inhibited differentiation and allostimulatory capacity of immature DCs similar to defects in HCV infection. This was associated with elevated IL-10 and decreased IL-2 levels during T cell proliferation. Increased IL-10 was produced by HCV patients’ DCs and by core- or NS3-treated normal DCs, while IL-12 was decreased only in HCV DCs. Addition of anti-IL-10 Ab, not IL-12, ameliorated T cell proliferation with HCV core- or NS3-treated DCs. Reduced allostimulatory capacity in HCV core- and NS3-treated immature DCs, but not in DCs of HCV patients, was reversed by LPS maturation, suggesting more complex DC defects in vivo than those mediated by core or NS3 proteins. Our results reveal that HCV core and NS3 proteins activate monocytes and inhibit DC differentiation in the absence of the intact virus and mediate some of the immunoinhibitory effects of HCV via IL-10 induction.

Additive inhibition of dendritic cell allostimulatory capacity by alcohol and hepatitis C is not restored by DC maturation and involves abnormal IL-10 and IL-2 induction

BACKGROUND Excessive alcohol use results in impaired immunity, and it is associated with increased incidence and progression of chronic hepatitis C virus (HCV) infection. Here we investigated the effects of HCV infection and alcohol on myeloid dendritic cells (DC) that are critical in antiviral immunity. METHODS Immature and mature DCs were generated from monocytes of chronic HCV infected patients (HCV-DC) and controls (N-DC) with IL-4 plus granulocyte-macrophage colony stimulating factor (GM-CSF) in the presence or absence of alcohol (25 mM). DC allostimulatory capacity was tested in mixed lymphocyte reaction (MLR) and cytokine production by ELISA. RESULTS Allostimulatory capacity of HCV-DCs was reduced compared to N-DCs and it was further inhibited by alcohol treatment (p < 0.01). MLR was also decreased with alcohol-treated N-DCs. DC phenotypic markers and apoptosis were comparable between HCV-DCs and N-DCs irrespective of alcohol treatment. However, HCV-DCs and alcohol-treated N-DCs exhibited elevated IL-10 and reduced IL-12 production. Reduced MLR with HCV-DCs and its further inhibition by alcohol coexisted with decreasing IL-2 levels (p < 0.017). DC maturation partially improved but failed to fully restore the reduced allostimulatory function of either alcohol-treated or alcohol-naïve HCV-DCs (p < 0.018). CONCLUSIONS Alcohol and HCV independently and together inhibit DC allostimulatory capacity, increase IL-10, reduce IL-12 and IL-2 production that cannot be normalized by DC maturation. HCV and alcohol interact to modulate innate and adaptive immune responses via dendritic cells.

Timing of video capsule endoscopy relative to overt obscure GI bleeding: implications from a retrospective study

BACKGROUND Diagnostic yield of video capsule endoscopy (VCE) may be higher if it is performed closer to the time of overt obscure GI bleeding (OOGIB). OBJECTIVE To evaluate the diagnostic yield of VCE and rate of therapeutic intervention for OOGIB for inpatients and outpatients with respect to timing of the intervention. DESIGN Retrospective cohort study. SETTING Tertiary academic center. PATIENTS Patients who had VCE for OOGIB between August 2008 and August 2010. INTERVENTIONS VCE for inpatients versus outpatients. MAIN OUTCOME MEASURES Diagnostic yield and rate of therapeutic intervention for inpatients versus outpatients. RESULTS One hundred forty-four inpatients (65 women) and 116 outpatients (49 women) were included. Diagnostic yield was 65.9% for inpatients versus 53.4% for outpatients (P = .054). Inpatients were divided into those who had VCE within 3 days (<3 days; n = 90) of admission versus after 3 days (>3 days; n = 54). Active bleeding and/or an angioectasia was found in 44.4% of the <3-day group compared with 27.8% of the >3-day group (P = .046) versus 25.8% of the outpatients. Therapeutic intervention was performed in 18.9% of the <3-day group versus 7.4% of the >3-day group (P = .046) versus 10.3% of outpatients. Diagnostic yield and therapeutic intervention rate between the >3-day group and outpatients were not significantly different. Length of stay (days) was less in the <3-day cohort, at 6.1 versus 10.3 in the >3-day cohort (P < .0001). LIMITATIONS Long-term outcomes were not studied. This was a retrospective study. CONCLUSIONS Early deployment of VCE within 3 days of admission results in a higher diagnostic yield and therapeutic intervention rate and an associated reduction of length of stay.

Type III Interferons, IL-28 and IL-29, Are Increased in Chronic HCV Infection and Induce Myeloid Dendritic Cell-Mediated FoxP3+ Regulatory T Cells

Background & Aims Hepatitis C virus (HCV) is difficult to eradicate and type III interferons (IFN-λ, composed of IL-28A, IL-28B and IL-29) are novel therapeutic candidates. We hypothesized that IFN-λ have immunomodulatory effects in HCV- infected individuals. Materials and Methods We analyzed the expression of IFN-λ and its receptor (composed of IL-10R2 and IFN-λR subunits) in the blood and livers of patients with chronic (c)HCV infection compared to controls (those who cleared HCV by sustained virological response, SVR, and those with liver inflammation of non-viral origin, non-alcoholic steatohepatitis, NASH). We also compared the proliferative capacity of dendritic cells (DCs) obtained from healthy individuals and those with chronic HCV using a mixed leukocyte reaction combined with 3H-Td incorporation. In addition, the composition of the IFN-λ receptor (IFN-λR) on myeloid DCs, plasmacytoid DCs, PBMCs, and T cells was determined by FACS analysis. Results We report that the expression of IFN-λ protein in serum and mRNA in liver is increased in cHCV patients, but not in those with HCV SVR or NASH, compared to controls. Liver level of IFN-λR mirrored the expression of serum IFN-λ and was higher in cHCV, compared to controls and HCV-SVR patients, suggesting that elevation of IFN-λ and IFN-λR are HCV-dependent. We further identified that innate immune cell populations expressed complete IFN-λ receptor. In vitro, recombinant IFN-λ promoted differentiation of monocyte-derived dendritic cells (DCs) into a phenotype with low T cell stimulatory capacity and high PD-L1 expression, which further promoted expansion of existing regulatory T cells. IFN-λ-DCs failed to induce de novo generation of regulatory T cells. The inhibitory capacity of IFN-λ-DCs was counteracted by recombinant IL-12 and by neutralization of the PD-1/PD-L1 system. Conclusions Our novel findings of the immunomodulatory effect of IFN-λ contribute to the understanding of the anti-inflammatory and/or anti-viral potential of IFN-λ in cHCV.

Microscopic colitis: A review of etiology, treatment and refractory disease

Microscopic colitis is a common cause of chronic, nonbloody diarrhea. Microscopic colitis is more common in women than men and usually affects patients in their sixth and seventh decade. This article reviews the etiology and medical management of microscopic colitis. The etiology of microscopic colitis is unknown, but it is associated with autoimmune disorders, such as celiac disease, polyarthritis, and thyroid disorders. Smoking has been identified as a risk factor of microscopic colitis. Exposure to medications, such as non-steroidal anti-inflammatory drugs, proton pump inhibitors, and selective serotonin reuptake inhibitors, is suspected to play a role in microscopic colitis, although their direct causal relationship has not been proven. Multiple medications, including corticosteroids, anti-diarrheals, cholestyramine, bismuth, 5-aminosalicylates, and immunomodulators, have been used to treat microscopic colitis with variable response rates. Budesonide is effective in inducing and maintaining clinical remission but relapse rate is as high as 82% when budesonide is discontinued. There is limited data on management of steroid-dependent microscopic colitis or refractory microscopic colitis. Immunomodulators seem to have low response rate 0%-56% for patients with refractory microscopic colitis. Response rate 66%-100% was observed for use of anti-tumor necrosis factor (TNF) therapy for refractory microscopic colitis. Anti-TNF and diverting ileostomy may be an option in severe or refractory microscopic colitis.

Monitoring the response of large (>3 cm) and locally advanced (T3-4, N0-2) breast cancer to neoadjuvant chemotherapy using 99mTc-Sestamibi uptake

Background and aim99mTc-Sestamibi (MIBI) scintimammography has an established role in the diagnosis of breast cancer. As a functional imaging technique, it may also be useful in assessing the response to chemotherapy. The aim of this study was to assess the effectiveness of the technique for this purpose. MethodsTwenty-six patients undergoing neoadjuvant chemotherapy for large or locally advanced breast cancer were monitored using the tumour to background ratio measured on MIBI scintimammograms. Patients were assessed and the size of the tumour was measured by callipers and ultrasonography before and at the end of treatment. Patients were assessed as complete, partial or non-responders. Following chemotherapy, patients proceeded to surgery with pathological evaluation of the operative specimen. ResultsTwenty-four of the 26 patients showed a reduction in MIBI uptake on completion of chemotherapy. Residual tumour was demonstrated on the scintimammogram in four patients and all had significant residual disease on histology. In the remaining 22 patients, the final scintimammogram was negative, although three patients were assessed as non-responders and had large residual tumours on histology. ConclusionA positive MIBI scan is highly predictive of the presence of significant residual disease on completion of chemotherapy. However, a negative MIBI scan does not rule out the presence of considerable residual tumour. Whereas ultrasound and clinical assessment may underestimate the response to chemotherapy, MIBI imaging tends to overestimate the response.

Comparison between Tc-HMPAO labelled white cells and Tc LeukoScan in the investigation of inflammatory bowel disease.

Objectives99mTc-HMPAO labelled leukocyte imaging (white cell scan) is used in the investigation and assessment of patients with inflammatory bowel disease (IBD), while 99mTc LeukoScan has been used to demonstrate the presence of orthopaedic infections and appendicitis. The single-injection technique used for the administration of LeukoScan removes the need for handling blood and the complex separation and labelling techniques that are required for white cell imaging. This study set out to investigate whether LeukoScan could be used to image IBD. MethodsTwenty-two patients under investigation for suspected IBD agreed to undergo LeukoScan imaging within 7 days of their routine planar white cell scan. Following injection of 650 MBq 99mTc LeukoScan planar images were acquired at 1, 2 and 4 h and single photon emission computed tomography (SPECT) images at 4 h post-injection. ResultsWhen both sets of images were compared to the clinical diagnosis, the sensitivity and specificity were 0.88 and 0.83 for white cell imaging and 0.88 and 0.50 for LeukoScan SPECT at 4 h. The figures for the specificity and sensitivity of the LeukoScan 2 h and 4 h planar images were 0.44 and 1.00, and 0.75 and 0.50, respectively. When the LeukoScan and white cell images were compared, the 4 h LeukoScan SPECT images showed additional areas of uptake in 10 patients. ConclusionsThe study demonstrates that LeukoScan locates IBD. However, the sensitivity and specificity preclude its use for the routine detection of this disease. Comparisons with white cell scan images show that LeukoScan is taken up slowly in IBD and also appears in the normal bowel by 4 h.

Endoscopic Removal of Small Gastrointestinal Stromal Tumors: Can We GIST-ify the Risk?

GISTs are the most common subepithelial tumor of the stomach with a prevalence of 129 per one million [1]. Nearly 500–600 new cases are diagnosed per year in the United States according to the Surveillance, Epidemiology, and End Results (SEER) program of the National Cancer Institute’s (NCI) report in 1995 [2]. More recently, including incidental findings, it is estimated that there are 3,000–6,000 new cases per year in the United States [3]. As more of these lesions are found, a strategy to manage these will need to be developed. A recent AGA technical review suggests observation for GISTs less than 3 cm without high risk features [4]. In this month’s issue of Digestive Diseases and Sciences, Bai et al. present an interesting article on the endoscopic removal of small GIST tumors (\30 mm) [5]. While the authors report a technical success with removal, we should be asking ourselves ‘‘should we,’’ rather than ‘‘can we.’’ To answer this question, we need to have a better understanding of the diagnosis, prognosis, and management of these lesions.

Optimization of %retention and %washout measurements of 99mTc sestamibi from breast tumours

Aim:Quantifying the %retention and %washout (%R&%W) of 99mTc sestamibi (MIBI) in breast cancer may allow prediction of response to chemotherapy. Many authors have investigated these parameters utilizing a wide variety of methods of quantification. The aim of this study is to determine the most accurate method of quantification. Method:Using an anthropomorphic phantom and a variety of tumour efflux rates, %R&%W were calculated using tumour only counts (T), tumour-to-background ratios (TBR), normalized TBR (nTBR) and background corrected counts (T - B) using images acquired at 0, 60, 120, 180 and 240 min post-injection. Results:T - B was the most accurate method of quantifying both %R&%W whilst TBR was the most inaccurate. The use of T was accurate when the tumour efflux rate was close to that of the normal breast and the inaccuracy increased as the difference between the tumour and normal breast efflux rates increased. The reverse was true when using nTBR as the accuracy increased as the difference between the efflux rates of tumour and normal breast increased. Time of image acquisition did not significantly alter accuracy. Conclusion:Background subtracted tumour counts must be used when quantifying the %retention or %washout of MIBI in breast cancer.

Preparation for video capsule endoscopy: A clear choice?

When we address the topic of appropriate preparation for VCE, we need to consider several factors: indication, timing, and tolerance. Since its introduction in 2001, wireless video capsule endoscopy (VCE) has proved to be an important tool for the examination of the small intestine. With its expanded on-label and off-label uses, VCE has become a part of both community and academic practices. Despite its widespread use, there are still unanswered questions regarding preparation. Society guidelines on the use of VCE have failed to conclusively recommend for and against the use of purgative preparation as a prelude to VCE, whereas 2 meta-analyses are supportive of its use. When we address the topic of appropriate preparation for VCE, we need to consider several factors: indication, timing, and tolerance. We must first ask ourselves, what is the indication for the VCE? Are we looking for active bleeding, Crohn’s disease, a tumor, or something subtle such as an angioectasia or erosion? The need to ask these questions has been overlooked, largely because we are in the shadow of colonoscopy, which uniformly requires a good preparation for the detection of small polyps. Whether or not preparation of the small intestine is useful is a problem compounded by the difficulties of objectivity and consistency in the assessment of what constitutes a good preparation. Given the small intestinal length of 5 to 6 meters with obligatory contents of gastric, biliary, and small-bowel secretions, it is not possible to completely clean the small bowel under any conditions. The best solution to minimizing the problem of consistency and objectivity is the use of computerized algorithms to assess the quality of preparation, as suggested by Van Weyenberg et al. This approach uses the coloration in the color bar to assess the preparation frame by frame. To date, the general question of whether preparation improves visualization of the small bowel is the only one that has been addressed, but little is known about whether preparation alters outcomes or whether there are benefits for selected patients. If the objective is detection of active bleeding, then small amounts of blood can clearly be washed away by purgative preparation, whereas large amounts of bleeding can readily be visualized under any preparatory conditions. Typically, in this group, fixed lesions such as angioectasia and erosions are much less of an issue, where the goal is planning deep enteroscopy. In fact, evolving data suggest