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Simple Summary: The standard interpretation methods of germline variants in cancer are limited due to overlapping features in constitutional and sporadically derived forms of cancers and the unavailability of key differentiating details in public settings. To address this challenge, we established INT2GRATE (INTegrated INTerpretation of GeRmline And Tumor gEnomes), a multi-institution oncology consortium to advance the integrated application of constitutional and tumor data and share the integrated variant data in a publicly accessible knowledgebase. The aim of our study is to introduce INT2GRATE|HPPGL, a platform for the integrated interpretation of hereditary paraganglioma–pheochromocytoma syndromes (HPPGL). We describe the details of the INT2GRATE|HPPGL Variant Evidence Framework for succinate dehydrogenase (SDHx) genes using key HPPGL personal and family history, as well as tumor-derived evidence. We applied the INT2GRATE|HPPGL Variant Evidence Framework to 8600 variants to programmatically process and share the integrated variant data in ClinVar using a custom-made INT2GRATE variant submission pipeline. This novel integrated variant assessment and data sharing in hereditary cancers is essential to help improve the clinical interpretation of genomic variants and advance precision oncology. Standard methods of variant assessment in hereditary cancer susceptibility genes are limited by the lack of availability of key supporting evidence. In cancer, information derived from tumors can serve as a useful source in delineating the tumor behavior and the role of germline variants in tumor progression. We have previously demonstrated the value of integrating tumor and germline findings to comprehensively assess germline variants in hereditary cancer syndromes. Building on this work, herein, we present the development and application of the INT2GRATE|HPPGL platform. INT2GRATE (INTegrated INTerpretation of GeRmline And Tumor gEnomes) is a multi-institution oncology consortium that aims to advance the integrated application of constitutional and tumor data and share the integrated variant information in publicly accessible repositories. The INT2GRATE|HPPGL platform enables automated parsing and integrated assessment of germline, tumor, and genetic findings in hereditary paraganglioma–pheochromocytoma syndromes (HPPGLs). Using INT2GRATE|HPPGL, we analyzed 8600 variants in succinate dehydrogenase (SDHx) genes and their associated clinical evidence. The integrated evidence includes germline variants in SDHx genes; clinical genetics evidence: personal and family history of HPPGL-related tumors; tumor-derived evidence: somatic inactivation of SDHx alleles, KIT and PDGFRA status in gastrointestinal stromal tumors (GISTs), multifocal or extra-adrenal tumors, and metastasis status; and immunohistochemistry staining status for SDHA and SDHB genes. After processing, 8600 variants were submitted programmatically from the INT2GRATE|HPPGL platform to ClinVar via a custom-made INT2GRATE|HPPGL variant submission schema and an application programming interface (API). This novel integrated variant assessment and data sharing in hereditary cancers aims to improve the clinical assessment of genomic variants and advance precision oncology. [ABSTRACT FROM AUTHOR]

The presence of variants of uncertain significance (VUS) in DNA mismatch repair (MMR) genes leads to uncertainty in the clinical management of patients being evaluated for Lynch syndrome (LS). Currently, there is no platform to systematically use tumor-derived evidence alongside germline data for the assessment of VUS in relation to LS. We developed INT2GRATE (INTegrated INTerpretation of GeRmline And Tumor gEnomes) to leverage information from the tumor genome to inform the potential role of constitutional VUS in MMR genes. INT2GRATE platform has two components: a comprehensive evidencebased decision tree that integrates well-established clinico-genomic data from both the tumor and constitutional genomes to help inform the potential relevance of germline VUS in LS; and a web-based user interface (UI). With the INT2GRATE decision tree operating in the backend, INT2GRATE UI enables the front-end collection of comprehensive clinical genetics and tumor-derived evidence for each VUS to facilitate INT2GRATE assessment and data sharing in the publicly accessible ClinVar database. The performance of the INT2GRATE decision tree was assessed by qualitative retrospective analysis of genomic data from 5057 cancer patients with MMR alterations which included 52 positive control cases. Of 52 positive control cases with LS and pathogenic MMR alterations, 23 had all the testing parameters for the evaluation by INT2GRATE. All these variants were correctly categorized as INT2GRATE POSITIVE. The stringent INT2GRATE decision tree flagged 29 of positive cases by identifying the absence or unusual presentation of specific evidence, highlighting the conservative INT2GRATE logic in favor of a higher degree of confidence in the results. The remaining 99% of cases were correctly categorized as INCONCLUSIVE due to the absence of LS criteria and =1 tumor parameters. INT2GRATE is an effective platform for clinical and genetics professionals to collect and assess clinical genetics and complimentary tumor-derived information for each germline VUS in suspected LS patients. Furthermore, INT2GRATE enables the collation of integrated tumor-derived evidence relevant to germline VUS in LS, and sharing them with a large community, a practice that is needed in precision oncology. [ABSTRACT FROM AUTHOR]

Background: Gastrointestinal immune‐related adverse events are frequently caused by immune checkpoint inhibitors (ICIs) and often require interruption of cancer treatment. Compared with ICI colitis and enteritis, limited information exists about ICI gastritis. This study characterized clinical features and treatment outcomes of ICI gastritis. Methods: Consecutive cancer patients who received ICIs and underwent endoscopy with gastric biopsies while on ICIs from 2011 to 2021 were retrospectively assessed. Specific histopathologic features identified ICI gastritis. Results: Of 6450 ICI‐treated patients, 162 (2.5%) underwent endoscopy with gastric biopsies. ICI gastritis was identified in 54 (33%) biopsied patients; 38 (70%) had concurrent ICI enteritis/colitis and 16 (30%) had isolated ICI gastritis. Dyspepsia (38%) and bloating (25%) were the most frequent symptoms of isolated ICI gastritis. Compared with patients with concomitant enteritis/colitis, patients with isolated gastritis were less likely to have diarrhea (13% vs 68%; p <.001) or abdominal pain (19% vs 47%; p =.07). Patients with isolated ICI gastritis less frequently required glucocorticoids (69% vs 92%; p =.04) and had lower incidence of ICI hold/withdrawal (13% vs 42%; p =.06). There was no association between severity or extent of luminal inflammation and antitumor response (p =.85 and p =.44, respectively). Endoscopically, gastric mucosa appeared normal in 11 (20%) patients with biopsy‐proven ICI gastritis. Conclusion: ICI gastritis may present alone or more commonly with concurrent enteritis/colitis, which may differentiate its clinical course. Gastric biopsies are required to diagnose a substantial minority of endoscopically normal, clinically significant cases. Most patients with isolated gastritis can continue ICI therapy uninterrupted, but a notable proportion require glucocorticoids. Plain language summary: Immune checkpoint inhibitors are effective anticancer treatments, but can cause inflammatory toxicities, including of the stomach (gastritis), intestine, and colon. Limited information is available on gastritis triggered by these agents.Adult patients with cancer who were treated with immune checkpoint inhibitors and had an upper gastrointestinal endoscopy with biopsies of the stomach were examined. More than two‐thirds (70%) of people with checkpoint inhibitor gastritis also had inflammatory changes of the small intestine and/or colon.Compared with patients with isolated checkpoint gastritis, the subgroup with concomitant enteritis/colitis more frequently had abdominal pain, diarrhea, needed steroids, and/or needed to pause or stop antitumor therapy. Immune checkpoint inhibitor (ICI) gastritis frequently presents with concomitant enteritis/colitis, which differentiates its clinical presentation and management from patients with isolated gastric involvement. A substantial minority of ICI gastritis cases are diagnosed histologically despite normal endoscopic appearance, highlighting the importance of gastric biopsies in patients with suspicious symptoms of or risk factors for checkpoint inhibitor gastritis. [ABSTRACT FROM AUTHOR]

Genomic profiles of tumors are often unique and represent characteristic mutational signatures defined by DNA damage or DNA repair response processes. The tumor-derived somatic information has been widely used in therapeutic applications, but it is grossly underutilized in the assessment of germline genetic variants. Here, we present a comprehensive approach for evaluating the pathogenicity of germline variants in cancer using an integrated interpretation of somatic and germline genomic data. We have previously demonstrated the utility of this integrated approach in the reassessment of pathogenic germline variants in selected cancer patients with unexpected or non-syndromic phenotypes. The application of this approach is presented in the assessment of rare variants of uncertain significance (VUS) in Lynch-related colon cancer, hereditary paraganglioma-pheochromocytoma syndrome, and Li-Fraumeni syndrome. Using this integrated method, germline VUS in PMS2, MSH6, SDHC, SHDA, and TP53 were assessed in 16 cancer patients after genetic evaluation. Comprehensive clinical criteria, somatic signature profiles, and tumor immunohistochemistry were used to re-classify VUS by upgrading or downgrading the variants to likely or unlikely actionable categories, respectively. Going forward, collation of such germline variants and creation of cross-institutional knowledgebase datasets that include integrated somatic and germline data will be crucial for the assessment of these variants in a larger cancer cohort. [ABSTRACT FROM AUTHOR]

Aim: Gastrointestinal stromal tumours (GISTs) typically show uniform nuclear morphology, with spindle cell, epithelioid or mixed histology. Risk of progression in GIST is estimated based on anatomical site, tumour size and mitotic index. Pleomorphic GISTs are rare and have not been systematically investigated. We evaluated the prognostic significance of pleomorphism in GIST. Methods and results: In total, 108 of 2517 (4.3%) GISTs reviewed between 2000 and 2021 were reported to show pleomorphism. Seventeen cases underwent mutational testing. Pleomorphism was noted prior to therapy in 37 GISTs, affecting 18 males and 19 females, with a mean age of 55.4 years. Most tumours arose in the stomach (n = 15) or small intestine (n = 19), with a mean size of 9.2 cm and a median mitotic rate of seven per 5 mm2; the median follow‐up was 5.7 years. Immunohistochemistry for KIT was positive in 36 (97.3%) tumours. Mutational testing revealed KIT and PDGFRA mutations in 68.4 and 21.0% of cases, respectively; no SDHX, KIT or PDGFRA alterations were found in two cases (one of which was succinate dehydrogenase‐deficient). According to standard risk assessment criteria for progressive disease, 18 tumours were high‐risk, five were moderate‐risk, four were low‐risk, five were very low‐risk and one had no risk. Disease progression was exclusive to high‐risk tumours (79%; P < 0.001). Conclusions: Pleomorphism is present prior to therapy in approximately 2% of GISTs and is most prevalent in high‐risk gastric and small intestinal GISTs. Pleomorphism appears to have no prognostic significance beyond conventional risk stratification. [ABSTRACT FROM AUTHOR]

Aims: Oesophageal verrucous carcinoma (VSCC) is a rare and morphologically distinct type of oesophageal squamous cell carcinoma (SCC). Diagnosing VSCC on biopsy material is challenging, given the lack of significant atypia and the presence of keratinising epithelium and exophytic growth. The molecular pathogenesis of VSCC remains unclear. The aim of this study was to characterise the genomic landscape of VSCC in comparison to conventional oesophageal SCC. Methods and results: Three cases of VSCC from the Brigham and Women's Hospital pathology archive were identified. Formalin‐fixed, paraffin‐embedded (FFPE) tumour tissue was used for p16 immunohistochemistry (IHC), high‐risk human papillomavirus (HPV) in‐situ mRNA hybridisation (ISH) and DNA isolation. Tumour DNA was sequenced using a targeted massively parallel sequencing assay enriched for cancer‐associated genes. Three additional cases of VSCC were identified by image review of The Cancer Genome Atlas (TCGA) oesophageal SCC cohort. VSCC cases were negative for p16 IHC and high‐risk HPV ISH. TP53 mutations (P < 0.001) and copy number variants (CNVs) for CDKN2A (P < 0.001), CDKN2B (P < 0.01) and CCND1 (P < 0.01) were absent in VSCC and significantly less frequent in comparison to conventional SCC. Five VSCC cases featured SMARCA4 missense mutations or in‐frame deletions compared to only four of 88 conventional SCC cases (P < 0.001). VSCC featured driver mutations in PIK3CA, HRAS and GNAS. Recurrent CNVs were rare in VSCC. Conclusions: VSCC is not only morphologically but also genetically distinct from conventional oesophageal SCC, featuring frequent SMARCA4 mutations and infrequent TP53 mutations or CDKN2A/B CNVs. Molecular findings may aid in establishing the challenging diagnosis of VSCC. [ABSTRACT FROM AUTHOR]

Aims: A varied spectrum of histopathological changes has been associated with immune checkpoint inhibitor (ICI) colitis. This study was performed to evaluate the prevalence of different histopathological patterns of injury in patients with ICI colitis and their association with specific immune check‐point inhibitors. Methods and results: Biopsies from patients with clinically and histologically confirmed ICI colitis were reviewed blindly to determine the predominant pattern of injury and to quantitate discrete histological parameters using the Geboes score. Paneth cell metaplasia, intraepithelial lymphocytes, abnormal subepithelial collagen and degree of crypt epithelial apoptosis was also recorded. A total of 86 patients with ICI colitis (ipilimumab, n = 14; ipilimumab + nivolumab, n = 29; nivolumab, n = 20 and pembrolizumab, n = 23) were included. The patterns of injury identified included diffuse active colitis (n = 22), chronic active colitis (n = 22), lymphocytic colitis (LC, n = 16), collagenous colitis (CC, n = 14), graft‐versus‐host disease‐like colitis (n = 7) and mixed colitis (n = 5). Patients on ipilimumab were more likely to have a diffuse active colitis pattern without features of chronicity (P < 0.01) and less likely to have LC (P < 0.05) compared to other ICIs. LC and CC were more common in patients on nivolumab and pembrolizumab relative to other groups (P < 0.05). Chronic active colitis was most frequent in nivolumab patients (P < 0.05), and these patients had received more ICI doses and had been on ICI treatment longer compared to other treatment groups. Conclusions: ICI colitis should be considered in the differential diagnosis of all the common inflammatory patterns of colitis and shows medication specific differences in patterns of injury. [ABSTRACT FROM AUTHOR]

Even though HIV-1 replication can be suppressed by combination antiretroviral therapy (cART) inflammatory processes still occur, contributing to comorbidities. Comorbidities are attributed to variety of factors, including HIV-1 mediated inflammation. Several HIV-1 proteins mediate central nervous system (CNS) inflammation, including Nef. Nef is an early HIV-1 protein, toxic to neurons and glia and is sufficient to cause learning impairment similar to some deficits observed in HIV-1 associated neurocognitive disorders. To determine whether hippocampal Nef expression by astrocytes contributes to comorbidities, specifically peripheral inflammation, we infused Sprague Dawley rats with GFP- (control) or Nef-transfected astrocytes into the right hippocampus. Brain, lung, and ileum were collected postmortem for the measurement of inflammatory markers. Increased blood-brain-barrier permeability and serum IL-1β levels were detected in the Nef-treated rats. The lungs of Nef-treated rats demonstrated leukocyte infiltration, macrophage upregulation, and enhanced vascular permeability. Ileal tissue showed reactive follicular lymphoid hyperplasia, increased permeability and macrophage infiltration. The intracerebroventricular application of IL-1 receptor antagonist reduced infiltration of immune cells into ileum and lung, indicating the important role of IL-1β in mediating the spread of inflammation from the brain to other tissues. This suggests that localized expression of a single viral protein, HIV-1 Nef, can contribute to a broader inflammatory response by upregulation of IL-1β. Further, these results suggest that Nef contributes to the chronic inflammation seen in HIV patients, even in those whose viremia is controlled by cART. [ABSTRACT FROM AUTHOR]

A 37-year-old woman with a history of inflammatory bowel disease on mercaptopurine presented with a week of recurrent fever, headache, myalgias and mildly elevated serum transaminases and leucopenia. Her workup revealed primary cytomegalovirus (CMV) infection with atypical lymphocytosis, elevated viral load, positive IgM and negative IgG. Two weeks after her initial presentation, she developed odynophagia and diarrhoea prompting endoscopic evaluation with biopsies, which demonstrated CMV disease of the gastrointestinal tract. Her fever and systemic symptoms improved rapidly with initiation of intravenous ganciclovir. She was transitioned to and maintained on oral valganciclovir until two and half months after discharge when her symptoms and lab abnormalities had fully subsided. [ABSTRACT FROM AUTHOR]

The probiotic mixture VSL#3 attenuates colitis in patients with Inflammatory Bowel Disease (IBD) and in animal models of this condition, but the mechanisms involved are incompletely understood. VSL#3 alters macrophage morphology and secretory profile in vitro in a polarization-dependent manner. We examined the effect of VSL#3 on macrophages in acute trinitrobenzene sulfonic acid-induced colitis. Rats were randomized to normal, colitis, or colitis+VSL#3 groups. After sacrifice, the colons were evaluated for macroscopic and microscopic damage. Serum cytokine levels were measured, and microbiome analysis undertaken. Total and M1 colonic macrophages, and total and proliferating hepatic macrophages were assessed by double immunofluorescence staining. Colitis+VSL#3 rats had lower macroscopic damage, with less microscopic damage in the proximal colon, compared with colitis alone. Colitis significantly increased colonic macrophage infiltration, which was significantly reduced by VSL#3 treatment. VSL#3 did not decrease the colitis-induced surge of colonic M1 macrophages or hepatic macrophages. VSL#3 reduced colitis-induced serum cytokine levels, and induced restoration of colonic transcript levels for pro-inflammatory, anti-inflammatory, and barrier proteins to, or past, normal levels. Fecal bacteria distribution changed between groups. In summary, the probiotic VSL#3 reduces colitis severity, colonic macrophage infiltration, and serum cytokine levels, but does not dampen the pro-inflammatory phenotype of M1 macrophages. [ABSTRACT FROM AUTHOR]

Our review focuses on the colonic macrophage, a monocyte-derived, tissue-resident macrophage, and the role it plays in health and disease, specifically in inflammatory conditions such as inflammatory bowel disease and cancer of the colon and rectum. We give special emphasis to macrophage polarization, or phenotype, in these different states. We focus on macrophages because they are one of the most numerous leukocytes in the colon, and because they normally contribute to homeostasis through an anti-inflammatory phenotype. However, in conditions such as inflammatory bowel disease, proinflammatory macrophages are increased in the colon and have been linked to disease severity and progression. In colorectal cancer, tumor cells may employ anti-inflammatory macrophages to promote tumor growth and dissemination, whereas proinflammatory macrophages may antagonize tumor growth. Given the key roles that this cell type plays in homeostasis, inflammation, and cancer, the colonic macrophage is an intriguing therapeutic target. As such, potential macrophage-targeting strategies are discussed. [ABSTRACT FROM AUTHOR]

The presence of variants of uncertain significance (VUS) in DNA mismatch repair (MMR) genes leads to uncertainty in the clinical management of patients being evaluated for Lynch syndrome (LS). Currently, there is no platform to systematically use tumor-derived evidence alongside germline data for the assessment of VUS in relation to LS. We developed INT 2 GRATE (INTegrated INTerpretation of GeRmline And Tumor gEnomes) to leverage information from the tumor genome to inform the potential role of constitutional VUS in MMR genes. INT 2 GRATE platform has two components: a comprehensive evidence-based decision tree that integrates well-established clinico-genomic data from both the tumor and constitutional genomes to help inform the potential relevance of germline VUS in LS; and a web-based user interface (UI). With the INT 2 GRATE decision tree operating in the backend, INT 2 GRATE UI enables the front-end collection of comprehensive clinical genetics and tumor-derived evidence for each VUS to facilitate INT 2 GRATE assessment and data sharing in the publicly accessible ClinVar database. The performance of the INT 2 GRATE decision tree was assessed by qualitative retrospective analysis of genomic data from 5057 cancer patients with MMR alterations which included 52 positive control cases. Of 52 positive control cases with LS and pathogenic MMR alterations, 23 had all the testing parameters for the evaluation by INT 2 GRATE. All these variants were correctly categorized as INT 2 GRATE POSITIVE. The stringent INT 2 GRATE decision tree flagged 29 of positive cases by identifying the absence or unusual presentation of specific evidence, highlighting the conservative INT 2 GRATE logic in favor of a higher degree of confidence in the results. The remaining 99% of cases were correctly categorized as INCONCLUSIVE due to the absence of LS criteria and ≥1 tumor parameters. INT 2 GRATE is an effective platform for clinical and genetics professionals to collect and assess clinical genetics and complimentary tumor-derived information for each germline VUS in suspected LS patients. Furthermore, INT 2 GRATE enables the collation of integrated tumor-derived evidence relevant to germline VUS in LS, and sharing them with a large community, a practice that is needed in precision oncology., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Isidro, Chittenden, Walker, Schwartz, Koeller, Hayes, Unal, Manam, Buehler, Manning, Sholl, Redston, Yurgelun, Rana, Garber and Ghazani.)

A 37-year-old woman with a history of inflammatory bowel disease on mercaptopurine presented with a week of recurrent fever, headache, myalgias and mildly elevated serum transaminases and leucopenia. Her workup revealed primary cytomegalovirus (CMV) infection with atypical lymphocytosis, elevated viral load, positive IgM and negative IgG. Two weeks after her initial presentation, she developed odynophagia and diarrhoea prompting endoscopic evaluation with biopsies, which demonstrated CMV disease of the gastrointestinal tract. Her fever and systemic symptoms improved rapidly with initiation of intravenous ganciclovir. She was transitioned to and maintained on oral valganciclovir until two and half months after discharge when her symptoms and lab abnormalities had fully subsided., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Aim: To determine serum vitamin D levels and colonic vitamin D receptor (VDR) expression in inflammatory bowel disease (IBD) and non-IBD patients and correlate these with histopathology., Methods: Puerto Rican IBD (n = 10) and non-IBD (n = 10) patients ≥ 21 years old scheduled for colonoscopy were recruited. Each patient completed a questionnaire and provided a serum sample and a colonic biopsy of normal-appearing mucosa. For IBD patients, an additional biopsy was collected from visually diseased mucosa. Serum vitamin D levels were measured by ultra-performance liquid chromatography and mass spectrometry. Hematoxylin and eosin stained tissue sections from colonic biopsies were classified histologically as normal or colitis (active/inactive), and scored for the degree of inflammation present (0-3, inactive/absent to severe). Tissue sections from colonic biopsies were also stained by immunohistochemistry for VDR, for which representative diagnostic areas were photographed and scored for staining intensity using a 4-point scale., Results: The IBD cohort was significantly younger (40.40 ± 5.27, P < 0.05) than the non-IBD cohort (56.70 ± 1.64) with a higher prevalence of vitamin D deficiency (40% vs 20%, respectively) and insufficiency (70% vs 50%, respectively). Histologic inflammation was significantly higher in visually diseased mucosa from IBD patients (1.95 ± 0.25) than in normal-appearing mucosa from control patients (0.25 ± 0.08, P < 0.01) and from IBD patients (0.65 ± 0.36, P < 0.05) and correlated inversely with VDR expression in visually diseased colonic tissue from IBD patients (r = -0.44, P < 0.05) and from IBD patients with Crohn's disease (r = -0.69, P < 0.05), but not in normal-appearing colonic tissue from control patients or IBD patients. Control and IBD patient serum vitamin D levels correlated positively with VDR expression in normal colon from control and IBD patients (r = 0.38, P < 0.05) and with patient age (r = 0.54, P < 0.01)., Conclusion: Levels of serum vitamin D correlate positively with colonic VDR expression in visually normal mucosa whereas inflammation correlates negatively with colonic VDR expression in visually diseased mucosa in Puerto Rican patients.

Background: Inflammatory bowel diseases (IBD) are chronic relapsing inflammatory conditions of unknown cause and likely result from the loss of immunological tolerance, which leads to over-activation of the gut immune system. Gut macrophages and dendritic cells (DCs) are essential for maintaining tolerance, but can also contribute to the inflammatory response in conditions such as IBD. Current therapies for IBD are limited by high costs and unwanted toxicities and side effects. The possibility of reducing intestinal inflammation with DCs genetically engineered to over-express the apoptosis-inducing FasL (FasL-DCs) has not yet been explored., Objective: Investigate the immunomodulatory effect of administering FasL-DCs in the rat trinitrobenzene sulfonic acid (TNBS) model of acute colitis., Methods: Expression of FasL on DCs isolated from the mesenteric lymph nodes (MLNs) of normal and TNBS-colitis rats was determined by flow cytometry. Primary rat bone marrow DCs were transfected with rat FasL plasmid (FasL-DCs) or empty vector (EV-DCs). The effect of these DCs on T cell IFNγ secretion and apoptosis was determined by ELISPOT and flow cytometry for Annexin V, respectively. Rats received FasL-DCs or EV-DCs intraperitoneally 96 and 48 hours prior to colitis induction with TNBS. Colonic T cell and neutrophil infiltration was determined by immunohistochemistry for CD3 and myeloperoxidase activity assay, respectively. Macrophage number and phenotype was measured by double immunofluorescence for CD68 and inducible Nitric Oxide Synthase., Results: MLN dendritic cells from normal rats expressed more FasL than those from colitic rats. Compared to EV-DCs, FasL-DCs reduced T cell IFNγ secretion and increased T cell apoptosis in vitro . Adoptive transfer of FasL-DCs decreased macroscopic and microscopic damage scores and reduced colonic T cells, neutrophils, and proinflammatory macrophages when compared to EV-DC adoptive transfer., Conclusion: FasL-DCs are effective at treating colonic inflammation in this model of IBD and represent a possible new treatment for patients with IBD.

Aim: To determine the expression of neurokinin-1 receptor (NK-1R), phosphorylated epidermal growth factor receptor (pEGFR), cyclooxygenase-2 (Cox-2), and vitamin D receptor (VDR) in normal, inflammatory bowel disease (IBD), and colorectal neoplasia tissues from Puerto Ricans., Methods: Tissues from patients with IBD, colitis-associated colorectal cancer (CAC), sporadic dysplasia, and sporadic colorectal cancer (CRC), as well as normal controls, were identified at several centers in Puerto Rico. Archival formalin-fixed, paraffin-embedded tissues were de-identified and processed by immunohistochemistry for NK-1R, pEGFR, Cox-2, and VDR. Pictures of representative areas of each tissues diagnosis were taken and scored by three observers using a 4-point scale that assessed intensity of staining. Tissues with CAC were further analyzed by photographing representative areas of IBD and the different grades of dysplasia, in addition to the areas of cancer, within each tissue. Differences in the average age between the five patient groups were assessed with one-way analysis of variance and Tukey-Kramer multiple comparisons test. The mean scores for normal tissues and tissues with IBD, dysplasia, CRC, and CAC were calculated and statistically compared using one-way analysis of variance and Dunnett's multiple comparisons test. Correlations between protein expression patterns were analyzed with the Pearson's product-moment correlation coefficient. Data are presented as mean ± SE., Results: On average, patients with IBD were younger (34.60 ± 5.81) than normal (63.20 ± 6.13, P < 0.01), sporadic dysplasia (68.80 ± 4.42, P < 0.01), sporadic cancer (74.80 ± 4.91, P < 0.001), and CAC (57.50 ± 5.11, P < 0.05) patients. NK-1R in cancer tissue (sporadic CRC, 1.73 ± 0.34; CAC, 1.57 ± 0.53) and sporadic dysplasia (2.00 ± 0.45) were higher than in normal tissues (0.73 ± 0.19). pEGFR was significantly increased in sporadic CRC (1.53 ± 0.43) and CAC (2.25 ± 0.47) when compared to normal tissue (0.07 ± 0.25, P < 0.05, P < 0.001, respectively). Cox-2 was significantly increased in sporadic colorectal cancer (2.20 ± 0.23 vs 0.80 ± 0.37 for normal tissues, P < 0.05). In comparison to normal (2.80 ± 0.13) and CAC (2.50 ± 0.33) tissues, VDR was significantly decreased in sporadic dysplasia (0.00 ± 0.00, P < 0.001 vs normal, P < 0.001 vs CAC) and sporadic CRC (0.47 ± 0.23, P < 0.001 vs normal, P < 0.001 vs CAC). VDR levels negatively correlated with NK-1R (r = -0.48) and pEGFR (r = -0.56) in normal, IBD, sporadic dysplasia and sporadic CRC tissue, but not in CAC., Conclusion: Immunohistochemical NK-1R and pEGFR positivity with VDR negativity can be used to identify areas of sporadic colorectal neoplasia. VDR immunoreactivity can distinguish CAC from sporadic cancer.

Background: Patients with Inflammatory Bowel Disease (IBD), most commonly Crohn's disease (CD) or ulcerative colitis (UC), suffer from chronic intestinal inflammation of unknown etiology. Increased proinflammatory macrophages (M1) have been documented in tissue from patients with CD. Anti-inflammatory macrophages (M2) may play a role in UC given the preponderance of Th2 cytokines in this variant of IBD. Animal and clinical studies have shown that the probiotic VSL#3 can ameliorate signs and symptoms of IBD. Although animal data suggests a modulatory effect on macrophage phenotype, the effect of VSL#3 on human macrophages remains unknown., Objective: To determine the effect of the probiotic VSL#3 on the phenotype of polarized (M1/M2) and unpolarized (MΦ) human macrophages., Methods: Human monocyte-derived macrophages, generated by culturing monocytes with M-CSF, were left unpolarized or were polarized towards an M1 or an M2 phenotype by culture with LPS and IFN-γ or IL-4, respectively, and were then cultured in the presence or absence of VSL#3 for 3 days. Changes in macrophage morphology were assessed. Cytokine and chemokine levels in supernatants were determined by multiplex assay., Results: VSL#3 decreased the granuloma-like aggregates of M1 macrophages, increased fibroblast-like M2 macrophages, and decreased fibroblast-like MΦ macrophages. VSL#3 increased the secretion of IL-1β, IL-6, IL-10, and G-CSF by M1, M2, and MΦ macrophages. VSL#3 exposure maintained the proinflammatory phenotype of M1 macrophages, sustaining IL-12 secretion, increasing IL-23 secretion, and decreasing MDC secretion. Both VSL#3-treated M2 and MΦ macrophages secreted higher levels of anti-inflammatory and pro-healing factors such as IL-1Ra, IL-13, EGF, FGF-2, TGF-α, and VEGF, as well as proinflammatory cytokines, including IL-12 and TNF-α., Conclusion: Under our experimental conditions VSL#3 induced a mixed proinflammatory and anti-inflammatory phenotype in polarized and unpolarized macrophages. This differential effect could explain why patients with CD do not respond to probiotic therapy as well as patients with UC.