Histologic Remission in Inflammatory Bowel Disease and Female Fertility: A Nationwide Study.

BACKGROUND & AIMS: Inflammatory bowel disease (IBD) is linked to reduced female fertility, but it is unclear how fertility rates vary by histologic disease activity. METHODS: Nationwide IBD cohort of Swedish women aged 15 to 44 years. We examined fertility rates during periods with vs without histologic inflammation (n = 21,046; follow-up, 1990-2016) and during periods with vs without clinical activity (IBD-related hospitalization, surgery, or treatment escalation) (n = 24,995; follow-up, 2006-2020). Accounting for sociodemographics and comorbidities, we used Poisson regression to estimate adjusted fertility rate ratios (aFRRs) for live births conceived during 12-month periods of histologic inflammation (vs histologic remission) and 3-month periods of clinically active IBD (vs quiescent IBD). RESULTS: During periods with vs without histologic inflammation, there were 6.35 (95% confidence interval [CI], 5.98-6.73) and 7.09 (95% CI, 6.48-7.70) live births conceived per 100 person-years of follow-up, respectively, or 1 fewer child per 14 women with 10 years of histologic inflammation (aFRR, 0.90; 95% CI, 0.81-1.00). In women with histologic inflammation, fertility was similarly reduced in ulcerative colitis (UC) (aFRR, 0.89 [95% CI, 0.78-1.02]) and Crohn's disease (CD) (aFRR, 0.86 [95% CI, 0.72-1.04]). Clinical IBD activity was associated with an aFRR of 0.76 (95% CI, 0.72-0.79) or 1 fewer child per 6 women with 10 years of clinical activity. Fertility was reduced in clinically active UC (aFRR, 0.75 [95% CI, 0.70-0.81]) and CD (aFRR, 0.76 [95% CI, 0.70-0.82]). Finally, among women with clinically quiescent IBD, histologic inflammation (vs histologic remission) was associated with reduced fertility (aFRR, 0.85 [95% CI, 0.73-0.98]). CONCLUSIONS: An association between histologic and clinical activity and reduced female fertility in CD and UC was found. Notably, histologic inflammation was also linked to reduced fertility in women with clinically quiescent IBD.

Hyaluronic Acid-Conjugated Thermoresponsive Polymer-Based Bioformulation Enhanced Wound Healing and Gut Barrier Repair of a TNBS-Induced Colitis Injury Ex Vivo Model in a Dynamic Perfusion Device.

Impairment of intestinal epithelium is a typical feature of inflammatory bowel disease (IBD) that causes leakage of bacteria and antigens from the intestinal lumen and thus results in persistent immune activation. Hence, healing and regeneration of the damaged gut mucosa is a promising therapeutic approach to achieve deep remission in IBD. Currently, available systemic therapies have moderate effects and are often associated with numerous side effects and malignancies. In this study, we aimed to develop a topical therapy by chemically conjugating a temperature-responsive polymer, i.e., poly(N-isopropylacrylamide), along with hyaluronic acid to obtain a sprayable therapeutic formulation that upon colon instillation adheres to the damaged gut mucosa due to its temperature-induced phase transition and mucoadhesive properties. An ex vivo adhesion experiment demonstrates that this therapeutic formulation forms a thin physical coating on the mucosal lining at a physiological temperature within 5 min. Physicochemical characterization of (P(NIPAM-co-NTBAM)-HA) established this formulation to be biocompatible, hemo-compatible, and non-immunogenic. Prednisolone was encapsulated within the polymer formulation to achieve maximum therapeutic efficacy in the case of IBD-like conditions as assessed in a custom-fabricated perfusion-based ex vivo model system. Histological analysis suggests that the prednisolone-encapsulated polymer formulation nearly restored the mucosal architecture after 2,4,6-trinitrobenzenesulfonic acid-induced damage. Furthermore, a significant (p ≤ 0.001) increase in mRNA levels of Muc-2 and ZO-1 in treated groups further confirmed the mucosal epithelial barrier restoration.

[New solutions for an old problem : Current developments in oral contrast agents]. / Neue Lösung für ein altes Problem : Aktuelle Entwicklungen bei oralen Kontrastmitteln.

BACKGROUND: Several radiological societies in Europe have so far spoken out in favor of MRI of the small intestine over CT, since MRI is said to provide more detailed image information. Due to the limited availability of MRI machines, long waiting times arise for many patients who clinically require small bowel imaging. OBJECTIVES: These circumstances guided our search for an improved CT technique that would provide scans that mimic the image impression of T1 sequence in MRI, i.e. with IV contrast-enhanced intestinal wall versus low/no signal lumen. MATERIAL AND METHODS: The oral use of fat or oil is poorly tolerated by patients, as is the placement of a naso-duodenal tube for air insufflation. We have now succeeded in creating a foamy drink with 44% air content, which is kept stable by protein together with buffers, and can be easily administered orally. CT with the Lumentin® beverage as bowel filling agent has been tested on healthy adult volunteers; oncology patients; and Crohn's disease patients who, for comparison, also underwent MRI of the small bowel with conventional oral contrast. RESULTS: The results so far with Lumentin® show a very good distribution throughout the entire small intestine with good lumen distension, images with strongly enhanced contrast of the intestinal mucosa, and lesions that are detected at the same or increased frequency as compared with MRI. Side effects were few and mild, and overall fewer than with commonly used oral agents. Lumentin's foamy consistency was unfamiliar to a few patients, but it wasn't difficult to drink. CONCLUSIONS: The new and innovative luminal HU-negative contrast agent Lumentin® improves the diagnostic CT image quality. In addition, Lumentin® experimental MRI tests have provided promising results, which are currently leading to further clinical MRI studies.

Emergency subtotal colectomy rates in relation to anti-TNF therapy in inflammatory bowel disease patients: comparison of retrospective cohorts.

OBJECTIVES: The introduction of anti-TNF therapy has revolutionized the management of inflammatory bowel disease (IBD). This study aimed to investigate the impact of anti-TNF therapy on emergency subtotal colectomy rates in patients with IBD (ulcerative colitis, Crohn's disease or indeterminate colitis) during two time periods. METHODS: IBD patients treated with emergency subtotal colectomy for acute severe colitis at Skåne University Hospital, Sweden, during two six-year windows were included. The two time-windows represented pre- and introductory (2004-2009) versus post-introductory (2012-2017) anti-TNF therapy usage. The two periods were compared in terms of the proportion of patients who received anti-TNF therapy, rates of emergency subtotal colectomy, and local IBD prevalence numbers. RESULTS: In total 91 patients were included, 42 (2004-2009) and 49 (2012-2017) patients, respectively. The proportion of patients that received anti-TNF therapy prior to admission was increased from 29% (2004-2009) to 63% (2012-2017) (p = .001). Despite this no significant difference was found in the emergency subtotal colectomy rates between the time periods considering the population at risk (IBD patients living in Skåne county); 1.44/1000 person years versus 1.37/1000 person years (p = .83). CONCLUSION: Despite an increased proportion of patients with IBD that received anti-TNF therapy prior to admission, no significant decrease in emergency subtotal colectomy rates was noted. Further research is warranted, and a prospective study design would facilitate a better causal understanding.

Histological remission in inflammatory bowel disease and risk of adverse pregnancy outcomes: A nationwide study.

Background: Inflammatory bowel disease (IBD) has been linked to adverse pregnancy outcomes, but it is unclear how risks vary by histological activity. Methods: We performed a nationwide study of Swedish women diagnosed with IBD 1990-2016 and a pre-pregnancy (<12 months) colorectal biopsy with vs. without histological inflammation (1223 and 630 births, respectively). We also examined pregnancy outcomes in 2007-2016 of women with vs. without clinically active IBD (i.e., IBD-related hospitalization, surgery, or medication escalation) <12 months before pregnancy (2110 and 4993 births, respectively). Accounting for smoking, socio-demographics, and comorbidities, generalized linear models estimated adjusted risk ratios (aRRs) for preterm birth (<37 gestational weeks) and small-for-gestational age (SGA, <10th percentile weight for age). Findings: Of infants to women with vs. without histological inflammation, 9.6% (n = 117) and 6.5% (n = 41) were preterm, respectively (aRR = 1.46; 95%CI = 1.03-2.06). Histological inflammation was associated with preterm birth in ulcerative colitis (UC) (aRR = 1.64; 95%CI = 1.07-2.52), especially extensive colitis (aRR = 2.37; 95%CI = 1.12-5.02), but not in Crohn's disease (aRR = 0.99; 95%CI = 0.55-1.78). Of infants to women with vs. without histological inflammation, 116 (9.6%) and 56 (8.9%), respectively, were SGA (aRR = 1.09; 95%CI = 0.81-1.47). Clinically active disease before pregnancy was linked to preterm birth (aRR = 1.42; 95%CI = 1.20-1.69), but not to SGA birth (aRR = 1.13; 95%CI = 0.96-1.32). Finally, of infants to women without clinical activity, histological inflammation was not significantly associated with preterm birth (aRR = 1.20; 95%CI = 0.68-2.13). Interpretation: Histological and clinical activity in IBD, especially in UC, were risk factors for preterm birth. Further research is needed to determine the importance of pre-pregnancy histological activity in women without clinically-defined disease activity. Funding: The Swedish Society of Medicine.

Management of Non-response and Loss of Response to Anti-tumor Necrosis Factor Therapy in Inflammatory Bowel Disease.

Anti-tumor necrosis factor (anti-TNF) therapy has been successfully used as first-line biologic treatment for moderate-to-severe inflammatory bowel disease (IBD), in both "step-up" and "top-down" approaches, and has become a cornerstone of IBD management. However, in a proportion of patients the effectiveness of anti-TNF therapy is sub-optimal. Either patients do not achieve adequate initial response (primary non-response) or they lose response after initial success (loss of response). Therapeutic drug monitoring determines drug serum concentrations and the presence of anti-drug antibodies (ADAbs) and can help guide treatment optimization to improve patient outcomes. For patients with low drug concentrations who are ADAb-negative or display low levels of ADAbs, dose escalation is recommended. Should response remain unchanged following dose optimization the question whether to switch within class (anti-TNF) or out of class (different mechanism of action) arises. If ADAb levels are high and the patient has previously benefited from anti-TNF therapy, then switching within class is a viable option as ADAbs are molecule specific. Addition of an immunomodulator may lead to a decrease in ADAbs and a regaining of response in a proportion of patients. If a patient does not achieve a robust therapeutic response with an initial anti-TNF despite adequate drug levels, then switching out of class is appropriate. In conjunction with the guidance above, other factors including patient preference, age, comorbidities, disease phenotype, extra-intestinal manifestations, and treatment costs need to be factored into the treatment decision. In this review we discuss current evidence in this field and provide guidance on therapeutic decision-making in clinical situations.

Real-world data on switching from intravenous to subcutaneous vedolizumab treatment in patients with inflammatory bowel disease.

BACKGROUND AND AIMS: Vedolizumab is a gut-selective treatment approved for Crohn's disease (CD) and ulcerative colitis (UC). Recently, a subcutaneous formulation of vedolizumab was approved. The aims of this study were to evaluate efficacy, safety, pharmacokinetics, patient experience and costs following a switch from intravenous to subcutaneous vedolizumab treatment. METHODS: Patients were switched from intravenous to subcutaneous vedolizumab maintenance treatment and followed prospectively for 6 months and a subgroup for 12 months. The primary endpoint was change in faecal calprotectin levels. Furthermore, we evaluated clinical disease activity, remission rates, plasma CRP, drug persistence, adverse events, local injection reactions, serum drug concentrations, patient satisfaction, quality-of-life and treatment costs. RESULTS: Eighty-nine patients were included (48 CD; 41 UC). Faecal calprotectin decreased significantly in CD but not in UC. Clinical indices, remission rates, plasma CRP levels and quality-of-life scores remained unchanged. Patients that had been on standard compared to optimised IV vedolizumab dosing displayed similar outcomes on standard SC dosing. Drug persistence at 6 and 12 months was 95.5% and 88.5%, respectively. Frequencies of adverse events were similar before and after the switch. No serious adverse events occurred. Transient severe local injection reactions were experienced by 1.2% of patients. Median vedolizumab trough levels were 2.3 times higher on subcutaneous compared to intravenous treatment. Patient satisfaction was generally high. Annualised treatment costs were reduced by 15% following the switch. CONCLUSIONS: The switch from intravenous to subcutaneous vedolizumab could be done with preserved therapeutic effectiveness, safety, high patient satisfaction and low discontinuation rate, at a reduced cost.

A novel food-based negative oral contrast agent compared with two conventional oral contrast agents in abdominal CT: a three-arm parallel blinded randomised controlled single-centre trial.

BACKGROUND: A negative oral contrast agent (OCA) has been long sought for, to better delineate the bowel and visualise surrounding structures. Lumentin® 44 (L44) is a new OCA formulated to fill the entire small bowel. The aim of this study was to compare L44 with positive and neutral conventional OCA in abdominal computed tomography (CT). METHODS: Forty-five oncologic patients were randomised to receive either L44 or one of the two comparators (MoviPrep® or diluted Omnipaque®). Abdominal CT examinations with intravenous contrast agent were acquired according to standard protocols. The studies were read independently by two senior radiologists. RESULTS: The mean intraluminal Hounsfield units (HU)-values of regions-of-interest (ROIs) for each subsegment of small bowel and treatment group were -404.0 HU for L44, 166.1 HU for Omnipaque®, and 16.7 HU for MoviPrep® (L44 versus Omnipaque, p < 0.001: L44 versus MoviPrep p < 0.001; Omnipaque versus MoviPrep, p = 0.003). Adverse events, only mild, using L44 were numerically fewer than for using conventional oral contrast agents. Visualisation of abdominal structures beyond the small bowel was similar to the comparators. CONCLUSIONS: L44 is a negative OCA with luminal radiodensity at approximately -400 HU creating a unique small bowel appearance on CT scans. The high bowel wall-to-lumen contrast may enable improved visualisation in a range of pathologic conditions. L44 showed a good safety profile and was well accepted by patients studied. TRIAL REGISTRATION: EudraCT (2017-002368-42) and in ClinicalTrials.gov (NCT03326518).

Discontinuation of Infliximab Therapy in Patients with Crohn's Disease.

BACKGROUND: Whether infliximab therapy can be successfully discontinued after patients with Crohn's disease have attained sustained, clinical, biochemical, and endoscopic remission is unknown. METHODS: We conducted a multicenter, randomized, double-blind, placebo-controlled withdrawal study of infliximab in patients with Crohn's disease who were in clinical, biochemical, and endoscopic remission after standard infliximab maintenance therapy for at least 1 year. Patients were randomly assigned 1:1 to continue infliximab therapy or to receive matching placebo for 48 weeks. The primary end point was time to relapse. RESULTS: This study randomly assigned 115 patients to either the infliximab-continuation group or to the infliximab-discontinuation group. No relapses were observed among the 59 patients continuing infliximab, whereas 23 of 56 patients discontinuing infliximab experienced relapse. Time to relapse was significantly shorter among patients who discontinued infliximab than among those who continued infliximab (hazard ratio, 0.080; 95% confidence interval [CI], 0.035 to 0.186; P<0.001). At the end of the trial at week 48, relapse-free survival was 100% in the infliximab-continuation group and 51% in the infliximab-discontinuation group. The key secondary end point, time to loss of remission, was significantly shorter among patients discontinuing infliximab therapy than those continuing infliximab (hazard ratio, 0.025; 95% CI, 0.003 to 0.187; P<0.001). No unexpected adverse events were reported. CONCLUSIONS: Discontinuation of infliximab for patients with Crohn's disease receiving long-term infliximab therapy and in clinical, biochemical, and endoscopic remission leads to a considerable risk of relapse. (Funded by the Nordic Trial Alliance [NordForsk], the Medical Fund of the Danish Regions [Regionernes Medicin og Behandlingspulje], the Danish Colitis-Crohn Association, and the A.P. Moller Foundation; ClinicalTrials.gov number, NCT01817426; EudraCT number, 2012-002702-51.)

Short and long-term efficacy of adalimumab in ulcerative colitis: a real-life study.

Objective: Randomized controlled trials have shown the effectiveness of Adalimumab in ulcerative colitis. However, real-life data is scarce. We aimed to assess the effectiveness and predictive factors of effectiveness in a large Swedish cohort.Methods: Retrospective capture of data from local registries at five Swedish IBD centers. Clinical response and remission rates were assessed at three months after starting adalimumab treatment and patients were followed until colectomy or need for another biological. Bio-naive patients were compared to bio experienced patients. Factors associated with short term responses were assessed using logistic regression model. Failure on drug was assessed using a Cox proportional hazards regression model.Results: 118 patients (59 males, 59 females) with median age 34.4 years (IQR 27.0-51.4) were included. Median disease duration was 4.3 years (IQR 2.0-9.0) and follow-up 1.27 years (IQR 0.33-4.1). A clinical corticosteroid-free remission was achieved by 38/118 (32.2%) and response by 91/118 (77%) after three months. CRP >3 mg/l at baseline was predictive of short-term failure to reach corticosteroid-free remission. Factors associated with survival on the drug were male gender, CRP <3 mg/l and absence of primary sclerosing cholangitis. Patients >42 years of age at diagnosis were more likely to respond to adalimumab and remain on treatment compared to patients <20 years.Conclusions: An elevated CRP-level, primary sclerosing cholangitis and female gender were predictors of treatment failure. In contrast older age at diagnosis was a predictor of short-term clinical response and drug survival. Prior infliximab failure, regardless of cause, did not influence the outcome of adalimumab treatment.

Colorectal cancer-infiltrating T lymphocytes display a distinct chemokine receptor expression profile.

BACKGROUND: T lymphocytes exert important homeostatic functions in the healthy intestinal mucosa, whereas in case of colorectal cancer (CRC), infiltration of T lymphocytes into the tumor is crucial for an effective anti-tumor immune response. In both situations, the recruitment mechanisms of T lymphocytes into the tissues are essential for the immunological functions deciding the outcome. The recruitment of T lymphocytes is largely dependent on their expression of various chemokine receptors. The aim of this study was to identify potential chemokine receptors involved in the recruitment of T lymphocytes to normal human colonic mucosa and to CRC tissue, respectively, by examining the expression of 16 different chemokine receptors on T lymphocytes isolated from these tissues. METHODS: Tissues were collected from patients undergoing bowel resection for CRC. Lymphocytes were isolated through enzymatic tissue degradation of CRC tissue and nearby located unaffected mucosa, respectively. The expression of a broad panel of chemokine receptors on the freshly isolated T lymphocytes was examined by flow cytometry. RESULTS: In the normal colonic mucosa, the frequencies of cells expressing CCR2, CCR4, CXCR3, and CXCR6 differed significantly between CD4+ and CD8+ T lymphocytes, suggesting that the molecular mechanisms mediating T lymphocyte recruitment to the gut differ between CD4+ and CD8+ T lymphocytes. In CRC, the frequencies of cells expressing CCR2 and CXCR5 were significantly lower in both the CD4+ and CD8+ T lymphocyte populations compared to unaffected colonic mucosa, and the frequency of CCR9+ cytotoxic T lymphocytes was significantly decreased in CRC tissue. CONCLUSIONS: With regard to the normal gut mucosa, the results suggest that the molecular mechanisms mediating T lymphocyte recruitment differ between CD4+ and CD8+ T lymphocytes, which are important for understanding gut homeostasis. Importantly, T lymphocytes from CRC compared to normal colonic tissue displayed a distinct chemokine receptor expression profile, suggesting that mechanisms for recruitment of T lymphocytes to CRC tissue are skewed compared to normal colonic mucosa. Understanding these mechanisms could help in developing new strategies in cancer immunotherapy and to optimize already available alternatives such as immune checkpoint inhibitors.

Long-term effectiveness of vedolizumab in inflammatory bowel disease: a national study based on the Swedish National Quality Registry for Inflammatory Bowel Disease (SWIBREG).

OBJECTIVES: Clinical trials have demonstrated the efficacy of vedolizumab in inflammatory bowel disease (IBD). However, these findings may not reflect the clinical practice. Therefore, we aimed to describe a vedolizumab-treated patient population and assess long-term effectiveness. MATERIALS AND METHODS: Patients initiating vedolizumab between 1 June 2014 and 30 May 2015 were identified through the Swedish National Quality Registry for IBD. Prospectively collected data on treatment and disease activity were extracted. Clinical remission was defined as Patient Harvey Bradshaw index <5 in Crohn's disease (CD) and Patient Simple Clinical Colitis Activity index <3 in ulcerative colitis (UC). RESULTS: Two-hundred forty-six patients (147 CD, 92 UC and 7 IBD-Unclassified) were included. On study entry, 86% had failed TNF-antagonist and 48% of the CD patients had undergone ≥1 surgical resection. After a median follow-up of 17 (IQR: 14-20) months, 142 (58%) patients remained on vedolizumab. In total, 54% of the CD- and 64% of the UC patients were in clinical remission at the end of follow-up, with the clinical activity decreasing (p < .0001 in both groups). Faecal-calprotectin decreased in CD (p < .0001) and in UC (p = .001), whereas CRP decreased in CD (p = .002) but not in UC (p = .11). Previous anti-TNF exposure (adjusted HR: 4.03; 95% CI: 0.96-16.75) and elevated CRP at baseline (adjusted HR: 2.22; 95% CI: 1.10-4.35) seemed to be associated with discontinuation because of lack of response. Female sex was associated with termination because of intolerance (adjusted HR: 2.75; 95% CI: 1.16-6.48). CONCLUSION: Vedolizumab-treated patients represent a treatment-refractory group. A long-term effect can be achieved, even beyond 1 year of treatment.

Vedolizumab treatment for immune checkpoint inhibitor-induced enterocolitis.

Immune checkpoint inhibitors (ICPI), such as ipilimumab [anti-cytotoxic T-lymphocyte antigen-4 (CTLA-4) antibody] and nivolumab or pembrolizumab [anti-programmed cell death protein-1 (PD-1) antibodies], improve survival in several cancer types. Since inhibition of CTLA-4 or PD-1 leads to non-selective activation of the immune system, immune-related adverse events (irAEs) are frequent. Enterocolitis is a common irAE, currently managed with corticosteroids and, if necessary, anti-tumor necrosis factor-α therapy. Such a regimen carries a risk of serious side-effects including infections, and may potentially imply impaired antitumor effects. Vedolizumab is an anti-integrin α4ß7 antibody with gut-specific immunosuppressive effects, approved for Crohn's disease and ulcerative colitis. We report a case series of seven patients with metastatic melanoma or lung cancer, treated with vedolizumab off-label for ipilimumab- or nivolumab-induced enterocolitis, from June 2014 through October 2016. Clinical, laboratory, endoscopic, and histologic data were analyzed. Patients initially received corticosteroids but were steroid-dependent and/or partially refractory. One patient was administered infliximab but was refractory. The median time from onset of enterocolitis to start of vedolizumab therapy was 79 days. Following vedolizumab therapy, all patients but one experienced steroid-free enterocolitis remission, with normalized fecal calprotectin. This was achieved after a median of 56 days from vedolizumab start, without any vedolizumab-related side-effects noted. The patient in whom vedolizumab was not successful, due to active ulcerative colitis, received vedolizumab prophylactically. This is the first case series to suggest that vedolizumab is an effective and well-tolerated therapeutic for steroid-dependent or partially refractory ICPI-induced enterocolitis. A larger prospective study to evaluate vedolizumab in this indication is warranted.

CD8αß+ γδ T Cells: A Novel T Cell Subset with a Potential Role in Inflammatory Bowel Disease.

γδ T cells have been attributed a wide variety of functions, which in some cases may appear as contradictory. To better understand the enigmatic biology of γδ T cells it is crucial to define the constituting subpopulations. γδ T cells have previously been categorized into two subpopulations: CD8αα+ and CD8- cells. In this study we have defined and characterized a novel subset of human γδ T-cells expressing CD8αß. These CD8αß+ γδ T cells differed from the previously described γδ T cell subsets in several aspects, including the degree of enrichment within the gut mucosa, the activation status in blood, the type of TCRδ variant used in blood, and small but significant differences in their response to IL-2 stimulation. Furthermore, the novel subset expressed cytotoxic mediators and CD69, and produced IFN-γ and TNF-α. In patients with active inflammatory bowel disease the mucosal frequencies of CD8αß+ γδ T cells were significantly lower as compared with healthy controls, correlated negatively with the degree of disease activity, and increased to normal levels as a result of anti-TNF-α therapy. In conclusion, our results demonstrate that CD8αß+ γδ T cells constitute a novel lymphocyte subset, which is strongly enriched within the gut and may play an important role in gut homeostasis and mucosal healing in inflammatory bowel disease.

Involvement of CCR9 at multiple stages of adult T lymphopoiesis.

The chemokine CCL25 is constitutively expressed in the thymus, and its receptor CCR9 is expressed on subsets of developing thymocytes. Nevertheless, the function of CCL25/CCR9 in adult thymopoiesis remains unclear. Here, we demonstrate that purified CCR9(-/-) hematopoietic stem cells are deficient in their ability to generate all major thymocyte subsets including double-negative 1 (DN1) cells in competitive transfers. CCR9(-/-) bone marrow contained normal numbers of lineage(-) Sca-1+c-kit+, common lymphoid progenitors, and lymphoid-primed multipotent progenitors (LMPP), and CCR9(-/-) LMPP showed similar T cell potential as their wild-type (WT) counterparts when cultured on OP9-delta-like 1 stromal cells. In contrast, early thymic progenitor and DN2 thymocyte numbers were reduced in the thymus of adult CCR9(-/-) mice. In fetal thymic organ cultures (FTOC), CCR9(-/-) DN1 cells were as efficient as WT DN1 cells in generating double-positive (DP) thymocytes; however, under competitive FTOC, CCR9(-/-) DP cell numbers were reduced significantly. Similarly, following intrathymic injection into sublethally irradiated recipients, CCR9(-/-) DN cells were out-competed by WT DN cells in generating DP thymocytes. Finally, in competitive reaggregation thymic organ cultures, CCR9(-/-) preselection DP thymocytes were disadvantaged significantly in their ability to generate CD4 single-positive (SP) thymocytes, a finding that correlated with a reduced ability to form TCR-MHC-dependent conjugates with thymic epithelial cells. Together, these results highlight a role for CCR9 at several stages of adult thymopoiesis: in hematopoietic progenitor seeding of the thymus, in the DN-DP thymocyte transition, and in the generation of CD4 SP thymocytes.

Involvement of CCL25 (TECK) in the generation of the murine small-intestinal CD8alpha alpha+CD3+ intraepithelial lymphocyte compartment.

The CC chemokine CCL25 (TECK) is selectively expressed in the thymus and small intestine, indicating a potential role in T lymphocyte development. In the present study we examined the role of CCL25 in the generation of the small-intestinal CD8alpha alpha(+)CD3(+) intraepithelial lymphocyte (IEL) compartment. CCL25 mRNA expression in the murine small intestine increased at three weeks of age and corresponded with the appearance of CD8alpha alpha(+)CD3(+) lymphocytes in the small-intestinal epithelium. Administration of monoclonal neutralizing anti-CCL25 antibody to two-week-old mice led to a approximately 50% reduction in the total number of CD8alpha alpha(+)TCRgamma delta(+) and CD8alpha alpha(+)TCRalpha beta(+) IEL at four weeks of age. Freshly isolated murine CD8alpha alpha(+)CD3(+) IEL migrated in response to CCL25 and expressed the CCL25 receptor, CCR9. Analysis of CCR9 expression on putative IEL precursor populations demonstrated the presence of both CCR9(-) and CCR9(+) cells and indicated that up-regulation of this receptor occurred during IEL precursor differentiation. Finally, data from wild-type and RAG(-/-) mice suggested that the reduction in CD8alpha alpha(+)CD3(+) IEL in anti-CCL25 antibody treated mice resulted primarily from defective maintenance and/or development of IEL precursors rather than a direct effect on mature CD8alpha alpha(+)CD3(+) IEL.