Human small intestine contains 2 functionally distinct regulatory T-cell subsets.

BACKGROUND: Regulatory T (Treg) CD4 cells in mouse gut are mainly specific for intestinal antigens and play an important role in the suppression of immune responses against harmless dietary antigens and members of the microbiota. However, information about the phenotype and function of Treg cells in the human gut is limited. OBJECTIVE: We performed a detailed characterization of Foxp3+ CD4 Treg cells in human normal small intestine (SI) as well as from transplanted duodenum and celiac disease lesions. METHODS: Treg cells and conventional CD4 T cells derived from SI were subjected to extensive immunophenotyping and their suppressive activity and ability to produce cytokines assessed. RESULTS: SI Foxp3+ CD4 T cells were CD45RA-CD127-CTLA-4+ and suppressed proliferation of autologous T cells. Approximately 60% of Treg cells expressed the transcription factor Helios. When stimulated, Helios-negative Treg cells produced IL-17, IFN-γ, and IL-10, whereas Helios-positive Treg cells produced very low levels of these cytokines. By sampling mucosal tissue from transplanted human duodenum, we demonstrated that donor Helios-negative Treg cells persisted for at least 1 year after transplantation. In normal SI, Foxp3+ Treg cells constituted only 2% of all CD4 T cells, while in active celiac disease, both Helios-negative and Helios-positive subsets expanded 5- to 10-fold. CONCLUSION: The SI contains 2 subsets of Treg cells with different phenotypes and functional capacities. Both subsets are scarce in healthy gut but increase dramatically in active celiac disease.

Validating the US pancreas donor risk index in a Norwegian population, a retrospective observational study.

OBJECTIVES: Despite advances in immunosuppression and surgical technique, pancreas transplantation is still associated with a significant graft loss rate. The Pancreas Donor Risk Index (PDRI) is a pre-transplant scoring tool derived from a US population. We sought to validate the PDRI in a Norwegian population. METHODS: We retrospectively retrieved donor data for 344 pancreas transplants undertaken in Norway between 2000 and 2019, utilising the Scandiatransplant database, and matched these to the respective recipients. The PDRI score was calculated for each transplanted pancreas, these were then stratified into quintiles. The association between the PDRI quintiles and 1-year graft survival was calculated, and this was repeated for the different types of pancreas transplantation. The association between PDRI as a continuous variable, and graft survival was determined. Donor and recipient data were compared to the original US population. RESULTS: The overall 1-year graft survival was 82.7%. There were no significant differences in survival between the different PDRI quintiles. When viewed as a continuous variable, increased PDRI score was not associated with decreased graft survival. Significant differences between the Norwegian and US populations were found. CONCLUSIONS: When applied to a Norwegian population, the PDRI score was unable to predict 1-year graft survival.

The Presence of Activated T Cell Subsets prior to Transplantation Is Associated with Increased Rejection Risk in Pancreas Transplant Recipients.

Pancreas and islet transplantation (PTx) are currently the only curative treatment options for type 1 diabetes. CD4+ and CD8+ T cells play a pivotal role in graft function, rejection, and survival. However, characterization of immune cell status from patients with and without rejection of the pancreas graft is lacking. We performed multiparameter immune phenotyping of T cells from PTx patients prior to and 1 y post-PTx in nonrejectors and histologically confirmed rejectors. Our results suggest that rejection is associated with presence of elevated levels of activated CD4+ and CD8+ T cells with a gut-homing phenotype both prior to and 1 y post-PTx. The CD4+ and CD8+ T cells were highly differentiated, with elevated levels of type 1 inflammatory markers (T-bet and INF-γ) and cytotoxic components (granzyme B and perforin). Furthermore, we observed increased levels of activated FOXP3+ regulatory T cells in rejectors, which was associated with a hyporesponsive phenotype of activated effector T cells. Finally, activated T and B cell status was correlated in PTx patients, indicating a potential interplay between these cell types. In vitro treatment of healthy CD4+ and CD8+ T cells with tacrolimus abrogated the proliferation and cytokine (INF-γ, IL-2, and TNF-α) secretion associated with the type 1 inflammatory phenotype observed in pre- and post-PTx rejectors. Together, our results suggest the presence of activated CD4+ and CD8+ T cells prior to PTx confer increased risk for rejection. These findings may be used to identify patients that may benefit from more intense immunosuppressive treatment that should be monitored more closely after transplantation.

Early detection of complications in pancreas transplants by microdialysis catheters, an observational feasibility study.

BACKGROUND: Despite advances in immunosuppression and surgical technique, pancreas transplantation is encumbered with a high rate of complication and graft losses. Particularly, venous graft thrombi occur relatively frequently and are rarely detected before the transplant is irreversibly damaged. METHODS: To detect complications early, when the grafts are potentially salvageable, we placed microdialysis catheters anteriorly and posteriorly to the graft in a cohort of 34 consecutive patients. Glucose, lactate, pyruvate, and glycerol were measured at the bedside every 1-2 hours. RESULTS: Nine patients with graft venous thrombosis had significant lactate and lactate-to-pyruvate-ratio increases without concomitant rise in blood glucose or clinical symptoms. The median lactate in these patients was significantly higher in both catheters compared to non-events (n = 15). Out of the nine thrombi, four grafts underwent successful angiographic extraction, one did not require intervention and four grafts were irreversibly damaged and explanted. Four patients with enteric anastomosis leakages had significantly higher glycerol measurements compared to non-events. As with the venous thrombi, lactate and lactate-to-pyruvate ratio were also increased in six patients with graft surrounding hematomas. CONCLUSIONS: Bedside monitoring with microdialysis catheters is a promising surveillance modality of pancreatic grafts, but differentiating between the various pathologies proves challenging.

Liver transplantation as last-resort treatment for patients with bile duct injuries following cholecystectomy: a multicenter analysis.

BACKGROUND: Liver transplantation (LT) has been used as a last resort in patients with end-stage liver disease due to bile duct injuries (BDI) following cholecystectomy. Our study aimed to identify and evaluate factors that cause or contribute to an extended liver disease that requires LT as ultimate solution, after BDI during cholecystectomy. METHODS: Data from 8 high-volume LT centers relating to patients who underwent LT after suffering BDI during cholecystectomy were prospectively collected and retrospectively analyzed. RESULTS: Thirty-four patients (16 men, 18 women) with a median age of 45 (range 22-69) years were included in this study. Thirty of them (88.2%) underwent LT because of liver failure, most commonly as a result of secondary biliary cirrhosis. The median time interval between BDI and LT was 63 (range 0-336) months. There were 23 cases (67.6%) of postoperative morbidity, 6 cases (17.6%) of post-transplant 30-day mortality, and 10 deaths (29.4%) in total after LT. There was a higher probability that patients with concomitant vascular injury (hazard ratio 10.69, P=0.039) would be referred sooner for LT. Overall survival following LT at 1, 3, 5 and 10 years was 82.4%, 76.5%, 73.5% and 70.6%, respectively. CONCLUSION: LT for selected patients with otherwise unmanageable BDI following cholecystectomy yields acceptable long-term outcomes.

CD4+ T cells persist for years in the human small intestine and display a TH1 cytokine profile.

Studies in mice and humans have shown that CD8+ T cell immunosurveillance in non-lymphoid tissues is dominated by resident populations. Whether CD4+ T cells use the same strategies to survey peripheral tissues is less clear. Here, examining the turnover of CD4+ T cells in transplanted duodenum in humans, we demonstrate that the majority of CD4+ T cells were still donor-derived one year after transplantation. In contrast to memory CD4+ T cells in peripheral blood, intestinal CD4+ TRM cells expressed CD69 and CD161, but only a minor fraction expressed CD103. Functionally, intestinal CD4+ TRM cells were very potent cytokine producers; the vast majority being polyfunctional TH1 cells, whereas a minor fraction produced IL-17. Interestingly, a fraction of intestinal CD4+ T cells produced granzyme-B and perforin after activation. Together, we show that the intestinal CD4+ T-cell compartment is dominated by resident populations that survive for more than 1 year. This finding is of high relevance for the development of oral vaccines and therapies for diseases in the gut.

Portal vein stent placement after hepatobiliary and pancreatic surgery.

PURPOSE: To evaluate the long-term outcomes of percutaneous transhepatic stent placement for portal vein (PV) stenosis after liver transplantation (LT) and hepato-pancreato-biliary (HPB) surgery. METHODS: Retrospective study of 455 patients who underwent LT and 522 patients who underwent resection of the pancreatic head between June 2011 and February 2016. Technical success, clinical success, patency, and complications were evaluated for both groups. RESULTS: A total of 23 patients were confirmed to have postoperative PV stenosis and were treated with percutaneous transhepatic PV stent placement. The technical success rate was 100%, the clinical success rate was 80%, and the long-term stent patency was 91.3% for the entire study population. Two procedure-related hemorrhages and two early stent thromboses occurred in the HPB group while no complications occurred in the LT group. A literature review of selected studies reporting PV stent placement for the treatment of PV stenosis after HPB surgery and LT showed a technical success rate of 78-100%, a clinical success rate of 72-100%, and a long-term patency of 57-100%, whereas the procedure-related complication rate varied from 0-33.3%. CONCLUSIONS: Percutaneous transhepatic PV stent is a safe and effective treatment for postoperative PV stenosis/occlusion in patients undergoing LT regardless of symptoms. Due to increased risk of complications, the indication for percutaneous PV stent placement after HPB surgery should be limited to patients with clinical symptoms after an individual assessment.

Resident memory CD8 T cells persist for years in human small intestine.

Resident memory CD8 T (Trm) cells have been shown to provide effective protective responses in the small intestine (SI) in mice. A better understanding of the generation and persistence of SI CD8 Trm cells in humans may have implications for intestinal immune-mediated diseases and vaccine development. Analyzing normal and transplanted human SI, we demonstrated that the majority of SI CD8 T cells were bona fide CD8 Trm cells that survived for >1 yr in the graft. Intraepithelial and lamina propria CD8 Trm cells showed a high clonal overlap and a repertoire dominated by expanded clones, conserved both spatially in the intestine and over time. Functionally, lamina propria CD8 Trm cells were potent cytokine producers, exhibiting a polyfunctional (IFN-γ+ IL-2+ TNF-α+) profile, and efficiently expressed cytotoxic mediators after stimulation. These results suggest that SI CD8 Trm cells could be relevant targets for future oral vaccines and therapeutic strategies for gut disorders.

Liver transplantation as a lifesaving procedure for posthepatectomy liver failure and iatrogenic liver injuries.

BACKGROUND: Iatrogenic injuries to vital structures of the liver and posthepatectomy liver failure are associated with high mortality. The current donor situation in Norway allows liver transplantation of patients beyond conventional criteria. METHODS: From 1984 to 2017, a total of 1510 liver transplantations were performed. In this retrospective study, we report the results of 13 patients undergoing liver transplantation due to iatrogenic injuries to the liver vasculature or posthepatectomy liver failure. RESULTS: Twelve men and one woman with a median age of 55 years (range 22-69) were included. Seven patients underwent radical surgery for cancer prior to transplantation. The median follow-up time was 70.5 months (range 2.2-177). Three of the patients with malignant disease did not experience disease recurrence, whereas four patients had cancer recurrence and died 7, 24, 45, and 78 months after transplantation. Five of six patients with non-malignant disease fully recovered, but one patient died after 9 months due to infectious complications. CONCLUSIONS: Liver transplantation for liver failure due to portal vein and hepatic artery injury in patients with non-malignant disease seems justified. However, it may be questioned whether patients with malignant disease beyond established criteria should be offered liver transplantation.

Transcriptional and functional profiling defines human small intestinal macrophage subsets.

Macrophages (Mfs) are instrumental in maintaining immune homeostasis in the intestine, yet studies on the origin and heterogeneity of human intestinal Mfs are scarce. Here, we identified four distinct Mf subpopulations in human small intestine (SI). Assessment of their turnover in duodenal transplants revealed that all Mf subsets were completely replaced over time; Mf1 and Mf2, phenotypically similar to peripheral blood monocytes (PBMos), were largely replaced within 3 wk, whereas two subsets with features of mature Mfs, Mf3 and Mf4, exhibited significantly slower replacement. Mf3 and Mf4 localized differently in SI; Mf3 formed a dense network in mucosal lamina propria, whereas Mf4 was enriched in submucosa. Transcriptional analysis showed that all Mf subsets were markedly distinct from PBMos and dendritic cells. Compared with PBMos, Mf subpopulations showed reduced responsiveness to proinflammatory stimuli but were proficient at endocytosis of particulate and soluble material. These data provide a comprehensive analysis of human SI Mf population and suggest a precursor-progeny relationship with PBMos.

CD8+ T Cells That Coexpress RORγt and T-bet Are Functionally Impaired and Expand in Patients with Distal Bile Duct Cancer.

CD8+ T cells that express retinoic acid-related orphan receptor (ROR)γt (TC17 cells) have been shown to promote procarcinogenic inflammation and contribute to a tolerogenic microenvironment in tumors. We investigated their phenotype and functional properties in relationship to the pathogenesis of human distal bile duct cancer (DBDC). DBDC patients had an elevated level of type 17 immune responses and the frequency of CD8+RORγt+ T cells (TC17 cells) was increased in peripheral blood. The CD8+RORγt+ T cells represented a highly activated subset and produced IL-17A in equal amount as CD4+RORγt+ T cells (TH17 cells). Most CD8+RORγt+ T cells coexpressed T-bet, a lineage transcription factor for TH1 and TC1 development, suggesting that CD8+RORγt+ T cells undergo plasticity toward a TC17/1-like phenotype with coproduction of IL-17A and INF-γ. In comparison with CD8+RORγt- T cells, the CD8+RORγt+ T cells had a higher level of TCR signaling and were terminally differentiated and exhausted. These cells also had impaired ability to re-express perforin after degranulation and reduced cytotoxic immune function. A subset of CD8+RORγt+ T cells expressing a low level of programmed cell death protein 1 and a high level of OX40 were associated with reduced patient survival. In conclusion, CD8+RORγt+ T cells are proinflammatory and functionally impaired and may contribute to the pathogenesis of DBDC.

Antibody-secreting plasma cells persist for decades in human intestine.

Plasma cells (PCs) produce antibodies that mediate immunity after infection or vaccination. In contrast to PCs in the bone marrow, PCs in the gut have been considered short lived. In this study, we studied PC dynamics in the human small intestine by cell-turnover analysis in organ transplants and by retrospective cell birth dating measuring carbon-14 in genomic DNA. We identified three distinct PC subsets: a CD19+ PC subset was dynamically exchanged, whereas of two CD19- PC subsets, CD45+ PCs exhibited little and CD45- PCs no replacement and had a median age of 11 and 22 yr, respectively. Accumulation of CD45- PCs during ageing and the presence of rotavirus-specific clones entirely within the CD19- PC subsets support selection and maintenance of protective PCs for life in human intestine.

Transplantation With Livers From Deceased Donors Older Than 75 Years.

BACKGROUND: The availability of donor organs limits the number of patients in need who are offered liver transplantation. Measures to expand the donor pool are crucial to prevent on-list mortality. The aim of this study was to evaluate the use of livers from deceased donors who were older than 75 years. METHODS: Fifty-four patients who received a first liver transplant (D75 group) from 2001 to 2011 were included. Donor and recipient data were collected from the Nordic Liver Transplant Registry and medical records. The outcome was compared with a control group of 54 patients who received a liver graft from donors aged 20 to 49 years (D20-49 group). Median donor age was 77 years (range, 75-86 years) in the D75 group and 41 years (range, 20-49 years) in the D20-49 group. Median recipient age was 59 years (range, 31-73 years) in the D75 group and 58 years (range, 31-74 years) in the D20-49 group. RESULTS: The 1-, 3-, and 5-year patient/graft survival values were 87/87%, 81/81%, and 71/67% for the D75 group and 88/87%, 75/73%, and 75/73% for the D20-49 group, respectively. Patient (P = 0.89) and graft (P = 0.79) survival did not differ between groups. The frequency of biliary complications was higher in the D75 group (29.6/13%, P = 0.03). CONCLUSIONS: Selected livers from donors over age 75 years should not be excluded based on age, which does not compromise patient or graft survival despite a higher frequency of biliary complications.

Liver transplantation with deceased ABO-incompatible donors is life-saving but associated with increased risk of rejection and post-transplant complications.

ABO-incompatible (ABOi) liver transplantation (LT) with deceased donor organs is performed occasionally when no ABO-compatible (ABOc) graft is available. From 1996 to 2011, 61 ABOi LTs were performed in Oslo and Gothenburg. Median patient age was 51 years (range 13-75); 33 patients were transplanted on urgent indications, 13 had malignancy-related indications, and eight received ABOi grafts for urgent retransplantations. Median donor age was 55 years (range 10-86). Forty-four patients received standard triple immunosuppression with steroids, tacrolimus, and mycophenolate mofetil, and forty-four patients received induction with IL-2 antagonist or anti-CD20 antibody. Median follow-up time was 29 months (range 0-200). The 1-, 3-, 5-, and 10-year Kaplan-Meier estimates of patient survival (PS) and graft survival (GS) were 85/71%, 79/57%, 75/55%, and 59/51%, respectively, compared to 90/87%, 84/79%, 79/73%, and 65/60% for all other LT recipients in the same period. The 1-, 3-, 5-, and 10-year GS for A2 grafts were 81%, 67%, 62%, and 57%, respectively. In conclusion, ABOi LT performed with non-A2 grafts is associated with inferior graft survival and increased risk of rejection, vascular and biliary complications. ABOi LT with A2 grafts is associated with acceptable graft survival and can be used safely in urgent cases.

Activated regulatory and memory T-cells accumulate in malignant ascites from ovarian carcinoma patients.

Invasive ovarian cancer is associated with poor outcome. The presence of infiltrating regulatory T-cells (Tregs) suppresses protective anti-tumor immune responses, and their accumulation into the tumor microenvironment correlates with reduced survival in ovarian cancer patients. Here, we conducted a detailed characterization of CD4(+) T-cells, CD8(+) T-cells and Treg subsets in the peripheral blood and malignant ascites fluid from seventeen patients with ovarian carcinoma of epithelial origin. Cell distribution, activation status and proliferation status were assessed by multi-color flow cytometry. In ascites fluid, a significant accumulation of CD8(+) cytotoxic T-cells and Tregs was observed compared to peripheral blood. Furthermore, a skewing toward the CD45RA(-) effector/memory compartment was observed in all T-cell subsets in the ascites fluid, but was most pronounced in the Treg population. Regulatory T-cells in the malignant ascites were more activated and had a higher proliferation rate compared to blood-derived cells from the same patient, and their number in ascites was positively correlated with the number of epithelial cells in effusion. In summary, we demonstrate an accumulation of activated CD4(+), CD8(+) and regulatory T-cells in the cancer microenvironment of ovarian carcinoma.

Low use of surveillance and early diagnosis of hepatocellular carcinoma in Norway--a population-based cohort study.

BACKGROUND AND AIMS: Curative treatment of hepatocellular carcinoma (HCC) is dependent on early diagnosis. Surveillance of patients at high risk for HCC is a key determinant to achieve this goal, but may be an underutilized tool. The aim of this study was to determine the rate of pre-diagnosis surveillance in patients with HCC in a large population-based cohort and to assess to what extent cirrhosis was known prior to the diagnosis of HCC. METHODS: All patients diagnosed with HCC during 2000-2009 in The South-Eastern Regional Health Authority, representing 56% of the Norwegian population, were identified from The National Cancer Registry and the medical records were reviewed. RESULTS: Fifteen out of 486 patients (3%) were diagnosed by surveillance. Potential curative treatment was offered to 58% of the patients who underwent surveillance as opposed to 15% in the non-surveillance group. Only age ≤ 65 years was an independent predictor of screening in a multivariate model. Almost two thirds of the patients with cirrhosis were unrecognized prior to the HCC diagnosis. Two hundred and fourteen patients (44%) were non-cirrhotics. CONCLUSION: Regular HCC surveillance in at-risk populations is virtually not applied in Norway and this may contribute to inferior overall survival. Failure to recognize cirrhosis and a high rate of HCC in non-cirrhotic patients will be limiting factors for the overall effectiveness of a potential surveillance program.

Aggressive treatment of patients with metastatic colorectal cancer increases survival: a scandinavian single-center experience.

Background. We examined overall and disease-free survivals in a cohort of patients subjected to resection of liver metastasis from colorectal cancer (CRLM) in a 10-year period when new treatment strategies were implemented. Methods. Data from 239 consecutive patients selected for liver resection of CRLM during the period from 2002 to 2011 at a single center were used to estimate overall and disease-free survival. The results were assessed against new treatment strategies and established risk factors. Results. The 5-year cumulative overall and disease-free survivals were 46 and 24%. The overall survival was the same after reresection, independently of the number of prior resections and irrespectively of the location of the recurrent disease. The time intervals between each recurrence were similar (11 ± 1 months). Patients with high tumor load given neoadjuvant chemotherapy had comparable survival to those with less extensive disease without neoadjuvant chemotherapy. Positive resection margin or resectable extrahepatic disease did not affect overall survival. Conclusion. Our data support that one still, and perhaps to an even greater extent, should seek an aggressive therapeutic strategy to achieve resectable status for recurrent hepatic and extrahepatic metastases. The data should be viewed in the context of recent advances in the understanding of cancer biology and the metastatic process.

Kinetics and activation requirements of contact-dependent immune suppression by human regulatory T cells.

Naturally occurring regulatory T cells (Tregs) maintain self tolerance by dominant suppression of potentially self-reactive T cells in peripheral tissues. However, the activation requirements, the temporal aspects of the suppressive activity, and mode of action of human Tregs are subjects of controversy. In this study, we show that Tregs display significant variability in the suppressive activity ex vivo as 54% of healthy blood donors examined had fully suppressive Tregs spontaneously, whereas in the remaining donors, anti-CD3/CD2/CD28 stimulation was required for Treg suppressive activity. Furthermore, anti-CD3/CD2/CD28 stimulation for 6 h and subsequent fixation in paraformaldehyde rendered the Tregs fully suppressive in all donors. The fixation-resistant suppressive activity of Tregs operated in a contact-dependent manner that was not dependent on APCs, but could be fully obliterated by trypsin treatment, indicating that a cell surface protein is directly involved. By add-back of active, fixed Tregs at different time points after activation of responding T cells, the responder cells were susceptible to Treg-mediated immune suppression up to 24 h after stimulation. This defines a time window in which effector T cells are susceptible to Treg-mediated immune suppression. Lastly, we examined the effect of a set of signaling inhibitors that perturb effector T cell activation and found that none of the examined inhibitors affected Treg activation, indicating pathway redundancy or that Treg activation proceeds by signaling mechanisms distinct from those of effector T cells.

Regulatory T-cell-mediated inhibition of antitumor immune responses is associated with clinical outcome in patients with liver metastasis from colorectal cancer.

Adaptive regulatory T cells (Tregs) contribute to an immunosuppressive microenvironment in colorectal cancer (CRC). Here, we examined whether the level of Treg-mediated inhibition of antitumor immune responses in patients with metastatic CRC (metCRC) selected for liver resection is associated with clinical outcome. Preoperatively and at follow-ups, we did flow-based phenotyping, examined antitumor immunity using peptides from carcinoembryonic antigen (CEA) protein in the presence or absence of CD4(+)CD25(+)CD127(dim/-) cells (Tregs) and determined cytokine and PGE(2) levels in patient blood samples. At 18 months post-surgery, 8 patients were disease free (7 alive and 1 dead of unrelated cause) and 10 had experienced disease recurrence (7 alive and 3 dead of metCRC). Prior to surgery, the patients demonstrated Treg-mediated suppression of TNFα and IFNγ expression that could be perturbed through the PGE(2)/cAMP pathway and the immune suppression was significantly higher in the group that later developed disease recurrence (P = 0.046). Furthermore, the post-surgery plasma PGE(2) levels were related to the clinical outcome (PGE(2) levels of 280 ± 47 vs. 704 ± 153 pg/ml (mean ± SEM) for disease free and recurrent disease, respectively). T-cell phenotyping revealed higher frequencies of COX-2(+) cells in the patients with recurrent disease. These findings support the notion that the level of Treg-mediated suppression of adaptive antitumor immune responses at the time of surgery may influence later clinical outcome of metCRC and provide valuable prognostic information.

CD147 (Basigin/Emmprin) identifies FoxP3+CD45RO+CTLA4+-activated human regulatory T cells.

Human CD4(+)FoxP3(+) T cells are functionally and phenotypically heterogeneous providing plasticity to immune activation and regulation. To better understand the functional dynamics within this subset, we first used a combined strategy of subcellular fractionation and proteomics to describe differences at the protein level between highly purified human CD4(+)CD25(+) and CD4(+)CD25(-) T-cell populations. This identified a set of membrane proteins highly expressed on the cell surface of human regulatory T cells (Tregs), including CD71, CD95, CD147, and CD148. CD147 (Basigin or Emmprin) divided CD4(+)CD25(+) cells into distinct subsets. Furthermore, CD147, CD25, FoxP3, and in particular CTLA-4 expression correlated. Phenotypical and functional analyses suggested that CD147 marks the switch between resting (CD45RA(+)) and activated (CD45RO(+)) subsets within the FoxP3(+) T-cell population. Sorting of regulatory T cells into CD147(-) and CD147(+) populations demonstrated that CD147 identifies an activated and highly suppressive CD45RO(+) Treg subset. When analyzing CD4(+) T cells for their cytokine producing potential, CD147 levels grouped the FoxP3(+) subset into 3 categories with different ability to produce IL-2, TNF-α, IFN-γ, and IL-17. Together, this suggests that CD147 is a direct marker for activated Tregs within the CD4(+)FoxP3(+) subset and may provide means to manipulate cells important for immune homeostasis.