Hyparillums A and B: polycyclic polyprenylated acylphloroglucinols from Hypericum patulum.

Hyparillums A (1) and B (2), two previously unidentified polycyclic polyprenylated acylphloroglucinols (PPAPs) with intricate architectures, were isolated from Hypericum patulum Thunb. Hyparillum A was the first PPAP with eight-carbon rings based on an unprecedented 6/6/5/6/6/5/6/4 octocyclic system featuring a rare heptacyclo[10.8.1.11,10.03,8.08,21.012,19.014,17]docosane core. In contrast, hyparillum B featured a novel heptacyclic architecture (6/6/5/6/6/5/5) based on a hexacyclo[9.6.1.11,9.03,7.07,18.011,16]nonadecane motif. Furthermore, hyparillums A and B demonstrated promising inhibitory effects on the proliferation of murine splenocytes stimulated by anti-CD3/anti-CD28 monoclonal antibodies and lipopolysaccharide, exhibiting half-maximal inhibitory concentration (IC50) values ranging from 6.13 ± 0.86 to 12.69 ± 1.31 µmol·L-1.

The Age-Male-Albumin-Bilirubin-Platelets (aMAP) Risk Score Predicts Liver Metastasis Following Surgery for Breast Cancer in Chinese Population: A Retrospective Study.

Objective: The current study is conducted to investigate the potential prognostic value of the age-male-albumin-bilirubin-platelets (aMAP) score in breast cancer patients with liver metastasis after surgery. Methods: This is a retrospective study of 178 breast cancer patients who developed liver metastasis after surgery. These patients were treated and followed up from 2000 to 2018 at our hospital. The aMAP risk score was estimated in accordance with the following formula: . The optimal cutoff value of the aMAP was evaluated via X-tile. Kaplan-Meier, Log-rank and Cox proportional hazards regression models were applied to determine the clinical influence of the aMAP score on the survival outcomes. The nomogram models were established by multivariate analyses. The calibration curves and decision curve analysis were applied to evaluate the estimated performance of the nomogram models. Results: A total of 178 breast cancer patients were divided into low aMAP score group (<47.6) and high aMAP score group (≥47.6) via X-tile plots. The aMAP score was a potential prognostic factor in multivariate analysis. The median disease free survival (p=0.0013) and overall survival (p=0.0003) in low aMAP score group were longer than in high aMAP score group. The nomograms were constructed to predict the DFS with a C-index of 0.722 (95% CI, 0.673-0.771), and the OS with a C-index of 0.708 (95% CI, 0.661-0.755). The aMAP-based nomograms had good predictive performance. Conclusion: The aMAP score is a potential prognostic factor in breast cancer with liver metastasis after surgery. The aMAP score-based nomograms were conducive to discriminate patients at high risks of liver metastasis and develop adjuvant treatment and prevention strategies.

LRP11 promotes stem-like T cells via MAPK13-mediated TCF1 phosphorylation, enhancing anti-PD1 immunotherapy.

BACKGROUND: Tumor-infiltrating T cells enter an exhausted or dysfunctional state, which limits antitumor immunity. Among exhausted T cells, a subset of cells with features of progenitor or stem-like cells has been identified as TCF1+ CD8+ T cells that respond to immunotherapy. In contrast to the finding that TCF1 controls epigenetic and transcriptional reprogramming in tumor-infiltrating stem-like T cells, little is known about the regulation of TCF1. Emerging data show that elevated body mass index is associated with outcomes of immunotherapy. However, the mechanism has not been clarified. METHODS: We investigated the proliferation of splenic lymphocytes or CD8+ T cells induced by CD3/CD28 stimulation in vitro. We evaluated the effects of low-density lipoprotein (LDL) and LRP11 inhibitors, as well as MAPK13 inhibitors. Additionally, we used shRNA technology to validate the roles of LRP11 and MAPK13. In an in vivo setting, we employed male C57BL/6J injected with B16 cells or MC38 cells to build a tumor model to assess the effects of LDL and LRP11 inhibitors, LRP11 activators, MAPK13 inhibitors on tumor growth. Flow cytometry was used to measure cell proportions and activation status. Molecular interactions and TCF1 status were examined using Western blotting. Moreover, we employed RNA sequencing to investigate the effects of LDL stimulation and MAPK13 inhibition in CD8+ T cells. RESULTS: By using a tumor-bearing mouse model, we found that LDL-induced tumor-infiltrating TCF1+PD1+CD8+ T cells. Using a cell-based chimeric receptor screening system, we showed that LRP11 interacted with LDL and activated TCF1. LRP11 activation enhanced TCF1+PD1+CD8+ T-cell-mediated antitumor immunity, consistent with LRP11 blocking impaired T-cell function. Mechanistically, LRP11 activation induces MAPK13 activation. Then, MAPK13 phosphorylates TCF1, leading to increase of stem-like T cells. CONCLUSIONS: LRP11-MAPK13-TCF1 enhanced antitumor immunity and induced tumor-infiltrating stem-like T cells.

Assessment of the albumin-bilirubin score in breast cancer patients with liver metastasis after surgery.

Objective: This study aims to investigate the potential prognostic value of albumin-bilirubin (ALBI) score in breast cancer patients with liver metastasis after surgery. Methods: This was a retrospective study of 178 breast cancer patients with liver metastasis after surgery. ALBI score was calculated by the following formula: (log10 bilirubin × 0.66) - (albumin × 0.085). The optimal cutoff value of ALBI score was assessed by X-tile. The clinical influence of ALBI score on survival outcomes using Kaplan-Meier method, Log-rank test, Cox proportional hazards regression model. The calibration curves, decision curve analysis and time-dependent ROC curve were used to assess the predictive performance of the nomogram's models. Results: The classifications of 178 breast cancer patients with liver metastasis after surgery were as follows: low ALBI score group (＜-3.36) vs. high ALBI score group (≥-3.36). The Cox proportional hazards regression model indicated that ALBI score was a potential predictor. Kaplan-Meier survival curve performed that the median disease free survival (p = 0.0029) and overall survival (p＜0.0001) in low ALBI score group were longer than in high ALBI score group. The ALBI-based nomograms had good predictive performance. Conclusions: The ALBI score has high prognostic ability for survival time in breast cancer with liver metastasis after surgery. These models will be valuable in discriminating patients at high risks of liver metastasis.

Adoptive transfer of immature dendritic cells with high HO-1 expression delays the onset of T1DM in NOD mice.

AIMS: To investigate the potential of imDCs with high expression of HO-1 in preventing or delaying the onset of Type 1 diabetes mellitus (T1DM) in non-obese diabetic (NOD) mice. MATERIALS AND METHODS: The phenotypic features of DCs in each group were assessed using flow cytometry. Western blot analysis was used to confirm the high expression of HO-1 in imDCs induced with CoPP. Additionally, flow cytometry was used to evaluate the suppressive capacity of CoPP-induced imDCs on splenic lymphocyte proliferation. Finally, the preventive effect of CoPP-induced imDCs was tested in NOD mice. KEY FINDINGS: Compared to imDCs, CoPP-induced imDCs exhibited a reduced mean fluorescence intensity (MFI) of the co-stimulatory molecule CD80 on their surface (P < 0.05) and significantly increased HO-1 protein expression (P < 0.05). Following LPS stimulation, the MFI of co-stimulatory molecules CD80 and CD86 on the surface of CoPP-induced imDCs remained at a lower level (P < 0.05). Furthermore, there was a reduced proliferation rate of lymphocytes stimulated with anti-CD3/28 antibodies. The adoptive transfer of CoPP-imDCs significantly reduced the incidence of T1DM (16.66 % vs. control group: 66.67 %, P = 0.004). Furthermore, at 15 weeks of age, the insulitis score was also decreased in the CoPP-induced imDC treatment group (P < 0.05). There were no significant differences in serum insulin levels among all groups. SIGNIFICANCE: ImDCs induced with CoPP and exhibiting high expression of HO-1 demonstrate a robust ability to inhibit immune responses and effectively reduce the onset of diabetes in NOD mice. This finding suggests that CoPP-induced imDCs could potentially serve as a promising treatment strategy for T1DM.

Prenyllongnols A-D, New Prenylated Acylphloroglucinols that Fight Concanavalin A-Induced Autoimmune Hepatitis.

Autoimmune hepatitis is a serious hepatic disorder with unknown nosogenesis, and natural products have been deemed to be one of the most significant sources of new drugs against this disease. Prenyllongnols A-D (1-4), four undescribed prenylated acylphloroglucinols, were isolated from Hypericum longistylum. Compounds 1-4 exhibited remarkable immunosuppressive activities in murine splenocyte proliferation under the induction of concanavalin A (Con A), and IC50 values ranged from 2.98 ± 0.21 to 6.34 ± 0.72 µM. Furthermore, in a Con A-challenged autoimmune hepatitis mouse model, the mice in the group that were pretreated with isolate 2 significantly ameliorated liver injury and decreased proinflammatory cytokine production. Notably, natural product 2 was the first prenylated acylphloroglucinol to protect against concanavalin A-induced autoimmune hepatitis. This finding underscores the potential of prenylated acylphloroglucinol-type metabolites as promising candidates for designing novel immunosuppressors in the quest for new antiautoimmune hepatitis drugs.

Bioactive Indole Alkaloid from Aspergillus amoenus TJ507 That Ameliorates Hepatic Ischemia/Reperfusion Injury.

Hepatic ischemia/reperfusion injury (IRI) is a major factor contributing to the failure of hepatic resection and liver transplantation. As part of our ongoing investigation into bioactive compounds derived from fungi, we isolated eight indole alkaloids (1-8) from the endophytic fungus Aspergillus amoenus TJ507. Among these alkaloids, one previously undescribed compound, amoenamide D (1), was identified. The planar structure of 1 was elucidated by extensive spectroscopic analysis, including HRESIMS and NMR spectra. The absolute configuration of 1 was elucidated by using electronic circular dichroism calculations. Notably, in the CoCl2-induced hepatocyte damage model, notoamide Q (3) exhibited significant anti-hypoxia injury activity. Furthermore, in a murine hepatic ischemia/reperfusion injury model, treatment with 3 prevents IRI-induced liver damage and hepatocellular apoptosis. Consequently, 3 might serve as a potential lead compound to prevent hepatic ischemia/reperfusion injury.

ERK Inhibition Promotes Engraftment of Allografts by Reprogramming T-Cell Metabolism.

Extracellular regulated protein kinases (ERK) signaling is a master regulator of cell behavior, life, and fate. Although ERK pathway is shown to be involved in T-cell activation, little is known about its role in the development of allograft rejection. Here, it is reported that ERK signaling pathway is activated in allograft-infiltrating T cells. On the basis of surface plasmon resonance technology, lycorine is identified as an ERK-specific inhibitor. ERK inhibition by lycorine significantly prolongs allograft survival in a stringent mouse cardiac allotransplant model. As compared to untreated mice, lycorine-treated mice show a decrease in the number and activation of allograft-infiltrated T cells. It is further confirmed that lycorine-treated mouse and human T cells are less responsive to stimulation in vitro, as indicated by their low proliferative rates and decreased cytokine production. Mechanistic studies reveal that T cells treated with lycorine exhibit mitochondrial dysfunction, resulting in metabolic reprogramming upon stimulation. Transcriptome analysis of lycorine-treated T cells reveals an enrichment in a series of downregulated terms related to immune response, the mitogen-activated protein kinase cascade, and metabolic processes. These findings offer new insights into the development of immunosuppressive agents by targeting the ERK pathway involved in T-cell activation and allograft rejection.

Discovery of ergosterol derivative from Aspergillus sp. TJ507 that protects against hepatic ischemia/reperfusion injury.

Hepatic ischemia/reperfusion injury is a major cause of hypohepatia after surgical procedures such as hypovolemic shock, transplantation, and so on. In our continuous study of bioactive natural products from fungus, eight ergosterol-type sterides (1-8), including two undescribed compounds, sterolaspers A (1) and B (2), were isolated from Aspergillus sp. TJ507. Structure elucidation was accomplished by extensive spectroscopic analysis and comparison with the reported NMR data as well as X-Ray single crystal diffraction tests. Activity screen of these isolates showed 5α-stigmast-3,6-dione (3) possessing anti-hypoxia injury effects against CoCl2-induced hypoxia damage in hepatocytes. More importantly, compound 3 could improve liver function, alleviate liver damage, and restrain the hepatocellular apoptosis in hepatic ischemia/reperfusion injury murine model. As such, this ergosterol-type steride, 5α-stigmast-3,6-dione (3), might serve as lead structure for the development of novel hepatoprotective agents in the clinical treatment of hepatic ischemia/reperfusion injury.

New Insight into the Concanavalin A-Induced Apoptosis in Hepatocyte of an Animal Model: Possible Involvement of Caspase-Independent Pathway.

Concanavalin A (Con A) is known to be a T-cell mitogen and has been shown to induce hepatitis in mice through the triggering of conventional T cells and NKT cells. However, it remains unknown whether Con A itself can directly induce rapid hepatocyte death in the absence of a functional immune system. Here, by using an immunodeficient mouse model, we found Con A rapidly induced liver injury in vivo despite a lack of immunocyte involvement. We further observed in vitro that hepatocytes underwent a dose-dependent but caspase-independent apoptosis in response to Con A stimulation in vitro. Moreover, transcriptome RNA-sequencing analysis revealed that apoptosis pathways were activated in both our in vivo and in vitro models. We conclude that Con A can directly induce rapid but non-classical apoptosis in hepatocytes without the participation of immunocytes. These findings provide new insights into the mechanism of Con A-induced hepatitis.

Long-Chain Acylcarnitines Induce Senescence of Invariant Natural Killer T Cells in Hepatocellular Carcinoma.

CD1d-restricted invariant natural killer T (iNKT) cells actively patrol the liver and possess valuable antitumor potential. However, clinical trials evaluating administration of iNKT cell-specific agonist α-galactosylceramide (α-GalCer) have failed to achieve obvious tumor regression. Improving the efficacy of iNKT cell-based immunotherapy requires a better understanding of the factors restraining the clinical benefits. In the context of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC), we found circulating and hepatic iNKT cells were hyperactivated but demonstrated imbalances in ratio and defective α-GalCer responsiveness. Exogenous IL2 helped to expand residual α-GalCer-responsive clones with reduced T-cell receptor diversity. However, transcriptome-wide analysis revealed activation of the senescence-associated secretory phenotype and dampened cytotoxicity in iNKT cells, weakening their immune surveillance capacity. The senescent status of iNKT cells from the patients was further illustrated by cell-cycle arrest, impaired telomere maintenance, perturbed calcium transport-related biological processes, and altered metabolism. Lipidomic profiling revealed the accumulation of long-chain acylcarnitines (LCAC) and aberrant lipid metabolism in HCC tissue. Exogenous LCACs, especially palmitoyl-carnitine and stearoyl-carnitine, inhibited iNKT cell expansion and promoted senescence. Collectively, our results provide deeper insights into iNKT cell dysregulation and identify a cell senescence-associated challenge for iNKT cell-based immunotherapy in HBV-related HCC. The mechanistic links between iNKT cell senescence and accumulated LCACs suggest new targets for anti-HCC immunotherapies. SIGNIFICANCE: Patients with HBV-related HCC exhibit a cell senescence-associated dysregulation of invariant natural killer cells that is related to altered lipid metabolism and accumulated LCACs in tumor tissue.

Discovery of Undescribed Monoterpenoid Polyprenylated Acylphloroglucinols with Immunosuppressive Activities from Hypericum longistylum.

Acylphlorostylums A-G (1-7), seven undescribed monoterpenoid polyprenylated acylphloroglucinols, were isolated and identified from Hypericum longistylum. Significantly, acylphlorostylums A and B were the first monoterpenoid polyprenylated acylphloroglucinols possessing a dodecahydro-1H-benzo [b]cyclopenta [e]oxepine moiety bearing a 6/7/5 fused tricyclic ring system that assembled by the attack from 4-OH to C-13. In addition, acylphlorostylums A-G exhibited moderate in vitro immunosuppressive activity in anti-CD3/anti-CD28 monoclonal antibodies, lipopolysaccharide and concanavalin A-induced murine splenocyte proliferation, with IC50 values ranging from 1.51 ± 0.12 to 18.49 ± 1.67 µM, underscoring those isolates as novel chemical templates in the development of novel immunosuppressors.

Discovery of immunosuppressive Lupane-type Triterpenoids from Hypericum longistylum.

The phytochemistry study of Hypericum longistylum afforded one new degraded lupane-type triterpenoid, triterhyper A (1), and seven known congeners (2-8). The structures of those natural products were identified by extensive spectroscopic techniques and single crystal diffraction tests. Notably, triterhyper A (1) possessing an unusual 28-nor-lupane skeleton. More significantly, compounds 1-3 exhibited potential inhibitory effects on murine splenocytes proliferation stimulated by anti-CD3/anti-CD28 monoclonal antibodies (mAbs) and lipopolysaccharide (LPS), with IC50 values ranging from (4.56 ± 0.45) µM to (18.34 ± 2.34) µM, highlighting that those isolates as potential lead compounds in the development of immunosuppressive drugs for autoimmune disease treatment.

New secondary metabolites from the endophytic fungus Aspergillus sp. from Tripterygium wilfordii.

One new 3,5-dimethylorsellinic acid (DMOA)-based meroterpenoid (1), one prenylated tryptophan derivative (2), together with ten known compounds (3-12) were isolated from the endophytic fungus Aspergillus sp. from Tripterygium wilfordii. Their structures and absolute configurations were determined by NMR spectroscopic data, HRESIMS data, UV and IR data as well as electronic circular dichroism (ECD) calculation. In structure, compound 1 was a rare example of DMOA-based meroterpenoid with a cis-fused C/D ring system, and compound 2 possessed an unusual (E)-oxime group. In bioactivity, the lovastatin analogues 5, 6, 9 and 10 showed potential immunosuppressive activity against anti-CD3/anti-CD28 monoclonal antibodies (mAbs)-irritated murine splenocytes proliferation, with IC50 values ranging from (5.30 ± 0.51) µM to (16.51 ± 1.62) µM.

Liraglutide protects against lethal renal ischemia-reperfusion injury by inhibiting high-mobility group box 1 nuclear-cytoplasmic translocation and release.

Liraglutide, a glucagon-like peptide-1 receptor (GLP-1R) agonist, has been reported to exert protective effects against myocardial, hepatic, and gastric ischemia-reperfusion injury (IRI), but whether it can protect against renal IRI remains unknown. Here, a lethal renal IRI model was established with a 100% mortality rate in untreated mice. Treatment with liraglutide involving a regimen of multiple doses resulted in 100% survival, remarkable preservation of renal function, a significant reduction in pathological damage, and blunted upregulation of TNF-α, IL-1ß, IL-6, MCP-1, TLR-2, TLR-4, and RAGE mRNA. We found that liraglutide treatment dramatically inhibited ischemia-induced nucleocytoplasmic translocation and release of HMGB1. This inhibition was associated with a marked decrease (~ 60%) in nuclear histone acetyltransferase activity. In addition, the protective effects of liraglutide on renal IRI were largely abolished by the administration of exogenous HMGB1. When the GLP-1R antagonist exendin (9-39) was given to mice before each liraglutide administration, or GLP-1R-/- mice were used for the renal IRI experiments, the protective effect of liraglutide on renal IRI was partially reversed. Moreover, liraglutide pretreatment significantly inhibited HMGB1 nucleocytoplasmic translocation during hypoxic culture of HK-2 cells in vitro, but the addition of exendin (9-39) significantly eliminated this inhibition. We demonstrate here that liraglutide can exert a strong protective effect on lethal renal IRI in mice. This protection appears to be related to the inhibition of HMGB1 nuclear-cytoplasmic translocation and release and partially depends on GLP-1R. Thus, liraglutide may be therapeutically useful for the clinical prevention and treatment of organ IRI.

The Expansion and Cytotoxicity Detection of Human iNKT Cells.

Invariant natural killer T (iNKT) cell is a type of innate-like T cell subsets with both T and NK cell phenotype and functions. They recognize lipid antigens presented by CD1d molecules and can be specifically activated by alpha-galactosylceramide (α-GalCer) in vitro. After activation, iNKT cells expand efficiently and exert direct killing effects. Based on it, we mainly introduce the protocols of detection of human iNKT cell functions in vitro, including in vitro expansion and their cytotoxicity to tumor cells.

Bioassay-Guided Isolation of an Abetiane-Type Diterpenoid from Prunella vulgaris That Protects against Concanavalin A-Induced Autoimmune Hepatitis.

Prunella vulgaris is a widely used edible Chinese medicinal plant. In the present study, two new abietane-type diterpenoids, abietoquinones A (1) and B (2), were isolated from this plant by an immunosuppressive bioassay-guided isolation procedure. Their structures were elucidated unambiguously by NMR spectroscopic analysis, single-crystal X-ray crystallography, and electronic circular dichroism calculations. Compounds 1 and 2 bear a cyclohex-2-ene-1,4-dione moiety, which is uncommon among abietane diterpenes. Also, abietoquinone A (1) suppressed murine splenocyte proliferation and decreased the production of proinflammatory cytokines induced by concanavalin A (Con A) in vitro. In Con A-challenged mice, preinjection with 1 significantly ameliorated liver injury. Additionally, abietoquinone A (1) exhibited inhibitory activities against the proliferation of murine splenocytes and human T cells induced by anti-CD3/anti-CD28 monoclonal antibodies (mAbs).

New immunosuppressive secondary metabolites from the endophytic fungus Aspergillus sp.

Six new metabolites, including two diphenolic derivatives (1 and 2), one pseurotin (3), one butenolide derivative (4), one benzopyran (5) and one isochromane lactone (6), together with ten known compounds (7-16) were isolated from an endophytic fungus Aspergillus sp. Their planar structures and absolute configurations were established based on techniques of MS, NMR, IR, UV, [Rh2(OCOCF3)4] complex-induced ECD, quantum chemical electronic circular dichroism (ECD) calculations, and single crystal X-ray diffraction. Structurally, compound 2 represents the first example of diphenolic derivative possessing an unusual 1-oxaspiro[2.4]heptane core bearing a 5/3 bicyclic skeleton; compound 3 represents the first example of pseurotin type natural products that only one hydroxy group is substituted at side chain. In bioassay, compounds 3, 7 and 8 exhibited potential inhibitory effect on the proliferation of anti-CD3/anti-CD28 monoclonal antibodies (mAbs) induced murine T cells, with IC50 values of (7.81 ± 0.71), (8.25 ± 0.78) and (8.84 ± 0.81) µM, respectively.

Rabbit antithymocyte globulin induces human lymphocyte activation, proliferation, and apoptosis in the absence of complement: an experimental study.

Rabbit antithymocyte globulin (rATG) has become the first choice for induction therapy in HLA-presensitized patients undergoing organ transplantation. Meanwhile, complement inhibitors have been approved for preventing or treating antibody-mediated rejection in these patients. The biological effects of rATG on lymphocytes in cases of complement deficiency or significant inhibition are not yet clear. We measured lymphocyte activation, proliferation, and apoptosis in response to rATG treatment in the absence of complement. T-cell subsets were analyzed transcriptomically features to rATG stimulation. Activation-related phenotypes on T cells were determined in patients after rATG administration. We found that rATG treatment led to lymphocyte activation and proliferation in vitro without the addition of complement. A dose-dependent apoptosis in rATG-treated lymphocytes was detected, which was partially caspase-3-dependent but Fas/FasL-independent. T cells were more sensitive to rATG stimulation than were non-T cells. Both CD4+ T cells and CD8+ T cells upregulated a series of genes related to cell activation, cytokine production and apoptosis to rATG stimulation. CD69 and CD25 levels in surviving T cells were increased in patients after rATG administration. These findings indicate that rATG can stimulate lymphocyte activation, proliferation, and apoptosis in the absence of complement. Biologic effects of rATG other than complement-dependent cytotoxicity need to be concerned.

Mucosal-Associated Invariant T Cell Dysregulation Correlates With Conjugated Bilirubin Level in Chronic HBV Infection.

BACKGROUND AND AIMS: Mucosal-associated invariant T (MAIT) cells are nonconventional T cells restricted to major histocompatibility complex class I-related protein 1 (MR1). They are highly abundant in human liver and activated by T-cell receptor (TCR)-dependent and TCR-independent mechanisms to exhibit rapid, innate-like effector responses. However, the roles of MAIT cells in chronic HBV infection are still open for study. This study aims to test their antiviral potential and investigate their dynamic changes and regulating factors during chronic HBV infection. APPROACH AND RESULTS: Blood samples from 257 chronic HBV-infected patients were enrolled, and nontumor liver specimens were collected from 58 HBV-infected HCC patients. Combining cell-culture experiments and human data, we showed that MAIT cells had strong cytotoxicity against HBV-transfected hepatocytes in an MR1-dependent way. However, circulating and hepatic MAIT cells in HBV-infected patients decreased significantly compared to controls. Correlation analysis suggested that MAIT cell frequency was associated with disease progression and inversely correlated with serum-conjugated bilirubin level. In particular, conjugated bilirubin not only directly promoted MAIT cell activation and apoptosis, but also impaired TCR-induced proliferation and expansion of MAIT cells, which could be partially rescued by IL-2 in the absence of conjugated bilirubin. Despite that MAIT cells from patients with high conjugated bilirubin levels showed decreased cytokine-producing capacity, the increased TCR-dependent antiviral cytokine production suggested MAIT cells as an important guardian of chronic HBV with high conjugated bilirubin. CONCLUSIONS: We reveal the MR1-dependent, anti-HBV potential of MAIT cells and identify conjugated bilirubin as a major factor dysregulating its frequency and function in chronic HBV-infected patients, suggesting a therapeutic target for MAIT-cell-based immunity against chronic HBV infection.