A Lesson of Immunosuppression in Renal Transplant: Retreat or Hold?

Background: Aiming at understanding whether there are cases of near-tolerance among long-term surviving kidney transplant recipients in our center, or even operant tolerance can be attempted based on their immune status, we analyzed changes of immune cell subsets and cytokines in various groups, and evaluated immune status of long-term survival recipients. Methods: A real-world, observational, retrospective cohort study was conducted in our hospital. Twenty-eight long-term recipients were selected as study subjects, 15 recent postoperative stable recipients, and 15 healthy subjects as controls. T and B lymphocyte subsets, MDSCs, and cytokines were detected and analyzed. Results: Treg/CD4 T cells, total B and B10 cells in long-term and recent renal recipients were lower than healthy controls (HC). The level of IFN-γ and IL-17A in long-term survival patients was obviously higher than that in recent postoperative stable recipients and HC, while TGF-ß1 level was significantly lower in long-term survival group than in short-term postoperative group and HC. Notably, compared with short-term recipients, it has been found that the IL-6 level in both positive and negative HLA groups were obviously lower (all P<0.05). In the long-term survival group, 43% of recipients were positive for urinary protein and 50% were positive for HLA antibody. Conclusion: This "real-world" study validates the findings of real status of long-term survival recipients observed in clinical trials. Contrary to a state of proper tolerance as expected, the group recipients in long-term survival were accompanied by the increased indicators of immune response, while those related to immune tolerance were not significantly increased. Long-term survival recipients with stable renal function may be in an immune equilibrium state where immunosuppression and rejection coexist under the action of low-intensity immune agents. If immunosuppressive agents are reduced or even removed, rejection may occur.

The role of pyroptosis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the main histologic type of liver cancer. It accounts for the majority of all diagnoses and deaths due to liver cancer. The induction of tumor cell death is an effective strategy to control tumor development. Pyroptosis is an inflammatory programmed cell death caused by microbial infection, accompanied by activation of inflammasomes and release of pro-inflammatory cytokines, interleukin-1ß (IL-1ß), and interleukin-18 (IL-18). The cleavage of gasdermins (GSDMs) promotes the occurrence of pyroptosis leading to cell swelling, lysis, and death. Accumulating evidence has indicated that pyroptosis influences the progression of HCC by regulating immune-mediated tumor cell death. Currently, some researchers hold the view that inhibition of pyroptosis-related components may prevent the incidence of HCC, but more researchers have the view that activation of pyroptosis exerts a tumor-inhibitory effect. Growing evidence indicates that pyroptosis can prevent or promote tumor development depending on the type of tumor. In this review, pyroptosis pathways and pyroptosis-related components were discussed. Next, the role of pyroptosis and its components in HCC was described. Finally, the therapeutic significance of pyroptosis in HCC was discussed.

The Novel MyD88 Inhibitor TJ-M2010-5 Protects Against Hepatic Ischemia-reperfusion Injury by Suppressing Pyroptosis in Mice.

BACKGROUND: . With the development of medical technology and increased surgical experience, the number of patients receiving liver transplants has increased. However, restoration of liver function in patients is limited by the occurrence of hepatic ischemia-reperfusion injury (IRI). Previous studies have reported that the Toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88) signaling pathway and pyroptosis play critical roles in the development of hepatic IRI. METHODS: . A mouse model of segmental (70%) warm hepatic IRI was established using BALB/c mice in vivo. The mechanism underlying inflammation in mouse models of hepatic IRI was explored in vitro using lipopolysaccharide- and ATP-treated bone marrow-derived macrophages. This in vitro inflammation model was used to simulate inflammation and pyroptosis in hepatic IRI. RESULTS: . We found that a MyD88 inhibitor conferred protection against partial warm hepatic IRI in mouse models by downregulating the TLR4/MyD88 signaling pathway. Moreover, TJ-M2010-5 (a novel MyD88 inhibitor, hereafter named TJ-5) reduced hepatic macrophage depletion and pyroptosis induction by hepatic IRI. TJ-5 treatment inhibited pyroptosis in bone marrow-derived macrophages by reducing the nuclear translocation of nuclear factor kappa-light-chain-enhancer of activated B cells, decreasing the release of high-mobility group box-1, and promoting endocytosis of lipopolysaccharide-high-mobility group box-1 complexes. CONCLUSIONS: . Inhibition of MyD88 may protect the liver from partial warm hepatic IRI by reducing pyroptosis in hepatic innate immune cells. These results reveal the mechanism underlying the development of inflammation in partially warm hepatic IRI and the induction of cell pyroptosis.

TJ-M2010-5, A self-developed MyD88 inhibitor, attenuates liver fibrosis by inhibiting the NF-κB pathway.

Liver fibrosis is the result of most chronic inflammatory liver damage and seriously endangers human health. However, no drugs have been approved to treat this disease. Previous studies showed that the Toll-like receptors (TLRs)/myeloid differentiation factor-88 (MyD88)/nuclear factor-κB (NF-κB) pathway plays a key role in liver fibrosis. TJ-M2010-5 is a self-developed small molecule MyD88 inhibitor, which has been proven to have a good protective effect in a variety of inflammatory disease models. In the present study, to investigate the anti-fibrotic effect of TJ-M2010-5, mice were injected with carbon tetrachloride (CCl4) in vivo and LX2 cells (a human hepatic stellate cell line) were treated with TGF-ß1 in vitro to induce liver fibrosis. In vivo studies showed that TJ-M2010-5 attenuated the CCl4-induced liver damage, collagen accumulation, and the activation of hepatic stellate cells by inhibiting the nuclear transfer of NF-κB. Moreover, in vitro experiments of LX2 cells stimulated with TGF-ß1 further indicated that the NF-κB pathway is involved in the development of liver fibrosis. TJ-M2010-5 significantly inhibited the proliferation and activation of LX2 cells. In addition, TJ-M2010-5 upregulated the expression of bone morphogenetic protein and membrane-bound inhibitor (BAMBI) in LX2 cells by blocking the activation of MyD88/NF-κB, thereby inhibiting the phosphorylation of Smad2/3 and the expression of collagen I (COL1A1) induced by TGF-ß1. In conclusion, this study illustrates the anti-hepatic fibrosis effect of TJ-M2010-5 and provides a new treatment method for liver fibrosis.

TJ-M2010-5, a novel CNS drug candidate, attenuates acute cerebral ischemia-reperfusion injury through the MyD88/NF-κB and ERK pathway.

Background: Cerebral ischemia-reperfusion injury (CIRI) inevitably occurs after vascular recanalization treatment for ischemic stroke. The accompanying inflammatory cascades have a major impact on outcome and regeneration after ischemic stroke. Evidences have demonstrated that TLR/MyD88/NF-κB signaling contributes to CIRI. This study aimed to investigate the druggability of MyD88 in the central nervous system (CNS) and the neuroprotective and anti-neuroinflammatory effects of the MyD88 inhibitor TJ-M2010-5 on CIRI. Methods: A middle cerebral artery occlusion (MCAO) model was used to simulate CIRI in mice. BV-2 cells were stimulated with oxygen glucose deprivation/reoxygenation (OGD/R) or lipopolysaccharide, and SH-SY5Y cells were induced by OGD/R in vitro. Neurological deficit scores and cerebral infarction volumes were evaluated. Immunofluorescence staining was performed to measure neuronal damage and apoptosis in the brain. The anti-neuroinflammatory effect of TJ-M2010-5 was evaluated by analyzing the expression of inflammatory cytokines, activation of microglia, and infiltration of peripheral myeloid cells. The expression of proteins of the MyD88/NF-κB and ERK pathway was detected by Simple Western. The concentrations of TJ-M2010-5 in the blood and brain were analyzed by liquid chromatography-mass spectrometry. Results: The cerebral infarction volume decreased in mice treated with TJ-M2010-5, with the most prominent decrease being approximately 80% of the original infarction volume. Neuronal loss and apoptosis were reduced following TJ-M2010-5 treatment. TJ-M2010-5 inhibited the infiltration of peripheral myeloid cells and the activation of microglia. TJ-M2010-5 also downregulated the expression of inflammatory cytokines and inhibited the MyD88/NF-κB and ERK pathway. Furthermore, TJ-M2010-5 showed good blood-brain barrier permeability and no neurotoxicity. Conclusion: TJ-M2010-5 has an excellent therapeutic effect on CIRI as a novel CNS drug candidate by inhibiting excessive neuroinflammatory responses.

Lymphocyte function based on IFN-γ secretion assay may be a promising indicator for assessing different immune status in renal transplant recipients.

BACKGROUND: Immunological monitoring plays a crucial role in organ recipients for allowing tailoring of immunosuppression. However, there is still a paucity of promising indicators for detecting immune status in recipients. METHODS: We conducted a prospective study to characterize the immune status by detecting dynamically lymphocyte subsets and function (represented by the abilities to secrete IFN-γ) in the first 6 months posttransplant in renal recipients. Participants were classified into an immune stable group, infected group, and rejected group. RESULTS: In the stable group, our study suggested that the counts and function of CD4+ T, CD8+ T, and NK lymphocytes decreased to their nadir at week 2, and thereafter these indicators were gradually restored. The counts exceeded pre-operative levels, whereas function did not reach the pre-transplant levels by 6 months. We demonstrated that function of lymphocytes was considerably decreased in infected recipients compared with the stable group when infection occurred. By contrast, the function of lymphocytes was obviously increased at the point of rejection. Receiver operating characteristic (ROC) analysis in the combination of subsets and function of lymphocytes presented a superior clinical value with an area under the curve (AUC) of 0.903 in the diagnosis of infected receivers, and IFN-γ+CD8+ T cells% is the highest indicator with the auROC curve of 0.862. Another ROC analysis confirmed that IFN-γ+CD4  T cells% presented a preferable diagnostic value with an area of 0.887 for rejected recipients. CONCLUSIONS: In conclusion, the ability of lymphocyte subsets secreting IFN-γ may provide a promising assessment of immune status in recipients and allow timely modifying immunosuppression.

Evaluation of hepatic steatosis before liver transplantation in ex vivo by volumetric quantitative PDFF-MRI.

PURPOSE: Over the last two decades, extended criteria have promoted an increased number of donor livers available for liver transplantation. But posttransplant graft loss is still a major concern. Macrovesicular hepatic steatosis (MHS) is recognized as the most significant prognostic histologic parameter in predicting posttransplant graft loss. We aimed to evaluate the utility of ex vivo volumetric quantitative MRI for quantifying MHS before liver transplantation using proton density fat-fraction (PDFF-MRI) histogram analysis. METHODS: PDFF-MRI was performed at 3.0T in 40 livers. We obtained histogram parameters of whole-liver volume of interest, including the mean, median, 5th, 10th, 25th, 75th, 90th, and 95th percentile PDFF; skewness; kurtosis; entropy; and volume. RESULTS: Livers from 40 cadaveric donors were included, and histologic ex vivo fat quantification was available for 33 livers. Ten livers had MHS and 23 had normal fat content. The MHS group had higher mean, median, 5th, 10th, 25th, 75th, 90th, and 95th percentile PDFF, and entropy than the group with normal fat content (P < .05). Median PDFF had greater area under the curve value than other parameters. Mean PDFF showed an excellent correlation with entropy and a moderate correlation with MHS quantification on histology. CONCLUSIONS: Ex vivo volumetric quantitative PDFF-MRI histogram analysis is a very useful and noninvasive method to detect MHS before liver transplantation. Median PDFF was the best predictor of the presence of MHS. Entropy is a very promising parameter.

TJ-M2010-5, a novel MyD88 inhibitor, corrects R848-induced lupus-like immune disorders of B cells in vitro.

B cell hyperactivities are involved in the development of systemic lupus erythematosus (SLE). Toll-like receptor 7 (TLR7) in the B cells plays a pivotal role in the pathogenesis of SLE. Previous studies have focused on the intrinsic role of B cells in TLR7/MyD88 signaling and consequently on immune activation, autoantibody production, and systemic inflammation. However, a feasible treatment for this immune disorder remains to be discovered. The in vitro cellular response that have been studied likely plays a central role in the production of some important autoantibodies in SLE. We successfully used R848 to build a lupus-like B cell model in vitro; these B cells were overactivated, differentiated into plasma cells, escaped apoptosis, massively proliferated, and produced large amounts of autoantibodies and cytokines. In the present study, we found that TJ-M2010-5, a novel MyD88 inhibitor previously synthesized in our lab, seemed to inhibit the lupus-like condition of B cells, including overactivation, massive proliferation, differentiation into plasma cells, and overproduction of autoantibodies and cytokines. TJ-M2010-5 also induce B cells apoptosis. Furthermore, TJ-M2010-5 was found to remarkably inhibit NF-κB and MAPK signaling. In summary, TJ-M2010-5 might correct R848-induced lupus-like immune disorders of B cells by blocking the TLR7/MyD88/NF-κB and TLR7/MyD88/MAPK signaling pathways.

Combination of lymphocyte number and function in evaluating host immunity.

Accurate monitoring of host immunity is hampered by the flaws of conventional tests. The relationship between lymphocyte number and function is unknown. The function of lymphocytes was analyzed based on IFN-γ secretion assay. Lymphocyte number and function was investigated in individuals under various states. The number of CD4+ and CD8+ T cells was gradually decreased, whereas the function of them was gradually increased with increasing age. A significantly negative correlation existed between the number and function of both CD4+ and CD8+ T cells. Differently, both the number and function of NK cells are maintained at a high level after birth. Staying up all night was found to impair the function of CD4+, CD8+ T cells, or NK cells. Lymphocyte number and function were both decreased in patients with immunosuppressive conditions or opportunistic infections, while the opposite phenomenon was observed in patients with some autoimmune diseases (except for NK cells). In kidney transplant recipients, the number and function of CD4+ and CD8+ T cells were increased or decreased when rejection or infection occurred. We demonstrated that evaluation of host immunity based on combination of lymphocyte number and function plays an important role in the diagnosis, treatment, and prognosis of diseases.