Identification of two key biomarkers CD93 and FGL2 associated with survival of acute myeloid leukaemia by weighted gene co-expression network analysis.

Acute myeloid leukaemia (AML) is a biologically heterogeneous haematological malignancy. This study was performed to identify the potential biomarkers for the prognosis and treatment of AML. We applied weighted gene co-expression network analysis to identify key modules and hub genes related to the prognosis of AML using data from The Cancer Genome Atlas (TCGA). In total, 1581 differentially expressed genes (1096 upregulated and 485 downregulated) were identified between AML patients and healthy controls, with the blue module being the most significant among 14 modules associated with AML morphology. Through functional enrichment analysis, we identified 217 genes in the blue module significantly enriched in 'neutrophil degranulation' and 'neutrophil activation involved in immune response' pathways. The survival analysis revealed six genes (S100A9, S100A8, HK3, CD93, CXCR2 and FGL2) located in the significantly enriched pathway that were notably related to AML survival. We validated the expression of these six genes at gene and single-cell levels and identified methylation loci of each gene, except for S100A8. Finally, in vitro experiments were performed to demonstrate whether the identified hub genes were associated with AML survival. After knockdown of CD93 and FGL2, cell proliferation was significantly reduced in U937 cell line over 5 days. In summary, we identified CD93 and FGL2 as key hub genes related to AML survival, with FGL2 being a novel biomarker for the prognosis and treatment of AML.

Unravelling the potential of TIM-3 gene polymorphism in allogeneic hematopoietic stem cell transplantation - a preliminary study.

BACKGROUND: T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) molecule is a key regulator of the immune response by exerting an inhibitory effect on various types of immune cells. Understanding the role of TIM-3 in hematopoietic stem cell transplantation (HSCT) may improve transplant outcomes. Our study evaluated the potential association between TIM-3 polymorphisms, namely rs1036199 (A > C) or rs10515746 (C > A), changes which are located in exon 3 and the promoter region of the TIM-3 gene, and post-HSCT outcomes. METHODS: One-hundred and twenty allogeneic HSCT patients and their respective donors were enrolled and genotyped for TIM-3 single nucleotide polymorphisms (SNPs) using real-time PCR with TaqMan assays. RESULTS: We found that the presence of the rare alleles and heterozygous genotypes of studied SNP in recipients tended to protect against or increase the risk for acute graft-versus-host disease (aGvHD). For the rs1036199 polymorphism, recipients with the AC heterozygous genotype (p = 0.0287) or carrying the rarer C allele (p = 0.0334) showed a lower frequency of aGvHD development along all I-IV grades. A similar association was detected for the rs10515746 polymorphism as recipients with the CA genotype (p = 0.0095) or the recessive A allele (p = 0.0117) less frequently developed aGvHD. Furthermore, the rarer A allele of rs10515746 SNP was also associated with a prolonged aGvHD-free survival (p = 0.0424). Cytomegalovirus (CMV) infection was more common in patients transplanted with TIM-3 rs10515746 mismatched donors (p = 0.0229) and this association was also found to be independent of HLA incompatibility and pre-transplant CMV-IgG status. Multivariate analyses confirmed the role of these recessive alleles and donor-recipient TIM-3 incompatibility as an independent factor in aGvHD and CMV development. CONCLUSIONS: Polymorphism of TIM-3 molecule may affect the immune response in HSCT patients. The recessive alleles of rs1036199 and rs10515746 SNPs decreased the risk of developing aGvHD. TIM-3 donor-recipient genetic matching may also affect the risk of post-transplant CMV infection, indicating the potential value of genetic profiling in optimizing transplant strategies.

The effect of Toxoplasma gondii infection on galectin-9 expression in decidual macrophages contributing to dysfunction of decidual NK cells during pregnancy.

BACKGROUND: Toxoplasma gondii infection causes adverse pregnancy outcomes by affecting the expression of immunotolerant molecules in decidual immune cells. Galectin-9 (Gal-9) is widely expressed in decidual macrophages (dMφ) and is crucial for maintaining normal pregnancy by interacting with the immunomodulatory protein T-cell immunoglobulin and mucin domain-containing molecule 3 (Tim-3). However, the effects of T. gondii infection on Gal-9 expression in dMφ, and the impact of altered Gal-9 expression levels on the maternal-fetal tolerance function of decidual natural killer (dNK) cells, are still unknown. METHODS: Pregnancy outcomes of T. gondii-infected C57BL/6 and Lgals9-/- pregnant mice models were recorded. Expression of Gal-9, c-Jun N-terminal kinase (JNK), phosphorylated JNK (p-JNK), and Forkhead box protein O1 (FOXO1) was detected by western blotting, flow cytometry or immunofluorescence. The binding of FOXO1 to the promoter of Lgals9 was determined by chromatin immunoprecipitation-polymerase chain reaction (ChIP-PCR). The expression of extracellular signal-regulated kinase (ERK), phosphorylated ERK (p-ERK), cAMP-response element binding protein (CREB), phosphorylated CREB (p-CREB), T-box expressed in T cells (T-bet), interleukin 10 (IL-10), and interferon gamma (IFN-γ) in dNK cells was assayed by western blotting. RESULTS: Toxoplasma gondii infection increased the expression of p-JNK and FOXO1 in dMφ, resulting in a reduction in Gal-9 due to the elevated binding of FOXO1 with Lgals9 promoter. Downregulation of Gal-9 enhanced the phosphorylation of ERK, inhibited the expression of p-CREB and IL-10, and promoted the expression of T-bet and IFN-γ in dNK cells. In the mice model, knockout of Lgals9 aggravated adverse pregnancy outcomes caused by T. gondii infection during pregnancy. CONCLUSIONS: Toxoplasma gondii infection suppressed Gal-9 expression in dMφ by activating the JNK/FOXO1 signaling pathway, and reduction of Gal-9 contributed to dysfunction of dNK via Gal-9/Tim-3 interaction. This study provides new insights for the molecular mechanisms of the adverse pregnancy outcomes caused by T. gondii.

Gene expression profile of immune-check point in response to Trastuzumab therapy in patients with HER-2 positive breast cancer.

OBJECTIVE: Aim: To clarify the association between response to Trastuzumab and molecular expression of TIM-3 and FOXP-3 immune checkpoints. PATIENTS AND METHODS: Materials and Methods: FOXP-3 and TIM-3 expression in peripheral blood was analyzed using qPCR, and the serum level of Trastuzumab was estimated using an immune sorbent enzyme assay. RESULTS: Results: During treatment with Trastuzumab, the FOXP-3 gene expression showed a significant decline throughout one year of treatment, going from 0.85 at cycle 9 to 0.75 at cycle 17. While the TIM-3 gene expression showed a significant up regulation at cycle 9 to 2.8 fold, followed by a reduction in the fold change from 2.8 to 1.7 in the font of reference gene expression. CONCLUSION: Conclusions:FOXP-3 and TIM-3 have the potential to be suggestive markers that can anticipate the response to Trastuzumab, but they are not capable of predicting the likelihood of recurrence.

Single-cell transcriptomic analysis of the immune microenvironment in pediatric acute leukemia.

Relapse and treatment resistance pose significant challenges in the management of pediatric B cell acute lymphoblastic leukemia (B-ALL) and acute myeloid leukemia (AML). The efficacy of immunotherapy in leukemia remains limited due to factors such as the immunosuppressive tumor microenvironment (TME) and lack of suitable immunotherapeutic targets. Thus, an in-depth characterization of the TME in pediatric leukemia is warranted to improve the efficacy of immunotherapy. Here, we used single-cell RNA sequencing (scRNA-seq) to characterize the TME of pediatric B-ALL and AML, focusing specifically on bone-marrow-derived T cells. Moreover, we investigated the transcriptome changes during the initiation, remission, and relapse stages of pediatric AML. Our findings revealed that specific functional expression programs correlated with fluctuations in various T cell subsets, which may be associated with AML progression and relapse. Furthermore, our analysis of cellular communication networks led to the identification of VISTA, CD244, and TIM3 as potential immunotherapeutic targets in pediatric AML. Finally, we detected elevated proportions of γδ T cells and associated functional genes in samples from pediatric patients diagnosed with B-ALL and AML, which could inform the development of novel therapeutic approaches, potentially focusing on γδ T cells.

Simultaneous targeting of Tim3 and A2a receptors modulates MSLN-CAR T cell antitumor function in a human cervical tumor xenograft model.

Introduction: Chimeric antigen receptor (CAR) T cell therapy has transformed the treatment of hematological malignancies. However, its efficacy in solid tumors is limited by the immunosuppressive tumor microenvironment that compromises CAR T cell antitumor function in clinical settings. To overcome this challenge, researchers have investigated the potential of inhibiting specific immune checkpoint receptors, including A2aR (Adenosine A2 Receptor) and Tim3 (T cell immunoglobulin and mucin domain-containing protein 3), to enhance CAR T cell function. In this study, we evaluated the impact of genetic targeting of Tim3 and A2a receptors on the antitumor function of human mesothelin-specific CAR T cells (MSLN-CAR) in vitro and in vivo. Methods: Second-generation anti-mesothelin CAR T cells were produced using standard cellular and molecular techniques. A2aR-knockdown and/or Tim3- knockdown anti-mesothelin-CAR T cells were generated using shRNA-mediated gene silencing. The antitumor function of CAR T cells was evaluated by measuring cytokine production, proliferation, and cytotoxicity in vitro through coculture with cervical cancer cells (HeLa cell line). To evaluate in vivo antitumor efficacy of manufactured CAR T cells, tumor growth and mouse survival were monitored in a human cervical cancer xenograft model. Results: In vitro experiments demonstrated that knockdown of A2aR alone or in combination with Tim3 significantly improved CAR T cell proliferation, cytokine production, and cytotoxicity in presence of tumor cells in an antigen-specific manner. Furthermore, in the humanized xenograft model, both double knockdown CAR T cells and control CAR T cells could effectively control tumor growth. However, single knockdown CAR T cells were associated with reduced survival in mice. Conclusion: These findings highlight the potential of concomitant genetic targeting of Tim3 and A2a receptors to augment the efficacy of CAR T cell therapy in solid tumors. Nevertheless, caution should be exercised in light of our observation of decreased survival in mice treated with single knockdown MSLN-CAR T cells, emphasizing the need for careful efficacy considerations.

Integrated bioinformatic analysis and experimental validation for exploring the key immune checkpoint of COPD.

BACKGROUND: There is growing evidence indicating immune inflammation is a key factor in the progression of chronic obstructive pulmonary disease (COPD). Immune checkpoints (ICs) are crucial targets for modulating the functional activation and differentiation of immune cells, particularly in relation to immune inflammation and the regulation of T cell activation and exhaustion. However, the precise mechanisms of ICs in COPD remain understood. METHODS: COPD datasets were obtained from the Gene Expression Omnibus (GEO) and analyzed using GEO2R and Limma to identify differentially expressed genes. LASSO regression was then applied to screen ICs closely associated with COPD. Finally, target genes were selected based on gene expression profiles. Gene ontology (GO), immune infiltration analysis, and gene set enrichment analysis (GSEA) were utilized to assess the relationship between IC genes (ICGs) and immune cells. Subsequently, tobacco-exposed mice, anti-Tim3-treated mice, and HAVCR2-knockout mice were generated, with flow cytometry being used to confirm the results. RESULTS: Through the analysis of GSE38974 and LASSO regression, five ICGs were identified. Subsequent validation using GSE20257 and GSE76925 confirmed these findings. Gene expression profiling highlighted HAVCR2 as having the strongest correlation with COPD. Further investigation through immune infiltration analysis, GO, and GSEA indicated a link between HAVCR2 and CD8+ T cells in COPD. Flow cytometry experiments demonstrated high Tim3 expression in CD8+ T cells of mice exposed to tobacco, promoting Tc1 and inhibiting Tc17, thus affecting CD8+ Tem activation and CD8+ Tcm formation, leading to an immune imbalance within CD8+ T cells. CONCLUSION: Prolonged exposure to tobacco upregulates Tim3 in CD8+ T cells, triggering its regulatory effects on Tc1/Tc17. Knocking out HAVCR2 further upregulated the expression of CD8+ Tem while suppressing the expression of CD8+ Tcm, indicating that Tim3 plays a role in the activation and differentiation of CD8+ T cells in the context of tobacco exposure.

In Situ Synthesis of an Immune-Checkpoint Blocker from Engineered Bacteria Elicits a Potent Antitumor Response.

Immune-checkpoint blockade (ICB) reinvigorates T cells from exhaustion and potentiates T-cell responses to tumors. However, most patients do not respond to ICB therapy, and only a limited response can be achieved in a "cold" tumor with few infiltrated lymphocytes. Synthetic biology can be used to engineer bacteria as controllable bioreactors to synthesize biotherapeutics in situ. We engineered attenuated Salmonella VNP20009 with synthetic gene circuits to produce PD-1 and Tim-3 scFv to block immunosuppressive receptors on exhausted T cells to reinvigorate their antitumor response. Secreted PD-1 and Tim-3 scFv bound PD-1+ Tim-3+ T cells through their targeting receptors in vitro and potentiated the T-cell secretion of IFN-γ. Engineered bacteria colonized the hypoxic core of the tumor and synthesized PD-1 and Tim-3 scFv in situ, reviving CD4+ T cells and CD8+ T cells to execute an antitumor response. The bacteria also triggered a strong innate immune response, which stimulated the expansion of IFN-γ+ CD4+ T cells within the tumors to induce direct and indirect antitumor immunity.

Association of tuberculosis risk with genetic polymorphisms of the immune checkpoint genes PDCD1, CTLA-4, and TIM3.

The immune checkpoint proteins were reported to involve to host resistance to Mycobacteria tuberculosis (Mtb). Here, we evaluated 11 single nucleotide polymorphisms (SNPs) in PDCD1, CTLA4, and HAVCR2 genes between participants with and without TB infection. Genomic DNA isolated from 285 patients with TB and 270 controls without TB infection were used to perform the genotyping assay. Odds ratios were used to characterize the association of 11 SNPs with TB risk. In this study, the various genotypes of the 11 SNPs did not differ significantly in frequency between the non-TB and TB groups. When patients were stratified by sex, however, men differed significantly from women in genotype frequencies at HAVCR2 rs13170556. Odds ratios indicated that rs2227982, rs13170556, rs231775, and rs231779 were sex-specifically associated with TB risk. In addition, the combinations of rs2227982/rs13170556 GA/TC in men and the A-C-C haplotype of rs231775-rs231777-rs231779 in women were significantly associated with TB risk. Our results indicate that rs2227982 in PDCD1 and rs13170556 in HAVCR2 are associated with increased TB susceptibility in men and that the CTLA4 haplotype appears protective against TB in women.

Ruxolitinib Monotherapy for a Child With HAVCR2 Gene Mutation Associated Subcutaneous Panniculitis-like T-cell Lymphoma: A Case Report.

BACKGROUND: The occurrence of hemophagocytic lymphohistiocytosis (HLH) in patients with subcutaneous panniculitis-like T-cell lymphoma (SPTCL) may be due to HAVCR2 gene mutation, leading to T-cell immunoglobulin and mucin domain-containing molecule 3 deficiency, T-cell and macrophage activation, and proinflammatory cytokine production. OBSERVATION: We report a patient with SPTCL and HLH for whom ruxolitinib, used as a novel treatment, showed notable therapeutic effects. CONCLUSIONS: Remission of both HAVCR2 mutation-induced high inflammatory characteristics and significant symptoms post-ruxolitinib administration suggested that patients with SPTCL and HLH may not represent typical lymphoma cases. Ruxolitinib, with its relatively low toxic side effects, can provide favorable outcomes.

TIM-3, LAG-3, or 2B4 gene disruptions increase the anti-tumor response of engineered T cells.

Background: In adoptive T cell therapy, the long term therapeutic benefits in patients treated with engineered tumor specific T cells are limited by the lack of long term persistence of the infused cellular products and by the immunosuppressive mechanisms active in the tumor microenvironment. Exhausted T cells infiltrating the tumor are characterized by loss of effector functions triggered by multiple inhibitory receptors (IRs). In patients, IR blockade reverts T cell exhaustion but has low selectivity, potentially unleashing autoreactive clones and resulting in clinical autoimmune side effects. Furthermore, loss of long term protective immunity in cell therapy has been ascribed to the effector memory phenotype of the infused cells. Methods: We simultaneously redirected T cell specificity towards the NY-ESO-1 antigen via TCR gene editing (TCRED) and permanently disrupted LAG3, TIM-3 or 2B4 genes (IRKO) via CRISPR/Cas9 in a protocol to expand early differentiated long-living memory stem T cells. The effector functions of the TCRED-IRKO and IR competent (TCRED-IRCOMP) cells were tested in short-term co-culture assays and under a chronic stimulation setting in vitro. Finally, the therapeutic efficacy of the developed cellular products were evaluated in multiple myeloma xenograft models. Results: We show that upon chronic stimulation, TCRED-IRKO cells are superior to TCRED-IRCOMP cells in resisting functional exhaustion through different mechanisms and efficiently eliminate cancer cells upon tumor re-challenge in vivo. Our data indicate that TIM-3 and 2B4-disruption preserve T-cell degranulation capacity, while LAG-3 disruption prevents the upregulation of additional inhibitory receptors in T cells. Conclusion: These results highlight that TIM-3, LAG-3, and 2B4 disruptions increase the therapeutic benefit of tumor specific cellular products and suggest distinct, non-redundant roles for IRs in anti-tumor responses.

Germline HAVCR2/TIM-3 Checkpoint Inhibitor Receptor Deficiency in Recurrent Autoinflammatory Myocarditis.

Myocarditis can be caused by viral infection, drug reaction or general inflammatory condition. To provide understanding on inflammatory myocarditis, we describe clinical, genetic, and immunological properties of a young male patient who suffered from recurrent myocarditis episodes since the age of four years. Electrocardiography, troponin I/T, echocardiography, myocardial magnetic resonance imaging and histological findings were consistent with recurrent myocarditis episodes. Homozygous c.245 A > G p.Tyr82Cys pathogenic variant in Hepatitis A Virus Cellular Receptor 2 (HAVCR2) gene encoding T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3) receptor was found. Peripheral blood mononuclear cells were collected when the patient was asymptomatic; CD4+ and CD8+ T lymphoblasts, CD56+ natural killer cells and CD14+ monocytes were negative for surface TIM-3 expression. In vitro, TLR4 mediated interleukin-1ß (IL-1ß) response was high after LPS/ATP stimulation. Clinical symptoms responded to IL-1 receptor antagonist anakinra. TIM-3 p.Tyr82Cys CD4+ and CD8+ T cell proliferation in vitro was unrestrained. Findings on IL-2, interferon gamma, regulatory T cells, signal transducer and activator of transcription (STAT) 1, 3 and 4 phosphorylation, and PD-1 and LAG-3 checkpoint inhibitor receptor analyses were comparable to controls. We conclude that TIM-3 deficiency due to homozygous HAVCR2 c.245 A > G p.Tyr82Cys pathogenic variant in the patient described here is associated with autoinflammatory symptoms limited to early onset recurrent febrile myocarditis. Excessive IL-1ß production and defective regulation of T cell proliferation may contribute to this clinical condition responsive to anakinra treatment.

Evaluation of changes in exhausted T lymphocytes and miRNAs expression in the different trimesters of pregnancy in pregnant women.

BACKGROUND: Throughout the three trimesters of a typical pregnancy, we looked at changes in the expression of miRNAs and exhausted T lymphocytes for this study. METHODS AND RESULTS: Fifty healthy subjects were included in this study. The frequency of exhausted T lymphocytes was measured in isolated PBMCs using flow cytometry. PD-1, TIM-3, and related miRNAs gene expression were assessed using qRT-PCR. The analyses revealed a significant decline in PD-1 and Tim-3 expression in PBMCs from RPL women (p = 0.0003 and p = 0.001, respectively). In addition, PD-1 and TIM-3 expression increased significantly in the 2nd trimester compared with the 1st trimester of healthy pregnant women (p < 0.0001 and p = 0.0002, respectively). PD-1 and TIM-3 expression was down-regulated in the 3rd trimester compared with the 1st and 2nd trimesters. In the present study, we demonstrated that TIM-3+/CD4+, TIM-3+/CD8+, PD-1+/CD4+, and PD-1+/CD8 + exhausted T lymphocytes increased in the circulation of women in the 2nd trimester compared to the 1st and 3rd trimester. In the 3rd trimester, the expression of miR-16-5p increased significantly (p < 0.0001). miR-125a-3p expression was down and upregulated in 2nd (p < 0.0001) and 3rd (p = 0.0007) trimesters compared to 1st trimester, respectively. This study showed a significant elevation of miR-15a-5p in 3rd trimester compared to 1st trimester of pregnant women (p = 0.0002). CONCLUSIONS: Expression pattern of PD-1 and TIM3 in exhausted T lymphocytes is different not only between normal pregnant and RPL women but also in different trimesters of pregnancy. So, our results showed the role of these markers in the modulation lymphocytes activity in different stages of pregnancy.

Spatial transcriptomic analysis of tumour-immune cell interactions in melanoma arising from congenital melanocytic nevus.

BACKGROUND: Studies on the interaction between tumour-infiltrating immune cells (TIICs) and tumour cells in melanoma arising from congenital melanocytic nevus (CMN) are lacking. OBJECTIVE: The aim of this study was to determine the intratumoral immune landscape of TIICs and tumour cells during invasion and metastasis. METHODS: Tissue specimens were obtained from patients with melanoma originating from CMN. Differential gene expression in melanoma cells and TIICs during invasion and metastasis was determined using spatial transcriptomics. RESULTS: As invasion depth increased, the expression of LGALS3, known to induce tumour-driven immunosuppression, increased in melanoma cells. In T cells, the expression of genes that inhibit T-cell activation increased with increasing invasion depth. In macrophages, the expression of genes related to the anti-inflammatory M2 phenotype was upregulated with increasing invasion depth. Compared to primary tumour cells, melanoma cells in metastatic lesions showed upregulated expression of genes associated with cancer immune evasion, including AXL and EPHA2, which impede T-cell recruitment, and BST2, associated with M2 polarization. Furthermore, T cells showed increased expression of genes related to immunosuppression, and macrophages exhibited increased expression of genes associated with the M2 phenotype. CONCLUSIONS: The interaction between melanomas arising from CMN and TIICs may be important for tumour progression and metastasis.

Post-transplant Inflammatory Bowel Disease Associated with Donor-Derived TIM-3 Deficiency.

Inflammatory bowel disease (IBD) occurring following allogeneic stem cell transplantation (aSCT) is a very rare condition. The underlying pathogenesis needs to be better defined. There is currently no systematic effort to exclude loss- or gain-of-function mutations in immune-related genes in stem cell donors. This is despite the fact that more than 100 inborn errors of immunity may cause or contribute to IBD. We have molecularly characterized a patient who developed fulminant inflammatory bowel disease following aSCT with stable 100% donor-derived hematopoiesis. A pathogenic c.A291G; p.I97M HAVCR2 mutation encoding the immune checkpoint protein TIM-3 was identified in the patient's blood-derived DNA, while being absent in DNA derived from the skin. TIM-3 expression was much decreased in the patient's serum, and in vitro-activated patient-derived T cells expressed reduced TIM-3 levels. In contrast, T cell-intrinsic CD25 expression and production of inflammatory cytokines were preserved. TIM-3 expression was barely detectable in the immune cells of the patient's intestinal mucosa, while being detected unambiguously in the inflamed and non-inflamed colon from unrelated individuals. In conclusion, we report the first case of acquired, "transplanted" insufficiency of the regulatory TIM-3 checkpoint linked to post-aSCT IBD.

Tim-3 deficiency aggravates cadmium nephrotoxicity via regulation of NF-κB signaling and mitochondrial damage.

Kidney is the target organ of serious cadmium injury. Kidney damage caused by cadmium exposure is greatly influenced by the inflammatory response and mitochondrial damage. T cell immunoglobulin domain and mucin domain 3 (Tim-3) is an essential protein that functions as a negative immunological checkpoint to regulate inflammatory responses. Mice were given cadmium treatments at various dosages (0, 1.5, 3, 4.5 mg/kg) and times (0, 3, 5, 7 days) to assess the effects of cadmium on kidney damage. We found that the optimal way to induce kidney injury in mice was to inject 4.5 mg/kg of cadmium intraperitoneally for five days. It is interesting that giving mice 4.5 mg/kg of cadmium intravenously for seven days drastically lowered their survival rate. After cadmium exposure, Tim-3 knockout mice exhibited higher blood concentrations of urea nitrogen and creatinine compared to control mice. Tim-3 impacted the expression of oxidative stress-associated genes such as UDP glucuronosyltransferase family 1 member A9 (Ugt1a9), oxidative stress-induced growth inhibitor 2 (Osgin2), and S100 calcium binding protein A8 (S100a8), according to RNA-seq and real-time RT-PCR data. Tim-3 deficiency also resulted in activated nuclear factor-kappa B (NF-κB) signaling pathway. The NF-κB inhibitor 2-[(aminocarbonyl)amino]-5-(4-fluorophenyl)-3-thiophenecarboxamide (TPCA-1) significantly alleviated cell apoptosis, oxidative stress response, and renal tubule inflammation in Tim-3 knockout mice exposed to cadmium. Furthermore, cadmium caused obvious B-cell lymphoma protein 2 (Bcl-2)-associated X (Bax) translocation from cytoplasm to mitochondria, which can be inhibited by TPCA-1. In conclusion, Tim-3 prevented mitochondrial damage and NF-κB signaling activation, hence providing protection against cadmium nephrotoxicity.

The evaluation of PD-1 and Tim-3 expression besides their related miRNAs in PBMCs of women with recurrent pregnancy loss.

Recurrent pregnancy loss (RPL) is a multifactorial disorder, associated with immunologic abnormalities. During pregnancy, the maternal immune system uses different tolerance mechanisms to deal with a semi-allogenic fetus. The expression of immune checkpoints and their related miRNAs in immune cells can ensure pregnancy at the feto-maternal interface by modulating immune responses. This study aims to evaluate the expression of the immune checkpoint molecules PD-1 and Tim-3 on circulating T cells by flow cytometry, that of mir-138 and mir-155 in PBMCs by Real-time PCR, and the concentrations of TGF-ß and IP-10 in the sera of women suffering from RPL as well as of gestational age-matched healthy pregnant women by ELISA. The percentage of PD-1 or Tim-3 expressing CD8+ T cells was significantly lower in RPL patients compared to the controls, while there was no significant difference in Tim-3 expression of CD4+ T cells between the two groups. The mRNA of both the PD-1 and Tim-3 genes were downregulated in PBMCs of RPL patients compared to controls, however, the difference was not statistically significant for Tim-3. The concentration of TGF-ß was significantly lower and that of IP-10 was significantly higher in the sera of RPL patients than in those of the controls. The relative expression of mir-138 and miR-155 were significantly lower, in PBMCs of RPL patients than in those of healthy pregnant women. These data confirm that by affecting cytokine production, immune checkpoints, and microRNAs play a role in establishing the appropriate local immune environment for successful pregnancy. The wider analysis of immune checkpoints may also yield new biomarkers for the diagnosis and prevention of RPL.

Tim-3 Is Not Required for Establishment of CD8+ T Cell Memory to Lymphocytic Choriomeningitis Virus.

Tim-3 is a transmembrane protein that is best known for being highly expressed on terminally exhausted CD8+ T cells associated with chronic infection and tumors, although its expression is not limited to those settings. Tim-3 is also expressed by CD8+ T cells during acute infection and by multiple other immune cell types, including CD4+ Th1 and regulatory T cells, dendritic cells, and mast cells. In this study, we investigated the role of Tim-3 signaling on CD8+ T cell memory using a Tim-3 conditional knockout mouse model and mice lacking the signaling portion of the Tim-3 cytoplasmic domain. Together, our results indicate that Tim-3 has at most a modest effect on the formation and function of CD8+ memory T cells.

Activated Tim-3/Galectin-9 participated in the development of multiple myeloma by negatively regulating CD4 T cells.

The interaction between Tim-3 on T cells and its ligand Galectin-9 negatively regulates the cellular immune response. However, the regulation of Tim-3/Galectin-9 on CD4 T cell subsets in multiple myeloma (MM) remains unclear. The aim of this study was to investigate the relationship between the regulation of CD4 T cell subsets by the Tim-3/Galectin-9 pathway and clinical prognostic indicators in MM. Tim-3/Galectin-9 were detected by flow cytometry, PCR and ELISA in 60 MM patients and 40 healthy controls, and its correlation with clinical prognostic parameters was analyzed. The expressions of Tim-3 on CD4 T cells, Galectin-9 mRNA in PBMC and level of Galectin-9 protein in serum were significantly elevated in MM patients, especially those with poor prognostic indicators. In MM patients, Tim-3 was highly expressed on the surfaces of Th1, Th2, and Th17 cells, but lowly expressed on Treg. Moreover, level of cytokine IFN-γ in serum was negatively correlated with Tim-3+Th1 cell and Galectin-9mRNA, Galectin-9 protein level. In addition, cell culture experiments showed that the anti-tumor effect and the ability to secrete IFN-γ were restored by blocking the Tim-3/Galectin-9 pathway. In MM patients, Tim-3/Galectin-9 is elevated and associated with disease progression, by inhibiting the cytotoxic function of Th1, and also promoting Th2 and Th17 to be involved in immune escape of MM. Therefore, Tim-3/Galectin-9 may serve as a new immunotherapeutic target for MM patients.