Interferon Regulatory Factor 4: An Alternative Marker for Plasma Cells in Daratumumab-Treated Patients With Multiple Myeloma.

INTRODUCTION: Anti-CD38 therapeutic modalities (e.g., daratumumab) can impede classical CD38 and CD138 gating use for plasma cell (PC) detection in multiple myeloma (MM) patients with minimal residual disease (MRD). We assessed the applicability of CD229, CD269, and interferon regulatory factor (IRF-4) for PC detection in MM MRD patients. METHODS: Bone marrow samples were collected from patients with MM. Through multiparameter flow cytometry, we evaluated the suitability of CD229, CD269, and IRF-4 for distinguishing PCs from other hematopoietic cells and compared their expression pattern on normal PCs (nPCs) and aberrant PCs (aPCs). We also assessed IRF-4 expression stability after sample storage under different conditions. A 10-color MRD antibody panel was used to determine whether IRF-4 is an alternative primary PC-gating marker for MM MRD assessment. RESULTS: IRF-4 was expressed specifically on all PCs; its mean fluorescence intensity (MFI) was highest on PCs among all hematopoietic cells. This MFI did not decrease even after sample storage at 4°C or 25°C for 72 h. In all 42 MRD assessment samples, except for samples (n = 10) with no PCs, the use of IRF-4 enabled accurate nPC (n = 12), aPC (n = 13), and nPC + aPC (n = 7) identification. Even samples from daratumumab-treated patients had high IRF-4 MFI, with no difference between pre-treatment and post-treatment (n = 7; p = 0.610). CONCLUSIONS: IRF-4 demonstrates high MFI on PCs, and it is not expressed on other leukocytes. In MM patients with MRD, daratumumab treatment does not affect IRF-4 expression. IRF-4 is a promising marker for PC identification in MRD assessment of MM patients undergoing anti-CD38 therapy.

Dehydroepiandrosterone promotes ovarian angiogenesis and improves ovarian function in a rat model of premature ovarian insufficiency by up-regulating HIF-1α/VEGF signalling.

RESEARCH QUESTION: What impact does dehydroepiandrosterone (DHEA) have on ovarian angiogenesis and function in a rat model of with premature ovarian insufficiency (POI), and what are the potential mechanisms of action? DESIGN: DHEA was added to a culture of human microvascular endothelial cells (HMEC-1) to investigate its effects on cell proliferation, migration and tube formation. A rat model of POI was established by intraperitoneal injection of cyclophosphamide, followed by continuous oral administration of DHEA or vehicle for 28 days. Ovarian angiogenesis, follicular growth and granulosa cell survival in ovarian tissues were assessed through haematoxylin and eosin staining, immunohistochemistry and TdT (terminal deoxynucleotidyl transferase)-mediated dUTP nick-end labelling (TUNEL). The effect of DHEA on the fertility of rats with POI was evaluated in pregnant animals. The expression levels of characteristic genes and proteins in the hypoxia-inducible factor (HIF)-1α/vascular endothelial growth factor (VEGF) pathway was determined using quantitative reverse transcription PCR and western blotting. RESULTS: In-vitro experiments revealed that DHEA stimulated the proliferation, migration and tube formation of HMEC-1. In in-vivo studies, DHEA treatment improved the disruption of the oestrous cycle and hormone imbalances in POI rats. Key genes in the HIF-1α/VEGF pathway exhibited up-regulated expression, promoting ovarian angiogenesis in POI rats, and enhancing follicular development and granulosa cell survival, thereby restoring fertility in rats. CONCLUSIONS: DHEA can potentially restore ovarian function in rats with cyclophosphamide-induced POI by up-regulating HIF-1α/VEGF signalling, which promotes the growth of blood vessels in the ovaries.

The impact of thyroid autoimmunity on pregnancy outcomes in women with unexplained infertility undergoing intrauterine insemination: a retrospective single-center cohort study and meta-analysis.

Introduction: Infertility affects 8-12% of couples worldwide, with 15-30% classified as unexplained infertility (UI). Thyroid autoimmunity (TAI), the most common autoimmune disorder in women of reproductive age, may impact fertility and pregnancy outcomes. However, the underlying mechanism is unclear. This study focuses on intrauterine insemination (IUI) and its potential association with TAI in UI patients. It is the first meta-analysis following a comprehensive literature review to improve result accuracy and reliability. Methods: Retrospective cohort study analyzing 225 women with unexplained infertility, encompassing 542 cycles of IUI treatment. Participants were categorized into TAI+ group (N=47, N= 120 cycles) and TAI- group (N=178, N= 422 cycles). Additionally, a systematic review and meta-analyses following PRISMA guidelines were conducted, incorporating this study and two others up to June 2023, totaling 3428 IUI cycles. Results: Analysis revealed no significant difference in independent variables affecting reproductive outcomes. However, comparison based on TAI status showed significantly lower clinical pregnancy rates (OR: 0.43, P= 0.028, 95%CI: 0.20-0.93) and live birth rate (OR: 0.20, P= 0.014, 95%CI: 0.05 ~ 0.71) were significantly lower than TAI- group. There was no significant difference in pregnancy rate between the two groups (OR: 0.61, P= 0.135, 95%CI: 0.32-1.17). However, the meta-analysis combining these findings across studies did not show statistically significant differences in clinical pregnancy rates (OR:0.77, P=0.18, 95%CI: 0.53-1.13) or live birth rates (OR: 0.68, P=0.64, 95%CI: 0.13-3.47) between the TAI+ and TAI- groups. Discussion: Our retrospective cohort study found an association between TAI and reduced reproductive outcomes in women undergoing IUI for unexplained infertility. However, the meta-analysis incorporating other studies did not yield statistically significant associations. Caution is required in interpreting the relationship between thyroid autoimmunity and reproductive outcomes. Future studies should consider a broader population and a more rigorous study design to validate these findings. Clinicians dealing with women with unexplained infertility and TAI should be aware of the complexity of this field and the limitations of available evidence.

Impact of oil-based contrast agents in hysterosalpingography on fertility outcomes in endometriosis: a retrospective cohort study.

BACKGROUND: Previous studies have suggested that oil-based contrast agents used during hysterosalpingography (HSG) in infertile patients can enhance fertility. However, limited research has investigated the effect of oil-based contrast medium specifically in individuals with endometriosis-related infertility. OBJECTIVE: This study aims to explore the impact of oil-based contrast medium on fertility outcomes in women with endometriosis-related infertility. METHODS: Conducted at the First Affiliated Hospital of Guangxi Medical University (January 2020 to June 2022), the study included 512 patients undergoing HSG. Patients were categorized into oil-based and non-oil-based groups, and after propensity score matching, demographic characteristics were compared. Main outcomes included clinical pregnancy rates, live birth rates, early miscarriage rates, and ectopic pregnancy rates. RESULTS: In our analysis, the Oil-based group showed significantly better outcomes compared to the Non-oil-based group. Specifically, the Oil-based group had higher clinical pregnancy rates (51.39% vs. 27.36%) and increased live birth rates (31.48% vs. 19.93%). This trend held true for expectant treatment, IUI, and IVF/ICSI, except for surgical treatment where no significant difference was observed. After adjusting for various factors using propensity score matching, the Non-oil-based group consistently exhibited lower clinical pregnancy rates compared to the Oil-based group. The Odds Ratio (OR) was 0.38 (95%CI: 0.27-0.55) without adjustment, 0.34 (0.22-0.51) in multivariable analysis, 0.39 (0.27-0.57) using inverse probability of treatment weighting (IPTW), and 0.22 (0.14-0.35) in propensity score matching. CONCLUSION: Oil-based contrast medium used in HSG for women with endometriosis-related infertility is associated with higher clinical pregnancy rates and live birth rates compared to Non-oil-based contrast medium.

Association between vitamin D and endometriosis among American women: National Health and Nutrition Examination Survey.

Endometriosis is a multifactorial disease associated with inflammation. Vitamin D has anti-inflammatory, antiproliferative, anti-oxidative, and immunomodulatory effects. Whether vitamin D levels are correlated with endometriosis is a subject of ongoing debate. This study aimed to examine the association between endometriosis and serum vitamin D levels. From the National Health and Nutrition Examination Survey, this study examined the cross-sectional data of American women aged 20-54 years from 2001 to 2006. After adjusting for covariates, multivariable logistic regression analysis was used to assess correlations. A total of 3,232 women were included in this study. The multiple linear regression model demonstrated a negative correlation between the serum 25-hydroxyvitamin D3 (cholecalciferol) concentration and the risk of endometriosis after controlling for all confounding variables. The odds ratio was 0.73 with a 95% confidence interval of 0.54-0.97 in the adequate vitamin D level group compared with the insufficient vitamin D level group. Our results showed that endometriosis was inversely correlated with serum 25-hydroxyvitamin D3 levels. Further research is needed to establish a causal relationship and determine the potential benefits of maintaining sufficient vitamin D levels for endometriosis prevention.

Assessing the impact of transplant site on ovarian tissue transplantation: a single-arm meta-analysis.

BACKGROUND: Survival rates of young women undergoing cancer treatment have substantially improved, with a focus on post-treatment quality of life. Ovarian tissue transplantation (OTT) is a viable option to preserve fertility; however, there is no consensus on the optimal transplantation site. Most studies on OTT are nonrandomized controlled trials with limited sample sizes and uncontrolled statistical analyses, leaving the question of which transplant site yields the highest chance of achieving a live birth unanswered. OBJECTIVE: This meta-analysis aimed to assess the effect of different ovarian transplant sites on postoperative reproductive outcomes. METHODS: We adhered to the PRISMA Reporting Items for Systematic Reviews and Meta-Analyses recommendations. Systematic searches were conducted in PubMed, Embase, Web of Science, and the Cochrane Library from inception to September 17, 2023. The inclusion criteria were as follows: (1) women who underwent OTT with a desire for future childbirth, and (2) reports of specific transplant sites and corresponding pregnancy outcomes. The exclusion criteria included the inability to isolate or extract relevant outcome data, case reports, non-original or duplicate data, and articles not written in English. RESULTS: Twelve studies (201 women) were included in the meta-analysis of cumulative live birth rates (CLBR) after OTT. The CLBR, which encompasses both spontaneous pregnancies and those achieved through assisted reproductive technology (ART) following OTT to the ovarian site, was 21% (95% CI: 6-40, I2: 52.81%, random effect). For transplantation to the pelvic site, the live birth rate was 30% (95% CI: 20-40, I2: 0.00%, fixed effect). Combining transplantation to both the pelvic and ovarian sites resulted in a live birth rate of 23% (95% CI: 11-36, I2: 0.00%, fixed effect). Notably, heterotopic OTT yielded a live birth rate of 3% (95% CI: 0-17, I2: 0.00%, fixed effect). CONCLUSION: Pregnancy outcomes were not significantly different after orthotopic ovarian transplantation, and pregnancy and live birth rates after orthotopic OTT were significantly higher than those after ectopic transplantation. REGISTRATION NUMBER: INPLASY202390008.

Association of APC Expression with Its Promoter Methylation Status and the Prognosis of Hepatocellular Carcinoma.

OBJECTIVE: The present study was aimed to investigate the APC expression, its promoter methylation status, the expression of ß-Catenin, c-Myc and Cyclin D1 and further explore their prognostic value in Hepatocellular carcinoma (HCC). PATIENTS AND METHODS: Serum samples from 90 HCC patients and 27 healthy donors were collected in this study. The methylation-specific PCR (MSP) was performed to evaluate promoter methylation status of APC gene. RT-qPCR was used to detect the mRNA expression of APC, ß-Catenin, c-Myc and Cyclin D1, meanwhile the protein expression were analyzed by Western blot. RESULTS: The positive rate of APC gene methylation in HCC patients (46.67%) was higher than healthy donors (11.11%). APC gene exhibited marked hypermethylation in the patients of TNM III-IV stage when compared to the patients of TNM I-II stage , the methylation status of APC gene was correlated with tumor size and lymph node metastasis whereas the APC gene methylation showed no relationship with the patient's sex and age. APC methylation may be associated with APC expression level, APC expression in HCC cells is silenced by aberrant promoter hypermethylation. In HCC patients with methylated APC, the mRNA and protein expression of ß-Catenin, c-Myc and Cyclin D1 were higher than the unmethylated patient subgroup and healthy donors. CONCLUTIONS: The downregulation of APC in HCC samples was associated with promoter hypermethylation. APC methylation could be used as a novel diagnostic biomarker in HCC, which was associated with regulation of Wnt/ß-Catenin signal pathway.

Melatonin-pretreated human umbilical cord mesenchymal stem cells improved endometrium regeneration and fertility recovery through macrophage immunomodulation in rats with intrauterine adhesions†.

Intrauterine adhesions (IUA) are a common gynecological problem. Stem cell therapy has been widely used in the treatment of IUA. However, due to the complex and harsh microenvironment of the uterine cavity, the effectiveness of such therapy is greatly inhibited. This study aimed to investigate whether melatonin pretreatment enhances the efficacy of human umbilical cord mesenchymal stem cells (HucMSCs) in IUA treatment in rats. First, we explored the effect of melatonin on the biological activity of HucMSCs in vitro through a macrophage co-culture system, Cell Counting Kit 8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), flow cytometry, immunofluorescence staining, and qRT-PCR. Subsequently, we established the IUA rat model and tracked the distribution of HucMSCs in this model. In addition, we observed the number of M1 and M2 macrophages through immunofluorescence staining and detected the levels of inflammatory cytokines. Four weeks after cell transplantation, HE, Masson, and immunohistochemical staining were performed. In vitro experiments showed that melatonin pretreatment of HucMSCs promoted proliferation, reduced apoptosis, up-regulated the stemness gene, and regulated macrophage polarization. In vivo, melatonin pretreatment caused more HucMSCs to remain in the uterine cavity. Melatonin-pretreated HucMSCs recruited more macrophages, regulated macrophage polarization, and reduced inflammation. Melatonin-pretreated HucMSCs relieved fibrosis, increased endometrium thickness, and up-regulated CD34, vimentin, proliferating cell nuclear antigen (PCNA), and alpha small muscle antigen (α-SMA) expression. Fertility tests showed that melatonin-pretreated HucMSCs increased the number of embryos. In summary, pretreatment with melatonin was beneficial for HucMSC treatment because it enhanced the cell's ability to recruit macrophages and regulate macrophage polarization, which led to the regeneration of the endometrium and improved pregnancy outcomes.

Lymphatic endothelial transcription factor Tbx1 promotes an immunosuppressive microenvironment to facilitate post-myocardial infarction repair.

The heart is an autoimmune-prone organ. It is crucial for the heart to keep injury-induced autoimmunity in check to avoid autoimmune-mediated inflammatory disease. However, little is known about how injury-induced autoimmunity is constrained in hearts. Here, we reveal an unknown intramyocardial immunosuppressive program driven by Tbx1, a DiGeorge syndrome disease gene that encodes a T-box transcription factor (TF). We found induced profound lymphangiogenic and immunomodulatory gene expression changes in lymphatic endothelial cells (LECs) after myocardial infarction (MI). The activated LECs penetrated the infarcted area and functioned as intramyocardial immune hubs to increase the numbers of tolerogenic dendritic cells (tDCs) and regulatory T (Treg) cells through the chemokine Ccl21 and integrin Icam1, thereby inhibiting the expansion of autoreactive CD8+ T cells and promoting reparative macrophage expansion to facilitate post-MI repair. Mimicking its timing and implementation may be an additional approach to treating autoimmunity-mediated cardiac diseases.

DNA repair mechanisms that promote insertion-deletion events during immunoglobulin gene diversification.

Insertions and deletions (indels) are low-frequency deleterious genomic DNA alterations. Despite their rarity, indels are common, and insertions leading to long complementarity-determining region 3 (CDR3) are vital for antigen-binding functions in broadly neutralizing and polyreactive antibodies targeting viruses. Because of challenges in detecting indels, the mechanism that generates indels during immunoglobulin diversification processes remains poorly understood. We carried out ultra-deep profiling of indels and systematically dissected the underlying mechanisms using passenger-immunoglobulin mouse models. We found that activation-induced cytidine deaminase-dependent ±1-base pair (bp) indels are the most prevalent indel events, biasing deleterious outcomes, whereas longer in-frame indels, especially insertions that can extend the CDR3 length, are rare outcomes. The ±1-bp indels are channeled by base excision repair, but longer indels require additional DNA-processing factors. Ectopic expression of a DNA exonuclease or perturbation of the balance of DNA polymerases can increase the frequency of longer indels, thus paving the way for models that can generate antibodies with long CDR3. Our study reveals the mechanisms that generate beneficial and deleterious indels during the process of antibody somatic hypermutation and has implications in understanding the detrimental genomic alterations in various conditions, including tumorigenesis.

Loss of Lkb1 in CD11c+ myeloid cells protects mice from diet-induced obesity while enhancing glucose intolerance and IL-17/IFN-γ imbalance.

Adipose tissue CD11c+ myeloid cell is an independent risk factor associated with obesity and metabolic disorders. However, the underlying molecular basis remains elusive. Here, we demonstrated that liver kinase B1 (Lkb1), a key bioenergetic sensor, is involved in CD11c+ cell-mediated immune responses in diet-induced obesity. Loss of Lkb1 in CD11c+ cells results in obesity resistance but lower glucose tolerance, which accompanies tissue-specific immune abnormalities. The accumulation and CD80's expression of Lkb1 deficient adipose-tissue specific dendritic cells but not macrophages is restrained. Additionally, the balance of IL-17A and IFN-γ remarkably tips towards the latter in fat T cells and CD11c- macrophages. Mechanistically, IFN-γ promotes apoptosis of preadipocytes and inhibits their adipogenesis while IL-17A promotes the adipogenesis in vitro, which might account in part for the fat gain resistant phenotype. In summary, these findings reveal that Lkb1 is essential for fat CD11c+ dendritic cells responding to HFD exposure and provides new insights into the IL-17A/IFN-γ balance in HFD-induced obesity.

Systematic comparation of the biological and transcriptomic landscapes of human amniotic mesenchymal stem cells under serum-containing and serum-free conditions.

BACKGROUND: Human amniotic mesenchymal stem cells (hAMSCs) are splendid cell sources for clinical application in the administration of numerous refractory and relapse diseases. Despite the preferable prospect of serum-free (SF) condition for cell product standardization and pathogenic contamination remission, yet the systematic and detailed impact upon hAMSCs at both cellular and transcriptomic levels is largely obscure. METHODS: For the purpose, we preconditioned hAMSCs under serum-containing (SC) and SF medium for 48 h and compared the biological signatures and biofunctions from the view of cell morphology, immunophenotypes, multi-lineage differentiation in vitro, cell vitality, cytokine expression, and immunosuppressive effect upon the subpopulations of T lymphocytes, together with the PI3K-AKT-mTOR signaling reactivation upon cell vitality. Meanwhile, we took advantage of RNA-SEQ and bioinformatic analyses to verify the gene expression profiling and genetic variation spectrum in the indicated hAMSCs. RESULTS: Compared with those maintained in SC medium, hAMSCs pretreated in SF conditions manifested conservation in cell morphology, immunophenotypes, adipogenic differentiation, and immunosuppressive effect upon the proliferation and activation of most of the T cell subpopulations, but with evaluated cytokine expression (e.g., TGF-ß1, IDO1, NOS2) and declined osteogenic differentiation and cell proliferation as well as proapoptotic and apoptotic cells. The declined proliferation in the SF group was efficiently rescued by PI3K-AKT-mTOR signaling reactivation. Notably, hAMSCs cultured in SF and SC conditions revealed similarities in gene expression profiling and variations in genetic mutation at the transcriptome level. Instead, based on the differentially expressed genes and variable shear event analyses, we found those genes were mainly involved in DNA synthesis-, protein metabolism-, and cell vitality-associated biological processes and signaling pathways (e.g., P53, KRAS, PI3K-Akt-mTOR). CONCLUSIONS: Collectively, our data revealed the multifaceted cellular and molecular properties of hAMSCs under SC and SF conditions, which suggested the feasibility of serum-free culture for the preferable preparation of standardized cell products for hAMSC drug development and clinical application.

Diverging regulation of Bach2 protein and RNA expression determine cell fate in early B cell response.

During the early phase of primary humoral responses, activated B cells can differentiate into different types of effector cells, dependent on B cell receptor affinity for antigen. However, the pivotal transcription factors governing these processes remain to be elucidated. Here, we show that transcription factor Bach2 protein in activated B cells is transiently induced by affinity-related signals and mechanistic target of rapamycin complex 1 (mTORC1)-dependent translation to restrain their expansion and differentiation into plasma cells while promoting memory and germinal center (GC) B cell fates. Affinity-related signals also downregulate Bach2 mRNA expression in activated B cells and their descendant memory B cells. Sustained and higher concentrations of Bach2 antagonize the GC fate. Repression of Bach2 in memory B cells predisposes their cell-fate choices upon memory recall. Our study reveals that differential dynamics of Bach2 protein and transcripts in activated B cells control their cell-fate outcomes and imprint the fates of their descendant effector cells.

Bach2 regulates B cell survival to maintain germinal centers and promote B cell memory.

The transcription factor Bach2 serves as a crucial regulator of the germinal center (GC) reaction, which is required for production of high-affinity antibodies and establishment of long-lived B cell memory. However, the stage at which Bach2 controls the GC programs and the precise mechanism underlying these processes remain poorly understood. In this study, we show that genetic ablation of Bach2 in GC B cells of mice impairs their survival and maintenance, and memory B cell formation. These defects can be rescued by enforced expression of anti-apoptotic gene Bcl2. As expected, Bach2-deficient GC B cells are defective in antibody affinity maturation, but have normal somatic hyper mutation and class switch recombination of immunoglobulin genes. Mechanistically, Bach2 controls the GC programs by directly repressing pro-apoptotic gene Bim and a set of genes involved in cell stress response and metabolic processes. Thus, our work reveals the precise roles of Bach2 in the GC biology, and demonstrates that Bach2 acts as a crucial survival regulator of GC B cells, providing a key mechanism underlying GC B maintenance and B cell memory formation.

Bach2 attenuates IL-2R signaling to control Treg homeostasis and Tfr development.

Differentiation and homeostasis of Foxp3+ regulatory T cells (Tregs) are tightly controlled by the interleukin-2 receptor (IL-2R) signaling, yet the mechanisms governing these processes are incompletely understood. Here, we report that transcription factor Bach2 attenuates IL-2R signaling to coordinate Treg differentiation and homeostasis. Bach2 is required for the quiescence, survival, and maintenance of resting Treg cells (rTregs). Unexpectedly, Bach2 directly represses CD25 (IL-2Rα) and subsequently attenuates IL-2R signaling in Tregs. Upregulated CD25/IL-2R signaling in Bach2-deficient rTregs acts as a parallel pathway to partially counteract their poor survival and maintenance. Furthermore, Bach2 suppresses CD25/IL-2R signaling in T follicular regulatory (Tfr) cells. Bach2 deficiency in Tregs prevents the formation of highly differentiated Tfr cells, associated with aberrant GC response. Finally, a mild and late onset of autoimmune disease is observed in mice with Bach2-deficient Tregs. Thus, Bach2 balances IL-2R signaling to orchestrate development and homeostasis of various Treg subsets.

The ubiquitin-specific protease USP8 directly deubiquitinates SQSTM1/p62 to suppress its autophagic activity.

SQSTM1/p62 (sequestosome 1) is a critical macroautophagy/autophagy receptor that promotes the formation and degradation of ubiquitinated aggregates. SQSTM1 can be modified by ubiquitination, and this modification modulates its autophagic activity. However, the molecular mechanisms underpinning its reversible deubiquitination have never been described. Here we report that USP8 (ubiquitin specific peptidase 8) directly interacted with and deubiquitinated SQSTM1. USP8 preferentially removed the lysine 11 (K11)-linked ubiquitin chains from SQSTM1. Moreover, USP8 deubiquitinated SQSTM1 principally at K420 within its ubiquitin-association (UBA) domain. Finally, USP8 inhibited SQSTM1 degradation and autophagic influx in cells with wild-type SQSTM1, but not its mutant with substitution of K420 with an arginine. Taken together, USP8 acts as a negative regulator of autophagy by deubiquitinating SQSTM1 at K420.Abbreviations: BafA1: bafilomycin A1; BAP1: BRCA1 associated protein 1; DUB: deubiquitinating enzyme; ESCRT: endosomal sorting complex required for transport; HTT: huntingtin; K: lysine; KEAP1: kelch like ECH associated protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; shRNA: short hairpin RNA; SQSTM1: sequestosome 1; Ub: ubiquitin; UBA: ubiquitin-association; UBE2D2: ubiquitin conjugating enzyme E2 D2; UBE2D3: ubiquitin conjugating enzyme E2 D3; USP: ubiquitin specific peptidase; WT: wild-type.

Bach2 Deficiency Leads to Spontaneous Expansion of IL-4-Producing T Follicular Helper Cells and Autoimmunity.

The transcription factor Bach2 is a susceptible gene for numerous autoimmune diseases including systemic lupus erythematosus (SLE). Bach2-/- mice can develop a lupus-like autoimmune disease. However, the exact cellular and molecular mechanisms via which Bach2 protects the hosts from developing autoimmunity remains incompletely understood. Here, we report that Bach2 ablation on T cells, but not B cells, resulted in humoral autoimmunity, and this was associated with expansion of T follicular helper (Tfh) cells and abnormal germinal centers. Bach2 was down-regulated in Tfh cells and directly suppressed by the Tfh-defining transcription factor BCL6. Mechanistically, Bach2 directly suppresses the transcription of Cxcr5 and c-Maf, two key regulators of Tfh cell differentiation. Bach2-deficient Tfh cells were skewed toward the IL-4-producing subset, which induced IgG1 and IgE isotype switching of B cells. Heterozygous Bcl6 deficiency reduced the formation of germinal center and autoantibodies, and ameliorated the pathology in Bach2-deficient mice. Our findings identify Bach2 as a crucial negative regulator of Tfh cells at steady state and prove that Bach2 controls autoimmunity in part by restraining accumulation of pathogenic Tfh cells.

Acylglycerol Kinase Maintains Metabolic State and Immune Responses of CD8+ T Cells.

CD8+ T cell expansions and functions rely on glycolysis, but the mechanisms underlying CD8+ T cell glycolytic metabolism remain elusive. Here, we show that acylglycerol kinase (AGK) is required for the establishment and maintenance of CD8+ T cell metabolic and functional fitness. AGK deficiency dampens CD8+ T cell antitumor functions in vivo and perturbs CD8+ T cell proliferation in vitro. Activation of phosphatidylinositol-3-OH kinase (PI3K)-mammalian target of rapamycin (mTOR) signaling, which mediates elevated CD8+ T cell glycolysis, is tightly dependent on AGK kinase activity. Mechanistically, T cell antigen receptor (TCR)- and CD28-stimulated recruitment of PTEN to the plasma membrane facilitates AGK-PTEN interaction and AGK-triggered PTEN phosphorylation, thereby restricting PTEN phosphatase activity in CD8+ T cells. Collectively, these results demonstrate that AGK maintains CD8+ T cell metabolic and functional state by restraining PTEN activity and highlight a critical role for AGK in CD8+ T cell metabolic programming and effector function.

HDAC3 Inhibition Upregulates PD-L1 Expression in B-Cell Lymphomas and Augments the Efficacy of Anti-PD-L1 Therapy.

Programmed cell-death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1) pathway blockade is a promising therapy for the treatment of advanced cancers, including B-cell lymphoma. The clinical response to PD-1/PD-L1 immunotherapy correlates with PD-L1 levels on tumor cells and other cells in the tumor microenvironment. Hence, it is important to understand the molecular mechanisms that regulate PD-L1 expression. Here, we report that histone deacetylase 3 (HDAC3) is a crucial repressor of PD-L1 transcription in B-cell lymphoma. Pan-HDACs or selective HDAC3 inhibitors could rapidly increase histone acetylation and recruitment of bromodomain protein BRD4 at the promoter region of PD-L1 gene, leading to activation of its transcription. Mechanically, HDAC3 and its putative associated corepressor SMRT were recruited to the PD-L1 promoter by the transcriptional repressor BCL6. In addition, HDAC3 inhibition reduced DNA methyltransferase 1 protein levels to indirectly activate PD-L1 transcription. Finally, HDAC3 inhibition increased PD-L1 expression on dendritic cells in the tumor microenvironment. Combining selective HDAC3 inhibitor with anti-PD-L1 immunotherapy enhanced tumor regression in syngeneic murine lymphoma model. Our findings identify HDAC3 as an important epigenetic regulator of PD-L1 expression and implicate combination of HDAC3 inhibition with PD-1/PD-L1 blockade in the treatment of B-cell lymphomas.

BCL6-Mediated Silencing of PD-1 Ligands in Germinal Center B Cells Maintains Follicular T Cell Population.

The programmed cell death protein 1 (PD-1) ligands PD-L1 and PD-L2 on germinal center (GC) B cells deliver coinhibitory signals to follicular T cells. The PD-L1/L2-PD-1 axis modulates the quality and quantity of follicular T cells and has been shown to influence the GC responses. However, the transcriptional control of PD-1 ligands on GC B cells remains largely unknown. In this study, we report that the transcription factor BCL6 is a key negative regulator of the PD-1 ligands PD-L1 and PD-L2 in GC B cells. Acute deletion of Bcl6 in mature GC B cells resulted in marked upregulation of mRNA and protein abundance of PD-1 ligands. Moreover, the expression levels of BCL6 and PD-1 ligands were inversely correlated during GC B cell development and in human GC-derived lymphoma specimens. Mechanically, BCL6 directly bound to the promoter region of PD-L1 and intron 2 of PD-L2 to suppress their transcription. In addition, BCL6 indirectly inhibited the transcription of PD-1 ligands by repressing the expression of STAT1/STAT3 and IRF1. Moreover, BCL6 exerted these effects via its BTB domain. Finally, PD-1 blockade promoted cell survival to sustain the follicular T cell pool in the presence of Bcl6-deficinet GC B cells. In summary, B cell-specific expression of BCL6 dampens the PD-L1/L2-PD-1 signaling to maintain the size of follicular T cells during GC development.