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Although inflammation is critical for the clearance of pathogens, uncontrolled inflammation also contributes to the development of multiple diseases such as cancer and sepsis. Since NF-κB–mediated transactivation in the nucleus is pivotal downstream of various stimuli to induce inflammation, searching the nuclear-localized targets specifically regulating NF-κB activation will provide important therapeutic application. Here, we have identified that homeodomain-interacting protein kinase 2 (HIPK2), a nuclear serine/threonine kinase, increases its expression in inflammatory macrophages. Importantly, HIPK2 deficiency or overexpression could enhance or inhibit inflammatory responses in LPS-stimulated macrophages, respectively. HIPK2-deficient mice were more susceptible to LPS-induced endotoxemia and CLP-induced sepsis. Adoptive transfer of Hipk2 +/− bone marrow cells (BMs) also aggravated AOM/DSS-induced colorectal cancer. Mechanistically, HIPK2 bound and phosphorylated histone deacetylase 3 (HDAC3) at serine 374 to inhibit its enzymatic activity, thus reducing the deacetylation of p65 at lysine 218 to suppress NF-κB activation. Notably, the HDAC3 inhibitors protected wild-type or Hipk2−/− BMs-reconstituted mice from LPS-induced endotoxemia. Our findings suggest that the HIPK2-HDAC3-p65 module in macrophages restrains excessive inflammation, which may represent a new layer of therapeutic mechanism for colitis-associated colorectal cancer and sepsis.

Introduction: Renal interstitial fibrosis is a common pathophysiological change in the chronic kidney disease (CKD). Nicotinamide adenine dinucleotide (NAD)-dependent deacetylase sirtuin 6 (SIRT6) is demonstrated to protect against kidney injury. Vitamin B3 is the mostly used form of NAD precursors. However, the role of SIRT6 overexpression in renal interstitial fibrosis of CKD and the association between dietary vitamin B3 intake and renal function remain to be elucidated. Methods: Wild-type (WT) and SIRT6-transgene (SIRT6-Tg) mice were given with high-adenine diets to establish CKD model. HK2 cells were exposed to transforming growth factor β1 (TGF-β1) in vitro to explore related mechanism. Population data from Multi-Ethnic Study of Atherosclerosis (MESA) was used to examine the association between dietary vitamin B3 intake and renal function decline. Results: Compared to WT mice, SIRT6-Tg mice exhibited alleviated renal interstitial fibrosis as evidenced by reduced collagen deposit, collagen I and α-smooth muscle actin expression. Renal function was also improved in SIRT6-Tg mice. Homeodomain interacting protein kinase 2 (HIPK2) was induced during the fibrogenesis in CKD, while HIPK2 was downregulated after SIRT6 overexpression. Further assay in vitro confirmed that SIRT6 depletion exacerbated epithelial-to-mesenchymal transition of HK2 cells, which might be linked with HIPK2 upregulation. HIPK2 was inhibited by SIRT6 in the post-transcriptional level. Population study indicated that higher dietary vitamin B3 intake was independently correlated with a lower risk of estimate glomerular filtration rate decline in those ≥65 years old during follow-up. Conclusion: SIRT6/HIPK2 axis serves as a promising target of renal interstitial fibrosis in CKD. Dietary vitamin B3 intake is beneficial for renal function in the old people.

Background Diabetic foot ulcers (DFU) are a serious complication of diabetes that lead to significant morbidity and mortality. Recent studies reported that exosomes secreted by human adipose tissue-derived mesenchymal stem cells (ADSCs) might alleviate DFU development. However, the molecular mechanism of ADSCs-derived exosomes in DFU is far from being addressed. Methods Human umbilical vein endothelial cells (HUVECs) were induced by high-glucose (HG), which were treated with exosomes derived from nuclear factor I/C (NFIC)-modified ADSCs. MicroRNA-204-3p (miR-204-3p), homeodomain-interacting protein kinase 2 (HIPK2), and NFIC were determined using real-time quantitative polymerase chain reaction. Cell proliferation, apoptosis, migration, and angiogenesis were assessed using cell counting kit-8, 5-ethynyl-2′-deoxyuridine (EdU), flow cytometry, wound healing, and tube formation assays. Binding between miR-204-3p and NFIC or HIPK2 was predicted using bioinformatics tools and validated using a dual-luciferase reporter assay. HIPK2, NFIC, CD81, and CD63 protein levels were measured using western blot. Exosomes were identified by a transmission electron microscope and nanoparticle tracking analysis. Results miR-204-3p and NFIC were reduced, and HIPK2 was enhanced in DFU patients and HG-treated HUVECs. miR-204-3p overexpression might abolish HG-mediated HUVEC proliferation, apoptosis, migration, and angiogenesis in vitro. Furthermore, HIPK2 acted as a target of miR-204-3p. Meanwhile, NFIC was an upstream transcription factor that might bind to the miR-204-3p promoter and improve its expression. NFIC-exosome from ADSCs might regulate HG-triggered HUVEC injury through miR-204-3p-dependent inhibition of HIPK2. Conclusion Exosomal NFIC silencing-loaded ADSC sheet modulates miR-204-3p/HIPK2 axis to suppress HG-induced HUVEC proliferation, migration, and angiogenesis, providing a stem cell-based treatment strategy for DFU.

Glycolytic intermediary metabolites such as fructose-1,6-bisphosphate can serve as signals, controlling metabolic states beyond energy metabolism. However, whether glycolytic metabolites also play a role in controlling cell fate remains unexplored. Here, we find that low levels of glycolytic metabolite 3-phosphoglycerate (3-PGA) can switch phosphoglycerate dehydrogenase (PHGDH) from cataplerosis serine synthesis to pro-apoptotic activation of p53. PHGDH is a p53-binding protein, and when unoccupied by 3-PGA interacts with the scaffold protein AXIN in complex with the kinase HIPK2, both of which are also p53-binding proteins. This leads to the formation of a multivalent p53-binding complex that allows HIPK2 to specifically phosphorylate p53-Ser46 and thereby promote apoptosis. Furthermore, we show that PHGDH mutants (R135W and V261M) that are constitutively bound to 3-PGA abolish p53 activation even under low glucose conditions, while the mutants (T57A and T78A) unable to bind 3-PGA cause constitutive p53 activation and apoptosis in hepatocellular carcinoma (HCC) cells, even in the presence of high glucose. In vivo, PHGDH-T57A induces apoptosis and inhibits the growth of diethylnitrosamine-induced mouse HCC, whereas PHGDH-R135W prevents apoptosis and promotes HCC growth, and knockout of Trp53 abolishes these effects above. Importantly, caloric restriction that lowers whole-body glucose levels can impede HCC growth dependent on PHGDH. Together, these results unveil a mechanism by which glucose availability autonomously controls p53 activity, providing a new paradigm of cell fate control by metabolic substrate availability.

Upon massive DNA damage cells fail to undergo productive DNA repair and trigger the cell death response. Resistance to cell death is linked to cellular transformation and carcinogenesis as well as radio- and chemoresistance, making the underlying signaling pathways a promising target for therapeutic intervention. Diverse DNA damage-induced cell death pathways are operative in mammalian cells and finally culminate in the induction of programmed cell death via activation of apoptosis or necroptosis. These signaling routes affect nuclear, mitochondria- and plasma membrane-associated key molecules to activate the apoptotic or necroptotic response. In this review, we highlight the main signaling pathways, molecular players and mechanisms guiding the DNA damage-induced cell death response.

Serine/arginine-rich splicing factor 3 (SRSF3) likely has wide-ranging roles in gene expression and facilitation of tumor cell growth. SRSF3 knockdown induced G1 arrest and apoptosis in colon cancer cells (HCT116) in association with altered expression of 833 genes. Pathway analysis revealed ‘G1/S Checkpoint Regulation’ as the most highly enriched category in the affected genes. SRSF3 knockdown did not induce p53 or stimulate phosphorylation of p53 or histone H2A.X in wild-type HCT116 cells. Furthermore, the knockdown induced G1 arrest in p53-null HCT116 cells, suggesting that p53-dependent DNA damage responses did not mediate the G1 arrest. Real-time reverse transcription–polymerase chain reaction and western blotting confirmed that SRSF3 knockdown reduced mRNA and protein levels of cyclins (D1, D3 and E1), E2F1 and E2F7. The decreased expression of cyclin D and E2F1 likely impaired the G1-to-S-phase progression. Consequently, retinoblastoma protein remained hypophosphorylated in SRSF3 knockdown cells. The knockdown also induced apoptosis in association with reduction of BCL2 protein levels. We also found that SRSF3 knockdown facilitated skipping of 81 5′-nucleotides (27 amino acids) from exon 8 of homeodomain-interacting protein kinase-2 ( HIPK2 ) and produced a HIPK2 Δe8 isoform. Full-length HIPK2 (HIPK2 FL) is constantly degraded through association with an E3 ubiquitin ligase (Siah-1), whereas HIPK2 Δe8, lacking the 27 amino acids, lost Siah-1-binding ability and became resistant to proteasome digestion. Interestingly, selective knockdown of HIPK2 FL induced apoptosis in various colon cancer cells expressing wild-type or mutated p53. Thus, these findings disclose an important role of SRSF3 in the regulation of the G1-to-S-phase progression and alternative splicing of HIPK2 in tumor growth.

The thymus is responsible for generating a diverse yet self-tolerant pool of T cells 1 . Although the thymic medulla consists mostly of developing and mature AIRE + epithelial cells, recent evidence has suggested that there is far greater heterogeneity among medullary thymic epithelial cells than was previously thought 2 . Here we describe in detail an epithelial subset that is remarkably similar to peripheral tuft cells that are found at mucosal barriers 3 . Similar to the periphery, thymic tuft cells express the canonical taste transduction pathway and IL-25. However, they are unique in their spatial association with cornified aggregates, ability to present antigens and expression of a broad diversity of taste receptors. Some thymic tuft cells pass through an Aire -expressing stage and depend on a known AIRE-binding partner, HIPK2, for their development. Notably, the taste chemosensory protein TRPM5 is required for their thymic function through which they support the development and polarization of thymic invariant natural killer T cells and act to establish a medullary microenvironment that is enriched in the type 2 cytokine, IL-4. These findings indicate that there is a compartmentalized medullary environment in which differentiation of a minor and highly specialized epithelial subset has a non-redundant role in shaping thymic function. A comprehensive analysis of the thymic medulla identifies a tuft-cell-like thymic epithelial cell population that is necessary for shaping thymic function.

The prevalence of chronic kidney disease (CKD) is increasing worldwide, posing a significant healthcare challenge. Despite the immense burden of CKD, optimal therapies remain limited in impact. Kidney fibrosis is a common mediator of all CKD progression, characterized by excessive extracellular matrix deposition and scarring of kidney parenchyma. Transforming growth factor-β (TGF-β) is a potent pro-fibrotic cytokine that signals through canonical and non-canonical pathways to promote kidney cell damage and fibrosis progression, thus garnering much interest as an optimal therapeutic target for CKD. However, the clinical translation of TGF-β inhibition in CKD and other disease settings has faced substantial challenges, particularly due to the highly pleiotropic effects of TGF-β in organ homeostasis and disease. Here, we review the kidney cell-specific biological effects of TGF-β signaling, discuss the current challenges in therapeutic targeting TGF-β in CKD, and provide the rationale for alternative targeting strategies of TGF-β signaling as potential approaches in CKD therapy. Selective inhibition of TGF-β signaling modulators to fine-tune TGF-β inhibition without a broad blockade may lead to new and safer treatments for CKD.

Background Inflammasomes have been found to interact with the gut microbiota, and this effect is associated with depression, but the mechanisms underlying this interaction have not been elucidated in detail. Results The locomotor activity of NLRP3 KO mice was significantly greater than that of their WT littermates, while cohousing and transplantation of the NLRP3 KO gut microbiota avoid the effects of NLRP3 KO on the general locomotor activity at baseline. Meanwhile, transplantation of the NLRP3 KO microbiota alleviated the CUS-induced depressive-like behaviors. The compositions of the gut microbiota in NLRP3 KO mice and WT mice were significantly different in terms of the relative abundance of Firmicutes , Proteobacteria , and Bacteroidetes . Fecal microbiota transplantation (FMT) from NLRP3 KO mice significantly ameliorated the depressive-like behavior induced by chronic unpredictable stress (CUS) in recipient mice. Given the correlation between circular RNA HIPK2 (circHIPK2) and depression and the observation that the level of circHIPK2 expression was significantly increased in CUS-treated mice compared with that in the control group, further experiments were performed. FMT significantly ameliorated astrocyte dysfunction in recipient mice treated with CUS via inhibition of circHIPK2 expression. Conclusions Our study illustrates the involvement of the gut microbiota-circHIPK2-astrocyte axis in depression, providing translational evidence that transplantation of the gut microbiota from NLRP3 KO mice may serve as a novel therapeutic strategy for depression.

Sepsis is a life-threatening syndrome that can result in multi-organ dysfunction. MicroRNA (miR)-483-3p was previously demonstrated to be upregulated in sepsis patients; however, its specific functions in sepsis-triggered intestinal injury remain unclarified. Human intestinal epithelial NCM460 cell line was stimulated with lipopolysaccharide (LPS) to mimic sepsis-induced intestinal injury in vitro. Terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) staining was utilized for examining cell apoptosis. Western blotting and real time quantitative polymerase chain reaction (RT-qPCR) were used for detecting molecular protein and RNA levels. LPS-induced cytotoxicity was determined by measuring concentrations of lactate dehydrogenase (LDH), diamine oxidase (DAO) and fatty acid binding protein 2 (FABP2). Luciferase reporter assay was utilized for verifying the interaction between miR-483-3p and homeodomain interacting protein kinase 2 (HIPK2). Inhibiting miR-483-3p alleviates LPS-triggered NCM460 cell apoptosis and cytotoxicity. miR-483-3p targeted HIPK2 in LPS-stimulated NCM460 cells. Knockdown of HIPK2 reversed the above effects mediated by miR-483-3p inhibitor. Inhibiting miR-483-3p ameliorates LPS-triggered apoptosis and cytotoxicity by targeting HIPK2.