Dataset Information

Low-Dose Decitabine Augments the Activation and Anti-Tumor Immune Response of IFN-γ⁺ CD4⁺ T Cells Through Enhancing IκBα Degradation and NF-κB Activation.

ABSTRACT:

Background

CD4⁺ T cells play multiple roles in controlling tumor growth and increasing IFN-γ⁺ T-helper 1 cell population could promote cell-mediated anti-tumor immune response. We have previously showed that low-dose DNA demethylating agent decitabine therapy promotes CD3⁺ T-cell proliferation and cytotoxicity; however, direct regulation of purified CD4⁺ T cells and the underlying mechanisms remain unclear.

Methods

The effects of low-dose decitabine on sorted CD4⁺ T cells were detected both in vitro and in vivo. The activation, proliferation, intracellular cytokine production and cytolysis activity of CD4⁺ T cells were analyzed by FACS and DELFIA time-resolved fluorescence assays. In vivo ubiquitination assay was performed to assess protein degradation. Moreover, phosphor-p65 and IκBα levels were detected in sorted CD4⁺ T cells from solid tumor patients with decitabine-based therapy.

Results

Low-dose decitabine treatment promoted the proliferation and activation of sorted CD4⁺ T cells, with increased frequency of IFN-γ⁺ Th1 subset and enhanced cytolytic activity in vitro and in vivo. NF-κB inhibitor, BAY 11-7082, suppressed decitabine-induced CD4⁺ T cell proliferation and IFN-γ production. In terms of mechanism, low-dose decitabine augmented the expression of E3 ligase β-TrCP, promoted the ubiquitination and degradation of IκBα and resulted in NF-κB activation. Notably, we observed that in vitro low-dose decitabine treatment induced NF-κB activation in CD4⁺ T cells from patients with a response to decitabine-primed chemotherapy rather than those without a response.

Conclusion

These data suggest that low-dose decitabine potentiates CD4⁺ T cell anti-tumor immunity through enhancing IκBα degradation and therefore NF-κB activation and IFN-γ production.

SUBMITTER: Li X

PROVIDER: S-EPMC8005576 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:We have previously documented that in liver cells, the multifunctional protein scaffold p62/SQSTM1 is closely associated with IκBα, an inhibitor of the transcriptional activator NF-κB. Such an intimate p62-IκBα association we now document leads to a marked 18-fold proteolytic IκBα-stabilization, enabling its nuclear entry and termination of the NF-κB-activation cycle. In p62-/--cells, such termination is abrogated resulting in the nuclear persistence and prolonged activation of NF-κB following inflammatory stimuli. Utilizing various approaches both classic (structural deletion, site-directed mutagenesis) as well as novel (in-cell chemical crosslinking), coupled with proteomic analyses, we have defined the precise structural hotspots of p62-IκBα association. Accordingly, we have identified such IκBα hotspots to reside around N-terminal (K38, K47, and K67) and C-terminal (K238/C239) residues in its fifth ankyrin repeat domain. These sites interact with two hotspots in p62: One in its PB-1 subdomain around K13, and the other comprised of a positively charged patch (R183/R186/K187/K189) between its ZZ- and TB-subdomains. APEX proximity analyses upon IκBα-cotransfection of cells with and without p62 have enabled the characterization of the p62 influence on IκBα-protein-protein interactions. Interestingly, consistent with p62's capacity to proteolytically stabilize IκBα, its presence greatly impaired IκBα's interactions with various 20S/26S proteasomal subunits. Furthermore, consistent with p62 interaction with IκBα on an interface opposite to that of its NF-κB-interacting interface, p62 failed to significantly affect IκBα-NF-κB interactions. These collective findings together with the known dynamic p62 nucleocytoplasmic shuttling leads us to speculate that it may be involved in "piggy-back" nuclear transport of IκBα following its NF-κB-elicited transcriptional activation and de novo synthesis, required for termination of the NF-κB-activation cycle. Consequently, mice carrying a liver-specific deletion of p62-residues 68 to 252 reveal age-dependent-enhanced liver inflammation. Our findings reveal yet another mode of p62-mediated pathophysiologically relevant regulation of NF-κB.

Project description:Rationale: Inflammation plays a crucial role in the progression of nonalcoholic steatohepatitis (NASH). Protein tyrosine phosphatase receptor type O truncated isoform (PTPROt) is an integral membrane protein that has been identified in osteoclasts, macrophages, and B lymphocytes. However, its relationship between inflammation and NASH is largely unknown. Herein, we aimed to study the function of PTPROt in NASH progression. Methods: We established a NASH mouse model in wild-type (WT), PTPRO knockout mice by western diet (WD) and methionine-choline-deficient diet (MCD). In addition, MCD-induced NASH model was established in BMT mice. Moreover, we determined the expression of PTPROt in liver macrophages in human subjects without steatosis, with simple steatosis, and with NASH to confirm the relationship between PTPROt and NASH. In vitro assays were also performed to study the molecular role of PTPROt in NASH progression. Results: Human samples and animal model results illustrated that PTPROt is increased in liver macrophages during NASH progression and is positively correlated with the degree of NASH. Our animal model also showed that PTPROt in liver macrophages can enhance the activation of the NF-κB signaling pathway, which induces the transcription of genes involved in the inflammatory response. Moreover, PTPROt promotes the transcription of pro-oxidant genes and inhibits antioxidant and protective genes via increased activation of the NF-κB signaling pathway, thereby causing an increased level of reactive oxygen species (ROS) and damaged mitochondria. This triggers the NLRP3-IL1β axis and causes a heightened inflammatory response. Notably, PTPROt partially limits inflammation and ROS production by promoting mitophagy, which participates in a negative feedback loop in this model. Conclusions: Our data strongly indicate that PTPROt plays a dual role in inflammation via the NF-κB signaling pathway in liver macrophages during NASH. Further studies are required to explore therapeutic strategies and prevention of this common liver disease through PTPROt.

Dataset Information

Low-Dose Decitabine Augments the Activation and Anti-Tumor Immune Response of IFN-γ⁺ CD4⁺ T Cells Through Enhancing IκBα Degradation and NF-κB Activation.

Background

Methods

Results

Conclusion

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Dataset Information

Low-Dose Decitabine Augments the Activation and Anti-Tumor Immune Response of IFN-γ+ CD4+ T Cells Through Enhancing IκBα Degradation and NF-κB Activation.

Background

Methods

Results

Conclusion

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Low-Dose Decitabine Augments the Activation and Anti-Tumor Immune Response of IFN-γ⁺ CD4⁺ T Cells Through Enhancing IκBα Degradation and NF-κB Activation.