Anh Tran Tram Voh

Intracerebral hemorrhage (ICH) poses acute fatality and long-term neurological risks, in part due to hemin and iron accumulation from hemoglobin breakdown. We observed that hemin induces DNA double-strand breaks (DSBs), prompting a senescence-like phenotype in neurons, necessitating a deeper exploration of cellular responses. Using experimental ICH models and human ICH patient tissue, we elucidate hemin-mediated DNA damage response (DDR) inducing transient senescence and delayed expression of heme oxygenase (HO-1). HO-1 co-localizes with senescence-associated β-Galactosidase (SA-β-Gal) in ICH patient tissues, emphasizing the clinical relevance of inducible HO-1 expression in senescent cells. We reveal a reversible senescence state protective against acute cell death by hemin, while repeat exposure leads to long-lasting senescence. Inhibiting early senescence expression increases cell death, supporting the protective role of senescence against hemin toxicity. Hemin-induced senescence is attenuated by a pleiotropic carbon nanoparticle that is a catalytic mimic of superoxide dismutase, but this treatment increased lipid peroxidation, consistent with ferroptosis from hemin breakdown released iron. When coupled with iron chelator deferoxamine (DEF), the nanoparticle reduces hemin-induced senescence and upregulates factors protecting against ferroptosis. Our study suggests transient senescence induced by DDR as an early potential neuroprotective mechanism in ICH, but the risk of iron-related toxicity supports a multi-pronged therapeutic approach.

Our group has synthesized a pleiotropic synthetic nanozyme redox mediator we term a “pleozyme” that displays multiple enzymatic characteristics, including acting as a superoxide dismutase mimetic, oxidizing NADH to NAD⁺, and oxidizing H₂S to polysulfides and thiosulfate. Benefits have been seen in acute and chronic neurological disease models. The molecule is sourced from coconut-derived activated charcoal that has undergone harsh oxidization with fuming nitric acid, which alters the structure and chemical characteristics, yielding 3–8 nm discs with broad redox potential. Prior work showed pleozymes localize to mitochondria and increase oxidative phosphorylation and glycolysis. Here, we measured cellular NAD⁺ and NADH levels after pleozyme treatment and observed increased total cellular NADH levels but not total NAD⁺ levels. A ¹³C-glucose metabolic flux analysis suggested pleozymes stimulate the generation of pyruvate and lactate glycolytically and from the tricarboxylic acid (TCA) cycle, pointing to malate decarboxylation. Analysis of intracellular fatty acid abundances suggests pleozymes increased fatty acid β-oxidation, with a concomitant increase in succinyl- and acetyl-CoA. Pleozymes increased total ATP, potentially via flexible enhancement of NAD⁺-dependent catabolic pathways such as glycolysis, fatty acid β-oxidation, and metabolic flux through the TCA cycle. These effects may be favorable for pathologies that compromise metabolism such as brain injury.

Harsh acid oxidation of activated charcoal transforms an insoluble carbon-rich source into water-soluble, disc structures of graphene decorated with multiple oxygen-containing functionalities. We term these pleiotropic nano-enzymes as “pleozymes”. A broad redox potential spans many crucial redox reactions including the oxidation of hydrogen sulfide (H₂S) to polysulfides and thiosulfate, dismutation of the superoxide radical (O₂⁻*), and oxidation of NADH to NAD⁺. The oxidation of H₂S is predicted to enhance protein persulfidation—the attachment of sulfur to cysteine residues. Persulfidated proteins act as redox intermediates, and persulfidation protects proteins from irreversible oxidation and ubiquitination, providing an important means of signaling. Protein persulfidation is believed to decline in several neurological disorders and aging. Importantly, and consistent with the role of persulfidation in signaling, the master antioxidant transcription factor Nrf2 is regulated by Keap1’s persulfidation. Here, we demonstrate that pleozymes increased overall protein persulfidation in cells from apparently healthy individuals and from individuals with the mitochondrial protein mutation responsible for Friedreich’s ataxia. We further find that pleozymes specifically enhanced Keap1 persulfidation, with subsequent increased accumulation of Nrf2 and Nrf2’s antioxidant targets.

Intracerebral hemorrhage (ICH) poses acute fatality and long-term neurological risks due to hemin and iron accumulation from hemoglobin breakdown. Our observation that hemin induces DNA double-strand breaks (DSBs), prompting a senescence-like phenotype in neurons, necessitating deeper exploration of cellular responses. Using experimental ICH models and human ICH patient tissue, we elucidate hemin-mediated DNA damage response (DDR) inducing transient senescence and delayed expression of heme oxygenase (HO-1). HO-1 co-localizes with senescence-associated β-Galactosidase (SA-β-Gal) in ICH patient tissues, emphasizing clinical relevance of inducible HO-1 expression in senescent cells. We reveal a reversible senescence state protective against acute cell death by hemin, while repeat exposure leads to long-lasting senescence. Inhibiting early senescence expression increases cell death, supporting the protective role of senescence against hemin toxicity. Hemin-induced senescence is attenuated by a pleiotropic carbon nanoparticle that is a catalytic mimic of superoxide dismutase, but this treatment increased lipid peroxidation, consistent with ferroptosis from hemin breakdown released iron. When coupled with iron chelator deferoxamine (DEF), the nanoparticle reduces hemin-induced senescence and upregulates factors protecting against ferroptosis. Our study suggests transient senescence induced by DDR as an early potential neuroprotective mechanism in ICH, but the risk or iron-related toxicity supports a multi-pronged therapeutic approach.

Inactivation of tumor-infiltrating lymphocytes (TIL) is one of the mechanisms mitigating antitumor immunity during tumor onset and progression. Epigenetic abnormalities are regarded as a major culprit contributing to the dysfunction of TILs within tumor microenvironments. In this study, we used a murine model of melanoma to discover that Tet2 inactivation significantly enhances the antitumor activity of TILs with an efficacy comparable to immune checkpoint inhibition imposed by anti-PD-L1 treatment. Single-cell RNA-sequencing analysis suggested that Tet2-deficient TILs exhibit effector-like features. Transcriptomic and ATAC-sequencing analysis showed that Tet2 ablation reshapes chromatin accessibility and favors binding of transcription factors geared toward CD8+ T-cell activation. Furthermore, the ETS family of transcription factors contributed to augmented CD8+ T-cell function following Tet2 depletion. Overall, our study establishes that Tet2 constitutes one of the epigenetic barriers that account for dysfunction of TILs and that Tet2 inactivation could promote antitumor immunity to suppress tumor growth.Significance:This study suggests that ablation of TET2+ from TILs could promote their antitumor function by reshaping chromatin accessibility for key transcription factors and enhancing the transcription of genes essential for antitumor activity.