Valery V. Khramtsov

Tissue pH and oxygenation are fundamental determinants of cellular metabolism, enzyme activity, and cell survival, and their dysregulation is a hallmark of many pathophysiological processes and diseases including cancer. Concurrent multiparametric mapping of the partial pressure of oxygen (pO2) and pH in a tumor model provides more biologically informative insights. However, such a mapping technique needs to be explored in potentially applicable imaging modalities. This article presents the proof of concept for dynamic concurrent monitoring of pO2 and pH using rapid-scan electron paramagnetic resonance (RS-EPR) and a monophosphonated triarylmethyl radical, HOPE spin probe. A home-built 800-MHz RS-EPR spectrometer/imager was used for four-dimensional spectral-spatial EPR imaging. The time-resolved mapping of pO2 and pH was demonstrated for a human monocyte cell culture system over 7 h. Temporal shifts in pO2 and pH were detected in the three-dimensional parametric data. From these results, it was shown that dynamic concurrent monitoring of pO2 and pH is feasible using RS-EPR and a monophosphonated trityl radical probe.

Ozone (O ₃ )-driven pulmonary inflammation is partly regulated by damage associated molecular patterns (DAMPs) binding to scavenging receptors (SRs). However, how SRs and DAMPs regulate O ₃ -induced pulmonary inflammation remains incompletely understood. CD163 is a SR responsible for clearing cell free hemoglobin (CFH), a DAMP which accumulates during acute pulmonary injury and is associated with worsening respiratory outcomes. We hypothesized that increased CD163 is necessary for reducing CFH levels and resolving O ₃ -induced pulmonary injury. To test this hypothesis, we defined CD163 and CFH responses to O ₃ exposure in C57BL/6N (WT) and CD163 deficient ( Cd163 ^-/- ) mice, as well as in human bronchoalveolar lavage fluid (BALF). In WT mice, lung Cd163 expression was significantly increased by O ₃ during peak inflammation and declined 24 hours post exposure. Human exposure studies revealed a diversity of Cd163 expression and a reduction of CFH following O ₃ exposure, suggesting regulation of this pathway in humans. When compared to WT mice, Cd163 ^-/- mice had augmented O ₃ -induced pulmonary injury, inflammation, and oxidative stress. Further, the antioxidant EUK-134 did not reduce O ₃ -induced pulmonary oxidative stress in Cd163 ^-/- mice, suggesting a role for CD163 in the pulmonary response to oxidative insults. Furthermore, compared to WT controls, Cd163 ^-/- mice receiving an oropharyngeal aspiration of CFH had a significant increase in airspace inflammation. Combined, these findings suggest that CD163 mediated clearance of CFH is involved in resolving O ₃ -induced pulmonary injury, inflammation, and oxidative stress. New & Noteworthy: Ozone (O ₃ ) is known to induce damage associated molecular patterns (DAMPs) which drive lung inflammation. The scavenging receptor, CD163, binds and clears the DAMP cell free hemoglobin (CFH), which accumulates during sterile lung injury. Our findings indicate that O ₃ exposure alters CD163 expression in the lung and that mice lacking Cd163 expression have more lung inflammation. Our data indicate that CD163 serves a protective role in response to acute O ₃ exposure perhaps through CFH clearance.

Precise control of mitochondrial electron transport is essential to maintain mitochondrial coupling and efficiency in ATP production. Furthermore, disruptions to ETC complex function can drive increased oxidant production, resulting in oxidative damage to the mitochondrion and bioenergetic inefficiency. This is highly relevant in the aging heart, as increased cardiac oxidative stress and mitochondrial dysfunction are hallmarks of age-related cardiovascular disease. Lysine acetylation has recently been characterized as a novel regulator of mitochondrial metabolic and bioenergetic function in the aging heart. In the present study, we investigated how lysine acetylation regulates oxidant production and redox milieu through mitochondrial acetyltransferase GCN5L1. Using a cardiac-specific GCN5L1 knockout mouse model, we observed that age-associated lipid peroxidation and semiquinone radicals were decreased with GCN5L1 KO. RNA sequencing analysis identified mitochondrial bioenergetic and respiratory pathways revolving around the respiratory chain to be enriched in the old KO group. Further, we showed the old KO group to exhibit reduced acetylation of ETC complex and antioxidant proteins, improved ETC complex and antioxidant protein activity. Overall, GCN5L1 regulates redox homeostasis in the aged heart by regulating mitochondrial ETC complex activity, oxidative stress, and mitochondrial bioenergetics. These findings identify GCN5L1 and acetylation as potential therapeutic targets in aging and age-related diseases.

Enzyme catalytic activities are critical biomarkers of tissue states under physiological and pathophysiological conditions. However, the direct measurement and imaging of enzyme activity in vivo remains extremely challenging. We report the synthesis and characterization of the first stable triarylmethyl (TAM) radical substrate of alkaline phosphatase (TAM-ALPs). The enzymatic dephosphorylation of TAM-ALPs results in a drastic change in its electron paramagnetic resonance (EPR) spectrum that can be used to image enzyme activity using EPR-based technologies. TAM-ALPs and their enzyme products were fully characterized using EPR and HPLC-MS techniques. A proof of concept of imaging enzyme activity using Overhauser-enhanced magnetic resonance imaging was demonstrated in vitro. This study clearly demonstrates the potential of EPR-based imaging technologies associated with TAM spin probes to map enzyme activity in vivo in future studies.

These data add to the growing body of literature demonstrating that electronic cigarette (Ecig) use during pregnancy (even without nicotine) is not safe, and primes offspring to have worse cardiovascular health outcomes in early and adult life. A key finding from this work is that a second insult from direct vaping in offspring with prior in utero exposure induces greater vascular dysfunction, increased oxidative stress, and shows evidence of neuronal dysfunction compared with either direct- or maternal-only exposure.

The A-site high-entropy perovskite oxide (La_1/6Pr_1/6Nd_1/6Gd_1/6Sr_1/6Ba_1/6)MnO₃ with enhanced hydrogen production, phase stability, and surface oxygen exchange kinetics, offering the potential for tailoring properties in the STCH application.

Recently, aqueous zinc-ion batteries (ZIBs) have become increasingly attractive as grid-scale energy storage solutions due to their safety, low cost, and environmental friendliness. However, severe dendrite growth, self-corrosion, hydrogen evolution, and irreversible side reactions occurring at Zn anodes often cause poor cyclability of ZIBs. This work develops a synergistic strategy to stabilize the Zn anode by introducing a molybdenum dioxide coating layer on Zn (MoO₂@Zn) and Tween 80 as an electrolyte additive. Due to the redox capability and high electrical conductivity of MoO₂, the coating layer can not only homogenize the surface electric field but also accommodate the Zn²⁺ concentration field in the vicinity of the Zn anode, thereby regulating Zn²⁺ ion distribution and inhibiting side reactions. MoO₂ coating can also significantly enhance surface hydrophilicity to improve the wetting of electrolyte on the Zn electrode. Meanwhile, Tween 80, a surfactant additive, acts as a corrosion inhibitor, preventing Zn corrosion and regulating Zn²⁺ ion migration. Their combination can synergistically work to reduce the desolvation energy of hydrated Zn ions and stabilize the Zn anodes. Therefore, the symmetric cells of MoO₂@Zn∥MoO₂@Zn with optimal 1 mM Tween 80 additive in 1 M ZnSO₄ achieve exceptional cyclability over 6000 h at 1 mA cm^-2 and stability (>700 h) even at a high current density (5 mA cm^-2). When coupling with the VO₂ cathode, the full cell of MoO₂@Zn∥VO₂ shows a higher capacity retention (82.4%) compared to Zn∥VO₂ (57.3%) after 1000 cycles at 5 A g^-1. This study suggests a synergistic strategy of combining surface modification and electrolyte engineering to design high-performance ZIBs.

GM-CSF has been employed as an adjuvant to cancer immunotherapy with mixed results based on dosage. We previously showed that GM-CSF regulated tumor angiogenesis by stimulating soluble vascular endothelial growth factor (VEGF) receptor-1 from monocytes/macrophages in a dose-dependent manner that neutralized free VEGF, and intratumoral injections of high-dose GM-CSF ablated blood vessels and worsened hypoxia in orthotopic polyoma middle T Ag (PyMT) triple-negative breast cancer (TNBC). In this study, we assessed both immunoregulatory and oxygen-regulatory components of low-dose versus high-dose GM-CSF to compare effects on tumor oxygen, vasculature, and antitumor immunity. We performed intratumoral injections of low-dose GM-CSF or saline controls for 3 wk in FVB/N PyMT TNBC. Low-dose GM-CSF uniquely reduced tumor hypoxia and normalized tumor vasculature by increasing NG2+ pericyte coverage on CD31+ endothelial cells. Priming of “cold,” anti-PD1–resistant PyMT tumors with low-dose GM-CSF (hypoxia reduced) sensitized tumors to anti-PD1, whereas high-dose GM-CSF (hypoxia exacerbated) did not. Low-dose GM-CSF reduced hypoxic and inflammatory tumor-associated macrophage (TAM) transcriptional profiles; however, no phenotypic modulation of TAMs or tumor-infiltrating lymphocytes were observed by flow cytometry. In contrast, high-dose GM-CSF priming increased infiltration of TAMs lacking the MHC class IIhi phenotype or immunostimulatory marker expression, indicating an immunosuppressive phenotype under hypoxia. However, in anti-PD1 (programmed cell death 1)–susceptible BALB/c 4T1 tumors (considered hot versus PyMT), high-dose GM-CSF increased MHC class IIhi TAMs and immunostimulatory molecules, suggesting disparate effects of high-dose GM-CSF across PyMT versus 4T1 TNBC models. Our data demonstrate a (to our knowledge) novel role for low-dose GM-CSF in reducing tumor hypoxia for synergy with anti-PD1 and highlight why dosage and setting of GM-CSF in cancer immunotherapy regimens require careful consideration.

Purpose Hypoxia and acidosis are recognized tumor microenvironment (TME) biomarkers of cancer progression. Alterations in cancer redox status and metabolism are also associated with elevated levels of intracellular glutathione (GSH) and interstitial inorganic phosphate (Pi). This study aims to evaluate the capability of these biomarkers to discriminate between stages and inform on a switch to malignancy. Procedures These studies were performed using MMTV-PyMT( +) female transgenic mice that spontaneously develop breast cancer and emulate human tumor staging. In vivo assessment of oxygen concentration (pO₂), extracellular acidity (pH_e), Pi, and GSH was performed using L-band electron paramagnetic resonance spectroscopy and multifunctional trityl and GSH-sensitive nitroxide probes. Results Profiling of the TME showed significant deviation of measured biomarkers upon tumor progression from pre-malignancy (pre-S4) to the malignant stage (S4). For the combined marker, HOP: (pH_e × pO₂)/Pi, a value > 186 indicated that the tumors were pre-malignant in 85% of the mammary glands analyzed, and when < 186, they were malignant 42% of the time. For GSH, a value < 3 mM indicated that the tumors were pre-malignant 74% of the time, and when > 3 mM, they were malignant 80% of the time. The only marker that markedly deviated as early as stage 1 (S1) from its value in pre-S1 was elevated Pi, followed by a decrease of pH_e and pO₂ and increase in GSH at later stages. Conclusion Molecular TME profiling informs on alteration of tumor redox and metabolism during tumor staging. Early elevation of interstitial Pi at S1 may reflect tumor metabolic alterations that demand elevated phosphorus supply in accordance with the high rate growth hypothesis. These metabolic changes are supported by the following decrease of pH_e due to a high tumor reliance on glycolysis and increase of intracellular GSH, a major intracellular redox buffer. The appreciable decrease in TME pO₂ was observed only at malignant S4, apparently as a consequence of tumor mass growth and corresponding decrease in perfusion efficacy and increase in oxygen consumption as the tumor cells proliferate.