Age-associated sex difference in the expression of mitochondria-based redox sensitive proteins and effect of pioglitazone in nonhuman primate brain.

BACKGROUND: Paraoxonase 2 (PON2) and neuronal uncoupling proteins (UCP4 and UCP5) possess antioxidant, anti-apoptotic activities and minimize accumulation of reactive oxygen species in mitochondria. While age and sex are risk factors for several disorders that are linked with oxidative stress, no study has explored the age- and sex-dependent expression of PON2 isoforms, UCP4 and UCP5 in primate brain or identified a drug to activate UCP4 and UCP5 in vivo. Preclinical studies suggest that the peroxisome proliferator-activated receptor gamma agonist, pioglitazone (PIO), can be neuroprotective, although the mechanism responsible is unclear. Our previous studies demonstrated that pioglitazone activates PON2 in primate brain and we hypothesized that pioglitazone also induces UCP4/5. This study was designed to elucidate the age- and sex-dependent expression of PON2 isoforms, UCP4 and UCP5, in addition to examining the impact of systemic PIO treatment on UCP4 and UCP5 expression in primate brain. METHODS: Western blot technique was used to determine the age- and sex-dependent expression of UCP4 and UCP5 in substantia nigra and striatum of African green monkeys. In addition, we tested the impact of daily oral pioglitazone (5 mg/kg/day) or vehicle for 1 or 3 weeks on expression of UCP4 and UCP5 in substantia nigra and striatum in adult male monkeys. PIO levels in plasma and cerebrospinal fluid (CSF) were determined using LC-MS. RESULTS: We found no sex-based difference in the expression of PON2 isoforms, UCP4 and UCP5 in striatum and substantia nigra of young monkeys. However, we discovered that adult female monkeys exhibit greater expression of PON2 isoforms than males in substantia nigra and striatum. Our data also revealed that adult male monkeys exhibit greater expression of UCP4 and UCP5 than females in substantia nigra but not in striatum. PIO increased UCP4 and UCP5 expression in substantia nigra and striatum at 1 week, but after 3 weeks of treatment this activation had subsided. CONCLUSIONS: Our findings demonstrate a sex-, age- and region-dependent profile to the expression of PON2, UCP4 and UCP5. These data establish a biochemical link between PPARγ, PON2, UCP4 and UCP5 in primate brain and demonstrate that PON2, UCP4 and UCP5 can be pharmacologically stimulated in vivo, revealing a novel mechanism for observed pioglitazone-induced neuroprotection. We anticipate that these outcomes will contribute to the development of novel neuroprotective treatments for Parkinson's disease and other CNS disorders.

Parkinson's disease (PD) is less common in women than men, which may be related to the protective effect of high levels of estrogens in women that maintain the activity of neuroprotective proteins in brain mitochondria. Our previous work suggests that paraoxonase-2 (PON2), uncoupling protein-4 (UCP4) and uncoupling protein-5 (UCP5) play vital roles in maintaining the health of brain dopamine neurons that are lost in PD. This work tested the hypothesis that female primate brains expresses higher levels of these proteins than males. In addition, this research investigated whether estrogen regulates the expression these factors and whether they can be pharmacologically activated later in life to protect dopamine neurons at a time when symptoms of PD typically emerge. The results indicate that before puberty when estrogen levels in females are relatively low, there is no difference in PON2, UCP4, UCP5 brain levels between males and females, but in adults PON2 is up to 3 × higher in females compared with males in regions relevant to PD, consistent with estrogen activation of PON2. Earlier studies have shown that pioglitazone can be neuroprotective in several adverse brain conditions, although the mechanism is not clear. The current research demonstrates that pioglitazone transiently activates by about twofold the expression of PON2, UCP4, UCP5 in vivo in primate brain, suggesting their involvement in the neuroprotective properties of the drug. Overall, the current data provides impetus for further work on activating protective factors that alter mitochondrial dynamics and function, leading to improved understanding and treatment of multiple diseases.

"Essential One Day and Forgotten the Next": Perceptions of Ohio's Early Childhood Workforce on Their Exclusion from the Educator Phase of COVID-19 Vaccine Distribution.

This study explores Ohio Early Childhood and Care (ECEC) workers' perspectives about different prioritization for COVID-19 vaccine distribution between Ohio educators employed in ECEC and prek-12 settings. Days after Ohio's shutdown, ECEC programs began reopening for children of essential workers, and by June 2020 all ECEC programs could reopen with enhanced mitigation strategies, while the prek-12 workforce remained remote as they cautiously returned in-person ranging from 2 to 9 months later. Ohio was 1 of 4 states that, despite contrary Center for Disease Control guidance, excluded ECEC workers from the phase of vaccine distribution in which prek-12 workers were eligible. Data on ECEC employee perceptions of this difference were collected via anonymous online questionnaire from 194 ECEC workers. Qualitative analysis revealed six themes: 1. Participants compared themselves to prek-12, 2. believe they are valuable, 3. disagreed with the decision, 4. felt undervalued compared to prek-12, 5. felt exploited, and 6. suffered mental health effects. ECEC workers' perspectives are valuable and should be included in decisions impacting them.

Pioglitazone transiently stimulates paraoxonase-2 expression in male nonhuman primate brain: Implications for sex-specific therapeutics in neurodegenerative disorders.

Paraoxonase-2 (PON2) enhances mitochondria function and protects against oxidative stress. Stimulating its expression has therapeutic potential for diseases where oxidative stress plays a significant role in the pathology, such as Parkinson's disease. Clinical and preclinical evidence suggest that the anti-diabetic drug pioglitazone may provide neuroprotection in Parkinson's disease, Alzheimer's disease, and stroke, but the biochemical pathway(s) responsible has not been fully elucidated. To determine the effect of pioglitazone on PON2 expression we treated male African green monkeys with oral pioglitazone (5 mg/kg/day) for 1 and 3 weeks. We found that pioglitazone increased PON2 mRNA and protein expression in brain following 1 week of treatment, however, by 3 weeks of treatment PON2 expression had returned to baseline. This transient increase was detected in substantia nigra, striatum, hippocampus, and dorsolateral prefrontal cortex The short-term impact of pioglitazone on PON2 expression in striatum may contribute to the discrepancy in the potency of the drug between short-term animal models and clinical trials for Parkinson's disease. Both PON2 and pioglitazone's receptor, peroxisome proliferator-activated receptor gamma (PPARγ), possess sex- and brain region-dependent expression, which may play a role in the short-term effect of pioglitazone and provide clues to extending the beneficial effects of PON2 activation.

Expression of PON2 isoforms varies among brain regions in male and female African green monkeys.

Mitochondrial dysfunction and oxidative stress contribute to the neuropathology of neurodegenerative disorders such as Parkinson's disease (PD). Paraoxonase-2 (PON2) is a mitochondrial protein that mitigates oxidative stress, enhances mitochondrial function and exhibits anti-inflammatory properties. Previously, we have documented sex-based variation in PON2 with higher brain PON2 expression in female (2-fold) as compared to male African green monkeys. This aim of this study is to identify PON2 isoforms and explore the region-based variations in the protein level of PON2 in brain of African green monkeys. Male and female brain tissue samples (striatum, hippocampus, occipital cortex, dorsolateral prefrontal cortex) from African green monkeys (Chlorocebus sabaeus) were analyzed by western blotting technique for PON2 expression. We found two PON2 isoforms (39 and 41 kDa) in each examined brain region of male and female monkeys. Male monkeys showed no significant difference in the expression level of PON2 isoforms among different brain regions whereas female monkeys showed a significant difference in the expression level of PON2 isoforms in all examined regions except dorsolateral prefrontal cortex. In addition, the result revealed highest expression of PON2 protein in striatum compared to other brain regions in both male and female monkeys. This report is the first to quantify expression of PON2 isoforms in different brain regions and it also establishes the existence of sex as well as region-based variation in PON2 protein expression in primate brain. Since PON2 serves a protective role for dopaminergic neurons it should be considered as a druggable target for oxidative stress-related neurodegenerative disorders like PD. We anticipate that the outcome of this study will contribute to the development of neuroprotective strategies in PD.

Sex-based disparity in paraoxonase-2 expression in the brains of African green monkeys.

The development of several neurodegenerative disorders, such as Parkinson's disease, has been linked with decreased mitochondrial performance, leading to oxidative stress as a result of increased production of reactive oxygen species (ROS). Previous studies have established that the mitochondrial enzyme, paraoxonase-2 (PON2), possesses potent antioxidant and anti-inflammatory properties, with its expression linked with lower ROS levels. The aim of this study was to explore the sex-based variations in the protein level of PON2 in different brain regions (striatum, hippocampus, occipital cortex, and dorsolateral prefrontal cortex) of African green monkeys. Our results revealed that the PON2 expression in females was significantly higher than in males, in each of the examined brain regions. As Parkinson's disease is more prevalent in males compared with females and is characterized by oxidative stress in the nigrostriatal system, these findings add to the growing evidence for PON2 as a target for development of therapeutics to combat this disorder.

PPARγ/PGC1α signaling as a potential therapeutic target for mitochondrial biogenesis in neurodegenerative disorders.

Neurodegenerative diseases represent some of the most devastating neurological disorders, characterized by progressive loss of the structure and function of neurons. Current therapy for neurodegenerative disorders is limited to symptomatic treatment rather than disease modifying interventions, emphasizing the desperate need for improved approaches. Abundant evidence indicates that impaired mitochondrial function plays a crucial role in pathogenesis of many neurodegenerative diseases and so biochemical factors in mitochondria are considered promising targets for pharmacological-based therapies. Peroxisome proliferator-activated receptors-γ (PPARγ) are ligand-inducible transcription factors involved in regulating various genes including peroxisome proliferator-activated receptor gamma co-activator-1 alpha (PGC1α). This review summarizes the evidence supporting the ability of PPARγ-PGC1α to coordinately up-regulate the expression of genes required for mitochondrial biogenesis in neurons and provide directions for future work to explore the potential benefit of targeting mitochondrial biogenesis in neurodegenerative disorders. We have highlighted key roles of NRF2, uncoupling protein-2 (UCP2), and paraoxonase-2 (PON2) signaling in mediating PGC1α-induced mitochondrial biogenesis. In addition, the status of PPARγ modulators being used in clinical trials for Parkinson's disease (PD), Alzheimer's disease (AD) and Huntington's disease (HD) has been compiled. The overall purpose of this review is to update and critique our understanding of the role of PPARγ-PGC1α-NRF2 in the induction of mitochondrial biogenesis together with suggestions for strategies to target PPARγ-PGC1α-NRF2 signaling in order to combat mitochondrial dysfunction in neurodegenerative disorders.

Pioglitazone activates paraoxonase-2 in the brain: A novel neuroprotective mechanism.

Paraoxonase-2 regulates reactive oxygen species production in mitochondria. Stimulating its expression has therapeutic potential for diseases where oxidative stress plays a significant role in the pathology. Evidence suggests that the anti-diabetic drug pioglitazone may provide neuroprotection in Parkinson's disease, Alzheimer's disease, brain trauma and ischemia, but the biochemical pathway(s) responsible has not been fully elucidated. Here we report that pioglitazone (10 mg/kg/day) for 5 days significantly increased paraoxonase-2 expression in mouse striatum. Thus, this result highlights paraoxonase-2 as a target for neuroprotective strategies and identifies pioglitazone as a tool to study the role of paraoxonase-2 in brain.

C-terminal phosphorylation of latrophilin-1/ADGRL1 affects the interaction between its fragments.

Latrophilin-1 is an adhesion G protein-coupled receptor that mediates the effect of α-latrotoxin, causing massive release of neurotransmitters from nerve terminals and endocrine cells. Autoproteolysis cleaves latrophilin-1 into two parts: the extracellular N-terminal fragment (NTF) and the heptahelical C-terminal fragment (CTF). NTF and CTF can exist as independent proteins in the plasma membrane, but α-latrotoxin binding to NTF induces their association and G protein-mediated signaling. We demonstrate here that CTF in synapses is phosphorylated on multiple sites. Phosphorylated CTF has a high affinity for NTF and copurifies with it on affinity columns and sucrose density gradients. Dephosphorylated CTF has a lower affinity for NTF and can behave as a separate protein. α-Latrotoxin (and possibly other ligands of latrophilin-1) binds both to the NTF-CTF complex and receptor-like protein tyrosine phosphatase σ, bringing them together. This leads to CTF dephosphorylation and facilitates CTF release from the complex. We propose that ligand-dependent phosphorylation-dephosphorylation of latrophilin-1 could affect the interaction between its fragments and functions as a G protein-coupled receptor.

Paper-Based Magneto-Resistive Sensor: Modeling, Fabrication, Characterization, and Application.

In this work, we developed and fabricated a paper-based anisotropic magneto-resistive sensor using a sputtered permalloy (Ni 81 Fe 19 ) thin film. To interpret the characteristics of the sensor, we proposed a computational model to capture the influence of the stochastic fiber network of the paper surface and to explain the physics behind the empirically observed difference in paper-based anisotropic magneto-resistance (AMR). Using the model, we verified two main empirical observations: (1) The stochastic fiber network of the paper substrate induces a shift of 45 ∘ in the AMR response of the paper-based Ni 81 Fe 19 thin film compared to a Ni 81 Fe 19 film on a smooth surface as long as the fibrous topography has not become buried. (2) The ratio of magnitudes of AMR peaks at different anisotropy angles and the inverted AMR peak at the 90 ∘ -anisotropy angle are explained through the superposition of the responses of Ni 81 Fe 19 inheriting the fibrous topography and smoother Ni 81 Fe 19 on buried fibrous topographies. As for the sensitivity and reproducibility of the sensor signal, we obtained a maximum AMR peak of 0 . 4 % , min-max sensitivity range of [ 0 . 17 , 0 . 26 ] % , average asymmetry of peak location of 2 . 7 kA m within two consecutive magnetic loading cycles, and a deviation of 250⁻850 A m of peak location across several anisotropy angles at a base resistance of ∼100 Ω . Last, we demonstrated the usability of the sensor in two educational application examples: a textbook clicker and interactive braille flashcards.

Proteolytically released Lasso/teneurin-2 induces axonal attraction by interacting with latrophilin-1 on axonal growth cones.

A presynaptic adhesion G-protein-coupled receptor, latrophilin-1, and a postsynaptic transmembrane protein, Lasso/teneurin-2, are implicated in trans-synaptic interaction that contributes to synapse formation. Surprisingly, during neuronal development, a substantial proportion of Lasso is released into the intercellular space by regulated proteolysis, potentially precluding its function in synaptogenesis. We found that released Lasso binds to cell-surface latrophilin-1 on axonal growth cones. Using microfluidic devices to create stable gradients of soluble Lasso, we show that it induces axonal attraction, without increasing neurite outgrowth. Using latrophilin-1 knockout in mice, we demonstrate that latrophilin-1 is required for this effect. After binding latrophilin-1, Lasso causes downstream signaling, which leads to an increase in cytosolic calcium and enhanced exocytosis, processes that are known to mediate growth cone steering. These findings reveal a novel mechanism of axonal pathfinding, whereby latrophilin-1 and Lasso mediate both short-range interaction that supports synaptogenesis, and long-range signaling that induces axonal attraction.

Expression of functional neuronal receptor latrophilin 1 in human acute myeloid leukaemia cells.

Acute myeloid leukaemia (AML) is a blood cancer affecting cells of myeloid lineage. It is characterised by rapid growth of malignant leukocytes that accumulate in the bone marrow and suppress normal haematopoiesis. This systemic disease remains a serious medical burden worldwide. Characterisation of protein antigens specifically expressed by malignant cells, but not by healthy leukocytes, is vital for the diagnostics and targeted treatment of AML. Here we report, for the first time, that the neuronal receptor latrophilin-1 is expressed in human monocytic leukaemia cell lines and in primary human AML cells. However, it is absent in healthy leukocytes. Latrophilin-1 is functional in leukaemia cells tested, and its biosynthesis is controlled through the mammalian target of rapamycin (mTOR), a master regulator of myeloid cell translational pathways. Our findings demonstrate that latrophilin-1 could be considered as a novel biomarker of human AML, which offers potential new avenues for AML diagnosis and treatment.

Blood ordering from the operating room: turnaround time as a quality indicator.

BACKGROUND: Quality indicators in transfusion medicine are necessary for patient safety and customer satisfaction. The turnaround time (TAT) of issuing red blood cells (RBCs) has emerged as a quality indicator but is not an established benchmark. We examined the TAT for issuing RBCs from the blood bank to the operating rooms (ORs) at Vanderbilt University Medical Center (VUMC) and Stanford University Medical Center (SUMC). STUDY DESIGN AND METHODS: TAT was defined from time of request to when RBCs exited the blood bank. Cases eligible for analysis had completed type-and-screen results with requests for four or fewer RBC units. Patients with a positive antibody screen had serologically crossmatched units prepared and reserved for intraoperative use. We also e-mailed surveys to academic institutions to establish the current state of TAT monitoring and to anesthesiologists at VUMC to gauge the TAT expectations of the OR. RESULTS: The mean TATs at the two institutions were comparable (VUMC, 10 ± 3.8 min; SUMC, 14 ± 7.2 min) for orders of RBCs. The most common reasons for delayed TAT were overlapping orders, medical technologists occupied by phone calls, and oversaturation of pneumatic tube stations. Only 3 of 24 surveyed institutions actively monitored RBC TAT. Surveyed anesthesiologists (n = 7) reported an expectation for RBC TAT of 5 to 15 minutes for urgent cases. Established internal TAT policies were 15 and 20 minutes at VUMC and SUMC, respectively, for crossmatched RBC requests for patients with complete diagnostic testing. CONCLUSION: Many of the surveyed institutions do not monitor stat RBC issue TAT as a quality indicator. This study serves as a starting point for establishing a benchmark for TAT for issuing RBCs from the blood bank to ORs.

Conformational studies of N(3)-substituted [1,3,4]-oxadiazinan-2-ones.

Pseudoephedrine-based [1,3,4]-oxadiazinan-2-ones acylated at the N(3)-position with either acetyl (2a), propionyl (2b), or phenylacetyl (2c) substituents are known to undergo conformational changes that are observable by (13)C NMR spectroscopy. The conformational properties of new [1,3,4]-oxadiazinan-2-one derivatives 2d-k are examined by X-ray crystallography and variable-temperature (13)C NMR spectroscopy and further evaluated by semiempirical AM1 calculations. The collected data reveal that the conformational changes of the overall ring system are dependent upon the stereoelectronic factors of the N(3)-substituent.