[Cytogenetic and Clinical Characteristics of Newly Diagnosed Elderly Patients with Acute Myeloid Leukemia].

OBJECTIVE: To investigate the cytogenetic characteristics and prognostic risk factors for elderly patients with newly diagnosed elderly acute myeloid leukemia(AML). METHODS: Cytogenetic test results of 76 elderly patients with AML admitted to the First Affiliated Hospital of the University of Science and Technology of China (Anhui Provincial Hospital) from April 2015 to December 2021 were retrospectively analyzed, and analyzed clinical characteristics of patients and risk factors influencing prognosis. RESULTS: According to cytogenetic risk stratification, 76 newly treated elderly AML patients were divided into the favorable, intermediate, and unfavorable groups with 6(7.9%), 58(76.3%), and 12(15.8%) cases, respectively. There was no significant difference in the patient's clinical characteristics and prognosis with the cytogenetics-risk classification groups. Correlation analysis showed that patients' objective response rate (ORR) was related to the age of onset and the mutation status of the CEBPA gene. Logistic regression analysis found that age ≥70 years was an independent risk factor for patients' ORR (OR=0.110, P=0.005). Remission determined the 1-year OS rate (OR=0.049, P=0.005). CONCLUSION: There is no significant difference in clinical characteristics among aged AML patients treated at initial treatment in different cytogenetic risk groups. The age of onset ≥70 years is the determinant of whether patients can obtain ORR, and the rate of ORR is closely related to the 1-year OS rate.

The Composite Immune Risk Score predicts overall survival after allogeneic hematopoietic stem cell transplantation: A retrospective analysis of 1838 cases.

There has been little consensus on how to quantitatively assess immune reconstitution after hematopoietic stem cell transplantation (HSCT) as part of the standard of care. We retrospectively analyzed 11 150 post-transplant immune profiles of 1945 patients who underwent HSCT between 2012 and 2020. 1838 (94.5%) of the cases were allogeneic HSCT. Using the training set of patients (n = 729), we identified a composite immune signature (integrating neutrophil, total lymphocyte, natural killer, total T, CD4+ T, and B cell counts in the peripheral blood) during days 91-180 after allogeneic HSCT that was predictive of early mortality and moreover simplified it into a formula for a Composite Immune Risk Score. When we verified the Composite Immune Risk Score in the validation (n = 284) and test (n = 391) sets of patients, a high score value was found to be associated with hazard ratios (HR) of 3.64 (95% C.I. 1.55-8.51; p = .0014) and 2.44 (95% C.I., 1.22-4.87; p = .0087), respectively, for early mortality. In multivariate analysis, a high Composite Immune Risk Score during days 91-180 remained an independent risk factor for early mortality after allogeneic HSCT (HR, 1.80; 95% C.I., 1.28-2.55; p = .00085). In conclusion, the Composite Immune Risk Score is easy to compute and could identify the high-risk patients of allogeneic HSCT who require targeted effort for prevention and control of infection.

GARP-mediated active TGF-ß1 induces bone marrow NK cell dysfunction in AML patients with early relapse post-allo-HSCT.

Relapse is a leading cause of death after allogeneic hematopoietic stem cell transplantation (allo-HSCT) for acute myeloid leukemia (AML). However, the underlying mechanisms remain poorly understood. Natural killer (NK) cells play a crucial role in tumor surveillance and cancer immunotherapy, and NK cell dysfunction has been observed in various tumors. Here, we performed ex vivo experiments to systematically characterize the mechanisms underlying the dysfunction of bone marrow-derived NK (BMNK) cells isolated from AML patients experiencing early relapse after allo-HSCT. We demonstrated that higher levels of active transforming growth factor ß1 (TGF-ß1) were associated with impaired effector function of BMNK cells in these AML patients. TGF-ß1 activation was induced by the overexpression of glycoprotein A repetitions predominant on the surface of CD4+ T cells. Active TGF-ß1 significantly suppressed mTORC1 activity, mitochondrial oxidative phosphorylation, the proliferation, and cytotoxicity of BMNK cells. Furthermore, pretreatment with the clinical stage TGF-ß1 pathway inhibitor, galunisertib, significantly restored mTORC1 activity, mitochondrial homeostasis, and cytotoxicity. Importantly, the blockade of the TGF-ß1 signaling improved the antitumor activity of NK cells in a leukemia xenograft mouse model. Thus, our findings reveal a mechanism explaining BMNK cell dysfunction and suggest that targeted inhibition of TGF-ß1 signaling may represent a potential therapeutic intervention to improve outcomes in AML patients undergoing allo-HSCT or NK cell-based immunotherapy.

Human-Induced CD49a+ NK Cells Promote Fetal Growth.

CD49a+ natural killer (NK) cells play a critical role in promoting fetal development and maintaining immune tolerance at the maternal-fetal interface during the early stages of pregnancy. However, given their residency in human tissue, thorough studies and clinical applications are difficult to perform. It is still unclear as to how functional human CD49a+ NK cells can be induced to benefit pregnancy outcomes. In this study, we established three no-feeder cell induction systems to induce human CD49a+ NK cells from umbilical cord blood hematopoietic stem cells (HSCs), bone marrow HSCs, and peripheral blood NK cells in vitro. These induced NK cells (iNKs) from three cell induction systems display high levels of CD49a, CD9, CD39, CD151 expression, low levels of CD16 expression, and no obvious cytotoxic capability. They are phenotypically and functionally similar to decidual NK cells. Furthermore, these iNKs display a high expression of growth-promoting factors and proangiogenic factors and can promote fetal growth and improve uterine artery blood flow in a murine pregnancy model in vivo. This research demonstrates the ability of human-induced CD49a+ NK cells to promote fetal growth via three cell induction systems, which could eventually be used to treat patients experiencing adverse pregnancy outcomes.

Dynamic modeling and optimal control of cystic echinococcosis.

BACKGROUND: Cystic echinococcosis is one of the most severe helminth zoonosis with a drastic impact on human health and livestock industry. Investigating optimal control strategy and assessing the crucial factors are essential for developing countermeasures to mitigate this disease. METHODS: Two compartment models were formulated to study the dynamics of cystic echinococcosis transmission, to evaluate the effectiveness of various control measures, and to find the optimal control strategy. Sensitive analyses were conducted by obtaining PRCCs and contour plot was used to evaluate the effect of key parameters on the basic reproduction number. Based on forward-backward sweep method, numerical simulations were employed to investigate effects of key factors on the transmission of cystic echinococcosis and to obtain the optimal control strategy. RESULTS: The food resources of stray dog and invalid sheep vaccination rate, which are always neglected, were significant to the transmission and control of cystic echinococcosis. Numerical simulations suggest that, the implementation of optimal control strategy can significantly reduce the infections. Improving the cost of health education and domestic dog deworming could not decrease human infections. CONCLUSIONS: Our study showed that only a long-term use of the optimal control measures can eliminate the disease. Meanwhile, during the intervention, sheep vaccination and stray dogs disposing should be emphasized ahead of domestic dogs deworming to minimize the control cost. Simultaneously reducing other wild intermediate hosts and strengthening the sheep vaccination as well as disposing the stray dogs would be most effective.

Peroxynitrite-Mediated SIRT (Sirtuin)-1 Inactivation Contributes to Nicotine-Induced Arterial Stiffness in Mice.

Objective- Inhibition of SIRT (sirtuin)-1, a nicotinamide adenine dinucleotide-dependent protein deacetylase, is linked to cigarette smoking-induced arterial stiffness, but the underlying mechanisms remain largely unknown. The aim of the present study was to determine the effects and mechanisms of nicotine, a major component of cigarette smoke, on SIRT1 activity and arterial stiffness. Approach and Results- Arterial stiffness, peroxynitrite (ONOO-) formation, SIRT1 expression and activity were monitored in mouse aortas of 8-week-old C57BL/6 mice (wild-type) or Sirt1-overexpressing ( Sirt1 Super) mice with or without nicotine for 4 weeks. In aortas of wild-type mice, nicotine reduced SIRT1 protein and activity by ≈50% without affecting its mRNA levels. In those from Sirt1 Super mice, nicotine also markedly reduced SIRT1 protein and activity to the levels that were comparable to those in wild-type mice. Nicotine infusion significantly induced collagen I, fibronectin, and arterial stiffness in wild-type but not Sirt1 Super mice. Nicotine increased the levels of iNOS (inducible nitric oxide synthase) and the co-staining of SIRT1 and 3-nitrotyrosine, a footprint of ONOO- in aortas. Tempol, which ablated ONOO- by scavenging superoxide anion, reduced the effects of nicotine on SIRT1 and collagen. Mutation of zinc-binding cysteine 395 or 398 in SIRT1 into serine (C395S) or (C398S) abolished SIRT1 activity. Furthermore, ONOO- dose-dependently inhibited the enzyme and increased zinc release in recombinant SIRT1. Finally, we found SIRT1 inactivation by ONOO- activated the YAP (Yes-associated protein) resulting in abnormal ECM (extracellular matrix) remodeling. Conclusions- Nicotine induces ONOO-, which selectively inhibits SIRT1 resulting in a YAP-mediated ECM remodeling. Visual Overview- An online visual overview is available for this article.

Impact of disposing stray dogs on risk assessment and control of Echinococcosis in Inner Mongolia.

Echinococcosis has been recognized as one of the most important helminth zoonosis in China. Available models always consider dogs as the mainly definitive hosts. However, such models ignore the distinctions between domestic dogs and stray dogs. In this study, we propose a 10-dimensional dynamic model distinguishing stray dogs from domestic dogs to explore the special role of stray dogs and potential effects of disposing stray dogs on the transmission of Echinococcosis. The basic reproduction number R0, which measures the impact of both domestic dogs and stray dogs on the transmission, is determined to characterize the transmission dynamics. Global dynamic analysis of the model reveals that, without disposing the stray dogs, the Echinococcosis becomes endemic even the domestic dogs are controlled. Moreover, due to the difficulties in estimating the parameters involved in R0 with real data and the limitation of R0 in real-world applications, a new risk assessment tool called relative risk index Irisk is defined for the control of zoonotic diseases, and the studies of the risk assessment for Echinococcosis infection show that it is essential to distinguish stray dogs from domestic dogs in applications.

Tryptophan-Derived 3-Hydroxyanthranilic Acid Contributes to Angiotensin II-Induced Abdominal Aortic Aneurysm Formation in Mice In Vivo.

BACKGROUND: Abnormal amino acid metabolism is associated with vascular disease. However, the causative link between dysregulated tryptophan metabolism and abdominal aortic aneurysm (AAA) is unknown. METHODS: Indoleamine 2,3-dioxygenase (IDO) is the first and rate-limiting enzyme in the kynurenine pathway of tryptophan metabolism. Mice with deficiencies in both apolipoprotein e (Apoe) and IDO (Apoe-/-/IDO-/-) were generated by cross-breeding IDO-/- mice with Apoe-/- mice. RESULTS: The acute infusion of angiotensin II markedly increased the incidence of AAA in Apoe-/- mice, but not in Apoe-/-/IDO-/- mice, which presented decreased elastic lamina degradation and aortic expansion. These features were not altered by the reconstitution of bone marrow cells from IDO+/+ mice. Moreover, angiotensin II infusion instigated interferon-γ, which induced the expression of IDO and kynureninase and increased 3-hydroxyanthranilic acid (3-HAA) levels in the plasma and aortas of Apoe-/- mice, but not in IDO-/- mice. Both IDO and kynureninase controlled the production of 3-HAA in vascular smooth muscle cells. 3-HAA upregulated matrix metallopeptidase 2 via transcription factor nuclear factor-κB. Furthermore, kynureninase knockdown in mice restrained 3-HAA, matrix metallopeptidase 2, and resultant AAA formation by angiotensin II infusion. Intraperitoneal injections of 3-HAA into Apoe-/- and Apoe-/-/IDO-/- mice for 6 weeks increased the expression and activity of matrix metallopeptidase 2 in aortas without affecting metabolic parameters. Finally, human AAA samples had stronger staining with the antibodies against 3-HAA, IDO, and kynureninase than those in adjacent nonaneurysmal aortic sections of human AAA samples. CONCLUSIONS: These data define a previously undescribed causative role for 3-HAA, which is a product of tryptophan metabolism, in AAA formation. Furthermore, these findings suggest that 3-HAA reduction may be a new target for treating cardiovascular diseases.

Macrophage Liver Kinase B1 Inhibits Foam Cell Formation and Atherosclerosis.

RATIONALE: LKB1 (liver kinase B1) is a serine/threonine kinase and tumor suppressor, which regulates the homeostasis of hematopoietic cells and immune responses. Macrophages transform into foam cells upon taking-in lipids. No role for LKB1 in foam cell formation has previously been reported. OBJECTIVE: We sought to establish the role of LKB1 in atherosclerotic foam cell formation. METHODS AND RESULTS: LKB1 expression was examined in human carotid atherosclerotic plaques and in western diet-fed atherosclerosis-prone Ldlr-/- and ApoE-/- mice. LKB1 expression was markedly reduced in human plaques when compared with nonatherosclerotic vessels. Consistently, time-dependent reduction of LKB1 levels occurred in atherosclerotic lesions in western diet-fed Ldlr-/- and ApoE-/- mice. Exposure of macrophages to oxidized low-density lipoprotein downregulated LKB1 in vitro. Furthermore, LKB1 deficiency in macrophages significantly increased the expression of SRA (scavenger receptor A), modified low-density lipoprotein uptake and foam cell formation, all of which were abolished by blocking SRA. Further, we found LKB1 phosphorylates SRA resulting in its lysosome degradation. To further investigate the role of macrophage LKB1 in vivo, ApoE-/-LKB1fl/flLysMcre and ApoE-/-LKB1fl/fl mice were fed with western diet for 16 weeks. Compared with ApoE-/-LKB1fl/fl wild-type control, ApoE-/-LKB1fl/flLysMcre mice developed more atherosclerotic lesions in whole aorta and aortic root area, with markedly increased SRA expression in aortic root lesions. CONCLUSIONS: We conclude that macrophage LKB1 reduction caused by oxidized low-density lipoprotein promotes foam cell formation and the progression of atherosclerosis.

A novel role for myeloid cell-specific neuropilin 1 in mitigating sepsis.

Sepsis-typically caused by an uncontrolled and amplified host systemic inflammatory response to microbial infection-is a life-threatening complex clinical disorder and remains a major cause of infection-related deaths in the intensive care unit. Emerging evidence suggests that neuropilin 1 (Nrp1), an originally defined coreceptor for class 3 semaphorins and VEGF, plays important roles in the immune system; however, the function and regulation of macrophage Nrp1 in host immune defense against bacterial infection remain unknown. To address this problem, we generated myeloid cell-specific Nrp1-knockout (Nrp1myel-KO) mice and applied 2 stringent animal models of sepsis: cecal ligation and puncture as well as intraperitoneal injection of LPS. Here, we reported that myeloid cell-specific Nrp1-deficient mice exhibited enhanced susceptibility to cecal ligation and puncture- and LPS-induced sepsis, which correlated with significantly decreased survival rates and heightened levels of proinflammatory cytokines in both peritoneal lavage and serum. Mechanistically, LPS specifically attenuated the expression of Nrp1 in macrophages, which was mediated by TLR4-NF-κB p50 and -65 pathways. By using isolated primary macrophages, loss of Nrp1 consistently resulted in increased production of proinflammatory cytokines, including iNOS, TNF-α, and IL-6. Together, these findings demonstrate a novel role of macrophage Nrp1 in sepsis.-Dai, X. Okon, I., Liu, Z., Wu, Y., Zhu, H., Song, P., Zou, M.-H. A novel role for myeloid cell-specific neuropilin 1 in mitigating sepsis.

AMP-activated protein kinase α1 promotes atherogenesis by increasing monocyte-to-macrophage differentiation.

Monocyte-to-macrophage differentiation, which can be initiated by physiological or atherogenic factors, is a pivotal process in atherogenesis, a disorder in which monocytes adhere to endothelial cells and subsequently migrate into the subendothelial spaces, where they differentiate into macrophages and macrophage-derived foam cells and cause atherosclerotic lesions. However, the monocyte-differentiation signaling pathways that are activated by atherogenic factors are poorly defined. Here we report that the AMP-activated protein kinase α1 (AMPKα1) in monocytes promotes atherosclerosis by increasing monocyte differentiation and survival. Exposure of monocytes to oxidized low-density lipoprotein, 7-ketocholesterol, phorbol 12-myristate 13-acetate, or macrophage colony-stimulated factor (M-CSF) significantly activated AMPK and promoted monocyte-to-macrophage differentiation. M-CSF-activated AMPK is via M-CSF receptor-dependent reactive oxygen species production. Consistently, genetic deletion of AMPKα1 or pharmacological inhibition of AMPK blunted monocyte-to-macrophage differentiation and promoted monocyte/macrophage apoptosis. Compared with apolipoprotein E knock-out (ApoE-/-) mice, which show impaired clearing of plasma lipoproteins and spontaneously develop atherosclerosis, ApoE-/-/AMPKα1-/- mice showed reduced sizes of atherosclerotic lesions and lesser numbers of macrophages in the lesions. Furthermore, aortic lesions were decreased in ApoE-/- mice transplanted with ApoE-/-/AMPKα1-/- bone marrow and in myeloid-specific AMPKα1-deficient ApoE-/- mice. Finally, rapamycin treatment, which abolished delayed monocyte differentiation in ApoE-/-/AMPKα1-/- mice, lost its atherosclerosis-lowering effects in these mice. Mechanistically, we found that AMPKα1 regulates FoxO3-dependent expression of both LC3 and ULK1, which are two important autophagy-related markers. Rapamycin treatment increased FoxO3 activity as well as LC3 and ULK1 expressions in macrophages from AMPKα1-/- mice. Our results reveal that AMPKα1 deficiency impairs autophagy-mediated monocyte differentiation and decreases monocyte/macrophage survival, which attenuates atherosclerosis in ApoE-/- mice in vivo.

AMP-Activated Protein Kinase α1 in Macrophages Promotes Collateral Remodeling and Arteriogenesis in Mice In Vivo.

OBJECTIVE: AMP-activated protein kinase (AMPK), an energy and redox sensor, is activated in response to various cellular stresses, including hypoxia, nutrient deprivation, oxidative stress, and fluid shear stress at the site of vessel blockade. The activation of AMPK is involved in angiogenesis. However, it is unknown whether AMPK can influence arteriogenesis. Here, we demonstrate the contribution of macrophage AMPK to arteriogenesis and collateral remodeling and their underlying mechanisms in well-characterized in vivo and in vitro models. APPROACH AND RESULTS: AMPKα1, AMPKα2 knockout and wild-type littermates underwent femoral artery ligation. Collateral arteriogenesis was monitored in wild-type, global AMPKα1 knockout, or macrophage-specific AMPKα1 knockout mice, with or without hindlimb ligation. Compared with wild-type mice with ligation, global AMPKα1 knockout mice displayed significant reduction in blood flow recovery and impaired remodeling of collateral arterioles. Similar impairments were observed in macrophage-specific AMPK α1 knockout mice after hindlimb ligation. Mechanistically, we found that AMPKα1 promotes the production of growth factors, such as transforming growth factor ß, by directly phosphorylating the inhibitor of nuclear factor κB kinase alpha, resulting in an nuclear factor κB-dependent production of growth factors CONCLUSIONS: Our findings suggest a novel role for macrophage AMPKα1 in arteriogenesis and collateral remodeling and indicate that AMPKα1 activation might be beneficial for recovery from occlusive vascular disorders.

Ablation of Adenosine Monophosphate-Activated Protein Kinase α1 in Vascular Smooth Muscle Cells Promotes Diet-Induced Atherosclerotic Calcification In Vivo.

RATIONALE: Atherosclerotic calcification is highly linked with plaque rapture. How calcification is regulated is poorly characterized. OBJECTIVE: We sought to determine the contributions of AMP-activated protein kinase (AMPK) in atherosclerotic calcification. METHODS AND RESULTS: Aortic calcification was evaluated in aortic roots and brachiocephalic arteries of atherosclerotic prone ApoE(-/-) mice or in mice with dual deficiencies of ApoE and AMPKα isoforms in whole body (ApoE(-/-)/AMPKα1(-/-) and ApoE(-/-)/AMPKα2(-/-)) or vascular smooth muscle cell (VSMC)-specific or macrophage-specific knockout of AMPKα1 fed with Western diet for 24 weeks. Genetic deficiency of AMPKα1 but not of AMPKα2 promoted atherosclerotic calcification and the expression of Runx2 (Runt-related transcription factor). Conversely, chronic administration of metformin, which activated AMPK, markedly reduced atherosclerotic calcification and Runx2 expression in ApoE(-/-) mice but had less effects in ApoE(-/-)/AMPKα1(-/-) mice. Furthermore, VSMC-specific but not macrophage-specific ablation of AMPKα1 promoted aortic calcification in vivo. Ablation of AMPKα1 in VSMC prevented Runx2 from proteasome degradation in parallel with aberrant osteoblastic differentiation of VSMC, whereas AMPK activation promoted Runx2 post-translational modification by small ubiquitin-like modifier (SUMO, SUMOylation), which is associated with its instability. Mechanically, we found that AMPKα1 directly phosphorylated protein inhibitor of activated STAT-1 (PIAS1), the SUMO E3-ligase of Runx2, at serine 510, to promote its SUMO E3-ligase activity. Finally, mutation of protein inhibitor of activated STAT-1 at serine 510 suppressed metformin-induced Runx2 SUMOylation and subsequently prevented metformin's effect on reducing oxidized low-density lipoprotein-triggered Runx2 expression in VSMC. CONCLUSIONS: AMPKα1 phosphorylated protein inhibitor of activated STAT-1 to promote Runx2 SUMOylation and subsequently lead to its instability. AMPKα1 deficiency in VSMC increased Runx2 expression and promoted atherosclerotic calcification in vivo.

Lipopolysaccharides Promote S-Nitrosylation and Proteasomal Degradation of Liver Kinase B1 (LKB1) in Macrophages in Vivo.

LKB1 (liver kinase B1) plays important roles in tumor suppression, energy metabolism, and, recently, in innate immune responses. However, how LKB1 is regulated under physiological or pathological conditions is still unclear. Here, we report that LKB1 protein (but not mRNA) was decreased in both LPS-treated RAW 264.7 cells and peritoneal macrophages isolated from LPS-challenged mice. Additional LPS treatment promoted protein ubiquitination and degradation of LKB1. Pharmacological inhibition or gene silencing of inducible NOS abrogated LPS-induced LKB1 degradation, whereas exposure of RAW 264.7 cells to S-nitroso-l-glutathione, a NO donor, triggered LKB1 S-nitrosylation. Consistently, mutation of one cysteine (C430S) in LKB1 prevented LPS-induced S-nitrosylation, ubiquitination, and degradation. Moreover, S-nitrosylation and ubiquitination of LKB1 were confirmed in macrophages from LPS-challenged mice in vivo. Co-administration of the inducible NOS inhibitor S-methylisothiourea or the proteasome inhibitor MG132 prevented LPS-induced LKB1 degradation and improved the survival rate. Finally, mice lacking LKB1 in macrophages had significantly lower survival rates in response to LPS challenge compared with wild-type mice. Thus, we concluded that LKB1 is degraded by LPS treatment via S-nitrosylation-dependent proteasome pathways, and this had a protective role in LPS-induced septic shock.

Liver kinase B1 suppresses lipopolysaccharide-induced nuclear factor κB (NF-κB) activation in macrophages.

Liver kinase B1 (LKB1), a serine/threonine kinase, is a tumor suppressor and metabolic regulator. Recent data suggest that LKB1 is essential in regulating homeostasis of hematopoietic cells and immune responses. However, its role in macrophages and innate immune system remains unclear. Here we report that macrophage LKB1 inhibits pro-inflammatory signaling in response to LPS. LPS-induced pro-inflammatory cytokines and pro-inflammatory enzymes were monitored in bone marrow-derived macrophages isolated from myeloid cell-specific LKB1 knock out mice and their wild type littermate control mice. LPS induced higher levels of pro-inflammatory cytokines and pro-inflammatory enzymes in bone marrow-derived macrophages from LKB1 KO than those from wild type mice. Consistently, LPS induced higher levels of NF-κB activation in LKB1-deficient macrophages than those in wild type. Further, LPS stimulation significantly increased LKB1 phosphorylation at serine 428, which promoted its binding to IκB kinaseß (IKKß), resulting in the inhibition of NF-κB. Finally, LPS injection caused higher levels of cytokine release and more severe tissue injury in the lung tissues of LKB1 KO mice than in those of control mice. We conclude that LKB1 inhibits LPS-induced NF-κB activation in macrophages.

Myeloperoxidase deletion prevents high-fat diet-induced obesity and insulin resistance.

Activation of myeloperoxidase (MPO), a heme protein primarily expressed in granules of neutrophils, is associated with the development of obesity. However, whether MPO mediates high-fat diet (HFD)-induced obesity and obesity-associated insulin resistance remains to be determined. Here, we found that consumption of an HFD resulted in neutrophil infiltration and enhanced MPO expression and activity in epididymal white adipose tissue, with an increase in body weight gain and impaired insulin signaling. MPO knockout (MPO(-/-)) mice were protected from HFD-enhanced body weight gain and insulin resistance. The MPO inhibitor 4-aminobenzoic acid hydrazide reduced peroxidase activity of neutrophils and prevented HFD-enhanced insulin resistance. MPO deficiency caused high body temperature via upregulation of uncoupling protein-1 and mitochondrial oxygen consumption in brown adipose tissue. Lack of MPO also attenuated HFD-induced macrophage infiltration and expression of proinflammatory cytokines. We conclude that activation of MPO in adipose tissue contributes to the development of obesity and obesity-associated insulin resistance. Inhibition of MPO may be a potential strategy for prevention and treatment of obesity and insulin resistance.

PRKAA1/AMPKα1 is required for autophagy-dependent mitochondrial clearance during erythrocyte maturation.

AMP-activated protein kinase α1 knockout (prkaa1(-/-)) mice manifest splenomegaly and anemia. The underlying molecular mechanisms, however, remain to be established. In this study, we tested the hypothesis that defective autophagy-dependent mitochondrial clearance in prkaa1(-/-) mice exacerbates oxidative stress, thereby enhancing erythrocyte destruction. The levels of ULK1 phosphorylation, autophagical flux, mitochondrial contents, and reactive oxygen species (ROS) were examined in human erythroleukemia cell line, K562 cells, as well as prkaa1(-/-) mouse embryonic fibroblasts and erythrocytes. Deletion of Prkaa1 resulted in the inhibition of ULK1 phosphorylation at Ser555, prevented the formation of ULK1 and BECN1- PtdIns3K complexes, and reduced autophagy capacity. The suppression of autophagy was associated with enhanced damaged mitochondrial accumulation and ROS production. Compared with wild-type (WT) mice, prkaa1(-/-) mice exhibited a shortened erythrocyte life span, hemolytic destruction of erythrocytes, splenomegaly, and anemia, all of which were alleviated by the administration of either rapamycin to activate autophagy or Mito-tempol, a mitochondria-targeted antioxidant, to scavenge mitochondrial ROS. Furthermore, transplantation of WT bone marrow into prkaa1(-/-) mice restored mitochondrial removal, reduced intracellular ROS levels, and normalized hematologic parameters and spleen size. Conversely, transplantation of prkaa1 (-/-) bone marrow into WT mice recapitulated the prkaa1(-/-) mouse phenotypes. We conclude that PRKAA1-dependent autophagy-mediated clearance of damaged mitochondria is required for erythrocyte maturation and homeostasis.

Peroxynitrite-dependent zinc release and inactivation of guanosine 5'-triphosphate cyclohydrolase 1 instigate its ubiquitination in diabetes.

Aberrant degradation of guanosine 5'-triphosphate cyclohydrolase 1 (GTPCH1) with consequent deficiency of tetrahydrobiopterin is considered the primary cause for endothelial dysfunction in diabetes. How GTPCH1 becomes susceptible to the degradation remains unknown. We hypothesized that oxidation and release of the zinc ion by peroxynitrite (ONOO(-)), a potent oxidant generated by nitric oxide and superoxide anions, instigates GTPCH1 ubiquitination and degradation. Zinc contents, GTPCH1 ubiquitination, and GTPCH1 activity were assayed in purified GTPCH1, endothelial cells, and hearts from diabetic mice. Exogenous ONOO(-) dose-dependently released zinc, inhibited its activity, and increased the ubiquitin binding affinity of GTPCH1 in vitro and in endothelial cells. Consistently, high glucose (30 mmol/L) inhibited GTPCH1 activity with increased ubiquitination, which was inhibited by antioxidants. Furthermore, mutation of the zinc-binding cysteine (141) (C141R or C141A) significantly reduced GTPCH1 activity and reduced its half-life but increased GTPCH1 ubiquitination, indicating an essential role of the zinc ion in maintaining the catalytic activity and stability of GTPCH1. Finally, GTPCH1 ubiquitination and degradation markedly increased in parallel with decreased GTPCH1 activity in the aortas and hearts of diabetic mice, both of which were attenuated by the inhibitors of ONOO(-) in mice in vivo. Taken together, we conclude that ONOO(-) releases zinc and inhibits GTPCH1, resulting in its ubiquitination and degradation of the enzyme.