Waste not, want not

• Text
• PDF

Abstract

As of the writing of this Editorial, the current JCI Editorial Board has evaluated approximately 7,000 manuscripts over the past 22 months for their suitability for publication in our journal. While many of you have received a negative decision on your manuscript, I suspect few are aware of the changes we have made to our review process to limit reviewers’ requests for what is in our view unnecessary and excessive experimentation.

Authors

Howard A. Rockman

Bone marrow mesenchymal stem cells and TGF-β signaling in bone remodeling

• Text
• PDF

Abstract

During bone resorption, abundant factors previously buried in the bone matrix are released into the bone marrow microenvironment, which results in recruitment and differentiation of bone marrow mesenchymal stem cells (MSCs) for subsequent bone formation, temporally and spatially coupling bone remodeling. Parathyroid hormone (PTH) orchestrates the signaling of many pathways that direct MSC fate. The spatiotemporal release and activation of matrix TGF-β during osteoclast bone resorption recruits MSCs to bone-resorptive sites. Dysregulation of TGF-β alters MSC fate, uncoupling bone remodeling and causing skeletal disorders. Modulation of TGF-β or PTH signaling may reestablish coupled bone remodeling and be a potential therapy.

Authors

Janet L. Crane, Xu Cao

A conversation with Don Ganem

• Text
• PDF

Abstract

Authors

Ushma S. Neill

From ST segments to endothelial pathophysiology: hypercholesterolemia and endothelial superoxide production

• Text
• PDF

Abstract

As a young medical resident, I encountered a patient suffering from spontaneous coronary vasospasm and was puzzled by these dramatic alterations in vasomotion. This encounter piqued my interest in understanding the drivers of vascular reactivity. In a paper published in the JCI, my colleagues and I revealed a role for superoxide production in the vascular dysfunction associated with hypercholesterolemia. Subsequent work by our group and others has unveiled complex associations between ROS generation and vascular disease.

Authors

David G. Harrison

Osteogenesis imperfecta in adults

• Text
• PDF

Abstract

A 42-year-old premenopausal woman with osteogenesis imperfecta presents to the metabolic bone clinic. She has a daughter with osteogenesis imperfecta who is seen regularly in a specialist pediatric clinic, but the patient herself hasn’t had a clinical consultation in years. She has pain and stiffness in her back and is worried for her future bone health. The patient asks, “Am I going to fall apart?” She had numerous fractures in childhood, including fractures of her femur and wrist; fractured her ankles several times in her late teens; and had occasional fractures in adulthood. Her last fracture was a comminuted fracture of her humerus three years ago, when she stumbled and fell forward onto her hands and knees. The woman is hyperextensible and thinks her ankles feel weak. Her bone mineral density T scores are –2.6 at the lumbar spine and –1.9 at the total hip, and spine imaging shows several vertebral endplate deformities, but overall preservation of vertebral height. What are the available pharmacological and nonpharmacological strategies to preserve her skeletal health and function?

Authors

Nick J. Bishop, Jennifer S. Walsh

At last: classification of human mammary cells elucidates breast cancer origins

• Text
• PDF

Abstract

Current breast cancer classification systems are based on molecular evaluation of tumor receptor status and do not account for distinct morphological phenotypes. In other types of cancer, taxonomy based on normal cell phenotypes has been extremely useful for diagnosis and treatment strategies. In this issue of the JCI, Santagata and colleagues developed a breast cancer classification scheme based on characterization of healthy mammary cells. Reclassification of breast cancer cells and breast cancer tissue microarrays with this system correlated with prognosis better than the standard receptor status designation. This scheme provides a major advance toward our understanding of the origin of the cells in the breast and breast cancers.

Authors

Robert D. Cardiff, Alexander D. Borowsky

An unexpected role for platelets in blocking Th17 differentiation

• Text
• PDF

Abstract

It is well known that platelets interact with cells of the innate immune system to promote tissue repair. In contrast, it is less clear whether these links extend to cells of the adaptive immune system, such as T cells. In this issue of the JCI, Morrell and colleagues provide compelling evidence that platelets are required to limit CD4⁺ Th17 differentiation through the actions of the chemokine platelet factor 4 (PF4). Absence of PF4 in the host leads to exaggerated Th17 differentiation after transplantation and rapid graft rejection. The authors’ findings argue that platelets are not bit part players, but rather fully fledged, critical members of the adaptive immune system.

Authors

Ronjon Chakraverty

Lipids rule: resetting lipid metabolism restores T cell function in systemic lupus erythematosus

• Text
• PDF

Abstract

Systemic lupus erythematosus (SLE) is a devastating autoimmune disease characterized by chronic inflammation and systemic destruction of host organs or tissue. A key feature of SLE is T cell dysfunction characterized by hyperresponsive antigen receptor signaling. In this issue of the JCI, McDonald and colleagues provide evidence that homeostasis of a subset of lipids, the glycosphingolipids (GSLs), is severely perturbed in the membranes of T cells from SLE patients. Furthermore, normalization of GSLs restored TCR signaling and ameliorated T cell dysfunction. These data suggest that targeting host metabolism may be an effective means of reinforcing self-tolerance and attenuating autoimmunity.

Authors

Yoko Kidani, Steven J. Bensinger

Paradoxical insights into whole body metabolic adaptations following SGLT2 inhibition

• Text
• PDF

Abstract

It is well known that glycemic control over time reduces microvascular and macrovascular complications in human subjects with type 2 diabetes. In addition, preclinical models of type 2 diabetes have demonstrated that long-term hyperglycemia exacerbates insulin resistance and reduces β cell function; therefore, therapies that reduce blood glucose levels are of great interest in not only controlling complications, but for restoring known defects in the pathogenesis of type 2 diabetes. Pharmacological inhibition of the sodium-glucose cotransporter 2 (SGLT2) reduces plasma glucose by limiting glucose absorption in the kidney and increasing glucose excretion in the urine. In this issue of the JCI, Merovci and colleagues and Ferrannini and colleagues independently report a paradoxical increase in endogenous glucose production in patients with type 2 diabetes following SGLT2 inhibition, despite an overall decrease in fasting plasma glucose. Together, these studies provide a unique insight into the effects of SGLT2 inhibition on whole body metabolism.

Authors

William T. Cefalu

SMN-targeted therapeutics for spinal muscular atrophy: are we SMArt enough yet?

• Text
• PDF

Abstract

Spinal muscular atrophy (SMA) remains one of the most common and lethal autosomal recessive diseases. Homozygous deletion of survival of motor neuron 1 (SMN1) and resulting SMN protein deficiency manifests predominantly with motor neuron degeneration; however, a wealth of emerging data supports a broader influence of SMN deficiency in disease pathogenesis. In this issue of the JCI, Kariya and colleagues demonstrate the relatively selective impact of SMN depletion on the distal motor unit using a series of SMN2-expressing transgenic mice in which constitutive SMN knockdown follows variable periods of normal development. Their observations provide further insights regarding the temporal requirements for SMN in mice, renewing speculation about when and where repletion of SMN is necessary for optimal outcomes in SMA patients.

Authors

Kathryn J. Swoboda

Evaluation of teriparatide treatment in adults with osteogenesis imperfecta

• Text
• PDF

Abstract

Background. Adults with osteogenesis imperfecta (OI) have a high risk of fracture. Currently, few treatment options are available, and bone anabolic therapies have not been tested in clinical trials for OI treatment.

Methods. 79 adults with OI were randomized to receive 20 μg recombinant human parathyroid hormone (teriparatide) or placebo for 18 months in a double-blind, placebo-controlled trial. The primary endpoint was the percent change in areal bone mineral density (aBMD) of the lumbar spine (LS), as determined by dual-energy X-ray absorptiometry. Secondary endpoints included percent change in bone remodeling markers and vertebral volumetric BMD (vBMD) by quantitative computed tomography, estimated vertebral strength by finite element analysis, and self-reported fractures.

Results. Compared with the placebo group, the teriparatide group showed increased LS aBMD (6.1% ± 1.0% vs. 2.8% ± 1.0% change from baseline; P < 0.05) and total hip aBMD (2.6% ± 1.0% vs. –2.4% ± 1.0% change; P < 0.001). Vertebral vBMD and strength improved with teriparatide therapy (18% ± 6% and 15% ± 3% change, respectively), but declined with placebo (–5.0% ± 6% and –2.0% ± 3% change; P < 0.05 for both comparisons). Serum procollagen type 1 N-terminal propeptide (P1NP) and urine collagen N-telopeptide (NTx) levels increased with teriparatide therapy (135% ± 14% and 64% ± 10% change, respectively). Teriparatide-induced elevation of P1NP levels was less pronounced in severe forms of OI (type III/IV) compared with the milder form (type I). Type I OI patients exhibited robust BMD increases with teriparatide; however, there was no observed benefit for those with type III/IV OI. There was no difference in self-reported fractures between the 2 groups.

Conclusions. Adults with OI, particularly those with less severe disease (type I), displayed a teriparatide-induced anabolic response, as well as increased hip and spine aBMD, vertebral vBMD, and estimated vertebral strength.

Trial registration. Clinicaltrials.gov NCT00131469.

Funding. The Osteoporosis Imperfecta Foundation, Eli Lilly and Co., the National Center for Advancing Translational Science (NCATS) at the NIH (grant no. UL1RR024140), and the Baylor College of Medicine General Clinical Research Center (grant no. RR00188).

Authors

Eric S. Orwoll, Jay Shapiro, Sandra Veith, Ying Wang, Jodi Lapidus, Chaim Vanek, Jan L. Reeder, Tony M. Keaveny, David C. Lee, Mary A. Mullins, Sandesh C.S. Nagamani, Brendan Lee

Metabolic response to sodium-glucose cotransporter 2 inhibition in type 2 diabetic patients

• Text
• PDF

Abstract

Background. Sodium-glucose cotransporter 2 (SGLT2) inhibitors lower glycemia by enhancing urinary glucose excretion. The physiologic response to pharmacologically induced acute or chronic glycosuria has not been investigated in human diabetes.

Methods. We evaluated 66 patients with type 2 diabetes (62 ± 7 years, BMI = 31.6 ± 4.6 kg/m², HbA_1c = 55 ± 8 mmol/mol, mean ± SD) at baseline, after a single dose, and following 4-week treatment with empagliflozin (25 mg). At each time point, patients received a mixed meal coupled with dual-tracer glucose administration and indirect calorimetry.

Results. Both single-dose and chronic empagliflozin treatment caused glycosuria during fasting (median, 7.8 [interquartile range {IQR}, 4.4] g/3 hours and 9.2 [IQR, 5.2] g/3 hours) and after meal ingestion (median, 29.0 [IQR, 12.5] g/5 hours and 28.2 [IQR, 15.4] g/5 hours). After 3 hours of fasting, endogenous glucose production (EGP) was increased 25%, while glycemia was 0.9 ± 0.7 mmol/l lower (P < 0.0001 vs. baseline). After meal ingestion, glucose and insulin AUC decreased, whereas the glucagon response increased (all P < 0.001). While oral glucose appearance was unchanged, EGP was increased (median, 40 [IQR, 14] g and 37 [IQR, 11] g vs. 34 [IQR, 11] g, both P < 0.01). Tissue glucose disposal was reduced (median, 75 [IQR, 16] g and 70 [IQR, 21] g vs. 93 [IQR, 18] g, P < 0.0001), due to a decrease in both glucose oxidation and nonoxidative glucose disposal, with a concomitant rise in lipid oxidation after chronic administration (all P < 0.01). β Cell glucose sensitivity increased (median, 55 [IQR, 35] pmol•min^–1•m^–2•mM^–1 and 55 [IQR, 39] pmol•min^–1•m^–2•mM^–1 vs. 44 [IQR, 32] pmol•min^–1•m^–2•mM^–1, P < 0.0001), and insulin sensitivity was improved. Resting energy expenditure rates and those after meal ingestion were unchanged.

Conclusions. In patients with type 2 diabetes, empagliflozin-induced glycosuria improved β cell function and insulin sensitivity, despite the fall in insulin secretion and tissue glucose disposal and the rise in EGP after one dose, thereby lowering fasting and postprandial glycemia. Chronic dosing shifted substrate utilization from carbohydrate to lipid.

Trial registration. ClinicalTrials.Gov NCT01248364 (EudraCT no. 2010-018708-99).

Funding. This study was funded by Boehringer Ingelheim.

Authors

Ele Ferrannini, Elza Muscelli, Silvia Frascerra, Simona Baldi, Andrea Mari, Tim Heise, Uli C. Broedl, Hans-Juergen Woerle

Dapagliflozin improves muscle insulin sensitivity but enhances endogenous glucose production

• Text
• PDF

Abstract

Chronic hyperglycemia impairs insulin action, resulting in glucotoxicity, which can be ameliorated in animal models by inducing glucosuria with renal glucose transport inhibitors. Here, we examined whether reduction of plasma glucose with a sodium-glucose cotransporter 2 (SGLT2) inhibitor could improve insulin-mediated tissue glucose disposal in patients with type 2 diabetes. Eighteen diabetic men were randomized to receive either dapagliflozin (n = 12) or placebo (n = 6) for 2 weeks. We measured insulin-mediated whole body glucose uptake and endogenous glucose production (EGP) at baseline and 2 weeks after treatment using the euglycemic hyperinsulinemic clamp technique. Dapagliflozin treatment induced glucosuria and markedly lowered fasting plasma glucose. Insulin-mediated tissue glucose disposal increased by approximately 18% after 2 weeks of dapagliflozin treatment, while placebo-treated subjects had no change in insulin sensitivity. Surprisingly, following dapagliflozin treatment, EGP increased substantially and was accompanied by an increase in fasting plasma glucagon concentration. Together, our data indicate that reduction of plasma glucose with an agent that works specifically on the kidney to induce glucosuria improves muscle insulin sensitivity. However, glucosuria induction following SGLT2 inhibition is associated with a paradoxical increase in EGP. These results provide support for the glucotoxicity hypothesis, which suggests that chronic hyperglycemia impairs insulin action in individuals with type 2 diabetes.

Authors

Aurora Merovci, Carolina Solis-Herrera, Giuseppe Daniele, Roy Eldor, Teresa Vanessa Fiorentino, Devjit Tripathy, Juan Xiong, Zandra Perez, Luke Norton, Muhammad A. Abdul-Ghani, Ralph A. DeFronzo

Interplay between FGF21 and insulin action in the liver regulates metabolism

• Text
• PDF

Abstract

The hormone FGF21 regulates carbohydrate and lipid homeostasis as well as body weight, and increasing FGF21 improves metabolic abnormalities associated with obesity and diabetes. FGF21 is thought to act on its target tissues, including liver and adipose tissue, to improve insulin sensitivity and reduce adiposity. Here, we used mice with selective hepatic inactivation of the IR (LIRKO) to determine whether insulin sensitization in liver mediates FGF21 metabolic actions. Remarkably, hyperglycemia was completely normalized following FGF21 treatment in LIRKO mice, even though FGF21 did not reduce gluconeogenesis in these animals. Improvements in blood sugar were due in part to increased glucose uptake in brown fat, browning of white fat, and overall increased energy expenditure. These effects were preserved even after removal of the main interscapular brown fat pad. In contrast to its retained effects on reducing glucose levels, the effects of FGF21 on reducing circulating cholesterol and hepatic triglycerides and regulating the expression of key genes involved in cholesterol and lipid metabolism in liver were disrupted in LIRKO mice. Thus, FGF21 corrects hyperglycemia in diabetic mice independently of insulin action in the liver by increasing energy metabolism via activation of brown fat and browning of white fat, but intact liver insulin action is required for FGF21 to control hepatic lipid metabolism.

Authors

Brice Emanuelli, Sara G. Vienberg, Graham Smyth, Christine Cheng, Kristin I. Stanford, Manimozhiyan Arumugam, Mervyn D. Michael, Andrew C. Adams, Alexei Kharitonenkov, C. Ronald Kahn

Positive feedback between NF-κB and TNF-α promotes leukemia-initiating cell capacity

• Text
• PDF

Abstract

Acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy that originates from leukemia-initiating cells (LICs). The identification of common mechanisms underlying LIC development will be important in establishing broadly effective therapeutics for AML. Constitutive NF-κB pathway activation has been reported in different types of AML; however, the mechanism of NF-κB activation and its importance in leukemia progression are poorly understood. Here, we analyzed myeloid leukemia mouse models to assess NF-κB activity in AML LICs. We found that LICs, but not normal hematopoietic stem cells or non-LIC fractions within leukemia cells, exhibited constitutive NF-κB activity. This activity was maintained through autocrine TNF-α secretion, which formed an NF-κB/TNF-α positive feedback loop. LICs had increased levels of active proteasome machinery, which promoted the degradation of IκBα and further supported NF-κB activity. Pharmacological inhibition of the proteasome complex markedly suppressed leukemia progression in vivo. Conversely, enhanced activation of NF-κB signaling expanded LIC frequency within leukemia cell populations. We also demonstrated a strong correlation between NF-κB activity and TNF-α secretion in human AML samples. Our findings indicate that NF-κB/TNF-α signaling in LICs contributes to leukemia progression and provide a widely applicable approach for targeting LICs.

Authors

Yuki Kagoya, Akihide Yoshimi, Keisuke Kataoka, Masahiro Nakagawa, Keiki Kumano, Shunya Arai, Hiroshi Kobayashi, Taku Saito, Yoichiro Iwakura, Mineo Kurokawa

Platelet factor 4 limits Th17 differentiation and cardiac allograft rejection

• Text
• PDF

Abstract

Th cells are the major effector cells in transplant rejection and can be divided into Th1, Th2, Th17, and Treg subsets. Th differentiation is controlled by transcription factor expression, which is driven by positive and negative cytokine and chemokine stimuli at the time of T cell activation. Here we discovered that chemokine platelet factor 4 (PF4) is a negative regulator of Th17 differentiation. PF4-deficient and platelet-deficient mice had exaggerated immune responses to cardiac transplantation, including increased numbers of infiltrating Th17 cells and increased plasma IL-17. Although PF4 has been described as a platelet-specific molecule, we found that activated T cells also express PF4. Furthermore, bone marrow transplantation experiments revealed that T cell–derived PF4 contributes to a restriction in Th17 differentiation. Taken together, the results of this study demonstrate that PF4 is a key regulator of Th cell development that is necessary to limit Th17 differentiation. These data likely will impact our understanding of platelet-dependent regulation of T cell development, which is important in many diseases, in addition to transplantation.

Authors

Guanfang Shi, David J. Field, Kyung-ae Ko, Sara Ture, Kalyan Srivastava, Scott Levy, M. Anna Kowalska, Mortimer Poncz, Deborah J. Fowell, Craig N. Morrell

Erythropoietin promotes breast tumorigenesis through tumor-initiating cell self-renewal

• Text
• PDF

Abstract

Erythropoietin (EPO) is a hormone that induces red blood cell production. In its recombinant form, EPO is the one of most prescribed drugs to treat anemia, including that arising in cancer patients. In randomized trials, EPO administration to cancer patients has been associated with decreased survival. Here, we investigated the impact of EPO modulation on tumorigenesis. Using genetically engineered mouse models of breast cancer, we found that EPO promoted tumorigenesis by activating JAK/STAT signaling in breast tumor-initiating cells (TICs) and promoted TIC self renewal. We determined that EPO was induced by hypoxia in breast cancer cell lines, but not in human mammary epithelial cells. Additionally, we demonstrated that high levels of endogenous EPO gene expression correlated with shortened relapse-free survival and that pharmacologic JAK2 inhibition was synergistic with chemotherapy for tumor growth inhibition in vivo. These data define an active role for endogenous EPO in breast cancer progression and breast TIC self-renewal and reveal a potential application of EPO pathway inhibition in breast cancer therapy.

Authors

Bing Zhou, Jeffrey S. Damrauer, Sean T. Bailey, Tanja Hadzic, Youngtae Jeong, Kelly Clark, Cheng Fan, Laura Murphy, Cleo Y. Lee, Melissa A. Troester, C. Ryan Miller, Jian Jin, David Darr, Charles M. Perou, Ross L. Levine, Maximilian Diehn, William Y. Kim

Sustained activation of SMAD3/SMAD4 by FOXM1 promotes TGF-β–dependent cancer metastasis

• Text
• PDF

Abstract

A key feature of TGF-β signaling activation in cancer cells is the sustained activation of SMAD complexes in the nucleus; however, the drivers of SMAD activation are poorly defined. Here, using human and mouse breast cancer cell lines, we found that oncogene forkhead box M1 (FOXM1) interacts with SMAD3 to sustain activation of the SMAD3/SMAD4 complex in the nucleus. FOXM1 prevented the E3 ubiquitin-protein ligase transcriptional intermediary factor 1 γ (TIF1γ) from binding SMAD3 and monoubiquitinating SMAD4, which stabilized the SMAD3/SMAD4 complex. Loss of FOXM1 abolished TGF-β–induced SMAD3/SMAD4 formation. Moreover, the interaction of FOXM1 and SMAD3 promoted TGF-β/SMAD3–mediated transcriptional activity and target gene expression. We found that FOXM1/SMAD3 interaction was required for TGF-β–induced breast cancer invasion, which was the result of SMAD3/SMAD4-dependent upregulation of the transcription factor SLUG. Importantly, the function of FOXM1 in TGF-β–induced invasion was not dependent on FOXM1’s transcriptional activity. Knockdown of SMAD3 diminished FOXM1-induced metastasis. Furthermore, FOXM1 levels correlated with activated TGF-β signaling and metastasis in human breast cancer specimens. Together, our data indicate that FOXM1 promotes breast cancer metastasis by increasing nuclear retention of SMAD3 and identify crosstalk between FOXM1 and TGF-β/SMAD3 pathways. This study highlights the critical interaction of FOXM1 and SMAD3 for controlling TGF-β signaling during metastasis.

Authors

Jianfei Xue, Xia Lin, Wen-Tai Chiu, Yao-Hui Chen, Guanzhen Yu, Mingguang Liu, Xin-Hua Feng, Raymond Sawaya, René H. Medema, Mien-Chie Hung, Suyun Huang

Thrombocytopenia-associated mutations in the ANKRD26 regulatory region induce MAPK hyperactivation

• Text
• PDF

Abstract

Point mutations in the 5′ UTR of ankyrin repeat domain 26 (ANKRD26) are associated with familial thrombocytopenia 2 (THC2) and a predisposition to leukemia. Here, we identified underlying mechanisms of ANKRD26-associated thrombocytopenia. Using megakaryocytes (MK) isolated from THC2 patients and healthy subjects, we demonstrated that THC2-associated mutations in the 5′ UTR of ANKRD26 resulted in loss of runt-related transcription factor 1 (RUNX1) and friend leukemia integration 1 transcription factor (FLI1) binding. RUNX1 and FLI1 binding at the 5′ UTR from healthy subjects led to ANKRD26 silencing during the late stages of megakaryopoiesis and blood platelet development. We showed that persistent ANKRD26 expression in isolated MKs increased signaling via the thrombopoietin/myeloproliferative leukemia virus oncogene (MPL) pathway and impaired proplatelet formation by MKs. Importantly, we demonstrated that ERK inhibition completely rescued the in vitro proplatelet formation defect. Our data identify a mechanism for development of the familial thrombocytopenia THC2 that is related to abnormal MAPK signaling.

Authors

Dominique Bluteau, Alessandra Balduini, Nathalie Balayn, Manuela Currao, Paquita Nurden, Caroline Deswarte, Guy Leverger, Patrizia Noris, Silverio Perrotta, Eric Solary, William Vainchenker, Najet Debili, Remi Favier, Hana Raslova

Opioid receptor–triggered spinal mTORC1 activation contributes to morphine tolerance and hyperalgesia

• Text
• PDF

Abstract

The development of opioid-induced analgesic tolerance and hyperalgesia is a clinical challenge for managing chronic pain. Adaptive changes in protein translation in the nervous system are thought to promote opioid tolerance and hyperalgesia; however, how opioids drive such changes remains elusive. Here, we report that mammalian target of rapamycin (mTOR), which governs most protein translation, was activated in rat spinal dorsal horn neurons after repeated intrathecal morphine injections. Activation was triggered through μ opioid receptor and mediated by intracellular PI3K/Akt. Spinal mTOR inhibition blocked both induction and maintenance of morphine tolerance and hyperalgesia, without affecting basal pain perception or locomotor functions. These effects were attributed to the attenuation of morphine-induced increases in translation initiation activity, nascent protein synthesis, and expression of some known key tolerance-associated proteins, including neuronal NOS (nNOS), in dorsal horn. Moreover, elevating spinal mTOR activity by knocking down the mTOR-negative regulator TSC2 reduced morphine analgesia, produced pain hypersensitivity, and increased spinal nNOS expression. Our findings implicate the μ opioid receptor–triggered PI3K/Akt/mTOR pathway in promoting morphine-induced spinal protein translation changes and associated morphine tolerance and hyperalgesia. These data suggest that mTOR inhibitors could be explored for prevention and/or reduction of opioid tolerance in chronic pain management.

Authors

Ji-Tian Xu, Jian-Yuan Zhao, Xiuli Zhao, Davinna Ligons, Vinod Tiwari, Fidelis E. Atianjoh, Chun-Yi Lee, Lingli Liang, Weidong Zang, Dolores Njoku, Srinivasa N. Raja, Myron Yaster, Yuan-Xiang Tao

Orexin neurons suppress narcolepsy via 2 distinct efferent pathways

• Text
• PDF

Abstract

The loss of orexin neurons in humans is associated with the sleep disorder narcolepsy, which is characterized by excessive daytime sleepiness and cataplexy. Mice lacking orexin peptides, orexin neurons, or orexin receptors recapitulate human narcolepsy phenotypes, further highlighting a critical role for orexin signaling in the maintenance of wakefulness. Despite the known role of orexin neurons in narcolepsy, the precise neural mechanisms downstream of these neurons remain unknown. We found that targeted restoration of orexin receptor expression in the dorsal raphe (DR) and in the locus coeruleus (LC) of mice lacking orexin receptors inhibited cataplexy-like episodes and pathological fragmentation of wakefulness (i.e., sleepiness), respectively. The suppression of cataplexy-like episodes correlated with the number of serotonergic neurons restored with orexin receptor expression in the DR, while the consolidation of fragmented wakefulness correlated with the number of noradrenergic neurons restored in the LC. Furthermore, pharmacogenetic activation of these neurons using designer receptor exclusively activated by designer drug (DREADD) technology ameliorated narcolepsy in mice lacking orexin neurons. These results suggest that DR serotonergic and LC noradrenergic neurons play differential roles in orexin neuron–dependent regulation of sleep/wakefulness and highlight a pharmacogenetic approach for the amelioration of narcolepsy.

Authors

Emi Hasegawa, Masashi Yanagisawa, Takeshi Sakurai, Michihiro Mieda

Cardiotoxicity of doxorubicin is mediated through mitochondrial iron accumulation

• Text
• PDF

Abstract

Doxorubicin is an effective anticancer drug with known cardiotoxic side effects. It has been hypothesized that doxorubicin-dependent cardiotoxicity occurs through ROS production and possibly cellular iron accumulation. Here, we found that cardiotoxicity develops through the preferential accumulation of iron inside the mitochondria following doxorubicin treatment. In isolated cardiomyocytes, doxorubicin became concentrated in the mitochondria and increased both mitochondrial iron and cellular ROS levels. Overexpression of ABCB8, a mitochondrial protein that facilitates iron export, in vitro and in the hearts of transgenic mice decreased mitochondrial iron and cellular ROS and protected against doxorubicin-induced cardiomyopathy. Dexrazoxane, a drug that attenuates doxorubicin-induced cardiotoxicity, decreased mitochondrial iron levels and reversed doxorubicin-induced cardiac damage. Finally, hearts from patients with doxorubicin-induced cardiomyopathy had markedly higher mitochondrial iron levels than hearts from patients with other types of cardiomyopathies or normal cardiac function. These results suggest that the cardiotoxic effects of doxorubicin develop from mitochondrial iron accumulation and that reducing mitochondrial iron levels protects against doxorubicin-induced cardiomyopathy.

Authors

Yoshihiko Ichikawa, Mohsen Ghanefar, Marina Bayeva, Rongxue Wu, Arineh Khechaduri, Sathyamangla V. Naga Prasad, R. Kannan Mutharasan, Tejaswitha Jairaj Naik, Hossein Ardehali

OTX2 loss causes rod differentiation defect in CRX-associated congenital blindness

• Text
• PDF

Abstract

Leber congenital amaurosis (LCA) encompasses a set of early-onset blinding diseases that are characterized by vision loss, involuntary eye movement, and nonrecordable electroretinogram (ERG). At least 19 genes are associated with LCA, which is typically recessive; however, mutations in homeodomain transcription factor CRX lead to an autosomal dominant form of LCA. The mechanism of CRX-associated LCA is not understood. Here, we identified a spontaneous mouse mutant with a frameshift mutation in Crx (Crx^Rip). We determined that Crx^Rip is a dominant mutation that results in congenital blindness with nonrecordable response by ERG and arrested photoreceptor differentiation with no associated degeneration. Expression of LCA-associated dominant CRX frameshift mutations in mouse retina mimicked the Crx^Rip phenotype, which was rescued by overexpression of WT CRX. Whole-transcriptome profiling using deep RNA sequencing revealed progressive and complete loss of rod differentiation factor NRL in Crx^Rip retinas. Expression of NRL partially restored rod development in Crx^Rip/+ mice. We show that the binding of homeobox transcription factor OTX2 at the Nrl promoter was obliterated in Crx^Rip mice and ectopic expression of OTX2 rescued the rod differentiation defect. Together, our data indicate that OTX2 maintains Nrl expression in developing rods to consolidate rod fate. Our studies provide insights into CRX mutation-associated congenital blindness and should assist in therapeutic design.

Authors

Jerome E. Roger, Avinash Hiriyanna, Norimoto Gotoh, Hong Hao, Debbie F. Cheng, Rinki Ratnapriya, Marie-Audrey I. Kautzmann, Bo Chang, Anand Swaroop

Phenothiazines induce PP2A-mediated apoptosis in T cell acute lymphoblastic leukemia

• Text
• PDF

Abstract

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive cancer that is frequently associated with activating mutations in NOTCH1 and dysregulation of MYC. Here, we performed 2 complementary screens to identify FDA-approved drugs and drug-like small molecules with activity against T-ALL. We developed a zebrafish system to screen small molecules for toxic activity toward MYC-overexpressing thymocytes and used a human T-ALL cell line to screen for small molecules that synergize with Notch inhibitors. We identified the antipsychotic drug perphenazine in both screens due to its ability to induce apoptosis in fish, mouse, and human T-ALL cells. Using ligand-affinity chromatography coupled with mass spectrometry, we identified protein phosphatase 2A (PP2A) as a perphenazine target. T-ALL cell lines treated with perphenazine exhibited rapid dephosphorylation of multiple PP2A substrates and subsequent apoptosis. Moreover, shRNA knockdown of specific PP2A subunits attenuated perphenazine activity, indicating that PP2A mediates the drug’s antileukemic activity. Finally, human T-ALLs treated with perphenazine exhibited suppressed cell growth and dephosphorylation of PP2A targets in vitro and in vivo. Our findings provide a mechanistic explanation for the recurring identification of phenothiazines as a class of drugs with anticancer effects. Furthermore, these data suggest that pharmacologic PP2A activation in T-ALL and other cancers driven by hyperphosphorylated PP2A substrates has therapeutic potential.

Authors

Alejandro Gutierrez, Li Pan, Richard W.J. Groen, Frederic Baleydier, Alex Kentsis, Jason Marineau, Ruta Grebliunaite, Elena Kozakewich, Casie Reed, Francoise Pflumio, Sandrine Poglio, Benjamin Uzan, Paul Clemons, Lynn VerPlank, Frank An, Jason Burbank, Stephanie Norton, Nicola Tolliday, Hanno Steen, Andrew P. Weng, Huipin Yuan, James E. Bradner, Constantine Mitsiades, A. Thomas Look, Jon C. Aster

Neuronal ferritin heavy chain and drug abuse affect HIV-associated cognitive dysfunction

• Text
• PDF

Abstract

Interaction of the chemokine CXCL12 with its receptor CXCR4 promotes neuronal function and survival during embryonic development and throughout adulthood. Previous studies indicated that μ-opioid agonists specifically elevate neuronal levels of the protein ferritin heavy chain (FHC), which negatively regulates CXCR4 signaling and affects the neuroprotective function of the CXCL12/CXCR4 axis. Here, we determined that CXCL12/CXCR4 activity increased dendritic spine density, and also examined FHC expression and CXCR4 status in opiate abusers and patients with HIV-associated neurocognitive disorders (HAND), which is typically exacerbated by illicit drug use. Drug abusers and HIV patients with HAND had increased levels of FHC, which correlated with reduced CXCR4 activation, within cortical neurons. We confirmed these findings in a nonhuman primate model of SIV infection with morphine administration. Transfection of a CXCR4-expressing human cell line with an iron-deficient FHC mutant confirmed that increased FHC expression deregulated CXCR4 signaling and that this function of FHC was independent of iron binding. Furthermore, examination of morphine-treated rodents and isolated neurons expressing FHC shRNA revealed that FHC contributed to morphine-induced dendritic spine loss. Together, these data implicate FHC-dependent deregulation of CXCL12/CXCR4 as a contributing factor to cognitive dysfunction in neuroAIDS.

Authors

Jonathan Pitcher, Anna Abt, Jaclyn Myers, Rachel Han, Melissa Snyder, Alessandro Graziano, Lindsay Festa, Michele Kutzler, Fernando Garcia, Wen-Jun Gao, Tracy Fischer-Smith, Jay Rappaport, Olimpia Meucci

Targeting the cell cycle inhibitor p57^Kip2 promotes adult human β cell replication

• Text
• PDF

Abstract

Children with focal hyperinsulinism of infancy display a dramatic, non-neoplastic clonal expansion of β cells that have undergone mitotic recombination, resulting in paternal disomy of part of chromosome 11. This disomic region contains imprinted genes, including the gene encoding the cell cycle inhibitor p57^Kip2 (CDKN1C), which is silenced as a consequence of the recombination event. We hypothesized that targeting p57^Kip2 could stimulate adult human β cell replication. Indeed, when we suppressed CDKN1C expression in human islets obtained from deceased adult organ donors and transplanted them into hyperglycemic, immunodeficient mice, β cell replication increased more than 3-fold. The newly replicated cells retained properties of mature β cells, including the expression of β cell markers such as insulin, PDX1, and NKX6.1. Importantly, these newly replicated cells demonstrated normal glucose-induced calcium influx, further indicating β cell functionality. These findings provide a molecular explanation for the massive β cell replication that occurs in children with focal hyperinsulinism. These data also provided evidence that β cells from older humans, in which baseline replication is negligible, can be coaxed to re-enter and complete the cell cycle while maintaining mature β cell properties. Thus, controlled manipulation of this pathway holds promise for the expansion of β cells in patients with type 2 diabetes.

Authors

Dana Avrahami, Changhong Li, Ming Yu, Yang Jiao, Jia Zhang, Ali Naji, Seyed Ziaie, Benjamin Glaser, Klaus H. Kaestner

Disruption of vascular Ca²⁺-activated chloride currents lowers blood pressure

• Text
• PDF

Abstract

High blood pressure is the leading risk factor for death worldwide. One of the hallmarks is a rise of peripheral vascular resistance, which largely depends on arteriole tone. Ca²⁺-activated chloride currents (CaCCs) in vascular smooth muscle cells (VSMCs) are candidates for increasing vascular contractility. We analyzed the vascular tree and identified substantial CaCCs in VSMCs of the aorta and carotid arteries. CaCCs were small or absent in VSMCs of medium-sized vessels such as mesenteric arteries and larger retinal arterioles. In small vessels of the retina, brain, and skeletal muscle, where contractile intermediate cells or pericytes gradually replace VSMCs, CaCCs were particularly large. Targeted disruption of the calcium-activated chloride channel TMEM16A, also known as ANO1, in VSMCs, intermediate cells, and pericytes eliminated CaCCs in all vessels studied. Mice lacking vascular TMEM16A had lower systemic blood pressure and a decreased hypertensive response following vasoconstrictor treatment. There was no difference in contractility of medium-sized mesenteric arteries; however, responsiveness of the aorta and small retinal arterioles to the vasoconstriction-inducing drug U46619 was reduced. TMEM16A also was required for peripheral blood vessel contractility, as the response to U46619 was attenuated in isolated perfused hind limbs from mutant mice. Out data suggest that TMEM16A plays a general role in arteriolar and capillary blood flow and is a promising target for the treatment of hypertension.

Authors

Christoph Heinze, Anika Seniuk, Maxim V. Sokolov, Antje K. Huebner, Agnieszka E. Klementowicz, István A. Szijártó, Johanna Schleifenbaum, Helga Vitzthum, Maik Gollasch, Heimo Ehmke, Björn C. Schroeder, Christian A. Hübner

Irradiation and anti–PD-L1 treatment synergistically promote antitumor immunity in mice

• Text
• PDF

Abstract

High-dose ionizing irradiation (IR) results in direct tumor cell death and augments tumor-specific immunity, which enhances tumor control both locally and distantly. Unfortunately, local relapses often occur following IR treatment, indicating that IR-induced responses are inadequate to maintain antitumor immunity. Therapeutic blockade of the T cell negative regulator programmed death–ligand 1 (PD-L1, also called B7-H1) can enhance T cell effector function when PD-L1 is expressed in chronically inflamed tissues and tumors. Here, we demonstrate that PD-L1 was upregulated in the tumor microenvironment after IR. Administration of anti–PD-L1 enhanced the efficacy of IR through a cytotoxic T cell–dependent mechanism. Concomitant with IR-mediated tumor regression, we observed that IR and anti–PD-L1 synergistically reduced the local accumulation of tumor-infiltrating myeloid-derived suppressor cells (MDSCs), which suppress T cells and alter the tumor immune microenvironment. Furthermore, activation of cytotoxic T cells with combination therapy mediated the reduction of MDSCs in tumors through the cytotoxic actions of TNF. Our data provide evidence for a close interaction between IR, T cells, and the PD-L1/PD-1 axis and establish a basis for the rational design of combination therapy with immune modulators and radiotherapy.

Authors

Liufu Deng, Hua Liang, Byron Burnette, Michael Beckett, Thomas Darga, Ralph R. Weichselbaum, Yang-Xin Fu

Changes in neural network homeostasis trigger neuropsychiatric symptoms

• Text
• PDF

Abstract

The mechanisms that regulate the strength of synaptic transmission and intrinsic neuronal excitability are well characterized; however, the mechanisms that promote disease-causing neural network dysfunction are poorly defined. We generated mice with targeted neuron type–specific expression of a gain-of-function variant of the neurotransmitter receptor for glycine (GlyR) that is found in hippocampectomies from patients with temporal lobe epilepsy. In this mouse model, targeted expression of gain-of-function GlyR in terminals of glutamatergic cells or in parvalbumin-positive interneurons persistently altered neural network excitability. The increased network excitability associated with gain-of-function GlyR expression in glutamatergic neurons resulted in recurrent epileptiform discharge, which provoked cognitive dysfunction and memory deficits without affecting bidirectional synaptic plasticity. In contrast, decreased network excitability due to gain-of-function GlyR expression in parvalbumin-positive interneurons resulted in an anxiety phenotype, but did not affect cognitive performance or discriminative associative memory. Our animal model unveils neuron type–specific effects on cognition, formation of discriminative associative memory, and emotional behavior in vivo. Furthermore, our data identify a presynaptic disease–causing molecular mechanism that impairs homeostatic regulation of neural network excitability and triggers neuropsychiatric symptoms.

Authors

Aline Winkelmann, Nicola Maggio, Joanna Eller, Gürsel Caliskan, Marcus Semtner, Ute Häussler, René Jüttner, Tamar Dugladze, Birthe Smolinsky, Sarah Kowalczyk, Ewa Chronowska, Günter Schwarz, Fritz G. Rathjen, Gideon Rechavi, Carola A. Haas, Akos Kulik, Tengis Gloveli, Uwe Heinemann, Jochen C. Meier

Normalizing glycosphingolipids restores function in CD4⁺ T cells from lupus patients

• Text
• PDF

Abstract

Patients with the autoimmune rheumatic disease systemic lupus erythematosus (SLE) have multiple defects in lymphocyte signaling and function that contribute to disease pathogenesis. Such defects could be attributed to alterations in metabolic processes, including abnormal control of lipid biosynthesis pathways. Here, we reveal that CD4⁺ T cells from SLE patients displayed an altered profile of lipid raft–associated glycosphingolipids (GSLs) compared with that of healthy controls. In particular, lactosylceramide, globotriaosylceramide (Gb3), and monosialotetrahexosylganglioside (GM1) levels were markedly increased. Elevated GSLs in SLE patients were associated with increased expression of liver X receptor β (LXRβ), a nuclear receptor that controls cellular lipid metabolism and trafficking and influences acquired immune responses. Stimulation of CD4⁺ T cells isolated from healthy donors with synthetic and endogenous LXR agonists promoted GSL expression, which was blocked by an LXR antagonist. Increased GSL expression in CD4⁺ T cells was associated with intracellular accumulation and accelerated trafficking of GSL, reminiscent of cells from patients with glycolipid storage diseases. Inhibition of GSL biosynthesis in vitro with a clinically approved inhibitor (N-butyldeoxynojirimycin) normalized GSL metabolism, corrected CD4⁺ T cell signaling and functional defects, and decreased anti-dsDNA antibody production by autologous B cells in SLE patients. Our data demonstrate that lipid metabolism defects contribute to SLE pathogenesis and suggest that targeting GSL biosynthesis restores T cell function in SLE.

Authors

Georgia McDonald, Shantal Deepak, Laura Miguel, Cleo J. Hall, David A. Isenberg, Anthony I. Magee, Terry Butters, Elizabeth C. Jury

Human IgG Fc domain engineering enhances antitoxin neutralizing antibody activity

• Text
• PDF

Abstract

The effector activity of antibodies is dependent on engagement with Fcγ receptors (FcγRs) and activation of the associated intracellular signaling pathways. Preclinical evaluation of therapeutic humanized or chimeric mAbs to study the interactions of their Fc regions with FcγRs is hampered by substantial structural and functional FcγR diversity among species. In this report, we used mice expressing only human FcγRs to evaluate the contribution of FcγR-mediated pathways to the neutralizing activity of an anti-anthrax toxin chimeric mAb. We observed that the protective activity of this mAb was highly dependent upon FcγR engagement, with minimal protection against anthrax toxin observed in FcγR-deficient mice following mAb administration. We generated anti-anthrax toxin mAbs with specific Fc domain variants with selectively enhanced affinity for particular human FcγRs and assessed their activity in FcγR-humanized mice. We determined that Fc domain variants that were capable of selectively engaging activating FcγRs substantially enhanced the in vitro and in vivo activity of anthrax toxin-neutralizing antibodies. These findings indicate that the application of Fc domain engineering is a feasible strategy to enhance toxin-neutralizing activity and suggest that engineered antitoxin antibodies will have improved therapeutic efficacy.

Authors

Stylianos Bournazos, Siu-Kei Chow, Nareen Abboud, Arturo Casadevall, Jeffrey V. Ravetch

NRF2-mediated Notch pathway activation enhances hematopoietic reconstitution following myelosuppressive radiation

• Text
• PDF

Abstract

A nuclear disaster may result in exposure to potentially lethal doses of ionizing radiation (IR). Hematopoietic acute radiation syndrome (H-ARS) is characterized by severe myelosuppression, which increases the risk of infection, bleeding, and mortality. Here, we determined that activation of nuclear factor erythroid-2–related factor 2 (NRF2) signaling enhances hematopoietic stem progenitor cell (HSPC) function and mitigates IR-induced myelosuppression and mortality. Augmenting NRF2 signaling in mice, either by genetic deletion of the NRF2 inhibitor Keap1 or by pharmacological NRF2 activation with 2-trifluoromethyl-2′-methoxychalone (TMC), enhanced hematopoietic reconstitution following bone marrow transplantation (BMT). Strikingly, even 24 hours after lethal IR exposure, oral administration of TMC mitigated myelosuppression and mortality in mice. Furthermore, TMC administration to irradiated transgenic Notch reporter mice revealed activation of Notch signaling in HSPCs and enhanced HSPC expansion by increasing Jagged1 expression in BM stromal cells. Administration of a Notch inhibitor ablated the effects of TMC on hematopoietic reconstitution. Taken together, we identified a mechanism by which NRF2-mediated Notch signaling improves HSPC function and myelosuppression following IR exposure. Our data indicate that targeting this pathway may provide a countermeasure against the damaging effects of IR exposure.

Authors

Jung-Hyun Kim, Rajesh K. Thimmulappa, Vineet Kumar, Wanchang Cui, Sarvesh Kumar, Ponvijay Kombairaju, Hao Zhang, Joseph Margolick, William Matsui, Thomas Macvittie, Sanjay V. Malhotra, Shyam Biswal

MNK1 pathway activity maintains protein synthesis in rapalog-treated gliomas

• Text
• PDF

Abstract

High levels of mammalian target of rapamycin complex 1 (mTORC1) activity in malignant gliomas promote tumor progression, suggesting that targeting mTORC1 has potential as a therapeutic strategy. Remarkably, clinical trials in patients with glioma revealed that rapamycin analogs (rapalogs) have limited efficacy, indicating activation of resistance mechanisms. Targeted depletion of MAPK-interacting Ser/Thr kinase 1 (MNK1) sensitizes glioma cells to the mTORC1 inhibitor rapamycin through an indistinct mechanism. Here, we analyzed how MNK1 and mTORC1 signaling pathways regulate the assembly of translation initiation complexes, using the cap analog m⁷GTP to enrich for initiation complexes in glioma cells followed by mass spectrometry–based quantitative proteomics. Association of eukaryotic translation initiation factor 4E (eIF4E) with eIF4E-binding protein 1 (4EBP1) was regulated by the mTORC1 pathway, whereas pharmacological blocking of MNK activity by CGP57380 or MNK1 knockdown, along with mTORC1 inhibition by RAD001, increased 4EBP1 binding to eIF4E. Furthermore, combined MNK1 and mTORC1 inhibition profoundly inhibited 4EBP1 phosphorylation at Ser65, protein synthesis and proliferation in glioma cells, and reduced tumor growth in an orthotopic glioblastoma (GBM) mouse model. Immunohistochemical analysis of GBM samples revealed increased 4EBP1 phosphorylation. Taken together, our data indicate that rapalog-activated MNK1 signaling promotes glioma growth through regulation of 4EBP1 and indicate a molecular cross-talk between the mTORC1 and MNK1 pathways that has potential to be exploited therapeutically.

Authors

Michal Grzmil, Roland M. Huber, Daniel Hess, Stephan Frank, Debby Hynx, Gerald Moncayo, Dominique Klein, Adrian Merlo, Brian A. Hemmings

Tgfbr2 disruption in postnatal smooth muscle impairs aortic wall homeostasis

• Text
• PDF

Abstract

TGF-β is essential for vascular development; however, excess TGF-β signaling promotes thoracic aortic aneurysm and dissection in multiple disorders, including Marfan syndrome. Since the pathology of TGF-β overactivity manifests primarily within the arterial media, it is widely assumed that suppression of TGF-β signaling in vascular smooth muscle cells will ameliorate aortic disease. We tested this hypothesis by conditional inactivation of Tgfbr2, which encodes the TGF-β type II receptor, in smooth muscle cells of postweanling mice. Surprisingly, the thoracic aorta rapidly thickened, dilated, and dissected in these animals. Tgfbr2 disruption predictably decreased canonical Smad signaling, but unexpectedly increased MAPK signaling. Type II receptor–independent effects of TGF-β and pathological responses by nonrecombined smooth muscle cells were excluded by serologic neutralization. Aortic disease was caused by a perturbed contractile apparatus in medial cells and growth factor production by adventitial cells, both of which resulted in maladaptive paracrine interactions between the vessel wall compartments. Treatment with rapamycin restored a quiescent smooth muscle phenotype and prevented dissection. Tgfbr2 disruption in smooth muscle cells also accelerated aneurysm growth in a murine model of Marfan syndrome. Our data indicate that basal TGF-β signaling in smooth muscle promotes postnatal aortic wall homeostasis and impedes disease progression.

Authors

Wei Li, Qingle Li, Yang Jiao, Lingfeng Qin, Rahmat Ali, Jing Zhou, Jacopo Ferruzzi, Richard W. Kim, Arnar Geirsson, Harry C. Dietz, Stefan Offermanns, Jay D. Humphrey, George Tellides

Nutrient sensing by the mitochondrial transcription machinery dictates oxidative phosphorylation

• Text
• PDF

Abstract

Sirtuin 3 (SIRT3), an important regulator of energy metabolism and lipid oxidation, is induced in fasted liver mitochondria and implicated in metabolic syndrome. In fasted liver, SIRT3-mediated increases in substrate flux depend on oxidative phosphorylation (OXPHOS), but precisely how OXPHOS meets the challenge of increased substrate oxidation in fasted liver remains unclear. Here, we show that liver mitochondria in fasting mice adapt to the demand of increased substrate oxidation by increasing their OXPHOS efficiency. In response to cAMP signaling, SIRT3 deacetylated and activated leucine-rich protein 130 (LRP130; official symbol, LRPPRC), promoting a mitochondrial transcriptional program that enhanced hepatic OXPHOS. Using mass spectrometry, we identified SIRT3-regulated lysine residues in LRP130 that generated a lysine-to-arginine (KR) mutant of LRP130 that mimics deacetylated protein. Compared with wild-type LRP130 protein, expression of the KR mutant increased mitochondrial transcription and OXPHOS in vitro. Indeed, even when SIRT3 activity was abolished, activation of mitochondrial transcription and OXPHOS by the KR mutant remained robust, further highlighting the contribution of LRP130 deacetylation to increased OXPHOS in fasted liver. These data establish a link between nutrient sensing and mitochondrial transcription that regulates OXPHOS in fasted liver and may explain how fasted liver adapts to increased substrate oxidation.

Authors

Lijun Liu, Minwoo Nam, Wei Fan, Thomas E. Akie, David C. Hoaglin, Guangping Gao, John F. Keaney Jr., Marcus P. Cooper

Requirement of enhanced Survival Motoneuron protein imposed during neuromuscular junction maturation

• Text
• PDF

Abstract

Spinal muscular atrophy is a common motor neuron disease caused by low survival motoneuron (SMN), a key protein in the proper splicing of genes. Restoring the protein is therefore a promising therapeutic strategy. Implementation of this strategy, however, depends on defining the temporal requirements for SMN. Here, we used controlled knockdown of SMN in transgenic mice to determine the precise postnatal stage requirements for this protein. Reducing SMN in neonatal mice resulted in a classic SMA-like phenotype. Unexpectedly, depletion of SMN in adults had relatively little effect. Insensitivity to low SMN emerged abruptly at postnatal day 17, which coincided with establishment of the fully mature neuromuscular junction (NMJ). Mature animals depleted of SMN eventually exhibited evidence of selective neuromuscular pathology that was made worse by traumatic injury. The ability to regenerate the mature NMJ in aged or injured SMN-depleted mice was grossly impaired, a likely consequence of the inability to meet the surge in demand for motoneuronal SMN that was seen in controls. Our results demonstrate that relative maturity of the NMJ determines the temporal requirement for the SMN protein. These observations suggest that the use of potent but potentially deleterious SMN-enhancing agents could be tapered in human patients once the neuromuscular system matures and reintroduced as needed to enhance SMN for remodeling aged or injured NMJs.

Authors

Shingo Kariya, Teresa Obis, Caterina Garone, Turgay Akay, Fusako Sera, Shinichi Iwata, Shunichi Homma, Umrao R. Monani

Prenatal retinoid deficiency leads to airway hyperresponsiveness in adult mice

• Text
• PDF

Abstract

There is increasing evidence that vitamin A deficiency in utero correlates with abnormal airway smooth muscle (SM) function in postnatal life. The bioactive vitamin A metabolite retinoic acid (RA) is essential for formation of the lung primordium; however, little is known about the impact of early fetal RA deficiency on postnatal lung structure and function. Here, we provide evidence that during murine lung development, endogenous RA has a key role in restricting the airway SM differentiation program during airway formation. Using murine models of pharmacological, genetic, and dietary vitamin A/RA deficiency, we found that disruption of RA signaling during embryonic development consistently resulted in an altered airway SM phenotype with markedly increased expression of SM markers. The aberrant phenotype persisted postnatally regardless of the adult vitamin A status and manifested as structural changes in the bronchial SM and hyperresponsiveness of the airway without evidence of inflammation. Our data reveal a role for endogenous RA signaling in restricting SM differentiation and preventing precocious and excessive SM differentiation when airways are forming.

Authors

Felicia Chen, Hector Marquez, Youn-Kyung Kim, Jun Qian, Fengzhi Shao, Alan Fine, William W. Cruikshank, Loredana Quadro, Wellington V. Cardoso

Macrophages eliminate circulating tumor cells after monoclonal antibody therapy

• Text
• PDF

Abstract

The use of monoclonal antibodies (mAbs) as therapeutic tools has increased dramatically in the last decade and is now one of the mainstream strategies to treat cancer. Nonetheless, it is still not completely understood how mAbs mediate tumor cell elimination or the effector cells that are involved. Using intravital microscopy, we found that antibody-dependent phagocytosis (ADPh) by macrophages is a prominent mechanism for removal of tumor cells from the circulation in a murine tumor cell opsonization model. Tumor cells were rapidly recognized and arrested by liver macrophages (Kupffer cells). In the absence of mAbs, Kupffer cells sampled tumor cells; however, this sampling was not sufficient for elimination. By contrast, antitumor mAb treatment resulted in rapid phagocytosis of tumor cells by Kupffer cells that was dependent on the high-affinity IgG-binding Fc receptor (FcγRI) and the low-affinity IgG-binding Fc receptor (FcγRIV). Uptake and intracellular degradation were independent of reactive oxygen or nitrogen species production. Importantly, ADPh prevented the development of liver metastases. Tumor cell capture and therapeutic efficacy were lost after Kupffer cell depletion. Our data indicate that macrophages play a prominent role in mAb-mediated eradication of tumor cells. These findings may help to optimize mAb therapeutic strategies for patients with cancer by helping us to aim to enhance macrophage recruitment and activity.

Authors

Nuray Gül, Liane Babes, Kerstin Siegmund, Rianne Korthouwer, Marijn Bögels, Rens Braster, Gestur Vidarsson, Timo L.M. ten Hagen, Paul Kubes, Marjolein van Egmond

Tie1 deletion inhibits tumor growth and improves angiopoietin antagonist therapy

• Text
• PDF

Abstract

The endothelial Tie1 receptor is ligand-less, but interacts with the Tie2 receptor for angiopoietins (Angpt). Angpt2 is expressed in tumor blood vessels, and its blockade inhibits tumor angiogenesis. Here we found that Tie1 deletion from the endothelium of adult mice inhibits tumor angiogenesis and growth by decreasing endothelial cell survival in tumor vessels, without affecting normal vasculature. Treatment with VEGF or VEGFR-2 blocking antibodies similarly reduced tumor angiogenesis and growth; however, no additive inhibition was obtained by targeting both Tie1 and VEGF/VEGFR-2. In contrast, treatment of Tie1-deficient mice with a soluble form of the extracellular domain of Tie2, which blocks Angpt activity, resulted in additive inhibition of tumor growth. Notably, Tie1 deletion decreased sprouting angiogenesis and increased Notch pathway activity in the postnatal retinal vasculature, while pharmacological Notch suppression in the absence of Tie1 promoted retinal hypervasularization. Moreover, substantial additive inhibition of the retinal vascular front migration was observed when Angpt2 blocking antibodies were administered to Tie1-deficient pups. Thus, Tie1 regulates tumor angiogenesis, postnatal sprouting angiogenesis, and endothelial cell survival, which are controlled by VEGF, Angpt, and Notch signals. Our results suggest that targeting Tie1 in combination with Angpt/Tie2 has the potential to improve antiangiogenic therapy.

Authors

Gabriela D’Amico, Emilia A. Korhonen, Andrey Anisimov, Georgia Zarkada, Tanja Holopainen, René Hägerling, Friedemann Kiefer, Lauri Eklund, Raija Sormunen, Harri Elamaa, Rolf A. Brekken, Ralf H. Adams, Gou Young Koh, Pipsa Saharinen, Kari Alitalo

Inactivation of SAG/RBX2 E3 ubiquitin ligase suppresses Kras^G12D-driven lung tumorigenesis

• Text
• PDF

Abstract

Cullin-RING ligases (CRLs) are a family of E3 ubiquitin ligase complexes that rely on either RING-box 1 (RBX1) or sensitive to apoptosis gene (SAG), also known as RBX2, for activity. RBX1 and SAG are both overexpressed in human lung cancer; however, their contribution to patient survival and lung tumorigenesis is unknown. Here, we report that overexpression of SAG, but not RBX1, correlates with poor patient prognosis and more advanced disease. We found that SAG is overexpressed in murine Kras^G12D-driven lung tumors and that Sag deletion suppressed lung tumorigenesis and extended murine life span. Using cultured lung cancer cells, we showed that SAG knockdown suppressed growth and survival, inactivated both NF-κB and mTOR pathways, and resulted in accumulation of tumor suppressor substrates, including p21, p27, NOXA, and BIM. Importantly, growth suppression by SAG knockdown was partially rescued by simultaneous knockdown of p21 or the mTOR inhibitor DEPTOR. Treatment with MLN4924, a small molecule inhibitor of CRL E3s, also inhibited the formation of Kras^G12D-induced lung tumors through a similar mechanism involving inactivation of NF-κB and mTOR and accumulation of tumor suppressor substrates. Together, our results demonstrate that Sag is a Kras-cooperating oncogene that promotes lung tumorigenesis and suggest that targeting SAG-CRL E3 ligases may be an effective therapeutic approach for Kras-driven lung cancers.

Authors

Hua Li, Mingjia Tan, Lijun Jia, Dongping Wei, Yongchao Zhao, Guoan Chen, Jie Xu, Lili Zhao, Dafydd Thomas, David G. Beer, Yi Sun

Prion disease tempo determined by host-dependent substrate reduction

• Text
• PDF

Abstract

The symptoms of prion infection can take years or decades to manifest following the initial exposure. Molecular markers of prion disease include accumulation of the misfolded prion protein (PrP^Sc), which is derived from its cellular precursor (PrP^C), as well as downregulation of the PrP-like Shadoo (Sho) glycoprotein. Given the overlapping cellular environments for PrP^C and Sho, we inferred that PrP^C levels might also be altered as part of a host response during prion infection. Using rodent models, we found that, in addition to changes in PrP^C glycosylation and proteolytic processing, net reductions in PrP^C occur in a wide range of prion diseases, including sheep scrapie, human Creutzfeldt-Jakob disease, and cervid chronic wasting disease. The reduction in PrP^C results in decreased prion replication, as measured by the protein misfolding cyclic amplification technique for generating PrP^Sc in vitro. While PrP^C downregulation is not discernible in animals with unusually short incubation periods and high PrP^C expression, slowly evolving prion infections exhibit downregulation of the PrP^C substrate required for new PrP^Sc synthesis and as a receptor for pathogenic signaling. Our data reveal PrP^C downregulation as a previously unappreciated element of disease pathogenesis that defines the extensive, presymptomatic period for many prion strains.

Authors

Charles E. Mays, Chae Kim, Tracy Haldiman, Jacques van der Merwe, Agnes Lau, Jing Yang, Jennifer Grams, Michele A. Di Bari, Romolo Nonno, Glenn C. Telling, Qingzhong Kong, Jan Langeveld, Debbie McKenzie, David Westaway, Jiri G. Safar

Taxonomy of breast cancer based on normal cell phenotype predicts outcome

• Text
• PDF

Abstract

Accurate classification is essential for understanding the pathophysiology of a disease and can inform therapeutic choices. For hematopoietic malignancies, a classification scheme based on the phenotypic similarity between tumor cells and normal cells has been successfully used to define tumor subtypes; however, use of normal cell types as a reference by which to classify solid tumors has not been widely emulated, in part due to more limited understanding of epithelial cell differentiation compared with hematopoiesis. To provide a better definition of the subtypes of epithelial cells comprising the breast epithelium, we performed a systematic analysis of a large set of breast epithelial markers in more than 15,000 normal breast cells, which identified 11 differentiation states for normal luminal cells. We then applied information from this analysis to classify human breast tumors based on normal cell types into 4 major subtypes, HR0–HR3, which were differentiated by vitamin D, androgen, and estrogen hormone receptor (HR) expression. Examination of 3,157 human breast tumors revealed that these HR subtypes were distinct from the current classification scheme, which is based on estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. Patient outcomes were best when tumors expressed all 3 hormone receptors (subtype HR3) and worst when they expressed none of the receptors (subtype HR0). Together, these data provide an ontological classification scheme associated with patient survival differences and provides actionable insights for treating breast tumors.

Authors

Sandro Santagata, Ankita Thakkar, Ayse Ergonul, Bin Wang, Terri Woo, Rong Hu, J. Chuck Harrell, George McNamara, Matthew Schwede, Aedin C. Culhane, David Kindelberger, Scott Rodig, Andrea Richardson, Stuart J. Schnitt, Rulla M. Tamimi, Tan A. Ince

Intramuscular viral delivery of paraplegin rescues peripheral axonopathy in a model of hereditary spastic paraplegia

• Text
• PDF

Abstract

Authors

Marinella Pirozzi, Angelo Quattrini, Gennaro Andolfi, Giorgia Dina, Maria Chiara Malaguti, Alberto Auricchio, Elena I. Rugarli