Andrew Crowther

Hexokinase-2-mediated aerobic glycolysis is integral to cerebellar neurogenesis and pathogenesis of medulloblastoma

BackgroundWhile aerobic glycolysis is linked to unconstrained proliferation in cancer, less is known about its physiological role. Why this metabolic program that promotes tumor growth is preserved in the genome has thus been unresolved. We tested the hypothesis that aerobic glycolysis derives from developmental processes that regulate rapid proliferation.MethodsWe performed an integrated analysis of metabolism and gene expression in cerebellar granule neuron progenitors (CGNPs) with and without Sonic Hedgehog (Shh), their endogenous mitogen. Because our analysis highlighted Hexokinase-2 (Hk2) as a key metabolic regulator induced by Shh, we studied the effect of conditional genetic Hk2 deletion in CGNP development. We then crossed Hk2 conditional knockout mice with transgenic SmoM2 mice that develop spontaneous medulloblastoma and determined changes in SmoM2-driven tumorigenesis.ResultsWe show that Shh and phosphoinositide 3-kinase (PI3K) signaling combine to induce an Hk2-dependent glycolytic phenotype in CGNPs. This phenotype is recapitulated in medulloblastoma, a malignant tumor of CGNP origin. Importantly, cre-mediated ablation of Hk2 abrogated aerobic glycolysis, disrupting CGNP development and Smoothened-induced tumorigenesis. Comparing tumorigenesis in medulloblastoma-prone SmoM2 mice with and without functional Hk2, we demonstrate that loss of aerobic glycolysis reduces the aggressiveness of medulloblastoma, causing tumors to grow as indolent lesions and allowing long-term survival of tumor bearing mice.ConclusionsOur investigations demonstrate that aerobic glycolysis in cancer derives from developmental mechanisms that persist in tumorigenesis. Moreover, we demonstrate in a primary tumor model the anti-cancer potential of blocking aerobic glycolysis by targeting Hk2.See commentary article:http://www.biomedcentral.com/1741-7007/11/3

Resting-State Connectivity Predictors of Response to Psychotherapy in Major Depressive Disorder

Despite the heterogeneous symptom presentation and complex etiology of major depressive disorder (MDD), functional neuroimaging studies have shown with remarkable consistency that dysfunction in mesocorticolimbic brain systems are central to the disorder. Relatively less research has focused on the identification of biological markers of response to antidepressant treatment that would serve to improve the personalized delivery of empirically supported antidepressant interventions. In the present study, we investigated whether resting-state functional brain connectivity (rs-fcMRI) predicted response to Behavioral Activation Treatment for Depression, an empirically validated psychotherapy modality designed to increase engagement with rewarding stimuli and reduce avoidance behaviors. Twenty-three unmedicated outpatients with MDD and 20 matched nondepressed controls completed rs-fcMRI scans after which the MDD group received an average of 12 sessions of psychotherapy. The mean change in Beck Depression Inventory-II scores after psychotherapy was 12.04 points, a clinically meaningful response. Resting-state neuroimaging data were analyzed with a seed-based approach to investigate functional connectivity with four canonical resting-state networks: the default mode network, the dorsal attention network, the executive control network, and the salience network. At baseline, the MDD group was characterized by relative hyperconnectivity of multiple regions with precuneus, anterior insula, dorsal anterior cingulate cortex (dACC), and left dorsolateral prefrontal cortex seeds and by relative hypoconnectivity with intraparietal sulcus, anterior insula, and dACC seeds. Additionally, connectivity of the precuneus with the left middle temporal gyrus and connectivity of the dACC with the parahippocampal gyrus predicted the magnitude of pretreatment MDD symptoms. Hierarchical linear modeling revealed that response to psychotherapy in the MDD group was predicted by pretreatment connectivity of the right insula with the right middle temporal gyrus and the left intraparietal sulcus with the orbital frontal cortex. These results add to the nascent body of literature investigating pretreatment rs-fcMRI predictors of antidepressant treatment response and is the first study to examine rs-fcMRI predictors of response to psychotherapy.

Essential Function of Dicer in Resolving DNA Damage in the Rapidly Dividing Cells of the Developing and Malignant Cerebellum.

Maintenance of genomic integrity is critical during neurodevelopment, particularly in rapidly dividing cerebellar granule neuronal precursors that experience constitutive replication-associated DNA damage. As Dicer was recently recognized to have an unexpected function in the DNA damage response, we examined whether Dicer was important for preserving genomic integrity in the developing brain. We report that deletion of Dicer in the developing mouse cerebellum resulted in the accumulation of DNA damage leading to cerebellar progenitor degeneration, which was rescued with p53 deficiency; deletion of DGCR8 also resulted in similar DNA damage and cerebellar degeneration. Dicer deficiency also resulted in DNA damage and death in other rapidly dividing cells including embryonic stem cells and the malignant cerebellar progenitors in a mouse model of medulloblastoma. Together, these results identify an essential function of Dicer in resolving the spontaneous DNA damage that occurs during the rapid proliferation of developmental progenitors and malignant cells.

Bax deficiency prolongs cerebellar neurogenesis, accelerates medulloblastoma formation and paradoxically increases both malignancy and differentiation

Neurogenesis requires negative regulation through differentiation of progenitors or their programmed cell death (PCD). Growth regulation is particularly important in the postnatal cerebellum, where excessive progenitor proliferation promotes medulloblastoma, the most common malignant brain tumor in children. We present evidence that PCD operates alongside differentiation to regulate cerebellar granule neuron progenitors (CGNPs) and to prevent medulloblastoma. Here, we show that genetic deletion of pro-apoptotic Bax disrupts regulation of cerebellar neurogenesis and promotes medulloblastoma formation. In Bax−/− mice, the period of neurogenesis was extended into the third week of postnatal life, and ectopic neurons and progenitors collected in the molecular layer of the cerebellum and adjacent tectum. Importantly, genetic deletion of Bax in medulloblastoma-prone ND2:SmoA1 transgenic mice greatly accelerated tumorigenesis. Bax-deficient medulloblastomas exhibited strikingly distinct pathology, with reduced apoptosis, increased neural differentiation and tectal migration. Comparing Bax+/+ and Bax−/− medulloblastomas, we were able to identify upregulation of Bcl-2 and nuclear exclusion of p27 as tumorigenic changes that are required to mitigate the tumor suppressive effect of Bax. Studies on human tumors confirmed the importance of modulating Bax in medulloblastoma pathogenesis. Our results demonstrate that Bax-dependent apoptosis regulates postnatal cerebellar neurogenesis, suppresses medulloblastoma formation and imposes selective pressure on tumors that form. Functional resistance to Bax-mediated apoptosis, required for medulloblastoma tumorigenesis, may be a tumor-specific vulnerability to be exploited for therapeutic benefit.

Sustained anterior cingulate cortex activation during reward processing predicts response to psychotherapy in major depressive disorder

BACKGROUND The purpose of the present investigation was to evaluate whether pre-treatment neural activation in response to rewards is a predictor of clinical response to Behavioral Activation Therapy for Depression (BATD), an empirically validated psychotherapy that decreases depressive symptoms by increasing engagement with rewarding stimuli and reducing avoidance behaviors. METHODS Participants were 33 outpatients with major depressive disorder (MDD) and 20 matched controls. We examined group differences in activation, and the capacity to sustain activation, across task runs using functional magnetic resonance imaging (fMRI) and the monetary incentive delay (MID) task. Hierarchical linear modeling was used to investigate whether pre-treatment neural responses predicted change in depressive symptoms over the course of BATD treatment. RESULT MDD and Control groups differed in sustained activation during reward outcomes in the right nucleus accumbens, such that the MDD group experienced a significant decrease in activation in this region from the first to second task run relative to controls. Pretreatment anhedonia severity and pretreatment task-related reaction times were predictive of response to treatment. Furthermore, sustained activation in the anterior cingulate cortex during reward outcomes predicted response to psychotherapy; patients with greater sustained activation in this region were more responsive to BATD treatment. LIMITATION The current study only included a single treatment condition, thus it unknown whether these predictors of treatment response are specific to BATD or psychotherapy in general. CONCLUSION Findings add to the growing body of literature suggesting that the capacity to sustain neural responses to rewards may be a critical endophenotype of MDD.

Aspm sustains postnatal cerebellar neurogenesis and medulloblastoma growth in mice

Alterations in genes that regulate brain size may contribute to both microcephaly and brain tumor formation. Here, we report that Aspm, a gene that is mutated in familial microcephaly, regulates postnatal neurogenesis in the cerebellum and supports the growth of medulloblastoma, the most common malignant pediatric brain tumor. Cerebellar granule neuron progenitors (CGNPs) express Aspm when maintained in a proliferative state by sonic hedgehog (Shh) signaling, and Aspm is expressed in Shh-driven medulloblastoma in mice. Genetic deletion of Aspm reduces cerebellar growth, while paradoxically increasing the mitotic rate of CGNPs. Aspm-deficient CGNPs show impaired mitotic progression, altered patterns of division orientation and differentiation, and increased DNA damage, which causes progenitor attrition through apoptosis. Deletion of Aspm in mice with Smo-induced medulloblastoma reduces tumor growth and increases DNA damage. Co-deletion of Aspm and either of the apoptosis regulators Bax or Trp53 (also known as p53) rescues the survival of neural progenitors and reduces the growth restriction imposed by Aspm deletion. Our data show that Aspm functions to regulate mitosis and to mitigate DNA damage during CGNP cell division, causes microcephaly through progenitor apoptosis when mutated, and sustains tumor growth in medulloblastoma. Summary: In the mouse cerebellum, Aspm depletion causes increased DNA damage and impaired mitosis, leading to granule cell progenitor apoptosis. In medulloblastoma, Aspm sustains tumor growth.

Attenuation of Frontostriatal Connectivity During Reward Processing Predicts Response to Psychotherapy in Major Depressive Disorder

There are few reliable predictors of response to antidepressant treatments. In the present investigation, we examined pretreatment functional brain connectivity during reward processing as a potential predictor of response to Behavioral Activation Treatment for Depression (BATD), a validated psychotherapy that promotes engagement with rewarding stimuli and reduces avoidance behaviors. Thirty-three outpatients with major depressive disorder (MDD) and 20 matched controls completed two runs of the monetary incentive delay task during functional magnetic resonance imaging after which participants with MDD received up to 15 sessions of BATD. Seed-based generalized psychophysiological interaction analyses focused on task-based connectivity across task runs, as well as the attenuation of connectivity from the first to the second run of the task. The average change in Beck Depression Inventory-II scores due to treatment was 10.54 points, a clinically meaningful response. Groups differed in seed-based functional connectivity among multiple frontostriatal regions. Hierarchical linear modeling revealed that improved treatment response to BATD was predicted by greater connectivity between the left putamen and paracingulate gyrus during reward anticipation. In addition, MDD participants with greater attenuation of connectivity between several frontostriatal seeds, and midline subcallosal cortex and left paracingulate gyrus demonstrated improved response to BATD. These findings indicate that pretreatment frontostriatal functional connectivity during reward processing is predictive of response to a psychotherapy modality that promotes improving approach-related behaviors in MDD. Furthermore, connectivity attenuation among reward-processing regions may be a particularly powerful endophenotypic predictor of response to BATD in MDD.

Activity-dependent signaling mechanisms regulating adult hippocampal neural stem cells and their progeny

Adult neural stem cells (NSCs) reside in a restricted microenvironment, where their development is controlled by subtle and presently underexplored cues. This raises a significant question: what instructions must be provided by this supporting niche to regulate NSC development and functions? Signaling from the niche is proposed to control many aspects of NSC behavior, including balancing the quiescence and proliferation of NSCs, determining the cell division mode (symmetric versus asymmetric), and preventing premature depletion of stem cells to maintain neurogenesis throughout life. Interactions between neurogenic niches and NSCs also govern the homeostatic regulation of adult neurogenesis under diverse physiological, environmental, and pathological conditions. An important implication from revisiting many previously-identifi ed regulatory factors is that most of them (e.g., the antidepressant fluoxetine and exercise) affect gross neurogenesis by acting downstream of NSCs at the level of intermediate progenitors and neuroblasts, while leaving the NSC pool unaffected. Therefore, it is critically important to address how various niche components, signaling pathways, and environmental stimuli differentially regulate distinct stages of adult neurogenesis.

Glymphatic fluid transport controls paravascular clearance of AAV vectors from the brain

Adeno-associated viruses (AAV) are currently being evaluated in clinical trials for gene therapy of CNS disorders. However, host factors that influence the spread, clearance, and transduction efficiency of AAV vectors in the brain are not well understood. Recent studies have demonstrated that fluid flow mediated by aquaporin-4 (AQP4) channels located on astroglial end feet is essential for exchange of solutes between interstitial and cerebrospinal fluid. This phenomenon, which is essential for interstitial clearance of solutes from the CNS, has been termed glial-associated lymphatic transport or glymphatic transport. In the current study, we demonstrate that glymphatic transport profoundly affects various aspects of AAV gene transfer in the CNS. Altered localization of AQP4 in aged mouse brains correlated with significantly increased retention of AAV vectors in the parenchyma and reduced systemic leakage following ventricular administration. We observed a similar increase in AAV retention and transgene expression upon i.c.v. administration in AQP4-/- mice. Consistent with this observation, fluorophore-labeled AAV vectors showed markedly reduced flux from the ventricles of AQP4-/- mice compared with WT mice. These results were further corroborated by reduced AAV clearance from the AQP4-null brain, as demonstrated by reduced transgene expression and vector genome accumulation in systemic organs. We postulate that deregulation of glymphatic transport in aged and diseased brains could markedly affect the parenchymal spread, clearance, and gene transfer efficiency of AAV vectors. Assessment of biomarkers that report the kinetics of CSF flux in prospective gene therapy patients might inform variable treatment outcomes and guide future clinical trial design.

Tonic Activation of Bax Primes Neural Progenitors for Rapid Apoptosis through a Mechanism Preserved in Medulloblastoma

Commitment to survival or apoptosis within expanding progenitor populations poses distinct risks and benefits to the organism. We investigated whether specialized mechanisms regulate apoptosis in mouse neural progenitors and in the progenitor-derived brain tumor medulloblastoma. Here, we identified constitutive activation of proapoptotic Bax, maintained in check by Bcl-xL, as a mechanism for rapid cell death, common to postnatal neural progenitors and medulloblastoma. We found that tonic activation of Bax in cerebellar progenitors, along with sensitivity to DNA damage, was linked to differentiation state. In cerebellar progenitors, active Bax localized to mitochondria, where it was bound to Bcl-xL. Disruption of Bax:Bcl-xL binding by BH3-mimetic ABT 737 caused rapid apoptosis of cerebellar progenitors and primary murine medulloblastoma cells. Conditional deletion of Mcl-1, in contrast, did not cause death of cerebellar progenitors. Our findings identify a mechanism for the sensitivity of brain progenitors to typical anticancer therapies and reveal that this mechanism persists in medulloblastoma, a malignant brain tumor markedly sensitive to radiation and chemotherapy.