[show abstract][hide abstract] ABSTRACT: Bacterial-derived lipopolysaccharide (LPS) stimulates naive B lymphocytes to differentiate into immunoglobulin (Ig)-secreting plasma cells. Differentiation of B lymphocytes is characterized by a proliferative phase followed by expansion of the intracellular membrane secretory network to support Ig production. A key question in lymphocyte biology is how naive B cells reprogram metabolism to support de novo lipogenesis necessary for proliferation and expansion of the endomembrane network in response to LPS. We report that extracellularly acquired glucose is metabolized, in part, to support de novo lipogenesis in response to LPS stimulation of splenic B lymphocytes. LPS stimulation leads to increased levels of endogenous ATP-citrate lyase (ACLY) and this is accompanied by increased ACLY enzymatic activity. ACLY produces cytosolic acetyl-CoA from mitochondrial-derived citrate. Inhibition of ACLY activity in LPS-stimulated B cells with the selective inhibitor 2-hydroxy-N-arylbenzenesulfonamide (C-9) blocks glucose incorporation into de novo lipid biosynthesis, including cholesterol, free fatty acids, neutral and acidic phospholipids. Moreover, inhibition of ACLY activity in splenic B cells results in inhibition of proliferation and defective endomembrane expansion and reduced expression of CD138 and Blimp-1, markers for plasma-like B cell differentiation. ACLY activity is also required for LPS-induced IgM production in CH12 B-lymphoma cells. These data demonstrate that ACLY mediates glucose-dependent de novo lipogenesis in response to LPS signaling and identify a role for ACLY in several phenotypic changes that define plasma cell differentiation.
Journal of Biological Chemistry 01/2014; · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: Emerging evidence indicates that cancer is primarily a metabolic disease involving disturbances in energy production through respiration and fermentation. The genomic instability observed in tumor cells and all other recognized hallmarks of cancer are considered downstream epiphenomena of the initial disturbance of cellular energy metabolism. The disturbances in tumor cell energy metabolism can be linked to abnormalities in the structure and function of the mitochondria. When viewed as a mitochondrial metabolic disease, the evolutionary theory of Lamarck can better explain cancer progression than can the evolutionary theory of Darwin. Cancer growth and progression can be managed following a whole-body transition from fermentable metabolites, primarily glucose and glutamine, to respiratory metabolites, primarily ketone bodies. As each individual is a unique metabolic entity, personalization of metabolic therapy as a broad-based cancer treatment strategy will require fine-tuning to match the therapy to an individual(')s unique physiology.
[show abstract][hide abstract] ABSTRACT: Cardiolipin is a complex polyglycerol phospholipid found almost exclusively in the inner mitochondrial membrane and regulates numerous enzyme activities especially those related to oxidative phosphorylation and coupled respiration. Abnormalities in cardiolipin can impair mitochondrial function and bioenergetics. We recently demonstrated that the ratio of shorter chain saturated and monounsaturated fatty acids (C16:0; C18:0; C18:1) to longer chain polyunsaturated fatty acids (C18:2; C20:4; C22:6) was significantly greater in the brains of adult VM/DK (VM) inbred mice than in the brains of C57BL/6 J (B6) mice. The cardiolipin fatty acid abnormalities in VM mice are also associated with alterations in the activity of mitochondrial respiratory complexes. In this study we found that the abnormal brain fatty acid ratio in the VM strain was inherited as an autosomal dominant trait in reciprocal B6 × VM F1 hybrids. To evaluate the potential influence of brain cardiolipin fatty acid composition on cognitive sensitivity, we placed the parental B6 and VM mice and their reciprocal male and female B6VMF1 hybrid mice (3-month-old) in a hypoxic chamber (5 % O2). Cognitive awareness (conscientiousness) under hypoxia was significantly lower in the VM parental mice and F1 hybrid mice (11.4 ± 0.4 and 11.0 ± 0.4 min, respectively) than in the parental B6 mice (15.3 ± 1.4 min), indicating an autosomal dominant inheritance like that of the brain cardiolipin abnormalities. These findings suggest that impaired cognitive awareness under hypoxia is associated with abnormalities in neural lipid composition.
[show abstract][hide abstract] ABSTRACT: Salutary responses to adeno-associated viral (AAV) gene therapy have been reported in the mouse model of Sandhoff disease (SD), a neurodegenerative lysosomal storage disease caused by deficiency of β-N-acetylhexosaminidase (Hex). While untreated mice reach the humane endpoint by 4.1 months of age, mice treated by a single intracranial injection of vectors expressing human hexosaminidase may live a normal life span of 2 years. When treated with the same therapeutic vectors used in mice, two cats with SD lived to 7.0 and 8.2 months of age, compared with an untreated life span of 4.5 ± 0.5 months (n = 11). Because a pronounced humoral immune response to both the AAV1 vectors and human hexosaminidase was documented, feline cDNAs for the hexosaminidase α- and β-subunits were cloned into AAVrh8 vectors. Cats treated with vectors expressing feline hexosaminidase produced enzymatic activity >75-fold normal at the brain injection site with little evidence of an immune infiltrate. Affected cats treated with feline-specific vectors by bilateral injection of the thalamus lived to 10.4 ± 3.7 months of age (n = 3), or 2.3 times as long as untreated cats. These studies support the therapeutic potential of AAV vectors for SD and underscore the importance of species-specific cDNAs for translational research.Molecular Therapy (2013); doi:10.1038/mt.2013.86.
[show abstract][hide abstract] ABSTRACT: Sandhoff disease is an incurable neurodegenerative disorder caused by mutations in the lysosomal hydrolase β-hexosaminidase. Deficiency in this enzyme leads to excessive accumulation of ganglioside GM2 and its asialo derivative, GA2, in brain and visceral tissues. Small molecule inhibitors of ceramide-specific glucosyltransferase, the first committed step in ganglioside biosynthesis, reduce storage of GM2 and GA2. Limited brain access or adverse effects have hampered the therapeutic efficacy of the clinically approved substrate reduction molecules, eliglustat tartrate and the imino sugar NB-DNJ (Miglustat). The novel eliglustat tartrate analog, 2-(2,3-dihydro-1H-inden-2-yl)-N-((1R,2R)-1-(2,3-dihydrobenzo[b][1, 4]dioxin-6-yl)-1-hydroxy-3-(pyrrolidin-1-yl)propan-2-yl)acetamide (EtDO-PIP2, CCG-203586 or "3h"), was recently reported to reduce glucosylceramide in murine brain. Here we assessed the therapeutic efficacy of 3h in juvenile Sandhoff (Hexb-/-) mice. Sandhoff mice received intraperitoneal injections of phosphate buffered saline (PBS) or 3h (60 mg/kg/day) from postnatal day 9 (p-9) to postnatal day 15 (p-15). Brain weight and brain water content was similar in 3h and PBS-treated mice. 3h significantly reduced total ganglioside sialic acid, GM2, and GA2 content in cerebrum, cerebellum and liver of Sandhoff mice. Data from the liver showed that 3h reduced the key upstream ganglioside precursor (glucosylceramide), providing evidence for an on target mechanism of action. No significant differences were seen in the distribution of cholesterol or of neutral and acidic phospholipids. These data suggest that 3h can be an effective alternative to existing substrate reduction molecules for ganglioside storage diseases.
Neurochemical Research 02/2013; · 2.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: Metastasis involves the spread of cancer cells from the primary tumor to surrounding tissues and to distant organs and is the primary cause of cancer morbidity and mortality. In order to complete the metastatic cascade, cancer cells must detach from the primary tumor, intravasate into the circulatory and lymphatic systems, evade immune attack, extravasate at distant capillary beds, and invade and proliferate in distant organs. Currently, several hypotheses have been advanced to explain the origin of cancer metastasis. These involve an epithelial mesenchymal transition, an accumulation of mutations in stem cells, a macrophage facilitation process, and a macrophage origin involving either transformation or fusion hybridization with neoplastic cells. Many of the properties of metastatic cancer cells are also seen in normal macrophages. A macrophage origin of metastasis can also explain the long-standing "seed and soil" hypothesis and the absence of metastasis in plant cancers. The view of metastasis as a macrophage metabolic disease can provide novel insight for therapeutic management.
Critical reviews in oncogenesis 01/2013; 18(1-2):43-73.
[show abstract][hide abstract] ABSTRACT: Abnormal cancer metabolism creates a glycolytic-dependency which can be exploited by lowering glucose availability to the tumor. The ketogenic diet (KD) is a low carbohydrate, high fat diet which decreases blood glucose and elevates blood ketones and has been shown to slow cancer progression in animals and humans. Abnormal tumor vasculature creates hypoxic pockets which promote cancer progression and further increase the glycolytic-dependency of cancers. Hyperbaric oxygen therapy (HBO2T) saturates tumors with oxygen, reversing the cancer promoting effects of tumor hypoxia. Since these non-toxic therapies exploit overlapping metabolic deficiencies of cancer, we tested their combined effects on cancer progression in a natural model of metastatic disease.
We used the firefly luciferase-tagged VM-M3 mouse model of metastatic cancer to compare tumor progression and survival in mice fed standard or KD ad libitum with or without HBO2T (2.5 ATM absolute, 90 min, 3x/week). Tumor growth was monitored by in vivo bioluminescent imaging.
KD alone significantly decreased blood glucose, slowed tumor growth, and increased mean survival time by 56.7% in mice with systemic metastatic cancer. While HBO2T alone did not influence cancer progression, combining the KD with HBO2T elicited a significant decrease in blood glucose, tumor growth rate, and 77.9% increase in mean survival time compared to controls.
KD and HBO2T produce significant anti-cancer effects when combined in a natural model of systemic metastatic cancer. Our evidence suggests that these therapies should be further investigated as potential non-toxic treatments or adjuvant therapies to standard care for patients with systemic metastatic disease.
PLoS ONE 01/2013; 8(6):e65522. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: Sandhoff Disease (SD) involves the CNS accumulation of ganglioside GM2 and asialo-GM2 (GA2) due to inherited defects in the β-subunit gene of β-hexosaminidase A and B (Hexb gene). Substrate reduction therapy, utilizing imino sugar N-butyldeoxygalactonojirimycin (NB-DGJ), reduces ganglioside biosynthesis and levels of stored GM2 in SD mice. Intracranial transplantation of Neural Stem Cells (NSCs) can provide enzymatic cross correction, to help reduce ganglioside storage and extend life. Here we tested the effect of NSCs and NB-DGJ, alone and together, on brain β-hexosaminidase activity, GM2, and GA2 content in juvenile SD mice. The SD mice received either cerebral NSC transplantation at post-natal day 0 (p-0), intraperitoneal injection of NB-DGJ (500 mg/kg/day) from p-9 to p-15, or received dual treatments. The brains were analyzed at p-15. β-galactosidase staining confirmed engraftment of lacZ-expressing NSCs in the cerebral cortex. Compared to untreated and sham-treated SD controls, NSC treatment alone provided a slight increase in Hex activity and significantly decreased GA2 content. However, NSCs had no effect on GM2 content when analyzed at p-15. NB-DGJ alone had no effect on Hex activity, but significantly reduced GM2 and GA2 content. Hex activity was slightly elevated in the NSC + drug-treated mice. GM2 and GA2 content in the dual treated mice were similar to that of the NB-DGJ treated mice. These data indicate that NB-DGJ alone was more effective in targeting storage in juvenile SD mice than were NSCs alone. No additive or synergistic effect between NSC and drug was found in these juvenile SD mice.
Neurochemical Research 02/2012; 37(6):1335-43. · 2.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mature vasculature contains an endothelial cell lining with a surrounding sheath of pericytes/vascular smooth muscle cells (VSMCs). Tumor vessels are immature and lack a pericyte sheath. Colocalization of vascular endothelial growth factor receptor 2 (VEGFR-2) and platelet-derived growth factor receptor beta (PDGF-Rβ) reduces pericyte ensheathment of tumor vessels. We found that a 30% dietary restriction (DR) enhanced vessel maturation in the mouse CT-2A astrocytoma. DR reduced microvessel density and VEGF expression in the astrocytoma, while increasing recruitment of pericytes, positive for alpha-smooth muscle actin (α-SMA). Moreover, DR reduced colocalization of VEGF-R2 and PDGF-Rβ, but did not reduce total PDGF-Rβ expression. These findings suggest that DR promoted vessel normalization by preventing VEGF-induced inhibition of the PDGF signaling axis in pericytes. DR appears to shift the tumor vasculature from a leaky immature state to a more mature state. We suggest that vessel normalization could improve delivery of therapeutic drugs to brain tumors.
[show abstract][hide abstract] ABSTRACT: Itaconic acid (ITA), or methylenesuccinic acid, is not generally classified as a mammalian metabolite. Using NMR-based metabolomics and (13)C-labeling, we have detected ITA in both macrophage-like VM-M3 and RAW 264.7 tumor cell lines as well as stimulated and unstimulated primary murine macrophages. Macrophage activation by addition of lipopolysaccharide and IFN-γ markedly increased ITA production and secretion. Crude cell extracts synthesize ITA via decarboxylation of cis-aconitate, indicative of a novel mammalian cis-aconitic decarboxylase activity. Our results highlight a previously unidentified biosynthetic pathway related to TCA cycle metabolism in mammalian cells and a novel metabolite that likely plays a role in macrophage-based immune response.
Journal of the American Chemical Society 09/2011; 133(41):16386-9. · 10.68 Impact Factor
[show abstract][hide abstract] ABSTRACT: Malignant brain cancer persists as a major disease of morbidity and mortality. The failure to recognize brain cancer as a disease of energy metabolism has contributed in large part to the failure in management. As long as brain tumor cells have access to glucose and glutamine, the disease will progress. The current standard of care provides brain tumors with access to glucose and glutamine. The high fat low carbohydrate ketogenic diet (KD) will target glucose availability and possibly that of glutamine when administered in carefully restricted amounts to reduce total caloric intake and circulating levels of glucose. The restricted KD (RKD) targets major signaling pathways associated with glucose and glutamine metabolism including the IGF-1/PI3K/Akt/Hif pathway. The RKD is anti-angiogenic, anti-invasive, anti-inflammatory, and pro-apoptotic when evaluated in mice with malignant brain cancer. The therapeutic efficacy of the restricted KD can be enhanced when combined with drugs that also target glucose and glutamine. Therapeutic efficacy of the RKD was also seen against malignant gliomas in human case reports. Hence, the RKD can be an effective non-toxic therapeutic option to the current standard of care for inhibiting the growth and invasive properties of malignant brain cancer.
Epilepsy research 08/2011; 100(3):310-26. · 2.48 Impact Factor
[show abstract][hide abstract] ABSTRACT: Most malignant brain tumours contain various numbers of cells with characteristics of activated or dysmorphic macrophages/microglia. These cells are generally considered part of the tumour stroma and are often described as TAM (tumour-associated macrophages). These types of cells are thought to either enhance or inhibit brain tumour progression. Recent evidence indicates that neoplastic cells with macrophage characteristics are found in numerous metastatic cancers of non-CNS (central nervous system) origin. Evidence is presented here suggesting that subpopulations of cells within human gliomas, specifically GBM (glioblastoma multiforme), are neoplastic macrophages/microglia. These cells are thought to arise following mitochondrial damage in fusion hybrids between neoplastic stem cells and macrophages/microglia.
[show abstract][hide abstract] ABSTRACT: Filipin is an antibiotic polyene widely used as a histochemical marker for cholesterol. We previously reported cholesterol/filipin-positive staining in brain of β-galactosidase (β-gal) knockout ((-/-)) mice (GM1 gangliosidosis). The content and distribution of cholesterol and gangliosides was analyzed in plasma membrane (PM) and microsomal (MS) fractions from whole-brain tissue of 15 week-old control (β-gal(+/-)) and GM1 gangliosidosis (β-gal(-/-)) mice. Total ganglioside content (μg sialic acid/mg protein) was 3-fold and 7-fold greater in the PM and MS fractions, respectively, in βgal(-/-) mice than in βgal(+/-) mice. GM1 content was 30-fold and 50-fold greater in the PM and MS fractions, respectively. In contrast, unesterified cholesterol content (μg/mg protein) was similar in the PM and the MS fractions of the βgal(-/-) and βgal(+/-) mice. Filipin is known to bind to various sterol derivatives and phospholipids on thin-layer chromatograms. Biochemical evidence is presented showing that filipin also binds to GM1 with an affinity similar to that for cholesterol, with a corresponding fluorescent reaction. Our data suggest that the GM1 storage seen in the β-gal(-/-) mouse contributes to the filipin ultraviolet fluorescence observed in GM1 gangliosidosis brain. The data indicate that in addition to cholesterol, filipin can also be useful for detecting GM1.
The Journal of Lipid Research 07/2011; 52(7):1345-51. · 4.39 Impact Factor
[show abstract][hide abstract] ABSTRACT: Malignant brain tumors are a significant health problem in children and adults. Conventional therapeutic approaches have been largely unsuccessful in providing long-term management. As primarily a metabolic disease, malignant brain cancer can be managed through changes in metabolic environment. In contrast to normal neurons and glia, which readily transition to ketone bodies (β-hydroxybutyrate) for energy under reduced glucose, malignant brain tumors are strongly dependent on glycolysis for energy. The transition from glucose to ketone bodies as a major energy source is an evolutionary conserved adaptation to food deprivation that permits the survival of normal cells during extreme shifts in nutritional environment. Only those cells with a flexible genome and normal mitochondria can effectively transition from one energy state to another. Mutations restrict genomic and metabolic flexibility thus making tumor cells more vulnerable to energy stress than normal cells. We propose an alternative approach to brain cancer management that exploits the metabolic flexibility of normal cells at the expense of the genetically defective and metabolically challenged tumor cells. This approach to brain cancer management is supported from recent studies in mice and humans treated with calorie restriction and the ketogenic diet. Issues of implementation and use protocols are presented for the metabolic management of brain cancer.
Biochimica et Biophysica Acta 06/2011; 1807(6):577-94. · 4.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: Autism is a multifactorial disorder that involves impairments in social interactions and communication, as well as restricted and repetitive behaviors. About 30% of individuals with autism develop epilepsy by adulthood. The EL mouse has long been studied as a natural model of multifactorial idiopathic generalized epilepsy with complex partial seizures. Because epilepsy is a comorbid trait of autism, we evaluated the EL mouse for behaviors associated with autism.
We compared the behavior of EL mice to age-matched control DDY mice, a genetically related nonepileptic strain. The mice were compared in the open field and in the light-dark compartment tests to measure activity, exploratory behavior, and restricted and repetitive behaviors. The social transmission of food preference test was employed to evaluate social communication. Home-cage behavior was also evaluated in EL and DDY mice as a measure of repetitive activity.
We found that EL mice displayed several behavioral abnormalities characteristic of autism. Impairments in social interaction and restricted patterns of interest were evident in EL mice. Activity, exploratory behavior, and restricted behavior were significantly greater in EL mice than in DDY mice. EL mice exhibited impairment in the social transmission of food preference assay. In addition, a stereotypic myoclonic jumping behavior was observed in EL mice, but was not seen in DDY mice. It is of interest to note that seizure activity within 24 h of testing exacerbated the autistic behavioral abnormalities found in EL mice.
These findings suggest that the EL mouse expresses behavioral abnormalities similar to those seen in persons with autism. We propose that the EL mouse can be utilized as a natural model of autism and epilepsy.
[show abstract][hide abstract] ABSTRACT: Technical advances in lipidomic analysis have generated tremendous amounts of quantitative lipid molecular species data, whose value has not been fully explored. We describe a novel computational method to infer mechanisms of de novo lipid synthesis and remodeling from lipidomic data. We focus on the mitochondrial-specific lipid cardiolipin (CL), a polyglycerol phospholipid with four acyl chains. The lengths and degree of unsaturation of these acyl chains vary across CL molecules, and regulation of these differences is important for mitochondrial energy metabolism. We developed a novel mathematical approach to determine mechanisms controlling the steady-state distribution of acyl chain combinations in CL . We analyzed mitochondrial lipids from 18 types of steady-state samples, each with at least 3 replicates, from mouse brain, heart, lung, liver, tumor cells, and tumors grown in vitro. Using a mathematical model for the CL remodeling mechanisms and a maximum likelihood approach to infer parameters, we found that for most samples the four chain positions have an independent and identical distribution, indicating they are remodeled by the same processes. Furthermore, for most brain samples and liver, the distribution of acyl chains is well-fit by a simple linear combination of the pools of acyl chains in phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylglycerol (PG). This suggests that headgroup chemistry is the key determinant of acyl donation into CL, with chain length/saturation less important. This canonical remodeling behavior appears damaged in some tumor samples, which display a consistent excess of CL molecules having particular masses. For heart and lung, the "proportional incorporation" assumption is not adequate to explain the CL distribution, suggesting additional acyl CoA-dependent remodeling that is chain-type specific. Our findings indicate that CL remodeling processes can be described by a small set of quantitative relationships, and that bioinformatic approaches can help determine these processes from high-throughput lipidomic data.
PLoS ONE 01/2011; 6(6):e21170. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: Many of the current standard therapies employed for the management of primary malignant brain cancers are largely viewed as palliative, ultimately because these conventional strategies have been shown, in many instances, to decrease patient quality of life while only offering a modest increase in the length of survival. We propose that caloric restriction (CR) is an alternative metabolic therapy for brain cancer management that will not only improve survival but also reduce the morbidity associated with disease. Although we have shown that CR manages tumor growth and improves survival through multiple molecular and biochemical mechanisms, little information is known about the role that CR plays in modulating inflammation in brain tumor tissue.
Phosphorylation and activation of nuclear factor κB (NF-κB) results in the transactivation of many genes including those encoding cycloxygenase-2 (COX-2) and allograft inflammatory factor-1 (AIF-1), both of which are proteins that are primarily expressed by inflammatory and malignant cancer cells. COX-2 has been shown to enhance inflammation and promote tumor cell survival in both in vitro and in vivo studies. In the current report, we demonstrate that the p65 subunit of NF-κB was expressed constitutively in the CT-2A tumor compared with contra-lateral normal brain tissue, and we also show that CR reduces (i) the phosphorylation and degree of transcriptional activation of the NF-κB-dependent genes COX-2 and AIF-1 in tumor tissue, as well as (ii) the expression of proinflammatory markers lying downstream of NF-κB in the CT-2A malignant mouse astrocytoma, [e.g. macrophage inflammatory protein-2 (MIP-2)]. On the whole, our date indicate that the NF-κB inflammatory pathway is constitutively activated in the CT-2A astrocytoma and that CR targets this pathway and inflammation.
CR could be effective in reducing malignant brain tumor growth in part by inhibiting inflammation in the primary brain tumor.
PLoS ONE 01/2011; 6(3):e18085. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: Hurler syndrome (MPS IH) is caused by a mutation in the gene encoding alpha-L-iduronidase (IDUA) and leads to the accumulation of partially degraded glycosaminoglycans (GAGs). Ganglioside content is known to increase secondary to GAG accumulation. Most studies in organisms with MPS IH have focused on changes in gangliosides GM3 and GM2, without the study of other lipids. We evaluated the total lipid distribution in the whole brain and cerebellum of MPS IH (Idua⁻/⁻) and control (Idua(+/?)) mice at 6 months and at 12 months of age. The content of total sialic acid and levels of gangliosides GM3, GM2, and GD3 were greater in the whole brains of Idua⁻/⁻ mice then in Idua (+/?) mice at 12 months of age. No other significant lipid differences were found in either whole brain or in cerebellum at either age. The accumulation of ganglioside GD3 suggests that neurodegeneration occurs in the Idua⁻/⁻) mouse brain, but not to the extent seen in human MPS IH brain.
Neurochemical Research 01/2011; 36(9):1669-76. · 2.13 Impact Factor