Antidepressant drugs and risk of developing glioma: a national registry-based case control study and a meta-analysis.

The aim of the present study was to investigate if use of antidepressants is related to the risk of developing lower (WHO grade 2-3) and higher grade (WHO grade 4) glioma. A registry-based case-control study was performed using 1283 glioma cases and 6400 age-, sex- and geographically matched controls, diagnosed in Sweden 2009-2013. Conditional logistic regression was used to analyze whether Selective Serotonin Reuptake Inhibitors (SSRIs) or non-SSRIs were associated with the risk of developing lower- or higher-grade glioma in the study population. Our results show that use of antidepressant medication was not associated with the risk of developing glioma. We also performed a meta-analysis in which the dataset from the present study was combined with results from two previous epidemiological studies to answer the same questions. The meta-analysis showed a modest risk reduction of developing glioma in relation to antidepressant treatment (OR 0.90 [95% CI 0.83-0.97]), when all glioma subgroups and all forms of antidepressant medications were combined. In conclusion, it remains possible that antidepressants may have common monoaminergic mechanism(s) that reduce the risk of developing glioma.

Metabolic response patterns in brain microdialysis fluids and serum during interstitial cisplatin treatment of high-grade glioma.

BACKGROUND: High-grade gliomas are associated with poor prognosis. Tumour heterogeneity and invasiveness create challenges for effective treatment and use of systemically administrated drugs. Furthermore, lack of functional predictive response-assays based on drug efficacy complicates evaluation of early treatment responses. METHODS: We used microdialysis to deliver cisplatin into the tumour and to monitor levels of metabolic compounds present in the tumour and non-malignant brain tissue adjacent to tumour, before and during treatment. In parallel, we collected serum samples and used multivariate statistics to analyse the metabolic effects. RESULTS: We found distinct metabolic patterns in the extracellular fluids from tumour compared to non-malignant brain tissue, including high concentrations of a wide range of amino acids, amino acid derivatives and reduced levels of monosaccharides and purine nucleosides. We found that locoregional cisplatin delivery had a strong metabolic effect at the tumour site, resulting in substantial release of glutamic acid, phosphate, and spermidine and a reduction of cysteine levels. In addition, patients with long-time survival displayed different treatment response patterns in both tumour and serum. Longer survival was associated with low tumour levels of lactic acid, glyceric acid, ketoses, creatinine and cysteine. Patients with longer survival displayed lower serum levels of ketohexoses, fatty acid methyl esters, glycerol-3-phosphate and alpha-tocopherol, while elevated phosphate levels were seen in both tumour and serum during treatment. CONCLUSION: We highlight distinct metabolic patterns associated with high-grade tumour metabolism, and responses to cytotoxic cisplatin treatment.

DJ-1 is a redox sensitive adapter protein for high molecular weight complexes involved in regulation of catecholamine homeostasis.

DJ-1 is an oxidation sensitive protein encoded by the PARK7 gene. Mutations in PARK7 are a rare cause of familial recessive Parkinson's disease (PD), but growing evidence suggests involvement of DJ-1 in idiopathic PD. The key clinical features of PD, rigidity and bradykinesia, result from neurotransmitter imbalance, particularly the catecholamines dopamine (DA) and noradrenaline. We report in human brain and human SH-SY5Y neuroblastoma cell lines that DJ-1 predominantly forms high molecular weight (HMW) complexes that included RNA metabolism proteins hnRNPA1 and PABP1 and the glycolysis enzyme GAPDH. In cell culture models the oxidation status of DJ-1 determined the specific complex composition. RNA sequencing indicated that oxidative changes to DJ-1 were concomitant with changes in mRNA transcripts mainly involved in catecholamine metabolism. Importantly, loss of DJ-1 function upon knock down (KD) or expression of the PD associated form L166P resulted in the absence of HMW DJ-1 complexes. In the KD model, the absence of DJ-1 complexes was accompanied by impairment in catecholamine homeostasis, with significant increases in intracellular DA and noraderenaline levels. These changes in catecholamines could be rescued by re-expression of DJ-1. This catecholamine imbalance may contribute to the particular vulnerability of dopaminergic and noradrenergic neurons to neurodegeneration in PARK7-related PD. Notably, oxidised DJ-1 was significantly decreased in idiopathic PD brain, suggesting altered complex function may also play a role in the more common sporadic form of the disease.

Pre-diagnostic serum levels of EGFR and ErbB2 and genetic glioma risk variants: a nested case-control study.

Genetic variants have been associated with the risk of developing glioma, but functional mechanisms on disease phenotypic traits remain to be investigated. One phenotypic trait of glioblastoma is the mutation and amplification of the epidermal growth factor receptor (EGFR) gene. We investigated associations between pre-diagnostic serum protein concentrations of EGFR and ErbB2, both members of the EGFR family, and future risk of glioma. Further, we studied if EGFR glioma risk variants were associated with EGFR and ErbB2 serum levels. We assessed the associations between genetic glioma risk variants and serum concentrations of EGFR and ErbB2, as measured in pre-diagnostic cohort serum samples of 593 glioma patients and 590 matched cancer-free controls. High serum EGFR and ErbB2 levels were associated with risk of developing glioblastoma (P = 0.008; OR = 1.58, 95 % CI = 1.13-2.22 and P = 0.017, OR = 1.63, 95 % CI = 1.09-2.44, respectively). High serum ErbB2 concentration was also associated with glioma risk overall (P = 0.049; OR = 1.39, 95 % CI = 1.00-1.93). Glioma risk variants were not associated with high serum protein abundance. In contrast, the EGFR risk variant rs4947986 (T) was correlated with decreased EGFR serum levels (study cohort P = 0.024 and controls P = 0.009). To our knowledge, this is the first study showing an association of EGFR and ErbB2 serum levels with glioma more than a decade before diagnosis, indicating that EGFR and ErbB2 serum proteins are important in early gliomagenesis. However, we did not find evidence that glioma risk variants were associated with high pre-diagnostic serum concentrations of EGFR and ErbB2.

Parkinson disease protein DJ-1 binds metals and protects against metal-induced cytotoxicity.

The progressive loss of motor control due to reduction of dopamine-producing neurons in the substantia nigra pars compacta and decreased striatal dopamine levels are the classically described features of Parkinson disease (PD). Neuronal damage also progresses to other regions of the brain, and additional non-motor dysfunctions are common. Accumulation of environmental toxins, such as pesticides and metals, are suggested risk factors for the development of typical late onset PD, although genetic factors seem to be substantial in early onset cases. Mutations of DJ-1 are known to cause a form of recessive early onset Parkinson disease, highlighting an important functional role for DJ-1 in early disease prevention. This study identifies human DJ-1 as a metal-binding protein able to evidently bind copper as well as toxic mercury ions in vitro. The study further characterizes the cytoprotective function of DJ-1 and PD-mutated variants of DJ-1 with respect to induced metal cytotoxicity. The results show that expression of DJ-1 enhances the cells' protective mechanisms against induced metal toxicity and that this protection is lost for DJ-1 PD mutations A104T and D149A. The study also shows that oxidation site-mutated DJ-1 C106A retains its ability to protect cells. We also show that concomitant addition of dopamine exposure sensitizes cells to metal-induced cytotoxicity. We also confirm that redox-active dopamine adducts enhance metal-catalyzed oxidation of intracellular proteins in vivo by use of live cell imaging of redox-sensitive S3roGFP. The study indicates that even a small genetic alteration can sensitize cells to metal-induced cell death, a finding that may revive the interest in exogenous factors in the etiology of PD.

Blood based metabolic markers of glioma from pre-diagnosis to surgery.

Gliomas are highly complex and metabolically active brain tumors associated with poor prognosis. Recent reports have found altered levels of blood metabolites during early tumor development, suggesting that tumor development could be detected several years before clinical manifestation. In this study, we performed metabolite analyses of blood samples collected from healthy controls and future glioma patients, up to eight years before glioma diagnosis, and on the day of glioma surgery. We discovered that metabolites related to early glioma development were associated with an increased energy turnover, as highlighted by elevated levels of TCA-related metabolites such as fumarate, malate, lactate and pyruvate in pre-diagnostic cases. We also found that metabolites related to glioma progression at surgery were primarily high levels of amino acids and metabolites of amino acid catabolism, with elevated levels of 11 amino acids and two branched-chain alpha-ketoacids, ketoleucine and ketoisoleucine. High amino acid turnover in glioma tumor tissue is currently utilized for PET imaging, diagnosis and delineation of tumor margins. By examining blood-based metabolic progression patterns towards disease onset, we demonstrate that this high amino acid turnover is also detectable in a simple blood sample. These findings provide additional insight of metabolic alterations during glioma development and progression.

Antidepressants and survival in glioma-A registry-based retrospective cohort study.

Background: Depression and treatment with antidepressant medication is common in patients with malignant glioma. However, the extent to which antidepressants may affect the disease is not fully understood. Therefore, the purpose of the present study was to investigate possible associations between treatment with antidepressant medication and survival in glioma patients. Methods: We performed a registry-based cohort study including 1231 patients with malignant glioma (WHO grades 2, 3, and 4) having undergone surgery, and 6400 matched controls without glioma. All data were extracted from the RISK North database, which contains information from multiple national population-based registries in Sweden. Results: Treatment with antidepressants is more common in patients with malignant glioma (27%), compared to controls (16%), P < .001. Treatment with antidepressants after surgery for glioma was significantly associated with poorer survival. These effects were observed both for selective serotonin reuptake inhibitors (SSRIs) and non-SSRIs. In grade 4 glioma, SSRI treatment was associated with a hazard ratio (HR) of 3.32 (95% CI 2.69-4.10, P < .001), and non-SSRI treatment a HR of 3.54 (95% CI 2.52-4.99, P < .001), compared to glioma patients without antidepressants. In grade 2-3 glioma, the HR for SSRI treatment was 3.26 (95% CI 2.19-4.85, P < .001), and for non-SSRI treatment was 7.71 (95% CI 4.22-14.12, P < .001). Conclusions: Our results demonstrate a negative association between antidepressant medication and survival in glioma. Further research will be needed to clarify causation.

LAPTM4B counteracts ferroptosis via suppressing the ubiquitin-proteasome degradation of SLC7A11 in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related deaths worldwide, necessitating the identification of novel therapeutic targets. Lysosome Associated Protein Transmembrane 4B (LAPTM4B) is involved in biological processes critical to cancer progression, such as regulation of solute carrier transporter proteins and metabolic pathways, including mTORC1. However, the metabolic processes governed by LAPTM4B and its role in oncogenesis remain unknown. In this study, we conducted unbiased metabolomic screens to uncover the metabolic landscape regulated by LAPTM4B. We observed common metabolic changes in several knockout cell models suggesting of a role for LAPTM4B in suppressing ferroptosis. Through a series of cell-based assays and animal experiments, we demonstrate that LAPTM4B protects tumor cells from erastin-induced ferroptosis both in vitro and in vivo. Mechanistically, LAPTM4B suppresses ferroptosis by inhibiting NEDD4L/ZRANB1 mediated ubiquitination and subsequent proteasomal degradation of the cystine-glutamate antiporter SLC7A11. Furthermore, metabolomic profiling of cancer cells revealed that LAPTM4B knockout leads to a significant enrichment of ferroptosis and associated metabolic alterations. By integrating results from cellular assays, patient tissue samples, an animal model, and cancer databases, this study highlights the clinical relevance of the LAPTM4B-SLC7A11-ferroptosis signaling axis in NSCLC progression and identifies it as a potential target for the development of cancer therapeutics.

Low-grade glioma risk SNP rs11706832 is associated with type I interferon response pathway genes in cell lines.

Genome-wide association studies (GWAS) have contributed to our understanding of glioma susceptibility. To date, 25 risk loci for development of any of the glioma subtypes are known. However, GWAS studies reveal little about the molecular processes that lead to increased risk, especially for non-coding single nucleotide polymorphisms (SNP). A particular SNP in intron 2 of LRIG1, rs11706832, has been shown to increase the susceptibility for IDH1 mutated low-grade gliomas (LGG). Leucine-rich repeats and immunoglobulin-like domains protein 1 (LRIG1) is important in cancer development as it negatively regulates the epidermal growth factor receptor (EGFR); however, the mechanism responsible for this particular risk SNP and its potential effect on LRIG1 are not known. Using CRISPR-CAS9, we edited rs11706832 in HEK293T cells. Four HEK293T clones with the risk allele were compared to four clones with the non-risk allele for LRIG1 and SLC25A26 gene expression using RT-qPCR, for global gene expression using RNA-seq, and for metabolites using gas chromatography-mass spectrometry (GC-MS). The experiment did not reveal any significant effect of the SNP on the expression levels or splicing patterns of LRIG1 or SLC25A26. The global gene expression analysis revealed that the risk allele C was associated with upregulation of several mitochondrial genes. Gene enrichment analysis of 74 differentially expressed genes in the genome revealed a significant enrichment of type I interferon response genes, where many genes were downregulated for the risk allele C. Gene expression data of IDH1 mutated LGGs from the cancer genome atlas (TCGA) revealed a similar under expression of type I interferon genes associated with the risk allele. This study found the expression levels and splicing patterns of LRIG1 and SLC25A26 were not affected by the SNP in HEK293T cells. However, the risk allele was associated with a downregulation of genes involved in the innate immune response both in the HEK293T cells and in the LGG data from TCGA.

Altered plasma metabolite levels can be detected years before a glioma diagnosis.

Genetic and metabolic changes in tissue and blood are reported to occur several years before glioma diagnosis. Since gliomas are currently detected late, a liquid biopsy for early detection could affect the quality of life and prognosis of patients. Here, we present a nested case-control study of 550 prediagnostic glioma cases and 550 healthy controls from the Northern Sweden Health and Disease study (NSHDS) and the European Prospective Investigation into Cancer and Nutrition (EPIC) study. We identified 93 significantly altered metabolites related to glioma development up to 8 years before diagnosis. Out of these metabolites, a panel of 20 selected metabolites showed strong disease correlation and a consistent progression pattern toward diagnosis in both the NSHDS and EPIC cohorts, and they separated future cases from controls independently of biological sex. The blood metabolite panel also successfully separated both lower-grade glioma and glioblastoma cases from controls, up to 8 years before diagnosis in patients within the NSHDS cohort and up to 2 years before diagnosis in EPIC. Pathway enrichment analysis detected metabolites related to the TCA cycle, Warburg effect, gluconeogenesis, and cysteine, pyruvate, and tyrosine metabolism as the most affected.

The Arabidopsis DJ-1a protein confers stress protection through cytosolic SOD activation.

Mutations in the DJ-1 gene (also known as PARK7) cause inherited Parkinson's disease, which is characterized by neuronal death. Although DJ-1 is thought to be an antioxidant protein, the underlying mechanism by which loss of DJ-1 function contributes to cell death is unclear. Human DJ-1 and its Arabidopsis thaliana homologue, AtDJ-1a, are evolutionarily conserved proteins, indicating a universal function. To gain further knowledge of the molecular features associated with DJ-1 dysfunction, we have characterized AtDJ-1a. We show that AtDJ-1a levels are responsive to stress treatment and that AtDJ-1a loss of function results in accelerated cell death in aging plants. By contrast, transgenic plants with elevated AtDJ-1a levels have increased protection against environmental stress conditions, such as strong light, H(2)O(2), methyl viologen and copper sulfate. We further identify superoxide dismutase 1 (SOD1) and glutathione peroxidase 2 (GPX2) as interaction partners of both AtDJ-1a and human DJ-1, and show that this interaction results in AtDJ-1a- and DJ-1-mediated cytosolic SOD1 activation in a copper-dependent fashion. Our data have highlighted a conserved molecular mechanism for DJ-1 and revealed a new protein player in the oxidative stress response of plants.

The prognostic value of MARCKS-like 1 in lymph node-negative breast cancer.

There is a need for new biomarkers to more correctly identify node-negative breast cancer patients with a good or bad prognosis. Myristoylated alanine-rich C kinase substrate like-1 (MARCKSL1) is a membrane-bound protein that is associated with cell spreading, integrin activation and exocytosis. Three hundred and five operable T(1,2)N(0)M(0) lymph node-negative breast cancer patients (median follow-up time 121 months, range 10-178 months) were evaluated for MARCKSL1 expression by immunohistochemistry and quantitative real-time PCR. The results were compared with classical prognosticators (age, tumor diameter, grade, estrogen receptor, and proliferation), using single (Kaplan-Meier) and multivariate survival analysis (Cox model). Forty-seven patients (15 %) developed distant metastases. With single and multivariate analysis of all features, MARCKSL1 protein expression was the strongest prognosticator (P < 0.001, HR = 5.1, 95 % CI = 2.7-9.8). Patients with high MARCKSL1 expression (n = 23) showed a 44 % survival versus 88 % in patients with low expression at 15-year follow-up. mRNA expression of MARCKSL1 in formalin fixed paraffin-embedded tissue was also prognostic (P = 0.002, HR = 3.6, 95 % CI = 1.5-8.3). However, the prognostic effect of high and low was opposite from the protein expression, i.e., low expression (relative expression ≤ 0.0264, n = 76) showed a 79 % survival versus 92 % in those with high expression of MARCKSL1 mRNA. Multivariate analysis of all features with distant metastases free survival as the end-point showed that the combination of MARCKSL1 protein and phosphohistone H3 (PPH3) has the strongest independent prognostic value. Patients with high expression (≥13) of PPH3 and high MARCKSL1 protein had 45 % survival versus 78 % survival for patients with low MARCKSL1 protein expression and high expression (≥13) of PPH3. In conclusion, MARCKSL1 has strong prognostic value in lymph node-negative breast cancer patients, especially in those with high proliferation.

Pre-diagnostic levels of sVEGFR2, sTNFR2, sIL-2Rα and sIL-6R are associated with glioma risk: A nested case-control study of repeated samples.

No strong aetiological factors have been established for glioma aside from genetic mutations and variants, ionising radiation and an inverse relationship with asthmas and allergies. Our aim was to investigate the association between pre-diagnostic immune protein levels and glioma risk. We conducted a case-control study nested in the Northern Sweden Health and Disease Study cohort. We analysed 133 glioma cases and 133 control subjects matched by age, sex and date of blood donation. ELISA or Luminex bead-based multiplex assays were used to measure plasma levels of 19 proteins. Conditional logistic regression models were used to estimate the odds ratios and 95% CIs. To further model the protein trajectories over time, the linear mixed-effects models were conducted. We found that the levels of sVEGFR2, sTNFR2, sIL-2Rα and sIL-6R were associated with glioma risk. After adjusting for the time between blood sample collection and glioma diagnosis, the odds ratios were 1.72 (95% CI = 1.01-2.93), 1.48 (95% CI = 1.01-2.16) and 1.90 (95% CI = 1.14-3.17) for sTNFR2, sIL-2Rα and sIL-6R, respectively. The trajectory of sVEGFR2 concentrations over time was different between cases and controls (p-value = 0.031), increasing for cases (0.8% per year) and constant for controls. Our findings suggest these proteins play important roles in gliomagenesis.

Prediagnostic biomarkers for early detection of glioma-using case-control studies from cohorts as study approach.

Background: Understanding the trajectory and development of disease is important and the knowledge can be used to find novel targets for therapy and new diagnostic tools for early diagnosis. Methods: Large cohorts from different parts of the world are unique assets for research as they have systematically collected plasma and DNA over long-time periods in healthy individuals, sometimes even with repeated samples. Over time, the population in the cohort are diagnosed with many different diseases, including brain tumors. Results: Recent studies have detected genetic variants that are associated with increased risk of glioblastoma and lower grade gliomas specifically. The impact for genetic markers to predict disease in a healthy population has been deemed low, and a relevant question is if the genetic variants for glioma are associated with risk of disease or partly consist of genes associated to survival. Both metabolite and protein spectra are currently being explored for early detection of cancer. Conclusions: We here present a focused review of studies of genetic variants, metabolomics, and proteomics studied in prediagnostic glioma samples and discuss their potential in early diagnostics.

Distinct metabolic hallmarks of WHO classified adult glioma subtypes.

BACKGROUND: Gliomas are complex tumors with several genetic aberrations and diverse metabolic programs contributing to their aggressive phenotypes and poor prognoses. This study defines key metabolic features that can be used to differentiate between glioma subtypes, with potential for improved diagnostics and subtype targeted therapy. METHODS: Cross-platform global metabolomic profiling coupled with clinical, genetic, and pathological analysis of glioma tissue from 224 tumors-oligodendroglioma (n = 31), astrocytoma (n = 31) and glioblastoma (n = 162)-were performed. Identified metabolic phenotypes were evaluated in accordance with the WHO classification, IDH-mutation, 1p/19q-codeletion, WHO-grading 2-4, and MGMT promoter methylation. RESULTS: Distinct metabolic phenotypes separate all six analyzed glioma subtypes. IDH-mutated subtypes, expressing 2-hydroxyglutaric acid, were clearly distinguished from IDH-wildtype subtypes. Considerable metabolic heterogeneity outside of the mutated IDH pathway were also evident, with key metabolites being high expression of glycerophosphates, inositols, monosaccharides, and sugar alcohols and low levels of sphingosine and lysoglycerophospholipids in IDH-mutants. Among the IDH-mutated subtypes, we observed high levels of amino acids, especially glycine and 2-aminoadipic acid, in grade 4 glioma, and N-acetyl aspartic acid in low-grade astrocytoma and oligodendroglioma. Both IDH-wildtype and mutated oligodendroglioma and glioblastoma were characterized by high levels of acylcarnitines, likely driven by rapid cell growth and hypoxic features. We found elevated levels of 5-HIAA in gliosarcoma and a subtype of oligodendroglioma not yet defined as a specific entity, indicating a previously not described role for the serotonin pathway linked to glioma with bimorphic tissue. CONCLUSION: Key metabolic differences exist across adult glioma subtypes.

JNK1 phosphorylation of SCG10 determines microtubule dynamics and axodendritic length.

c-Jun NH(2)-terminal kinases (JNKs) are essential during brain development, when they regulate morphogenic changes involving cell movement and migration. In the adult, JNK determines neuronal cytoarchitecture. To help uncover the molecular effectors for JNKs in these events, we affinity purified JNK-interacting proteins from brain. This revealed that the stathmin family microtubule-destabilizing proteins SCG10, SCLIP, RB3, and RB3' interact tightly with JNK. Furthermore, SCG10 is also phosphorylated by JNK in vivo on sites that regulate its microtubule depolymerizing activity, serines 62 and 73. SCG10-S73 phosphorylation is significantly decreased in JNK1-/- cortex, indicating that JNK1 phosphorylates SCG10 in developing forebrain. JNK phosphorylation of SCG10 determines axodendritic length in cerebrocortical cultures, and JNK site-phosphorylated SCG10 colocalizes with active JNK in embryonic brain regions undergoing neurite elongation and migration. We demonstrate that inhibition of cytoplasmic JNK and expression of SCG10-62A/73A both inhibited fluorescent tubulin recovery after photobleaching. These data suggest that JNK1 is responsible for regulation of SCG10 depolymerizing activity and neurite elongation during brain development.

Identification of Pre-Diagnostic Metabolic Patterns for Glioma Using Subset Analysis of Matched Repeated Time Points.

Here, we present a strategy for early molecular marker pattern detection-Subset analysis of Matched Repeated Time points (SMART)-used in a mass-spectrometry-based metabolomics study of repeated blood samples from future glioma patients and their matched controls. The outcome from SMART is a predictive time span when disease-related changes are detectable, defined by time to diagnosis and time between longitudinal sampling, and visualization of molecular marker patterns related to future disease. For glioma, we detect significant changes in metabolite levels as early as eight years before diagnosis, with longitudinal follow up within seven years. Elevated blood plasma levels of myo-inositol, cysteine, N-acetylglucosamine, creatinine, glycine, proline, erythronic-, 4-hydroxyphenylacetic-, uric-, and aceturic acid were particularly evident in glioma cases. We use data simulation to ensure non-random events and a separate data set for biomarker validation. The latent biomarker, consisting of 15 interlinked and significantly altered metabolites, shows a strong correlation to oxidative metabolism, glutathione biosynthesis and monosaccharide metabolism, linked to known early events in tumor development. This study highlights the benefits of progression pattern analysis and provide a tool for the discovery of early markers of disease.

Thrombospondin 2 inhibits microvascular endothelial cell proliferation by a caspase-independent mechanism.

The matricellular protein thrombospondin 2 (TSP2) regulates a variety of cell-matrix interactions. A prominent feature of TSP2-null mice is increased microvascular density, particularly in connective tissues synthesized after injury. We investigated the cellular basis for the regulation of angiogenesis by TSP2 in cultures of murine and human fibroblasts and endothelial cells. Fibroblasts isolated from murine and human dermis synthesize TSP2 mRNA and secrete significant amounts of immunoreactive TSP2, whereas endothelial cells from mouse lung and human dermis did not synthesize TSP2 mRNA or protein. Recombinant mouse TSP2 inhibited growth of human microvascular endothelial cells (HMVECs) mediated by basic fibroblast growth factor, insulin-like growth factor-1, epidermal growth factor, and vascular endothelial growth factor (VEGF). HMVECs exposed to TSP2 in the presence of these growth factors had a decreased proportion of cells in S and G2/M phases. HMVECs cultured with a combination of basic fibroblast growth factor, insulin-like growth factor-1, and epidermal growth factor displayed an increased proportion of nonviable cells in the presence of TSP2, but the addition of VEGF blocked this TSP2-mediated impairment of cell viability. TSP2-mediated inhibition of DNA synthesis by HMVECs in the presence of VEGF was not affected by the broad-spectrum caspase inhibitor zVAD-fmk. Similar findings were obtained with TSP1. Taken together, these observations indicate that either TSP2 or TSP1 can inhibit HMVEC proliferation by inhibition of cell cycle progression and induction of cell death, but the mechanisms responsible for TSP2-mediated inhibition of cell cycle progression are independent from those leading to cell death.

Constitutively active cytoplasmic c-Jun N-terminal kinase 1 is a dominant regulator of dendritic architecture: role of microtubule-associated protein 2 as an effector.

Normal functioning of the nervous system requires precise regulation of dendritic shape and synaptic connectivity. Here, we report a severe impairment of dendritic structures in the cerebellum and motor cortex of c-Jun N-terminal kinase 1 (JNK1)-deficient mice. Using an unbiased screen for candidate mediators, we identify the dendrite-specific high-molecular-weight microtubule-associated protein 2 (MAP2) as a JNK substrate in the brain. We subsequently show that MAP2 is phosphorylated by JNK in intact cells and that MAP2 proline-rich domain phosphorylation is decreased in JNK1-/- brain. We developed compartment-targeted JNK inhibitors to define whether a functional relationship exists between the physiologically active, cytosolic pool of JNK and dendritic architecture. Using these, we demonstrate that cytosolic, but not nuclear, JNK determines dendritic length and arbor complexity in cultured neurons. Moreover, we confirm that MAP2-dependent process elongation is enhanced after activation of JNK. Using JNK1-/- neurons, we reveal a dominant role for JNK1 over ERK in regulating dendritic arborization, whereas ERK only regulates dendrite shape under conditions in which JNK activity is low (JNK1-/- neurons). These results reveal a novel antagonism between JNK and ERK, potentially providing a mechanism for fine-tuning the dendritic arbor. Together, these data suggest that JNK phosphorylation of MAP2 plays an important role in defining dendritic architecture in the brain.