Expression of cannabinoid receptors and neurotrophins in human gliomas

Recent studies have shown an anti-tumour activity of cannabinoid receptors CB1 and CB2 in gliomas. This effect was mediated by neurotrophins in breast and prostate carcinoma, while in gliomas this relationship has not yet been considered. The aim of this study was to investigate the expression of cannabinoid receptors CB1 and CB2, neurotrophin NGF and NT-3 and their receptors TrkA and TrkC in glioma and endothelial cells. The analysis was performed in 14 gliomas and 2 non-tumour brain specimens by immunohistochemistry and real-time quantitative-polymerase chain reaction (RTQ-PCR). Gliomas showed a weak immunoreactivity for CB1 and CB2 in tumour and in endothelial cells, and for NGF/TrkA mainly in tumour cells, while a moderate/diffuse immunoreactivity was found for NT-3/TrkC. CB2 was expressed on 3 out of 6 low-grade gliomas and in all high-grade gliomas. Non-tumour brain tissues were weakly positive in astrocytes and endothelium for CB1, CB2, NT-3 and TrkC and negative for NGF and TrkA. By RTQ-PCR, gliomas showed low mRNA levels of NGF/TrkA and moderate levels of CB1, NT-3 and TrkC. CB2 mRNA expression was low or absent. A potential role of cannabinoids, particularly of CB2 agonists devoid of psychotropic side effects, in glioma therapy could have a basis in glioblastomas, because they were all positive, though weakly, to CB2. The presence of neurotrophins and their receptors, mainly NT-3 and TrkC, suggests a possible role of these pathways in glioma growth/invasion, but further investigations are required to verify this hypothesis and a potential relationship between cannabinoids and neurotrophins.     

Developmental changes in concentrations and distributions of neurotrophins in the monkey cerebellar cortex

Neurotrophins are involved in the survival, differentiation, migration and neurite outgrowth of various neuronal populations. Neurotrophins and their receptors are widely expressed in the developing cerebellum of various experimental animals. To gain some insight into the possible roles played by these molecules in monkey cerebellum, we examined the protein levels of BDNF, NT-4/5 and NT-3 and distributions of those neurotrophins and TrkC, a high affinity receptor for NT-3, in the cerebellum of developing macaque monkeys using ELISAs and immunohistochemical methods. We found that the level of BDNF increased during development, while the level of NT-3 was higher during embryonic stages and decreased toward adulthood. The level of NT-4/5 increased from embryonic stages to infant stages and gradually declined with age. Among the three neurotrophins, BDNF immunoreactivity was found in all kinds of cerebellar neurons, including all inhibitory interneurons, throughout the postnatal periods examined, indicating that BDNF may be an essential factor for the maintenance of cerebellar neural functions. The Bergmann glial fibers and the internal part of the external granule cell layer were strongly NT-3 immunopositive at the early postnatal stages, and more weakly immunoreactive toward adulthood. In addition, we found that the premigratory precursors of the granule cells were TrkC immunopositive at early postnatal stages. These findings suggest that NT-3 in Bergmann glial fibers may be involved in the migration of the premigratory granule cells.     

TrkC Is Essential for Nephron Function and Trans-Activates Igf1R Signaling

BackgroundInjury to kidney podocytes often results in chronic glomerular disease and consecutive nephron malfunction. For most glomerular diseases, targeted therapies are lacking. Thus, it is important to identify novel signaling pathways contributing to glomerular disease. Neurotrophic tyrosine kinase receptor 3 (TrkC) is expressed in podocytes and the protein transmits signals to the podocyte actin cytoskeleton.MethodsNephron-specific TrkC knockout (TrkC-KO) and nephron-specific TrkC-overexpressing (TrkC-OE) mice were generated to dissect the role of TrkC in nephron development and maintenance.ResultsBoth TrkC-KO and TrkC-OE mice exhibited enlarged glomeruli, mesangial proliferation, basement membrane thickening, albuminuria, podocyte loss, and aspects of FSGS during aging. Igf1 receptor (Igf1R)–associated gene expression was dysregulated in TrkC-KO mouse glomeruli. Phosphoproteins associated with insulin, erb-b2 receptor tyrosine kinase (Erbb), and Toll-like receptor signaling were enriched in lysates of podocytes treated with the TrkC ligand neurotrophin-3 (Nt-3). Activation of TrkC by Nt-3 resulted in phosphorylation of the Igf1R on activating tyrosine residues in podocytes. Igf1R phosphorylation was increased in TrkC-OE mouse kidneys while it was decreased in TrkC-KO kidneys. Furthermore, TrkC expression was elevated in glomerular tissue of patients with diabetic kidney disease compared with control glomerular tissue.ConclusionsOur results show that TrkC is essential for maintaining glomerular integrity. Furthermore, TrkC modulates Igf-related signaling in podocytes.     

Voluntary exercise increases neurotrophin-3 and its receptor TrkC in the spinal cord

We have evaluated changes in the expression of neurotrophin-3 (NT-3) and its tyrosine kinase C (TrkC) receptor in the neuromuscular system as a result of voluntary physical activity. We assessed changes in the mRNAs and proteins for NT-3 and TrkC in the lumbar spinal cord and associated soleus muscle following 3 and 7 days of voluntary wheel running. We used quantitative Taqman RT-PCR to measure mRNA and ELISA to assess protein levels. NT-3 mRNA and protein levels increased in the spinal cord to reach statistical significance after 7 days of exercise compared to sedentary control rats. Immunohistochemical analyses localized the elevated NT-3 to the substantia gelatinosa (SG) and nucleus of the dorsal horn. TrkC mRNA levels were significantly elevated in the spinal cord after 3 and 7 days of running. In the soleus muscle, NT-3 mRNA levels and its receptor TrkC were elevated after 3 days, while NT-3 protein levels remained unaffected. The results demonstrate that voluntary exercise has a differential effect on NT-3 as well as its receptor TrkC in the neural and muscular components of the neuromuscular system, and emphasize the role of voluntary activity on the spinal cord and muscle.     

Comparative study of the distribution of the alpha-subunits of voltage-gated sodium channels in normal and axotomized rat dorsal root ganglion neurons

We compared the distribution of the α‐subunit mRNAs of voltage‐gated sodium channels Nav1.1–1.3 and Nav1.6–1.9 and a related channel, Nax, in histochemically identified neuronal subpopulations of the rat dorsal root ganglia (DRG). In the naïve DRG, the expression of Nav1.1 and Nav1.6 was restricted to A‐fiber neurons, and they were preferentially expressed by TrkC neurons, suggesting that proprioceptive neurons possess these channels. Nav1.7, ‐1.8, and ‐1.9 mRNAs were more abundant in C‐fiber neurons compared with A‐fiber ones. Nax was evenly expressed in both populations. Although Nav1.8 and ‐1.9 were preferentially expressed by TrkA neurons, other α‐subunits were expressed independently of TrkA expression. Actually, all IB4⁺ neurons expressed both Nav1.8 and ‐1.9, and relatively limited subpopulations of IB4⁺ neurons (3% and 12%, respectively) expressed Nav1.1 and/or Nav1.6. These findings provide useful information in interpreting the electrophysiological characteristics of some neuronal subpopulations of naïve DRG. After L5 spinal nerve ligation, Nav1.3 mRNA was up‐regulated mainly in A‐fiber neurons in the ipsilateral L5 DRG. Although previous studies demonstrated that nerve growth factor (NGF) and glial cell‐derived neurotrophic factor (GDNF) reversed this up‐regulation, the Nav1.3 induction was independent of either TrkA or GFRα1 expression, suggesting that the induction of Nav1.3 may be one of the common responses of axotomized DRG neurons without a direct relationship to NGF/GDNF supply. J. Comp. Neurol. 510:188–206, 2008. © 2008 Wiley‐Liss, Inc.     

NT-3 and BDNF protect CNS neurons against metabolic/excitotoxic insults

Neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) were recently shown to have biological activity in central neurons. In the present study, NT-3 and BDNF attenuated glucose deprivation-induced neuronal damage dose-dependently in rat hippocampal, septal and cortical cultures. Direct measurements of intraneuronal free calcium levels ([Ca²⁺]_i) and manipulations of calcium inlux demonstrated that NT-3 and BDNF each prevented the elevation of [Ca²⁺]_i that mediated glucose deprivation-induced injury. Studies in cultures depleted of glia indicateda direct action of NT-3 and BDNF on neurons. Neurons pretreated with NT-3 or BDNF for 24 hr were more resistant to glutamate neurotoxicity, and showed attenuated [Ca²⁺]_i responses to glutamate. TrkB (BDNF receptor) and trkC (NT-3 receptor) proteins were present in hippocampal, cortical and septal cultures where they were localied to neuronal cell bodies and neurites. The data demonstrate that NT-3 and BDNF can protect neurons against metabolic and excitotoxic insults, and suggest that these neurotrophins may serve [Ca²⁺]_i-stabilizing and neuroprotective functions in the brain.     

酪氨酸激酶受体C在膀胱癌SCaBER细胞裸鼠种植瘤中的作用

【目的】通过酪氨酸激酶受体C(tyrosine receptor kinase,TrkC)siRNA干涉质粒下调膀胱癌细胞SCaBER裸鼠种植瘤中TrkC的表达,分析TrkC在膀胱癌中的作用。【方法】采用构建SCaBER细胞裸鼠种植瘤模型,以脂质体Lipofect Amine 2000为介质,采用瘤内注射的方法,用TrkCsiRNA干涉质粒实现下调SCaBER细胞中TrkC的表达,以空载体pSilencer作为阴性对照,干涉过程中记录裸鼠种植瘤的体积变化,绘制种植瘤生长曲线,干涉结束后处死裸鼠,剥除肿瘤,拍照,称重,并提取蛋白,western blotting检测干涉效果。【结果】Western blotting检测发现,构建的TrkCsiRNA干涉质粒顺利实现了在SCaBER细胞裸鼠种植瘤中对TrkC的表达的下调(F=7.46,P<0.05),种植瘤生长曲线表明下调TrkC的表达可以抑制SCaBER细胞在裸鼠活体内的增殖,实验组肿瘤肿瘤体积(F=11.50,P<0.05)和重量(F=19.32,P<0.01)明显低于对照组。【结论】下调TrkC表达可抑制膀胱癌细胞SCaBER在活体内增殖,TrkC可能参与膀胱癌发生发展过程。     

TrkC expression predicts favorable clinical outcome in invasive ductal carcinoma of breast independent of NT-3 expression

Background: TrkC, a member of neurotrophin receptor family, functions not only as an oncogene, but also act as a tumor suppressor via a manner of dependence receptor in human malignant tumors. Little is known on the action of TrkC for the clinical prognosis and the progression of breast cancer according to the availability of its ligand NT-3. We sought to investigate the prognostic relevance of NT-3-TrkC axis in breast cancer and estimate its role during the process of breast cancer progression. Methods: 236 cases of invasive ductal carcinoma (IDC), 60 pure ductal carcinoma in situ (DCIS) and 30 normal breast tissue (NBT) between 2004 and 2005 were included in the study. Spearman’s rank correlation test was used to analyze the association of NT-3-TrkC expression and breast cancer progression. The Kaplan-Meier method and Cox proportional hazards model were performed to identify the relevant prognostic factors. Results: 50.4% IDC tumors displayed absent or low TrkC expression, while 49.6% was high TrkC expression. TrkC expression was negatively associated with lymph node metastasis (P = 0.029) and tumor proliferation (P = 0.015). Patients with lower TrkC expressing tumors had a higher risk of recurrence (odds ratio, 0.401; 95% confidence interval, 0.207-0.778; P = 0.007). The layered analysis indicated that patients with high TrkC expression tumors had a favor disease-free survival whether NT-3 and TrkC were co-expressed or solitarily expressed in the tumor (P = 0.000). NT-3 was demonstrated to be not a predictor of IDC patients’ prognosis. But NT-3 expression was inversely correlated with the progression of breast cancer (r = -0.341, P = 0.000), since more IDC tumors showed high NT-3 expression than DCIS tumors (51.7% vs. 25.9%), while no NBT showed high NT-3 expression, as well. Conclusion: The study indicates TrkC expression reduces tumor relapse independent of NT-3 availability in the IDC. Elevated NT-3 expression contributes to the progression of breast cancer.     

蛴螬对激光损伤兔血-视网膜屏障后视网膜TekC、NGF表达的影响＊

目的 探讨蛴螬促进激光损伤兔血-视网膜屏障后的修复作用及机理。方法 30只实验性兔随机分为正常组、模型组、蛴螬组。采用激光损伤血-视网膜屏障动物模型，在屏障损伤后1周及2周两个时相，观察蛴螬对兔血-视网膜屏障损伤修复情况、血-视网膜屏障组织形态结构及神经营养因子受体TrkC、NGF表达及影响。结果 蛴螬能促进损伤的血-视网膜屏障修复，减轻激光导致的视网膜组织及细胞形态学损伤，促进TrkC、NGF的表达，抑制视网膜细胞增殖与变性，从而改善视网膜功能。结论 蛴螬可能通过改善视网膜组织能量代谢，清除氧自由基，促进神经营养因子受体的表达，抑制视细胞增殖，起到保护血-视网膜屏障组织结构的作用。