Previous investigations of retrograde survival signaling by nerve growth factor （NGF） and other neurotrophins have supported diverse mechanisms, but all proposed mechanisms have in common the generation of survival ...Previous investigations of retrograde survival signaling by nerve growth factor （NGF） and other neurotrophins have supported diverse mechanisms, but all proposed mechanisms have in common the generation of survival signals retrogradely transmitted to the neuronal cell bodies. We report the finding of a retrograde apoptotic signal in axons that is suppressed by local NGF signaling. NGF withdrawal from distal axons alone was sufficient to activate the pro-apoptotic transcription factor, c-jun, in the cell bodies. Providing NGF directly to cell bodies, thereby restoring a source of NGF-induced survival signals, could not prevent c-jun activation caused by NGF withdrawal from the distal axons. This is evidence that c-jun is not activated due to loss of survival signals at the cell bodies. Moreover, blocking axonal transport with colchicine inhibited c-jun activation caused by NGF deprivation suggesting that a retrogradely transported pro-apoptotic signal, rather than loss of a retrogradely transported survival signal, caused c-jun activation. Additional experiments showed that activation of c-jun, pro-caspase-3 cleavage, and apoptosis were blocked by the protein kinase C inhibitors, rottlerin and chelerythrine, only when applied to distal axons suggesting that they block the axon-specific pro-apoptotic signal. The rottlerin-sensitive mechanism was found to regulate glyco- gen synthase kinase 3 （GSK3） activity. The effect of siRNA knockdown, and pharmacological inhibition of GSK3 suggests that GSK3 is required for apoptosis caused by NGF deprivation and may function as a retrograde carrier of the axon apoptotic signal. The existence of a retrograde death signaling system in axons that is suppressed by neurotro- phins has broad implications for neurodevelopment and for discovering treatments for neurodegenerative diseases and neurotrauma.展开更多
With the changes of life style, diabetes and its complications have become a major cause of morbidity and mortality. It is reasonable to anticipate a continued rise in the incidence of diabetes and its complications a...With the changes of life style, diabetes and its complications have become a major cause of morbidity and mortality. It is reasonable to anticipate a continued rise in the incidence of diabetes and its complications along with the aging of the population, increase in adult obesity rate, and other risk factors. Diabetic en- cephalopathy is one of the severe microvascular complications of diabetes, characterized by impaired cogni- tive functions, and electrophysiological, neurochemical, and structural abnormalities. It may involve direct neuronal damage caused by intracellular glucose. However, the pathogenesis of this disease is complex and its diagnosis is not very clear. Previous researches have suggested that chronic metabolic alterations, vascular changes, and neuronal apoptosis may play important roles in neuronai loss and damaged cognitive functions. Multiple factors are responsible for neuronal apoptosis, such as disturbed insulin growth factor （IGF） system, hyperglycemia, and the aging process. Recent data suggest that insulin/C-peptide deficiency may exert a primary and key effect in diabetic encephalopathy. Administration of C-peptide partially improves the condition of the IGF system in the brain and prevents neuronal apoptosis in the hippocampus of diabetic patients. Those findings provide a basis for application of C-peptide as a potentially effective therapy for diabetes and diabetic encephalopathy.展开更多
To explore how the intrinsic apoptosis pathway is controlled in the spontaneous fog （forebrain overgrowth） mutant mice with an Apafl splicing deficiency, we examined spleen and bone marrow cells from Apafl＋/＋ （＋...To explore how the intrinsic apoptosis pathway is controlled in the spontaneous fog （forebrain overgrowth） mutant mice with an Apafl splicing deficiency, we examined spleen and bone marrow cells from Apafl＋/＋ （＋/＋） and Apafl^fog/fog （fog/fog） mice for initiator caspase-9 activation by cellular stresses. When the mitochondrial inner membrane potential （△ψm） was disrupted by staurosporine, ＋/＋ cells but not fog/fog cells activated caspase-9 to cause apoptosis, indicating the lack of apoptosome （apoptosis protease activating factor 1 （Apaf-l）/cytochrome c/（d）ATP/procaspase-9） function in fog/fog cells. However, when a marginal （-20%） decrease in △ψm was caused by hydrogen peroxide （0.1 mM）, peroxynitrite donor 3-morpholinosydnonimine （0.1 mM） and UV-C irradiation （20 J/m^2）, both ＋/＋ and fog/fog cells triggered procaspase-9 auto-processing and its downstream cascade activation. Supporting our previous results, procaspase-9 pre-existing in the mitochondria induced its auto-processing before the cytosolic caspase activation regardless of the genotypes. Cellular ATP concentration significantly decreased under the hypoactive △ψm condition. Furthermore, we detected accumulation of citrate, a kosmotrope known to facilitate procaspase-9 dimerization, probably due to a feedback control of the Krebs cycle by the electron transfer system. Thus, mitochondrial in situ caspase-9 activation may be caused by the major metabolic reactions in response to physiological stresses, which may represent a mode of Apaf-l-independent apoptosis hypothesized from recent genetic studies.展开更多
Objective: To study the role of p38MAPK in mediating TNF-α-induced apoptosis of rat glioma cell line C6. Methods: Effect of TNF-α on the proliferation of C6 cells was determined by MTT assay. The TNF-α induced apop...Objective: To study the role of p38MAPK in mediating TNF-α-induced apoptosis of rat glioma cell line C6. Methods: Effect of TNF-α on the proliferation of C6 cells was determined by MTT assay. The TNF-α induced apoptosis was detected by transmission electron microscopy and flow cytometry. The expression of p38MAPK was detected by SABC method and Western-blot. The effect of SB202190, a specific inhibitor of p38MAPK, on TNF-α-induced apoptosis was observed by flow cytometry and SABC method. Results: Inhibitory rate of TNF-α(2×105 U/L) on C6 cells was 43.75%. In the TNF-α treated group, apoptotic cells were observed by transmission electron microscopy and the apoptotic rate was 37.5% by flow cytometry. p38MAPK positive signals were detected by SABC method and Western-blot. In the SB202190 treated group, the apoptotic rate was 7.0% and no p38MAPK signals were found. Conclusion: Apoptosis of C6 cells and expression of p38MAPK can be induced by TNF-α. The activation of p38MAPK promotes the apoptosis of C6 cells.展开更多
Here, we explore the role of Cbl proteins in regulation of neuronal apoptosis. In two paradigms of neuron apoptosis -- nerve growth factor （NGF） deprivation and DNA damage -- cellular levels of c-Cbl and Cbl-b fell ...Here, we explore the role of Cbl proteins in regulation of neuronal apoptosis. In two paradigms of neuron apoptosis -- nerve growth factor （NGF） deprivation and DNA damage -- cellular levels of c-Cbl and Cbl-b fell well before the onset of cell death. NGF deprivation also induced rapid loss of tyrosine phosphorylation （and most likely, activation） of c-Cbl. Targeting c-Cbl and Cbl-b with siRNAs to mimic their loss/inactivation sensitized neuronal cells to death promoted by NGF deprivation or DNA damage. One potential mechanism by which Cbl proteins might affect neuronal death is by regulation of apoptotic c-Jun N-terminal kinase （JNK） signaling. We demonstrate that Cbl proteins interact with the JNK pathway components mixed lineage kinase （MLK） 3 and POSH and that knockdown of Cbl proteins is sufficient to increase JNK pathway activity. Furthermore, expression of c-Cbl blocks the ability of MLKs to signal to downstream components of the kinase cascade leading to JNK activation and protects neuronal cells from death induced by MLKs, but not from downstream JNK activators. On the basis of these findings, we propose that Cbls suppress cell death in healthy neurons at least in part by inhibiting the ability of MLKs to activate JNK signaling. Apoptotic stimuli lead to loss of Cbl protein/activity, thereby removing a critical brake on JNK activation and on cell death.展开更多
文摘Previous investigations of retrograde survival signaling by nerve growth factor （NGF） and other neurotrophins have supported diverse mechanisms, but all proposed mechanisms have in common the generation of survival signals retrogradely transmitted to the neuronal cell bodies. We report the finding of a retrograde apoptotic signal in axons that is suppressed by local NGF signaling. NGF withdrawal from distal axons alone was sufficient to activate the pro-apoptotic transcription factor, c-jun, in the cell bodies. Providing NGF directly to cell bodies, thereby restoring a source of NGF-induced survival signals, could not prevent c-jun activation caused by NGF withdrawal from the distal axons. This is evidence that c-jun is not activated due to loss of survival signals at the cell bodies. Moreover, blocking axonal transport with colchicine inhibited c-jun activation caused by NGF deprivation suggesting that a retrogradely transported pro-apoptotic signal, rather than loss of a retrogradely transported survival signal, caused c-jun activation. Additional experiments showed that activation of c-jun, pro-caspase-3 cleavage, and apoptosis were blocked by the protein kinase C inhibitors, rottlerin and chelerythrine, only when applied to distal axons suggesting that they block the axon-specific pro-apoptotic signal. The rottlerin-sensitive mechanism was found to regulate glyco- gen synthase kinase 3 （GSK3） activity. The effect of siRNA knockdown, and pharmacological inhibition of GSK3 suggests that GSK3 is required for apoptosis caused by NGF deprivation and may function as a retrograde carrier of the axon apoptotic signal. The existence of a retrograde death signaling system in axons that is suppressed by neurotro- phins has broad implications for neurodevelopment and for discovering treatments for neurodegenerative diseases and neurotrauma.
文摘With the changes of life style, diabetes and its complications have become a major cause of morbidity and mortality. It is reasonable to anticipate a continued rise in the incidence of diabetes and its complications along with the aging of the population, increase in adult obesity rate, and other risk factors. Diabetic en- cephalopathy is one of the severe microvascular complications of diabetes, characterized by impaired cogni- tive functions, and electrophysiological, neurochemical, and structural abnormalities. It may involve direct neuronal damage caused by intracellular glucose. However, the pathogenesis of this disease is complex and its diagnosis is not very clear. Previous researches have suggested that chronic metabolic alterations, vascular changes, and neuronal apoptosis may play important roles in neuronai loss and damaged cognitive functions. Multiple factors are responsible for neuronal apoptosis, such as disturbed insulin growth factor （IGF） system, hyperglycemia, and the aging process. Recent data suggest that insulin/C-peptide deficiency may exert a primary and key effect in diabetic encephalopathy. Administration of C-peptide partially improves the condition of the IGF system in the brain and prevents neuronal apoptosis in the hippocampus of diabetic patients. Those findings provide a basis for application of C-peptide as a potentially effective therapy for diabetes and diabetic encephalopathy.
文摘To explore how the intrinsic apoptosis pathway is controlled in the spontaneous fog （forebrain overgrowth） mutant mice with an Apafl splicing deficiency, we examined spleen and bone marrow cells from Apafl＋/＋ （＋/＋） and Apafl^fog/fog （fog/fog） mice for initiator caspase-9 activation by cellular stresses. When the mitochondrial inner membrane potential （△ψm） was disrupted by staurosporine, ＋/＋ cells but not fog/fog cells activated caspase-9 to cause apoptosis, indicating the lack of apoptosome （apoptosis protease activating factor 1 （Apaf-l）/cytochrome c/（d）ATP/procaspase-9） function in fog/fog cells. However, when a marginal （-20%） decrease in △ψm was caused by hydrogen peroxide （0.1 mM）, peroxynitrite donor 3-morpholinosydnonimine （0.1 mM） and UV-C irradiation （20 J/m^2）, both ＋/＋ and fog/fog cells triggered procaspase-9 auto-processing and its downstream cascade activation. Supporting our previous results, procaspase-9 pre-existing in the mitochondria induced its auto-processing before the cytosolic caspase activation regardless of the genotypes. Cellular ATP concentration significantly decreased under the hypoactive △ψm condition. Furthermore, we detected accumulation of citrate, a kosmotrope known to facilitate procaspase-9 dimerization, probably due to a feedback control of the Krebs cycle by the electron transfer system. Thus, mitochondrial in situ caspase-9 activation may be caused by the major metabolic reactions in response to physiological stresses, which may represent a mode of Apaf-l-independent apoptosis hypothesized from recent genetic studies.
文摘Objective: To study the role of p38MAPK in mediating TNF-α-induced apoptosis of rat glioma cell line C6. Methods: Effect of TNF-α on the proliferation of C6 cells was determined by MTT assay. The TNF-α induced apoptosis was detected by transmission electron microscopy and flow cytometry. The expression of p38MAPK was detected by SABC method and Western-blot. The effect of SB202190, a specific inhibitor of p38MAPK, on TNF-α-induced apoptosis was observed by flow cytometry and SABC method. Results: Inhibitory rate of TNF-α(2×105 U/L) on C6 cells was 43.75%. In the TNF-α treated group, apoptotic cells were observed by transmission electron microscopy and the apoptotic rate was 37.5% by flow cytometry. p38MAPK positive signals were detected by SABC method and Western-blot. In the SB202190 treated group, the apoptotic rate was 7.0% and no p38MAPK signals were found. Conclusion: Apoptosis of C6 cells and expression of p38MAPK can be induced by TNF-α. The activation of p38MAPK promotes the apoptosis of C6 cells.
基金Acknowledgments This work was supported by grants from the NIH/NINDS （NS33689） （L.A.G.） and from the National Science Foundation of China （NSFC） （30525007/30670663）, the Ministry of Science and Technology of China （2006AA02Z173/2007CB947202） and the Chinese Academy of Sciences （KSCX1-YW-R-59） （Z.X.）.
文摘Here, we explore the role of Cbl proteins in regulation of neuronal apoptosis. In two paradigms of neuron apoptosis -- nerve growth factor （NGF） deprivation and DNA damage -- cellular levels of c-Cbl and Cbl-b fell well before the onset of cell death. NGF deprivation also induced rapid loss of tyrosine phosphorylation （and most likely, activation） of c-Cbl. Targeting c-Cbl and Cbl-b with siRNAs to mimic their loss/inactivation sensitized neuronal cells to death promoted by NGF deprivation or DNA damage. One potential mechanism by which Cbl proteins might affect neuronal death is by regulation of apoptotic c-Jun N-terminal kinase （JNK） signaling. We demonstrate that Cbl proteins interact with the JNK pathway components mixed lineage kinase （MLK） 3 and POSH and that knockdown of Cbl proteins is sufficient to increase JNK pathway activity. Furthermore, expression of c-Cbl blocks the ability of MLKs to signal to downstream components of the kinase cascade leading to JNK activation and protects neuronal cells from death induced by MLKs, but not from downstream JNK activators. On the basis of these findings, we propose that Cbls suppress cell death in healthy neurons at least in part by inhibiting the ability of MLKs to activate JNK signaling. Apoptotic stimuli lead to loss of Cbl protein/activity, thereby removing a critical brake on JNK activation and on cell death.
