BACKGROUND: Some scholars believed that the neuronal injury after status epilepticus is apoptosis, the main evidence is the changes of expressions of various apoptosis related genes, such as immediate-early gene, p53 ...BACKGROUND: Some scholars believed that the neuronal injury after status epilepticus is apoptosis, the main evidence is the changes of expressions of various apoptosis related genes, such as immediate-early gene, p53 gene and genes of bcl-2 family, etc. But there is still no ultrastructural evidence for apoptosis. OBJECTIVE: To observe the ultrastructural damages of mitochondrion and nucleus and the changes of caspase expression in neurons of hippocampal CA3 region in rats with status epilepticus induced by kainic acid. DESIGN: A randomized controlled study. SETTING: Department of Anesthesiology and Department of Neurology, Qilu Hospital of Shandong University. MATERIALS: Seventy-five adult male Wistar rats of 250-300 g, clean degree, were provided by the experimental animal center of Shandong University. Kainic acid was purchased from Sigma Company (USA); rabbit anti-rat polyclonal antibody caspase-3 from Santa Cruz Company (USA). METHODS: The experiments were carried out in the Department of Anesthesiology, Qilu Hospital of Shandong University from October 2005 to February 2006. ① The 75 rats were randomly divided into experimental group (n =45) and control group (n =30). ② Model establishment, convulsion grading and the judging standards for status epilepticus: Rats in the experimental group were given intraperitoneal injection of kainic acid (10 mg/kg), and those in the control group were injected with saline of the same volume. The time of seizure was recorded and their behavioral manifestations were observed, and the seizure was terminated by intraperitoneal injection of diazepam (10 mg/kg). ③ Observation under electron microscope: At 3, 12 and 24 hours after status epilepticus respectively, bilateral hippocampal tissues were taken out, semithin sections of about 75 nm were prepared after fixation, dehydration and embedding, and then observed under H-800 transmission electron microscope. ④ Immunohistochemical detection: Bilateral hippocampi were removed at 3, 12 and 24 hours after status epilepticus respectively, the fixation, dehydration, transparence, wax immersion and embedding were performed, then serial sections of CA3 region were immunohistochemically determined by the SABC method. Leica QWinV3 image analytical software was applied, then the average number and average gray value of positive cells were calculated. MAIN OUTCOME MEASURES: Results of observation under electron microscope, that of immunohistochemical staining of neurons in hippocampal CA3 region; Comparison of number of caspase-3 positive cells and gray value. RESULTS: All the 75 Wistar rats were involved in the analysis of results. ① Results of observation under electron microscope: At 3 hours after status epilepticus, swelling crista and membranous disintegration were observed under electron microscope. At 24 hours, obvious nuclear changes occurred, and manifested as the side-aggegation of chromatins. ② Results of immunohistochemical detection: In the experimental group, the number of caspase-3 positive cells at 3 hours after status epilepticus had no obvious difference as compared with that in the control group (P > 0.05); At 12 hours, the number and gray value of caspase-3 positive cells in the experimental group were higher than those in the control group (10.49±0.68 vs. 5.33±0.43; 45.57±2.27 vs. 19.79±0.33, P < 0.05), the same results were also observed at 24 hours (37.36±0.57 vs. 5.12±0.47; 115.24±1.22 vs. 18.73±0.42, P < 0.01). CONCLUSION: In the rat models of status epilepticus induced by kainic acid, mitochondrial damage was earlier than the increase of caspase-3 expression and nuclear changes, which suggested that mitochondrion was the key link for the neuronal death after status epilepticus.展开更多
The clinical presentation of schizophrenia involves a variety of symptoms, which in many cases include hallucinations and delusions. Experimentally revealed alterations in both pre-pulse inhibition (PPI) and latent in...The clinical presentation of schizophrenia involves a variety of symptoms, which in many cases include hallucinations and delusions. Experimentally revealed alterations in both pre-pulse inhibition (PPI) and latent inhibition (LI) are also apparent in individuals afflicted with this disorder. Many have speculated that altered synaptic connections are, in part, responsible for this subset of behavioral abnormalities. We have previously reported that neonatal chronic low-dose injections of domoic acid (DOM) produce adult rats with deficits in PPI and LI. The current study was conducted to determine whether this toxin-treatment would alter the degree of apoptosis occurring in the developing brain. Results revealed significant decreases in caspase-3 within the right prelimbic cortex (PrL) in both male and female DOM-treated rats suggesting that even modest alterations in glutamate (Glu) signaling during critical periods of central nervous system (CNS) maturation will modify ontogenetic processes in the prefrontal cortex (PFC) of the juvenile rat.展开更多
In this study, primary cultured cerebral cortical neurons of Sprague-Dawley neonatal rats were treated with 0.25, 0.5, and 1.0 μM calmodulin-dependent protein kinase II inhibitor KN-93 after 50 μM N-methyI-D-asparti...In this study, primary cultured cerebral cortical neurons of Sprague-Dawley neonatal rats were treated with 0.25, 0.5, and 1.0 μM calmodulin-dependent protein kinase II inhibitor KN-93 after 50 μM N-methyI-D-aspartic acid-induced injury. Results showed that, compared with N-methyi-D- aspartic acid-induced injury neurons, the activity of cells markedly increased, apoptosis was significantly reduced, leakage of lactate dehydrogenase decreased, and intracellular Ca2＋ concentrations in neurons reduced after KN-93 treatment. The expression of caspase-3, phosphorylated calmodulin-dependent protein kinase II and total calmodulin-dependent protein kinase II protein decreased after KN-93 treatment. And the effect was apparent at a dose of 1.0 pM KN-93. Experimental findings suggest that KN-93 can induce a dose-dependent neuroprotective effect, and that the underlying mechanism may be related to the down-regulation of caspase-3 and calmodulin- dependent protein kinase II expression.展开更多
Objective: To study synergistic effect with Ascorbic acid(AA) on arsenic trioxide inducing human Hepatocarcinoma cell apoptosis, and provide theoretical basis for promoting human Hepatocarcinoma cell apoptosis induced...Objective: To study synergistic effect with Ascorbic acid(AA) on arsenic trioxide inducing human Hepatocarcinoma cell apoptosis, and provide theoretical basis for promoting human Hepatocarcinoma cell apoptosis induced by arsenic trioxide(AT). Methods: Human Hepatocarcinoma cell line BEL-7402 being cultured in vitro, the effect of AT and (or) AA on its growth inhibition and its two intracellular signal molecules was evaluated separately using MTT and Western blot. Results: AT at a few μmol/L concentration could suppress abnormal proliferation of human hepatocarcinoma cells, and initiate their apoptosis by activation of caspase-3, and activate extracellular-signal regulated kinases (ERKs), which were dependent on the dosage of AT conspicuously. The effect of AA on BEL-7402 was not significant; However, AA could effectively enhance AT-induced hepatocarcinoma cell apoptosis and lesion severity through activation of caspase-3 but not ERKs. Conclusion: Caspase-3 and ERKs proteins could involve in arsenic-induced hepatocarcinoma cell apoptosis and differentiation respectively as intracellular signaling molecules; The effect between AT and AA on hepatocarcinoma is synergistic, which further inhibits cell growth and induces apoptosis in human hepatocarcinoma cells through activation of caspase-3 but not ERKs.展开更多
Objective： To study the mechanisms in gambogic acid （GA） -induced JeKo-1 human Mantle Cell Lymphoma cell apoptosis in vitro. Methods： The proliferation of GA-treated JeKo-1 cells was measured by CCK-8 assay and Ki...Objective： To study the mechanisms in gambogic acid （GA） -induced JeKo-1 human Mantle Cell Lymphoma cell apoptosis in vitro. Methods： The proliferation of GA-treated JeKo-1 cells was measured by CCK-8 assay and Ki-67 immunocytochemical detection. Apopt0sis, cell cycle and mitochondrial membrane potential were measured by flow cytometric analysis. Caspase-3, -8 and -9 were detected by colorimetric assay. Bcl-2 and Bax were analyzed by Western blotting. Results： GA inhibited cell growth in a time- and dose- dependent manner. GA induces apoptosis in JeKo- 1 cells but not in normal bone marrow cells, which was involved in reducing the membrane potential of mitochondria, activating caspases-3, -8 and -9 and decreasing the ratio of Bd-2 and Bax without cell cycle arresting. Conclusions： GA induced apoptosis in human MCL JeKo-1 cells by regulating Bcl-2/Bax and activating caspase-3, -8 and -9 via mitochondrial pathway without affecting cell cycle.展开更多
AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro...AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro were observed by MTT colorimetric method. The morphological changes of apoptosis before and after the treatment of SalB were observed by Hoechst 33258 fluorescent staining method. Apoptosis rate and cell cycle changes of HXO-RB44 cells were detected by flow cytometer at 48 hours after treated by SalB. The expression changes of Caspase-3 protein in HXO-RB44 cells were detected by Western Blot. RESULTS: SalB significantly inhibited the growth of HXO-RB44 cells, while the inhibition was in a concentration-and time-dependent manner. The results of fluorescent staining method indicated that HXO-RB44 cells showed significant phenomenon of apoptosis including karyorrhexis, fragmentation and the formation of apoptotic bodies, etc. after 24, 48 and 72 hours co-culturing of SalB and HXO-RB44 cells. The results of flow cytometer showed that the apoptosis rate and the proportion of cells in S phase were gradually increased at 48 hours and 72 hours after treated by different concentrations of SalB. Western Blot strip showed that the expression of Caspase-3 protein in HXO-RB44 cells was gradually increased with the increase of the concentration of SalB. CONCLUSION: SalB can significantly affect on HXO-RB44 cells growth inhibition and apoptosis induction which may be achieved through the up-regulation of Caspase-3 expression and the induction of cell cycle arrest.展开更多
The effects of betulinic acid (BA), a pentacyclic lupane-type triterpene, on the cell viability, cell cycle and apoptosis in human leukemia K562 cells were investigated. The effects of BA on the growth of K562 cells w...The effects of betulinic acid (BA), a pentacyclic lupane-type triterpene, on the cell viability, cell cycle and apoptosis in human leukemia K562 cells were investigated. The effects of BA on the growth of K562 cells were studied by MTT assay. Apoptosis was assayed through Annexin V/propidium iodide (PI) double-labeled cytometry. The effects of BA on the cell cycle of K562 cells were studied by a PI method. The expression of Bax and capase-3 was detected by using Western blot. The results showed that BA was cytotoxic to K562 cells with an IC50 of 21.26 μg/mL at 24 h. After treating K562 cells with 10 μg/mL BA for 72 h, the number of cells was reduced by 58%. BA induced apoptosis of K562 cells in a time-and dose-dependent manner. The proportion of cells in G0/G1 and G2/M phases was decreased and that in S phase was increased after K562 cells were treated with BA for 24 h. BA treatment also increased the expression of the pro-apoptotic proteins Bax and caspase-3. It suggested that BA could inhibit the proliferation of K562 cells through the induction of cell cycle arrest and apoptosis. The antitumor effects of BA were related with up-regulation of the expression of Bax and caspase-3 proteins. BA may qualify for the development of new therapies for leukemia.展开更多
Ketamine exposure can lead to selective neuroapoptosis in the developing brain.p66ShcA,the cellular adapter protein expressed selectively in immature neurons,is a known pro-apoptotic molecule that triggers neuroapopto...Ketamine exposure can lead to selective neuroapoptosis in the developing brain.p66ShcA,the cellular adapter protein expressed selectively in immature neurons,is a known pro-apoptotic molecule that triggers neuroapoptosis when activated.Sprague-Dawley rats at postnatal day 7 were subcutaneously injected in the neck with ketamine 20 mg/kg,six times at 2-hour intervals.At 0,1,3,and 6 hours after final injection,western blot assay was used to detect the expression of cleaved caspase-3,p66ShcA,and phosphorylated p66ShcA.We found that the expression of activated p66ShcA and caspase-3 increased after ketamine exposure and peaked at 3 hours.The same procedure was performed on a different group of rats.At the age of 4 weeks,spatial learning and memory abilities were tested with the Morris water maze.Latency to find the hidden platform for these rats was longer than it was for control rats,although the residence time in the target quadrant was similar.These findings indicate that ketamine exposure resulted in p66ShcA being activated in the course of an apoptotic cascade during the neonatal period.This may have contributed to the deficit in spatial learning and memory that persisted into adulthood.The experimental protocol was approved by the Institutional Animal Care and Use Committee at the University of Texas at Arlington,USA (approval No.A13.008) on January 22,2013.展开更多
Hypoxia promotes proliferation and differentiation of neural stem cells from embryonic day 12 rat brain tissue, but the concentration and time of hypoxic preconditioning are controversial. To address this, we cultured...Hypoxia promotes proliferation and differentiation of neural stem cells from embryonic day 12 rat brain tissue, but the concentration and time of hypoxic preconditioning are controversial. To address this, we cultured neural stem cells isolated from embryonic day 14 rat cerebral cortex in 5% and 10% oxygen in vitro. MTT assay, neurosphere number, and immunofluorescent staining found that 5% or 10% oxygen preconditioning for 72 hours improved neural stem cell viability and proliferation. With prolonged hypoxic duration （120 hours）, the proportion of apoptotic cells increased. Thus, 5% oxygen preconditioning for 72 hours promotes neural stem cell prolif- eration and neuronal differentiation. Our findings indicate that the optimal concentration and duration of hypoxic preconditioning for promoting proliferation and differentiation of neural stem cells from the cerebral cortex are 5% oxygen for 72 hours.展开更多
Brain cell death after intracerebral hemorrhage may be mediated in part by an apoptotic mechanism Colostrum is the first milk produced by mammals for their young. It plays an important role in protection and developme...Brain cell death after intracerebral hemorrhage may be mediated in part by an apoptotic mechanism Colostrum is the first milk produced by mammals for their young. It plays an important role in protection and development by providing various antibodies, growth factors and nutrients, and has been used for various diseases in many countries. In the present study, we investigated the anti-apoptotic effects of bovine colostrum using organotypic hippocampal slice cultures and an intracerebral hemorrhage animal model. We performed densitometric measurements of propidium iodide uptake, a step-down avoidance task, Nissl staining, and caspase-3 immunohistochemistry. The present results revealed that colostrum treatment significantly suppressed N-methyI-D-aspartic acid-induced neuronal cell death in the rat hippocampus. Moreover, colostrum treatment improved short-term memory by suppressing hemorrhage-induced apoptotic neuronal cell death and decreasing the volume of the lesion induced by intracerebral hemorrhage in the rat hippocampus. These results suggest that colostrum may have a beneficial role in recovering brain function following hemorrhagic stroke by suppressing apoptotic cell death.展开更多
Fetal rat models with neural tube defects were established by injection with retinoic acid at 10 days after conception. The immunofluorescence assay and western blot analysis showed that the number of caspase-3 positi...Fetal rat models with neural tube defects were established by injection with retinoic acid at 10 days after conception. The immunofluorescence assay and western blot analysis showed that the number of caspase-3 positive cells in myeloid tissues for spina bifida manifesta was increased. There was also increased phosphorylation of c-Jun N-terminal kinase, a member of the mitogen activated protein kinase family. The c-Jun N-terminal kinase phosphorylation level was positively correlated with caspase-3 expression in myeloid tissues for spina bifida manifesta. Experimental findings indicate that abnormal apoptosis is involved in retinoic acid-induced dominant spina bifida formation in fetal rats, and may be associated with the c-Jun N-terminal kinase signal transduction pathway.展开更多
Objective： To explore the neuroprotective effects of baicalin against hypoxia and glucose deprivation- reperfusion （OGD/RO）-induced injury in SH-SY5Y cells. Methods： SH-SY5Y cells were divided into a control group...Objective： To explore the neuroprotective effects of baicalin against hypoxia and glucose deprivation- reperfusion （OGD/RO）-induced injury in SH-SY5Y cells. Methods： SH-SY5Y cells were divided into a control group, a OGD/RO group, which was subject to OGD/RO induction; and 3 baicalin groups subject to baicalin （1, 5, 25 μmol/L） for 2 h before induction of OGD/RO （low-, medium-, and high-dose baicalin groups）. Cell viability was detected by thiazolyl blue tetrazoliurn bromide （MTT） assay and flow cytometric analysis was used to detect cell apoptosis. Real-time polymerase chain reaction was performed to determine the mRNA expression of caspase-3 gene. Westem blot analysis was conducted to determine the expression of nuclear factor （NF）- κB and N-methyl-d- aspartic acid receptor-1 （NMDAR1）. Results： Baicalin could significantly attenuate OGD/RO mediated apoptotic cell death in SH-SY5Y cells; the apoptosis rates in the low-, medium- and high-dose groups were 12.1%, 7.9%, and 5.4%, respectively. Western blot and real-time PCR analysis revealed that significant decrease in caspase-3 expression in the baicalin group compared with the OGD/RO group （P〈0.01）. Additionally, down-regulation of NF-K B and NMDAR1 was observed in the baicalin group compared with those obtained from the OGD/RO group. Compared with the low-dose baicalin group, remarkable decrease was noted in the medium- and high-dose groups （P〈0.01）. Conclusion： Baicalin pre-treatment attenuates brain ischemia reperfusion injury by suppressing cellular apoptosis.展开更多
文摘BACKGROUND: Some scholars believed that the neuronal injury after status epilepticus is apoptosis, the main evidence is the changes of expressions of various apoptosis related genes, such as immediate-early gene, p53 gene and genes of bcl-2 family, etc. But there is still no ultrastructural evidence for apoptosis. OBJECTIVE: To observe the ultrastructural damages of mitochondrion and nucleus and the changes of caspase expression in neurons of hippocampal CA3 region in rats with status epilepticus induced by kainic acid. DESIGN: A randomized controlled study. SETTING: Department of Anesthesiology and Department of Neurology, Qilu Hospital of Shandong University. MATERIALS: Seventy-five adult male Wistar rats of 250-300 g, clean degree, were provided by the experimental animal center of Shandong University. Kainic acid was purchased from Sigma Company (USA); rabbit anti-rat polyclonal antibody caspase-3 from Santa Cruz Company (USA). METHODS: The experiments were carried out in the Department of Anesthesiology, Qilu Hospital of Shandong University from October 2005 to February 2006. ① The 75 rats were randomly divided into experimental group (n =45) and control group (n =30). ② Model establishment, convulsion grading and the judging standards for status epilepticus: Rats in the experimental group were given intraperitoneal injection of kainic acid (10 mg/kg), and those in the control group were injected with saline of the same volume. The time of seizure was recorded and their behavioral manifestations were observed, and the seizure was terminated by intraperitoneal injection of diazepam (10 mg/kg). ③ Observation under electron microscope: At 3, 12 and 24 hours after status epilepticus respectively, bilateral hippocampal tissues were taken out, semithin sections of about 75 nm were prepared after fixation, dehydration and embedding, and then observed under H-800 transmission electron microscope. ④ Immunohistochemical detection: Bilateral hippocampi were removed at 3, 12 and 24 hours after status epilepticus respectively, the fixation, dehydration, transparence, wax immersion and embedding were performed, then serial sections of CA3 region were immunohistochemically determined by the SABC method. Leica QWinV3 image analytical software was applied, then the average number and average gray value of positive cells were calculated. MAIN OUTCOME MEASURES: Results of observation under electron microscope, that of immunohistochemical staining of neurons in hippocampal CA3 region; Comparison of number of caspase-3 positive cells and gray value. RESULTS: All the 75 Wistar rats were involved in the analysis of results. ① Results of observation under electron microscope: At 3 hours after status epilepticus, swelling crista and membranous disintegration were observed under electron microscope. At 24 hours, obvious nuclear changes occurred, and manifested as the side-aggegation of chromatins. ② Results of immunohistochemical detection: In the experimental group, the number of caspase-3 positive cells at 3 hours after status epilepticus had no obvious difference as compared with that in the control group (P > 0.05); At 12 hours, the number and gray value of caspase-3 positive cells in the experimental group were higher than those in the control group (10.49±0.68 vs. 5.33±0.43; 45.57±2.27 vs. 19.79±0.33, P < 0.05), the same results were also observed at 24 hours (37.36±0.57 vs. 5.12±0.47; 115.24±1.22 vs. 18.73±0.42, P < 0.01). CONCLUSION: In the rat models of status epilepticus induced by kainic acid, mitochondrial damage was earlier than the increase of caspase-3 expression and nuclear changes, which suggested that mitochondrion was the key link for the neuronal death after status epilepticus.
文摘The clinical presentation of schizophrenia involves a variety of symptoms, which in many cases include hallucinations and delusions. Experimentally revealed alterations in both pre-pulse inhibition (PPI) and latent inhibition (LI) are also apparent in individuals afflicted with this disorder. Many have speculated that altered synaptic connections are, in part, responsible for this subset of behavioral abnormalities. We have previously reported that neonatal chronic low-dose injections of domoic acid (DOM) produce adult rats with deficits in PPI and LI. The current study was conducted to determine whether this toxin-treatment would alter the degree of apoptosis occurring in the developing brain. Results revealed significant decreases in caspase-3 within the right prelimbic cortex (PrL) in both male and female DOM-treated rats suggesting that even modest alterations in glutamate (Glu) signaling during critical periods of central nervous system (CNS) maturation will modify ontogenetic processes in the prefrontal cortex (PFC) of the juvenile rat.
基金supported by Liaoning Social Development Key Projects of Scientific and Technological Department of Liaoning Province, No. 2012225019
文摘In this study, primary cultured cerebral cortical neurons of Sprague-Dawley neonatal rats were treated with 0.25, 0.5, and 1.0 μM calmodulin-dependent protein kinase II inhibitor KN-93 after 50 μM N-methyI-D-aspartic acid-induced injury. Results showed that, compared with N-methyi-D- aspartic acid-induced injury neurons, the activity of cells markedly increased, apoptosis was significantly reduced, leakage of lactate dehydrogenase decreased, and intracellular Ca2＋ concentrations in neurons reduced after KN-93 treatment. The expression of caspase-3, phosphorylated calmodulin-dependent protein kinase II and total calmodulin-dependent protein kinase II protein decreased after KN-93 treatment. And the effect was apparent at a dose of 1.0 pM KN-93. Experimental findings suggest that KN-93 can induce a dose-dependent neuroprotective effect, and that the underlying mechanism may be related to the down-regulation of caspase-3 and calmodulin- dependent protein kinase II expression.
文摘Objective: To study synergistic effect with Ascorbic acid(AA) on arsenic trioxide inducing human Hepatocarcinoma cell apoptosis, and provide theoretical basis for promoting human Hepatocarcinoma cell apoptosis induced by arsenic trioxide(AT). Methods: Human Hepatocarcinoma cell line BEL-7402 being cultured in vitro, the effect of AT and (or) AA on its growth inhibition and its two intracellular signal molecules was evaluated separately using MTT and Western blot. Results: AT at a few μmol/L concentration could suppress abnormal proliferation of human hepatocarcinoma cells, and initiate their apoptosis by activation of caspase-3, and activate extracellular-signal regulated kinases (ERKs), which were dependent on the dosage of AT conspicuously. The effect of AA on BEL-7402 was not significant; However, AA could effectively enhance AT-induced hepatocarcinoma cell apoptosis and lesion severity through activation of caspase-3 but not ERKs. Conclusion: Caspase-3 and ERKs proteins could involve in arsenic-induced hepatocarcinoma cell apoptosis and differentiation respectively as intracellular signaling molecules; The effect between AT and AA on hepatocarcinoma is synergistic, which further inhibits cell growth and induces apoptosis in human hepatocarcinoma cells through activation of caspase-3 but not ERKs.
基金supported by a grant from the Key Project supported by medical science and technology development Foundation of Nanjing Department of Health (No. ZKX09016)
文摘Objective： To study the mechanisms in gambogic acid （GA） -induced JeKo-1 human Mantle Cell Lymphoma cell apoptosis in vitro. Methods： The proliferation of GA-treated JeKo-1 cells was measured by CCK-8 assay and Ki-67 immunocytochemical detection. Apopt0sis, cell cycle and mitochondrial membrane potential were measured by flow cytometric analysis. Caspase-3, -8 and -9 were detected by colorimetric assay. Bcl-2 and Bax were analyzed by Western blotting. Results： GA inhibited cell growth in a time- and dose- dependent manner. GA induces apoptosis in JeKo- 1 cells but not in normal bone marrow cells, which was involved in reducing the membrane potential of mitochondria, activating caspases-3, -8 and -9 and decreasing the ratio of Bd-2 and Bax without cell cycle arresting. Conclusions： GA induced apoptosis in human MCL JeKo-1 cells by regulating Bcl-2/Bax and activating caspase-3, -8 and -9 via mitochondrial pathway without affecting cell cycle.
基金National "Eleventh Five-year Plan" Science and Technology Support Project (No. 2006BAI06 A15-3)
文摘AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro were observed by MTT colorimetric method. The morphological changes of apoptosis before and after the treatment of SalB were observed by Hoechst 33258 fluorescent staining method. Apoptosis rate and cell cycle changes of HXO-RB44 cells were detected by flow cytometer at 48 hours after treated by SalB. The expression changes of Caspase-3 protein in HXO-RB44 cells were detected by Western Blot. RESULTS: SalB significantly inhibited the growth of HXO-RB44 cells, while the inhibition was in a concentration-and time-dependent manner. The results of fluorescent staining method indicated that HXO-RB44 cells showed significant phenomenon of apoptosis including karyorrhexis, fragmentation and the formation of apoptotic bodies, etc. after 24, 48 and 72 hours co-culturing of SalB and HXO-RB44 cells. The results of flow cytometer showed that the apoptosis rate and the proportion of cells in S phase were gradually increased at 48 hours and 72 hours after treated by different concentrations of SalB. Western Blot strip showed that the expression of Caspase-3 protein in HXO-RB44 cells was gradually increased with the increase of the concentration of SalB. CONCLUSION: SalB can significantly affect on HXO-RB44 cells growth inhibition and apoptosis induction which may be achieved through the up-regulation of Caspase-3 expression and the induction of cell cycle arrest.
基金supported by a grant from the National Natural Sciences Foundation of China (No. 30500686)
文摘The effects of betulinic acid (BA), a pentacyclic lupane-type triterpene, on the cell viability, cell cycle and apoptosis in human leukemia K562 cells were investigated. The effects of BA on the growth of K562 cells were studied by MTT assay. Apoptosis was assayed through Annexin V/propidium iodide (PI) double-labeled cytometry. The effects of BA on the cell cycle of K562 cells were studied by a PI method. The expression of Bax and capase-3 was detected by using Western blot. The results showed that BA was cytotoxic to K562 cells with an IC50 of 21.26 μg/mL at 24 h. After treating K562 cells with 10 μg/mL BA for 72 h, the number of cells was reduced by 58%. BA induced apoptosis of K562 cells in a time-and dose-dependent manner. The proportion of cells in G0/G1 and G2/M phases was decreased and that in S phase was increased after K562 cells were treated with BA for 24 h. BA treatment also increased the expression of the pro-apoptotic proteins Bax and caspase-3. It suggested that BA could inhibit the proliferation of K562 cells through the induction of cell cycle arrest and apoptosis. The antitumor effects of BA were related with up-regulation of the expression of Bax and caspase-3 proteins. BA may qualify for the development of new therapies for leukemia.
基金supported by the National Natural Science Foundation of China,No.81200851(to DL)the National Institutes of Health of the USA,No.NS 040723(to QL)
文摘Ketamine exposure can lead to selective neuroapoptosis in the developing brain.p66ShcA,the cellular adapter protein expressed selectively in immature neurons,is a known pro-apoptotic molecule that triggers neuroapoptosis when activated.Sprague-Dawley rats at postnatal day 7 were subcutaneously injected in the neck with ketamine 20 mg/kg,six times at 2-hour intervals.At 0,1,3,and 6 hours after final injection,western blot assay was used to detect the expression of cleaved caspase-3,p66ShcA,and phosphorylated p66ShcA.We found that the expression of activated p66ShcA and caspase-3 increased after ketamine exposure and peaked at 3 hours.The same procedure was performed on a different group of rats.At the age of 4 weeks,spatial learning and memory abilities were tested with the Morris water maze.Latency to find the hidden platform for these rats was longer than it was for control rats,although the residence time in the target quadrant was similar.These findings indicate that ketamine exposure resulted in p66ShcA being activated in the course of an apoptotic cascade during the neonatal period.This may have contributed to the deficit in spatial learning and memory that persisted into adulthood.The experimental protocol was approved by the Institutional Animal Care and Use Committee at the University of Texas at Arlington,USA (approval No.A13.008) on January 22,2013.
基金supported by the Science Foundation of Jining Science and Technology Bureau of China,No.2012jnjc07
文摘Hypoxia promotes proliferation and differentiation of neural stem cells from embryonic day 12 rat brain tissue, but the concentration and time of hypoxic preconditioning are controversial. To address this, we cultured neural stem cells isolated from embryonic day 14 rat cerebral cortex in 5% and 10% oxygen in vitro. MTT assay, neurosphere number, and immunofluorescent staining found that 5% or 10% oxygen preconditioning for 72 hours improved neural stem cell viability and proliferation. With prolonged hypoxic duration （120 hours）, the proportion of apoptotic cells increased. Thus, 5% oxygen preconditioning for 72 hours promotes neural stem cell prolif- eration and neuronal differentiation. Our findings indicate that the optimal concentration and duration of hypoxic preconditioning for promoting proliferation and differentiation of neural stem cells from the cerebral cortex are 5% oxygen for 72 hours.
文摘Brain cell death after intracerebral hemorrhage may be mediated in part by an apoptotic mechanism Colostrum is the first milk produced by mammals for their young. It plays an important role in protection and development by providing various antibodies, growth factors and nutrients, and has been used for various diseases in many countries. In the present study, we investigated the anti-apoptotic effects of bovine colostrum using organotypic hippocampal slice cultures and an intracerebral hemorrhage animal model. We performed densitometric measurements of propidium iodide uptake, a step-down avoidance task, Nissl staining, and caspase-3 immunohistochemistry. The present results revealed that colostrum treatment significantly suppressed N-methyI-D-aspartic acid-induced neuronal cell death in the rat hippocampus. Moreover, colostrum treatment improved short-term memory by suppressing hemorrhage-induced apoptotic neuronal cell death and decreasing the volume of the lesion induced by intracerebral hemorrhage in the rat hippocampus. These results suggest that colostrum may have a beneficial role in recovering brain function following hemorrhagic stroke by suppressing apoptotic cell death.
基金supported by the National Natural Science Foundation of China, No.30872705/HD426 and No.81070538/HD429
文摘Fetal rat models with neural tube defects were established by injection with retinoic acid at 10 days after conception. The immunofluorescence assay and western blot analysis showed that the number of caspase-3 positive cells in myeloid tissues for spina bifida manifesta was increased. There was also increased phosphorylation of c-Jun N-terminal kinase, a member of the mitogen activated protein kinase family. The c-Jun N-terminal kinase phosphorylation level was positively correlated with caspase-3 expression in myeloid tissues for spina bifida manifesta. Experimental findings indicate that abnormal apoptosis is involved in retinoic acid-induced dominant spina bifida formation in fetal rats, and may be associated with the c-Jun N-terminal kinase signal transduction pathway.
基金Supported by Seed Fund of the 2nd Hospital of Shandong University(No.S2013010002)
文摘Objective： To explore the neuroprotective effects of baicalin against hypoxia and glucose deprivation- reperfusion （OGD/RO）-induced injury in SH-SY5Y cells. Methods： SH-SY5Y cells were divided into a control group, a OGD/RO group, which was subject to OGD/RO induction; and 3 baicalin groups subject to baicalin （1, 5, 25 μmol/L） for 2 h before induction of OGD/RO （low-, medium-, and high-dose baicalin groups）. Cell viability was detected by thiazolyl blue tetrazoliurn bromide （MTT） assay and flow cytometric analysis was used to detect cell apoptosis. Real-time polymerase chain reaction was performed to determine the mRNA expression of caspase-3 gene. Westem blot analysis was conducted to determine the expression of nuclear factor （NF）- κB and N-methyl-d- aspartic acid receptor-1 （NMDAR1）. Results： Baicalin could significantly attenuate OGD/RO mediated apoptotic cell death in SH-SY5Y cells; the apoptosis rates in the low-, medium- and high-dose groups were 12.1%, 7.9%, and 5.4%, respectively. Western blot and real-time PCR analysis revealed that significant decrease in caspase-3 expression in the baicalin group compared with the OGD/RO group （P〈0.01）. Additionally, down-regulation of NF-K B and NMDAR1 was observed in the baicalin group compared with those obtained from the OGD/RO group. Compared with the low-dose baicalin group, remarkable decrease was noted in the medium- and high-dose groups （P〈0.01）. Conclusion： Baicalin pre-treatment attenuates brain ischemia reperfusion injury by suppressing cellular apoptosis.
