Induced Apoptosis In PC12 Cells Research Articles

The gut microbiota metabolite butyrate mitigates MPTP/MPP+ -induced Parkinson's disease by inhibiting the JAK2/STAT3 signaling pathway.

Butyrate (BU), a gut microbiota-derived metabolite, has been reported to play a neuroprotective role in Parkinson's disease (PD). However, the specific molecular mechanism of BU has not been fully interpreted. This work aimed to verify the protective effects of BU against MPTP/MPP+ -induced neurotoxicity and explore the mechanisms involved. The results showed that BU protected against MPTP-induced motor dysfunction and decreased tyrosine hydroxylase (TH) and dopamine transporter (DAT) levels. Additionally, BU pretreatment improved PC12 cell viability and reduced MPP+ -induced PC12 cell apoptosis. BU treatment also attenuated MPP+ -stimulated oxidative stress and inflammatory response in PC12 cells. Furthermore, BU inhibited MPTP/MPP+ -induced hyperactivation of the JAK2/STAT3 signaling in mice and PC12 cells. Besides, a JAK2 agonist, Coumermycin A1 (C-A1), substantially reversed BU-mediated inhibition on JAK2/STAT3 phosphorylation in MPP+ -challenged PC12 cells and abated BU-induced repression on MPP+ -triggered apoptosis, oxidative stress, and inflammatory response in PC12 cells. To sum up, BU might exert neuroprotective effects against MPP+ /MPTP-induced neurotoxicity by inactivating the JAK2/STAT3 signaling.

Microglia-induced autophagic death of neurons via IL-6/STAT3/miR-30d signaling following hypoxia/ischemia.

There is a relationship between autophagy and the occurrence, maintenance, and progression of several neurodegenerative diseases. The activation of microglia after ischemia contributes to neuronal injury via proinflammatory cytokines and neurotoxic elements. The purpose of this study was to evaluate the function of autophagy in the microglia-mediated death of neuronal cells. Microglial activation by oxygen/glucose deprivation induced both apoptosis and autophagy in neuron-like PC12 cells. Microglia-derived interleukin (IL)-6 induced PC12 cell apoptosis in vitro; however, this effect was inhibited by the autophagy inhibitor chloroquine. Further analysis demonstrated that miR-30d in PC12 cells suppressed microglia-induced PC12 apoptosis and autophagy by directly targeting autophagy protein 5. Moreover, microglia-derived IL-6 activated signal transducer and activator of transcription 3 (STAT3), which can then directly repress miR-30d genes via a conserved STAT3-binding site in its promoter, thereby promoting PC12 cell autophagy and apoptosis. Our study identified IL-6-dependent autophagy-related signaling between microglia and neurons, which contributed to neuronal apoptosis. Importantly, we also provided potential therapeutic targets for ischemic treatment via the interruption of proinflammatory signaling.

Protective Effects of Marine Alkaloid Neolamellarin A Derivatives against Glutamate Induced PC12 Cell Apoptosis.

Marine alkaloids obtained from sponges possess a variety of biological activities and potential medicinal value. The pyrrole-derived lamellarin-like alkaloids, especially their permethyl derivatives, show low cytotoxicity and potent MDR reversing activity. Neolamellarin A is a novel lamellarin-like alkaloid which was extracted from marine animal sponges. We reported the synthetic method of permethylated Neolamellarin A and its derivatives by a convergent strategy in 2015. In 2018, we reported the synthesis and the neuroprotective activity in PC12 cells of 3,4-bisaryl-N-alkylated permethylated Neolamellarin A derivatives. In this report, another series of 15 different 3,4-bisaryl-N-acylated permethylated Neolamellarin A derivatives were synthesized, and the outstanding protective effects of these compounds against glutamate induced PC12 cell apoptosis were presented and discussed. These Neolamellarin A derivatives which possessed low cytotoxicity and superior neuroprotective activity may have the potential to be developed into antagonists against glutamate induced nerve cell apoptosis.

Pleiotrophin affects the susceptibility of prostate cancer cells to cisplatin.

Drug resistance is a major problem in cancer therapy with cisplatin. It has not been reported that pleiotrophin, which is anti-apoptotic in some cancer cells, is associated with cisplatin resistance. Pleiotrophin was exogenously expressed in 293 cells. Viability and apoptosis of PC3 cells treated with different concentrations of cisplatin in the presence or absence of purified pleiotrophin were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, respectively. PC3 cells transfected with shRNAs were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and western blotting 24 h after transfection. MTT assay data indicated that the EC50 value of cisplatin for PC3 cells was significantly increased in the presence of pleiotrophin. Flow cytometry data demonstrated the pleiotrophin dose-dependent anti-apoptosis in PC3 cells treated with cisplatin. Knockdown of pleiotrophin with sh-RNA, as justified by RT-PCR and western blotting analysis, led to increased cisplatin induced-apoptosis in PC3 cells with an increased level of the cleaved poly ADP-ribose polymerase protein. Pleiotrophin may be a potential antiapoptotic protein associated with cisplatin susceptibility, which warrants further study on the role of pleiotrophin in cisplatin resistance.

Caffeine‐induced neurotoxicity mediated by Nrf2 pathway in PC12 cells and zebrafish larvae

Caffeine is one of the most widely used psychostimulants in the world and possesses central excitative, anti-depressive, and neuroprotective properties. However, excessive ingestion or abuse of caffeine can lead to intoxication. Many toxic effects are attributed to oxidative damage, and nuclear factor erythroid 2-related factor 2 (Nrf2) is a critical intracellular regulator of the oxidative stress response. Here, we investigated the neurotoxicity of caffeine in rat pheochromocytoma PC12 cells and zebrafish larvae. It was found that caffeine inhibited the viability of PC12 cells in a dose- and time-dependent manner. Furthermore, it induced PC12 cell apoptosis and elevated reactive oxygen species (ROS) production. Quantitative polymerase chain reaction (qPCR) and western blotting revealed that caffeine also inhibited the expression levels of Nrf2 mRNA and protein and its target genes (e.g., NADPH quinone oxidoreductase 1 [NQO1]). Furthermore, Nrf2 silencing attenuated the toxic effects of caffeine. In addition, zebrafish larvae were treated with different doses of caffeine. Behavioral experiments showed that a low dose of caffeine (0.05 to 0.3 mM) increased the average distance of movement and promoted excitation. Survivorship curves showed that caffeine (0.2 to 1.5 mM) caused lethality. Finally, qPCR revealed that a higher dose of caffeine inhibited mRNA levels in the Nrf2 pathway. Based on these results, this study identified for the first time that overuse of caffeine can induce neurotoxicity by inhibiting the Nrf2 pathway. These results will provide a new perspective for studies on caffeine toxicity.

Bone marrow mesenchymal stem cells-derived exosomes reduce apoptosis and inflammatory response during spinal cord injury by inhibiting the TLR4/MyD88/NF-κB signaling pathway.

Spinal cord injury (SCI) is one of the most common destructive injuries, which may lead to permanent neurological dysfunction. Currently, transplantation of bone marrow mesenchymal stem cells (BMSCs) in experimental models of SCI shows promise as effective therapies. BMSCs secrete various factors that can regulate the microenvironment, which is called paracrine effect. Among these paracrine substances, exosomes are considered to be the most valuable therapeutic factors. Our study found that BMSCs-derived exosomes therapy attenuated cell apoptosis and inflammation response in the injured spinal cord tissues. In in vitro studies, BMSCs-derived exosomes significantly inhibited lipopolysaccharide (LPS)-induced PC12 cell apoptosis, reduced the secretion of pro-inflammatory factors including tumor necrosis factor (TNF)-α and IL (interleukin)-1β and promoted the secretion of anti-inflammatory factors including IL-10 and IL-4. Moreover, we found that LPS-induced protein expression of toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear transcription factor-κB (NF-κB) was significantly downregulated after treatment with BMSCs-derived exosomes. In in vivo studies, we found that hindlimb motor function was significantly improved in SCI rats with systemic administration of BMSCs-derived exosomes. We also observed that the expression of pro-apoptotic proteins and pro-inflammatory factors was significantly decreased, while the expression of anti-apoptotic proteins and anti-inflammatory factors were upregulated in SCI rats after exosome treatment. In conclusion, BMSCs-derived exosomes can inhibit apoptosis and inflammation response induced by injury and promote motor function recovery by inhibiting the TLR4/MyD88/NF-κB signaling pathway, which suggests that BMSCs-derived exosomes are expected to become a new therapeutic strategy for SCI.

Protective Effects of Crataegus pinnatifida Extracts and Crataegolic Acid Against Neurotoxicity of Paraquat in PC12 Cells

The neuroprotective mechanisms of Crataegus pinnatifida extracts and crataegolic acid were studied using paraquat induced cytotoxicity in PC12 cells. C. pinnatifida extracts were prepared using hexane, ethyl acetate, and 95% ethanol. Additionally, crataegolic acid (also known as maslinic acid) was found in C. pinnatifida extracts. Assessment methods included the examinations of cytotoxicity, intracellular reactive oxygen species and calcium changes, activity of caspase-3 and α-synuclein, apoptotic cell death, and the expression levels of the B-cell lymphoma 2 (Bcl-2) and BCL2-associated X (Bax) proteins to investigate the neuroprotective mechanisms of C. pinnatifida extracts and its active component, crataegolic acid. The three extracts and crataegolic acid exhibited potent neuroprotective actions against paraquat induced PC12 cell apoptosis at 5–20µg/mL and 80–100µM concentrations, respectively. The key protective mechanisms included decreasing cell apoptosis, upregulating Bcl-2 protein levels, and downregulating Bax protein levels. The 95% ethanol extract also decreased paraquat induced reactive oxygen species production, calcium overloading, and caspase-3 and α-synuclein activities. The beneficial effects of these extracts could be explained by the active component, crataegolic acid that also inhibited paraquat-induced apoptosis through the suppression of reactive oxygen species generation and the caspase-3 signaling pathway.

Longxuetongluo Capsule protects against cerebral ischemia/reperfusion injury through endoplasmic reticulum stress and MAPK-mediated mechanisms

IntroductionLongxuetongluo Capsule (LTC) is wildly applied to treat ischemic stroke in clinical practice in China. However, the pharmacological mechanism of LTC on ischemic stroke is still unstated. ObjectiveOur research was designed to study the protective effect of LTC against cerebral ischemia–reperfusion (I/R) injury and reveal the underlying mechanism both in vivo and in vitro. MethodsPC12 cells treated with glucose deprivation/reperfusion (OGD/R) were used to simulate in vitro ischemia/reperfusion (I/R) injury. The cell viability, apoptosis rate, and protein expressions of PC12 cells were evaluated. In vivo validation of the protective effect of LTC was carried out by middle cerebral artery occlusion (MCAO)/reperfusion treatment, and the underlying mechanism of its anti-apoptosis ability was further revealed by immunohistochemistry staining and Western blotting. ResultsIn the current study, we observed that LTC effectively inhibited oxygen-glucose deprivation/reperfusion (OGD/R) induced apoptosis of PC12 cells through suppressing the cleavage of poly ADP-ribose polymerase (PARP), caspase-3, and caspase-9. Further investigation revealed that OGD/R insult remarkably triggered the endoplasmic reticulum stress responses (ER stress) to induce PC12 cell apoptosis. LTC treatment alleviated OGD/R induced ER stress by inhibiting the activation of protein kinase RNA (PKR)-like ER kinase (PERK)/eukaryotic translation initiation factor 2 (eIF2α) and inositol requiring enzyme 1 (IRE1)/tumor necrosis factor receptor-associated factor 2 (TRAF2) pathways. Additionally, LTC also restrained the OGD/R-induced PC12 cell apoptosis by reversing the activated mitogen-activated protein kinase (MAPK) through IRE1/TRAF2 pathway. Animal studies demonstrated LTC significantly restricted the infarct region induced by middle cerebral artery occlusion (MCAO)/reperfusion, the activation of ER stress and apoptosis of neuronal cells had also been suppressed by LTC in the penumbra region. ConclusionLTC protects the cerebral neuronal cell against ischemia/reperfusion injury through ER stress and MAPK-mediated mechanisms.

Plasma miR-409-3p promotes acute cerebral infarction via suppressing CTRP3.

Abnormal expression of miR-409-3p has been found in several neurodevelopmental disorders, but whether it is dysregulated in the patients with acute cerebral infarction (ACI) has not been evaluated. The current study mainly focused on the clinical significance and the underlying mechanism of plasma miR-409-3p in the progression of ACI. The level of plasma miR-409-3p was determined in ACI patients (n = 80) and healthy controls (n = 30). Pearson correlation assay was performed to evaluate the association and cardiovascular risk factors. A receiver operating characteristic curve (ROC) was used to evaluate the diagnostic value of plasma miR-409-3p levels in patients with ACI. Dual luciferase reporter assay and western blot were performed to determine the possible target gene of miR-409-3p. Our data showed that the expression of plasma miR-409-3p in the ACI group was higher than that in the healthy controls. Furthermore, Pearson correlation analysis indicated a positive correlation between plasma miR-409-3p and the NIHSS score. ROC analysis indicated that plasma miR-409-3p could differentiate plasma miR-409-3p in ACI patients from healthy controls. Then, we explored the possible target genes of miR-409-3p. Interestingly, C1q and TNF-related 3 (CTRP3), a novel adipose tissue-derived secreted factor, was found to be a target gene of miR-409-3p. We found that knockdown of CTRP3 significantly induced PC12 cell apoptosis, even in PC12 cells transfected with miR-409-3p inhibitor. These data suggested that miR-409-3p induced PC12 cell apoptosis by targeting CTRP3. Altogether, elevated plasma miR-409-3p is correlated with disease severity and may be efficient for the early diagnosis of ACI.

IncRNA NEAT1 aggravates cerebral ischemia/reperfusion injury by sponging miR-874-3p.

Many lncRNAs have been reported to affect cerebral ischemia/reperfusion (I/R) injury. The purpose of this study is to elucidate the role of lncRNA NEAT1 as well as the regulatory mechanism of lncRNA NEAT1/miR-874-3p in cerebral I/R injury. A cellular model of cerebral I/R injury was built. RT-qPCR was used to detect NEAT1 and miR-874-3p expression. Cell viability was detected by MTT assay. The expression of apoptosis-related proteins (Bcl-2 and Bax) was measured by Western blot analysis. The relationship between NEAT1 and miR-874-3p was confirmed by dual luciferase reporter assay. We found that LncRNA NEAT1 was upregulated in the PC12 cells treated by I/R and upregulation of lncRNA NEAT1 can aggravate I/R injury of PC12 cells. Additionally, lncRNA NEAT1 overexpression decreased cell viability and induced apoptosis in PC12 cells treated by I/R. Furthermore, miR-874-3p was confirmed to be a target of lncRNA NEAT1. mR-874-3p and NEAT1 expression are found to be reciprocally inhibited in PC12 cells. In summary, LncRNA NEAT1 aggravates cerebral I/R injury by suppressing miR-874-3p expression.

Formononetin induces apoptosis of PC-3 human prostate cancer cells via regulating long noncoding RNA H19 and the mitochondrial apoptosis pathway

Background: Prostate cancer is a life-threating disease with high incidence and mortality in male. Formononetin, the main active component of some natural products, has been hypothesized as a promising anticancer agent in previous studies. Objectives: We investigated the toxic effects and potential molecular mechanism of formononetin in PC-3 prostate cancer cells to further understand the pharmacological effects of formononetin and provide more references for intensive research. Materials and Methods: PC-3 cells were incubated with different doses of formononetin for 24 h or 48 h. After that, cell viability was measured by Cell Counting Kit-8, and apoptosis was analyzed by Hoechst 33258 stains. The expression levels of tumor-related factors such as long noncoding RNA (LncRNA) H19, Bax, and Bcl-2 were determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot methods. Subsequently, PC-3 cells were infected with a lentiviral vector to overexpress or knock down H19, and then, the expression of insulin-like growth factor-1 receptor (IGF-1R) mRNA was measured by RT-qPCR. Results: Formononetin significantly inhibited the viability of PC-3 cells and promoted apoptosis in a time-dose-dependent manner. We observed that the expressions of lncRNA H19 and Bcl-2 were significantly downregulated compared with the untreated group, while an opposite pattern was observed for Bax. According to the results of gene interaction experiments, IGF-1R may be a downstream target of H19 in PC-3 cells. Conclusion: Our results present evidence that formononetin induced apoptosis of PC-3 cells by regulating lncRNA H19 and the mitochondrial apoptosis pathway. Furthermore, we put forward the hypothesis that formononetin has an interference effect on the H19/IGF-1R pathway, which remains to be further confirmed.

Neuroprotective effect of green tea extractives against oxidative stress by enhancing the survival and proliferation of PC12 cells

Previous studies were mainly focused on the effect of catechins and caffeine, the main pharmacological ingredients of green tea. However, the ordinary way of taking green tea is by drinking the tea water. Hence the benefits of water extractives of green tea were investigated in the current study. H2O2 was applied to induce the oxidative stress on PC12 cells. The neuroprotective effects of green tea against oxidative stress were determined by observing the biological behaviors of PC12 cells. With the pre-treatment of light green tea, the survival and proliferation of oxidative stressed PC12 cells were significantly enhanced. H2O2 induced PC12 cell apoptosis was completely reversed. Consistently, the expression of JNK was up-regulated while the caspase-3 was down-regulated. These results suggested that drinking green tea, particularly the light green tea, performed neuroprotective effects against oxidative stress.

Cathepsin L induced PC-12 cell apoptosis via activation of B-Myb and regulation of cell cycle proteins

Cathepsin L (CTSL), a cysteine protease, is responsible for the degradation of a variety of proteins. It is known to participate in neuronal apoptosis associated with abnormal cell cycle. However, the mechanisms underlying CTSL-induced cell apoptosis remain largely unclear. We reported here that rotenone caused an activation of CTSL expression in PC-12 cells, while knockdown of CTSL by small interfering RNAs or its inhibitor reduced the rotenone-induced cell cycle arrest and apoptosis. Moreover, elevation of CTSL and increased-apoptosis were accompanied by induction of B-Myb, a crucial cell cycle regulator. We found that B-Myb was increased in rotenone-treated PC-12 cells and knockdown of B-Myb ameliorated rotenone-stimulated cell apoptosis. Further analysis demonstrated that CTSL influenced the expression of B-Myb as suppression of CTSL activity led to a decreased B-Myb expression, whereas overexpression of CTSL resulted in B-Myb induction. Reduction of B-Myb in CTSL-overexpressing cells revealed that regulation of cell cycle-related proteins, including cyclin A and cyclin B1, through CTSL was mediated by the transcription factor B-Myb. In addition, we demonstrated that the B-Myb target, Bim, and its regulator, Egr-1, which was also associated with CTSL closely, were both involved in rotenone-induced apoptosis in PC-12 cells. Our data not only revealed the role of CTSL in rotenone-induced neuronal apoptosis, but also indicated the involvement of B-Myb in CTSL-related cell cycle regulation.

Protective effects of ethyl gallate on H2O2-induced mitochondrial dysfunction in PC12 cells.

Oxidative stress has been suggested to play an important role in neuronal injury. Ethyl gallate (EG) is the ethyl ester of gallic acid which has been acknowledged as an antioxidant. We previously demonstrated that EG effectively inhibited H2O2-induced cytotoxicity and decreased the ROS levels in PC12 cells, while the relevant mechanisms of action of this compound remain largely uncharacterized. The present study was carried out in an attempt to clarify the underlying mechanisms of EG against H2O2-induced neurotoxicity in PC12 cells. EG pretreatment attenuated H2O2-induced mitochondrial dysfunction as indicated by the decreased caspase-9/-3 activation, PARP cleavage, mitochondrial membrane potential (MMP) depletion, Bax/Bcl-2 ratio, cytochrome c release and ROS overproduction. Furthermore, EG treatment resulted in nuclear translocation of Nrf2 along with increased expression of ARE-dependent cytoprotective genes, such as γ-GCS and NQO1, which indicated EG as an Nrf2 pathway activator. Silencing of Nrf2 signaling by siRNA abrogated the protective effects offered by EG on H2O2-induced PC12 cells injury, which suggested the important role of Nrf2 pathway in the protection of EG against oxidative stress induced PC12 cell apoptosis. These results taken together indicated that EG protects PC12 cells against H2O2-induced cell mitochondrial dysfunction possibly through activation of Nrf2 pathway. EG might be a potential candidate for further preclinical study aimed at the prevention and treatment of neurodegenerative diseases.

Tenofovir disoproxil fumarate induces pheochromocytoma cells apoptosis

Despite the triumph of highly active antiretroviral therapy (HAART) in anti-HIV infection, more than half of the HIV infection individuals receiving antiretroviral therapy acquire HIV-associated neurocognitive disorder (HAND). Previously researches had reported that the HAART neurotoxicity is implicated in HAND-related morbidity. The molecular mechanism of HAND is not clear. Tenofovir disoproxil fumarate (TDF) is a novel nucleotide reverse transcriptase inhibitor (NRTI), which was recommended as first-line therapeutic schedule for free AIDS antiviral drugs. Whether the neurotoxicity of TDF is associated with HAND is not well known. In this study, the cell viability of TDF-treated pheochromocytoma cells (PC-12) line was detected using MTT assay, while apoptosis was evaluated by Hoechst 33342 staining, TUNEL assay, as well as flow cytometry. In addition, the level of reactive oxygen species and BAX protein expression were evaluated using DCFH-DA staining and western blotting. The results showed that the proliferation of PC-12 cells was significantly inhibited by TDF. The morphological assay, TUNEL assay and flow cytometry showed that TDF efficiently triggered apoptosis in PC-12 cells. The reactive oxygen species levels were BAX expression was markedly up-regulated in PC-12 cells after treatment with TDF. These findings indicated that TDF may induce PC-12 cell apoptosis. TDF has neural toxicity effect that is relevant to the cell apoptosis, which may be related to the increasing prevalence of HAND.

Experimental study on the neurotoxic effect of β-amyloid on the cytoskeleton of PC12 cells.

The aim of the present study was to establish a cell model of Alzheimer's disease (AD) and investigate the neurotoxic effects of β-amyloid (Aβ) on the cytoskeleton. PC12 cells were cultured and treated with Aβ25-35, and cell survival was analyzed with the MTT assay. Cell apoptosis was visualized using 4′,6-diamidino-2-phenylindole staining and the terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. Immunocytochemistry and phalloidin staining were used to label the cytoskeleton of PC12 cells. Aβ25-35 was found to induce PC12 cell apoptosis in a dose-dependent manner (P<0.05). Moreover, Aβ25-35 also caused dose-dependent disintegration of the cytoskeleton (P<0.05). Therefore, the PC12 cell cytoskeleton was found to be sensitive to Aβ25-35 neurotoxicity. The disintegration of the cytoskeleton is likely an important pathological alteration in AD, and Aβ is a key molecule involved in AD pathogenesis.

Potential molecular mechanisms mediating the protective effects of tetrahydroxystilbene glucoside on MPP+-induced PC12 cell apoptosis.

Our previous work demonstrated that tetrahydroxystilbene glucoside (TSG) was able to effectively attenuate 1-methyl-4-phenylpyridinium (MPP+)-induced apoptosis in PC12 cells partially via inhibiting reactive oxygen species (ROS) generation. However, the precise molecular mechanisms of TSG responsible for suppressing neuronal apoptosis have not been fully elucidated. To investigate the possible mechanism, we studied the neuroprotective effects of TSG on MPP+-induced PC12 cells apoptosis and explored the molecular mechanisms that mediated the effects of TSG. Our results showed that treatment with TSG prior to MPP+ exposure effectively attenuated the cell viability decrease in PC12 cells, reversed the cell apoptosis, and further restored the mitochondria membrane potential (MMP). In addition, TSG remarkably enhanced the anti-oxidant enzyme activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px), and efficiently reduced the malondialdehyde (MDA) content in the PC12 cells. Meanwhile, TSG markedly upregulated the Bcl-2/Bax ratio, reversed release of Cytochrome c, and inhibited the activation of caspase-3 induced by MPP+. Furthermore, TSG significantly inhibited the activation of p38 mitogen-activated protein kinase (p38MAPK) signaling pathway, while extracellular signal-regulated protein kinases (ERK) phosphorylation was not affected. Together, these findings provide the basis for TSG clinical application as a new therapeutic strategy in the treatment of neurodegenerative diseases.

Betulinic Acid Induces Apoptosis in Differentiated PC12 Cells Via ROS-Mediated Mitochondrial Pathway.

Betulinic acid (BA), a pentacyclic triterpene of natural origin, has been demonstrated to have varied biologic activities including anti-viral, anti-inflammatory, and anti-malarial effects; it has also been found to induce apoptosis in many types of cancer. However, little is known about the effect of BA on normal cells. In this study, the effects of BA on normal neuronal cell apoptosis and the mechanisms involved were studied using differentiated PC12 cells as a model. Treatment with 50μM BA for 24h apparently induced PC12 cell apoptosis. In the early stage of apoptosis, the level of intracellular reactive oxygen species (ROS) increased. Afterwards, the loss of the mitochondrial membrane potential, the release of cytochrome c and the activation of caspase-3 occurred. Treatment with antioxidants could significantly reduce BA-induced PC12 cell apoptosis. In conclusion, we report for the first time that BA induced the mitochondrial apoptotic pathway in differentiated PC12 cells through ROS.

Lemur Tyrosine Kinase-3 Suppresses Growth of Prostate Cancer Via the AKT and MAPK Signaling Pathways

Background/Aims: Lemur tyrosine kinase (LMTK)-3 is a member of the receptor tyrosine kinase (RTK) family. Abnormal expression of LMTK-3 exists in various types of cancers, especially in endocrine-resistant breast cancers; however, the precise level of expression and the biological function in prostate cancer are poorly understood. Methods: In the present study, we determined the expression of LMTK-3 in prostate cancer using immunohistochemistry and Western blotting. We infected PC3 and LNCaP cells with lentivirus-LMTK-3 and observed the biologic characteristics of the PC3 and LNCaP cells in vitro with TUNEL, and migration and invasion assays, respectively. We also established a transplant tumor model of human prostate cancer with infected cells in 15 BALB/c-nu/nu nude mice. Results: LMTK-3 was expressed in prostate epithelial cells. There was a significant decline in the level of LMTK-3 expression in prostate cancers compared to normal tissues. LMTK-3 inhibited PC3 and LNCaP cell growth, migration, and invasion, and induced cell apoptosis in vitro. We also observed that LMTK-3 induced PC3 cell apoptosis in vivo. Further study showed that LMTK-3 inhibited phosphorylation of AKT and ERK, and promoted phosphorylation and activation of p38 kinase and Jun kinase (JNK). Conclusion: Recombinant lentivirus with enhanced expression of LMTK-3 inhibited prostate cancer cell growth and induced apoptosis in vitro and in vivo. AKT and MAPK signaling pathways may contribute to the process.

Epidermal growth factor prevents thallium(I)- and thallium(III)-mediated rat pheochromocytoma (PC12) cell apoptosis

We have reported recently that the proliferation of PC12 cells exposed to micromolar concentrations of Tl(I) or Tl(III) has different outcomes, depending on the absence (EGF- cells) or the presence (EGF+ cells) of epidermal growth factor (EGF) added to the media. In the current work, we investigated whether EGF supplementation could also modulate the extent of Tl(I)- or Tl(III)-induced cell apoptosis. Tl(I) and Tl(III) (25-100μM) decreased cell viability in EGF- but not in EGF+ cells. In EGF- cells, Tl(I) decreased mitochondrial potential, enhanced H2O2 generation, and activated mitochondrial-dependent apoptosis. In addition, Tl(III) increased nitric oxide production and caused a misbalance between the anti- and pro-apoptotic members of Bcl-2 family. Tl(I) increased ERK1/2, JNK, p38, and p53 phosphorylation in EGF- cells. In these cells, Tl(III) did not affect ERK1/2 and JNK phosphorylation but increased p53 phosphorylation that was related to the promotion of cell senescence. In addition, this cation significantly activated p38 in both EGF- and EGF+ cells. The specific inhibition of ERK1/2, JNK, p38, or p53 abolished Tl(I)-mediated EGF- cell apoptosis. Only when p38 activity was inhibited, Tl(III)-mediated apoptosis was prevented in EGF- and EGF+ cells. Together, current results indicate that EGF partially prevents the noxious effects of Tl by preventing the sustained activation of MAPKs signaling cascade that lead cells to apoptosis and point to p38 as a key mediator of Tl(III)-induced PC12 cell apoptosis.