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OBJECTIVE: To investigate the neuroprotective effect of Huangpu Tongqiao Capsule (HPTQ) in a rat model of Wilson disease (WD) and explore the underlying mechanisms. METHODS: SD rat models of WD were established by feeding of coppersupplemented chow diet and drinking water for 12 weeks, and starting from the 9th week, the rats were treated with low-, moderate- and high-dose HPTQ, penicillamine, or normal saline by gavage on a daily basis for 3 weeks. Copper levels in the liver and 24-h urine of the rats were detected, and their learning and memory abilities were evaluated using Morris water maze test. HE staining was used to observe morphological changes of CA1 region neurons in the hippocampus, and neuronal apoptosis was detected with TUNEL staining. Hippocampal expressions of endoplasmic reticulum stress (ERS)-mediated apoptosis pathway-related proteins GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 at both the mRNA and protein levels were detected using RT-qPCR, immunofluorescence assay or Western blotting. RESULTS: Compared with normal control rats, the rat models with copper overload-induced WD exhibited significantly increased copper levels in both the liver and 24-h urine, impaired learning and memory abilities, obvious hippocampal neuronal damage in the CA1 region and increased TUNEL-positive neurons (P<0.01), with also lowered mRNA and protein expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the hippocampus (all P<0.01). Treatments with HPTQ and penicillamine significantly lowered copper level in the liver but increased urinary copper level, improved learning and memory ability, alleviated neuronal damage and apoptosis in the hippocampus, and decreased hippocampal expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the rat models (P<0.01 or 0.05). CONCLUSION: HPTQ Capsule has neuroprotective effects in rat models of WD possibly by inhibiting ERS-mediated apoptosis pathway.
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Disfunción Cognitiva , Degeneración Hepatolenticular , Ratas , Animales , Ratas Sprague-Dawley , Degeneración Hepatolenticular/tratamiento farmacológico , Caspasa 3/metabolismo , Caspasa 9/metabolismo , Caspasa 12/metabolismo , Cobre/metabolismo , Cobre/farmacología , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico , Apoptosis , Hipocampo/metabolismo , Proteínas Reguladoras de la Apoptosis/metabolismo , Penicilamina/farmacología , Disfunción Cognitiva/tratamiento farmacológico , ARN MensajeroRESUMEN
OBJECTIVE: To investigate the pharmacological mechanism of Qili Qiangxin Capsule (QLQX) improvement of heart failure (HF) based on miR133a-endoplasmic reticulum stress (ERS) pathway. METHODS: A left coronary artery ligation-induced HF after myocardial infarction model was used in this study. Rats were randomly assigned to the sham group, the model group, the QLQX group [0.32 g/(kg·d)], and the captopril group [2.25 mg/(kg·d)], 15 rats per group, followed by 4 weeks of medication. Cardiac function such as left ventricular ejection fraction (EF), fractional shortening (FS), left ventricular systolic pressure (LVSP), left ventricular end diastolic pressure (LVEDP), the maximal rate of increase of left ventricular pressure (+dp/dt max), and the maximal rate of decrease of left ventricular pressure (-dp/dt max) were monitored by echocardiography and hemodynamics. Hematoxylin and eosin (HE) and Masson stainings were used to visualize pathological changes in myocardial tissue. The mRNA expression of miR133a, glucose-regulated protein78 (GRP78), inositol-requiring enzyme 1 (IRE1), activating transcription factor 6 (ATF6), X-box binding protein1 (XBP1), C/EBP homologous protein (CHOP) and Caspase 12 were detected by RT-PCR. The protein expression of GRP78, p-IRE1/IRE1 ratio, cleaved-ATF6, XBP1-s (the spliced form of XBP1), CHOP and Caspase 12 were detected by Western blot. TdT-mediated dUTP nick-end labeling (TUNEL) staining was used to detect the rate of apoptosis. RESULTS: QLQX significantly improved cardiac function as evidenced by increased EF, FS, LVSP, +dp/dt max, -dp/dt max, and decreased LVEDP (P<0.05, P<0.01). HE staining showed that QLQX ameliorated cardiac pathologic damage to some extent. Masson staining indicated that QLQX significantly reduced collagen volume fraction in myocardial tissue (P<0.01). Results from RT-PCR and Western blot showed that QLQX significantly increased the expression of miR133a and inhibited the mRNA expressions of GRP78, IRE1, ATF6 and XBP1, as well as decreased the protein expressions of GRP78, cleaved-ATF6 and XBP1-s and decreased p-IRE1/IRE1 ratio (P<0.05, P<0.01). Further studies showed that QLQX significantly reduced the expression of CHOP and Caspase12, resulting in a significant reduction in apoptosis rate (P<0.05, P<0.01). CONCLUSION: The pharmacological mechanism of QLQX in improving HF is partly attributed to its regulatory effect on the miR133a-IRE1/XBP1 pathway.
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Medicamentos Herbarios Chinos , Estrés del Retículo Endoplásmico , Insuficiencia Cardíaca , MicroARNs , Animales , MicroARNs/genética , MicroARNs/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/genética , Masculino , Ratas Sprague-Dawley , Cápsulas , Factor de Transcripción Activador 6/metabolismo , Factor de Transcripción Activador 6/genética , Chaperón BiP del Retículo Endoplásmico , Apoptosis/efectos de los fármacos , Caspasa 12/metabolismo , Caspasa 12/genética , Miocardio/patología , Miocardio/metabolismo , Proteínas de Choque Térmico/metabolismo , Proteínas de Choque Térmico/genética , Ratas , Proteína 1 de Unión a la X-Box/metabolismo , Proteína 1 de Unión a la X-Box/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/patología , Infarto del Miocardio/genética , Infarto del Miocardio/fisiopatologíaRESUMEN
Aflatoxin B1 (AFB1) is extremely carcinogenic and can cause liver cancer in humans and animals with continued ingestion. As a natural compound, curcumin (Cur) exhibits excellent anti-inflammatory, and anti-cancer properties with few side effects. In this study, a total of 60 male mice (6-week-olds, 15 per group). After one week of acclimatization feeding, the mice were divided into control group (Con), AFB1 group, curcumin group (Cur), and AF+Cur group. The mice were gavaged with curcumin (Cur, 100 mg/kg) and/or AFB1 (0.75 mg/kg). To identify a new therapeutic target for AFB1-induced pyroptosis, we performed proteomic profiling for curcumin alleviating liver injury caused by AFB1 to further validate the targets through volcano plot analysis, Venn analysis, heatmap analysis, correlation, cluster analysis, GO and KEGG enrichment. AFB1 exposure resulted in the loss of hepatocyte membrane, swelling of the endoplasmic reticulum, and a significant increase in transaminase (ALT and AST) contents, while curcumin greatly improved these changes. We found that differentially expressed proteins are enriched in the endoplasmic reticulum membrane and identified ITPR2 as a target of curcumin that alleviates AFB1-induced liver injury by proteomics. Furthermore, ITPR2 expression was detected by immunofluorescence, and qRT-PCR for mRNA expression of genes downstream of ITPR2 (calpain1, calpain2, caspase-12, caspase-3). ITPR2-activated endoplasmic reticulum stress-related proteins (calpain1, calpaini2, bcl-2, BAX, cl-caspase-12, cl-caspase-3), apoptosis (PARP) and pyroptosis (DFNA5) related proteins were examined by western blotting. The analysis showed that it effectively prevents AFB1-induced pyroptosis by lowering endoplasmic reticulum stress via interfering with ITPR2 and its downstream proteins (calpain1, calpain2, bcl-2, Bax) and inhibiting caspase-12/caspase-3 pathway. Conclusively, this study applied proteomic profiling to elucidate ITPR2 as a new target, which might give a new perspective on the mechanism of curcumin alleviating AFB1-induced pyroptosis.
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Curcumina , Piroptosis , Masculino , Ratones , Humanos , Animales , Caspasa 3/metabolismo , Aflatoxina B1 , Curcumina/farmacología , Proteína X Asociada a bcl-2/metabolismo , Proteómica , Caspasa 12/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptores de Inositol 1,4,5-TrifosfatoRESUMEN
OBJECTIVE: To investigate the protective effects and its possible mechanism of Wuzi Yanzong Pill (WYP) on Parkinson's disease (PD) model mice. METHODS: Thirty-six C57BL/6 male mice were randomly assigned to 3 groups including normal, PD, and PD+WYP groups, 12 mice in each group. One week of intraperitoneal injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was used to establish the classical PD model in mice. Meanwhile, mice in the PD+WYP group were administrated with 16 g/kg WYP, twice daily by gavage. After 14 days of administration, gait test, open field test and pole test were measured to evaluate the movement function. Tyrosine hydroxylase (TH) neurons in substantia nigra of midbrain and binding immunoglobulin heavy chain protein (GRP78) in striatum and cortex were observed by immunohistochemistry. The levels of TH, GRP78, p-PERK, p-eIF2α, ATF4, p-IRE1α, XBP1, ATF6, CHOP, ASK1, p-JNK, Caspase-12, -9 and -3 in brain were detected by Western blot. RESULTS: Compared with the PD group, WYP treatment ameliorated gait balance ability in PD mice (P<0.05). Similarly, WYP increased the total distance and average speed (P<0.05 or P<0.01), reduced rest time and pole time (P<0.05). Moreover, WYP significantly increased TH positive cells (P<0.01). Immunofluorescence showed WYP attenuated the levels of GRP78 in striatum and cortex. Meanwhile, WYP treatment significantly decreased the protein expressions of GRP78, p-PERK, p-eIF2α, ATF4, p-IRE1 α, XBP1, CHOP, Caspase-12 and Caspase-9 (P<0.05 or P<0.01). CONCLUSIONS: WYP ameliorated motor symptoms and pathological lesion of PD mice, which may be related to the regulation of unfolded protein response-mediated signaling pathway and inhibiting the endoplasmic reticulum stress-mediated neuronal apoptosis pathway.
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Fármacos Neuroprotectores , Enfermedad de Parkinson , Ratones , Masculino , Animales , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/metabolismo , Endorribonucleasas/metabolismo , Chaperón BiP del Retículo Endoplásmico , Caspasa 12/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Ratones Endogámicos C57BL , Estrés del Retículo Endoplásmico , Respuesta de Proteína Desplegada , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Modelos Animales de EnfermedadRESUMEN
This study aims to investigate the effect of atractylenolide â ¢(ATL-â ¢) on hydrogen peroxide(H_2O_2)-induced endoplasmic reticulum stress and apoptosis of H9 c2 cells via the ROS/GRP78/caspase-12 signaling pathway.The binding activity of ATL-â ¢ to GRP78 was determined by molecular docking.The result showed that ATL-â ¢ had a good binding activity to GRP78, and the binding activity of ATL-â ¢ was stronger than that of its specific inhibitor.The endoplasmic reticulum stress model of H9 c2 was established by H_2O_2(100 µmol·L~(-1)) treatment.Five groups were designed: blank control group, model group, and ATL-â ¢(15, 30, and 60 µmol·L~(-1)) groups.Apoptosis was detected by Hoechst/PI double staining and flow cytometry.The levels of superoxide dismutase(SOD), malondialdehyde(MDA), and lactate dehydrogenase(LDH) were measured by colorimetry.The levels of reactive oxygen species(ROS) and calcium(Ca~(2+)) in cytoplasm were determined by the fluorescence probe DCFH-DA and the calcium fluorescence probe Flou-4, respectively.The protein levels of GRP78, caspase-12, and caspase-3 were determined by Western blot, and the mRNA levels of GRP78 and caspase-12 by RT-qPCR.N-acetyl-L-cysteine(NAC) and 4-phenylbutyric acid(4-PBA) were respectively used to inhibit ROS and GRP78, and then the mechanism of ATL-â ¢ in protecting the cells from endoplasmic reticulum stress induced by H_2O_2 were deduced.ATL-â ¢(15, 30, and 60 µmol·L~(-1)) decreased the apoptosis rate and ROS, MDA, and LDH levels(P<0.01), increased the SOD activity(P<0.01), and down-regulated the protein levels of GRP78, caspase-12, and caspase-3 and the mRNA levels of GRP78 and caspase-12(P<0.05).The addition of NAC decreased the apoptosis rate and ROS, MDA, GRP78, caspase-12, and caspase-3 levels(P<0.01), while it elevated the SOD level(P<0.01).The addition of 4-PBA also decreased the apoptosis rate and the levels of GRP78, caspase-12, caspase-3, and Ca~(2+)(P<0.01).The effect of inhibitors were consistent with that of ATL-â ¢.In conclusion, ATL-â ¢ can protect H9 c2 cardiomyocytes by regulating ROS/GRP78/caspase-12 signaling pathway to inhibit H_2O_2-induced endoplasmic reticulum stress and apoptosis.
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Calcio , Chaperón BiP del Retículo Endoplásmico , Apoptosis , Calcio/farmacología , Caspasa 12/genética , Caspasa 12/metabolismo , Caspasa 3/genética , Caspasa 3/metabolismo , Estrés del Retículo Endoplásmico , Lactonas , Simulación del Acoplamiento Molecular , ARN Mensajero , Especies Reactivas de Oxígeno/metabolismo , Sesquiterpenos , Transducción de Señal , Superóxido Dismutasa/metabolismoRESUMEN
BACKGROUND: Reducing cerebral ischemia-reperfusion injury is crucial for improving survival and neurologic outcomes after cardiac arrest/cardiopulmonary resuscitation (CA/CPR). The purpose of this study is to investigate the neuroprotective effects of green tea polyphenols (GTPs) concern with the modulation of endogenous antioxidation and endoplasmic reticulum stress. METHODS: After subjecting to CA/CPR, rats were randomized into the saline group (NS, n = 40) and the GTPs group (GTPs, n = 40) and the GTPs group (GTPs, n = 40) and the GTPs group (GTPs. RESULTS: Comparing with that in NS group, GTPs increased the expression of SOD1 and SOD2 at 12 h, 24 h, 48 h, 72 h, and the expression of GRP78 at 24 h and 48 h (p < 0.05) butdecreased caspase-12, CHOP, caspase-3 level, and apoptotic number of neurons (p < 0.05) butdecreased caspase-12, CHOP, caspase-3 level, and apoptotic number of neurons (. CONCLUSION: GTPs exert neuroprotective effects via mechanisms that may be related to the enhancement of endogenous antioxidant capacity and inhibition of endoplasmic reticulum stress in CA/CPR rat models.
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Reanimación Cardiopulmonar/métodos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Paro Cardíaco/metabolismo , Fármacos Neuroprotectores/farmacología , Polifenoles/farmacología , Superóxido Dismutasa/metabolismo , Té/química , Animales , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Caspasa 12/metabolismo , Caspasa 3/metabolismo , Corteza Cerebral/metabolismo , Modelos Animales de Enfermedad , Proteínas de Choque Térmico/metabolismo , Masculino , Neuronas/metabolismo , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/metabolismo , Superóxido Dismutasa-1/metabolismo , Factor de Transcripción CHOP/metabolismoRESUMEN
Endoplasmic reticulum (ER) stress has been identified as a primary factor involved in brain ischemia-reperfusion injury progression. p21-activated kinase 2 (Pak2) is a novel ER function regulator. The aim of our study is to explore the influence of Pak2 on ER stress and determine whether melatonin attenuates ER stress-mediated cell death by modulating Pak2 expression in vitro using N2a cells. The results of our study demonstrated that hypoxia-reoxygenation (HR) injury repressed the levels of Pak2, an effect that was accompanied by activation of ER stress. In addition, decreased Pak2 was associated with oxidative stress, calcium overload, and caspase-12-mediated apoptosis activation in HR-treated N2a cells. Interestingly, melatonin treatment reversed the decreased Pak2 expression under HR stress. Knockdown of Pak2 abolished the protective effects of melatonin on ER stress, oxidative stress, and caspase-12-related N2a cells death. Additionally, we found that Pak2 was regulated by melatonin via the AMPK pathway; inhibition of AMPK prevented melatonin-mediated Pak2 upregulation, a result that was accompanied by an increase in N2a cell death. Altogether, these results identify the AMPK-Pak2 axis as a new signaling pathway responsible for ER stress and N2a cell viability under HR injury. Modulation of the AMPK-Pak2 cascade via supplementation of melatonin might be considered an effective approach to attenuate reperfusion-mediated N2a cell damage via repression of ER stress.
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Proteínas Quinasas Activadas por AMP/metabolismo , Hipoxia de la Célula/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Melatonina/farmacología , Quinasas p21 Activadas/metabolismo , Animales , Caspasa 12/metabolismo , Línea Celular Tumoral , Estrés Oxidativo/efectos de los fármacosRESUMEN
BACKGROUND: The clinical efficacy of electroacupuncture in treating diabetic peripheral neuropathy (DPN) is significant, but the underlying mechanism of action is not clear. Considering that glucose-regulated protein 78 (GRP78) and caspase-12 are major proteins participating in cell apoptosis, we investigated the effects of "adjusting internal organs and dredging channel" electroacupuncture therapy on GRP78 and caspase-12 levels in streptozotocin (STZ)-diabetic rats to elucidate the mechanism of action. METHODS: Rats were first divided into two groups: one group was rendered diabetic with a single injection of 50 mg/kg STZ, whereas the other normal control group was injected with an equivalent volume of citrate buffer. The STZ-diabetic rats were randomly divided into three groups: model control and electroacupuncture- and mecobalamin-treated groups. After 12 weeks treatment, the therapeutic efficacy of electroacupuncture was assessed using sciatic nerves isolated from rats. In the electroacupuncture group, rats were treated by electroacupuncture for 20 minutes once daily for 6 days each week, with 1 day off, for 12 consecutive weeks. The selected acupressure points include bilateral acupressure points of BL13 (Fehu), BL20 (Pishu), BL23 (Shenshu), LI4 (Hegu), LR3 (faichong), ST36 (Zusanli), and SP6 (Sanyiniiao). Acupressure points were stimulated using a HuaTuo SDZ-V Electric Acupuncture Therapy Apparatus. The acupressure points of BL13 and BL23, as well as SP6 and LR3, were connected on the same side with a dilatational wave of 3 Hz (frequency ratio of 1 : 5) to stimulate the parts of the body to the extent that could be tolerated by the rat. As for the mecobalamin-treated groups, mecobalamin was administrated to rats intragastrically at a dose of 20 mg/kg once daily for 12 consecutive weeks. Immunofluorescence and western blot analysis were used to determine GRP78 and caspase-12 levels in sciatic nerves. In addition, cell apoptosis in sciatic nerves was determined using the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labeling (TUNEL) assay. RESULTS: Electroacupuncture markedly reduced the pathological injury to sciatic nerves in STZ-diabetic rats. Moreover, electroacupuncture significantly downregulated GRP78 and caspase-12 and reduced cell apoptosis of sciatic nerves in DPN rats. CONCLUSIONS: Electroacupuncture improved DPN by downregulating GRP78 and caspase-12 and reducing cell apoptosis of sciatic nerves in STZ-diabetic rats, and further inhibited the occurrence of endoplasmic reticulum stress, thus preventing sciatic nerve injuries.
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Caspasa 12/metabolismo , Diabetes Mellitus Experimental/terapia , Neuropatías Diabéticas/prevención & control , Electroacupuntura , Proteínas de Choque Térmico/metabolismo , Nervio Ciático/enzimología , Estreptozocina , Puntos de Acupuntura , Animales , Apoptosis , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/enzimología , Diabetes Mellitus Experimental/patología , Neuropatías Diabéticas/inducido químicamente , Neuropatías Diabéticas/enzimología , Neuropatías Diabéticas/patología , Regulación hacia Abajo , Masculino , Ratas Sprague-Dawley , Nervio Ciático/ultraestructura , Transducción de SeñalRESUMEN
BACKGROUND: Skeletal muscle atrophy is a striking example of the multiple changes in the physiological state of humans and animals induced by microgravity. Previous studies have shown that a blood circulation disorder may be a cause of this atrophy, and traditional Chinese medicine has been regarded as a potential countermeasure to reverse the atrophy in China. This study was carried out to test the effects of Xuefuzhuyu capsules (XFZY) on the skeletal muscle atrophy induced by simulated microgravity. METHODS: The mass and cross-sectional area of the soleus muscle were compared in rats treated with XFZY that were hindlimb unloaded for 30 d (XFZY-TS group), untreated rats that were hindlimb unloaded for 30 d (TS group), and control rats (CON group). The expression and phosphorylation levels of key proteins of the sarcoplasmic reticulum stress system were also measured. RESULTS: Treatment with XFZY attenuated the loss of muscle mass and cross-sectional area induced by hindlimb unloading. Western blot analysis showed that the phosphatidyl-inositol-3-kinase/phospho-Akt (PI3K/p-Akt) pathways were down-regulated after 30 d in the TS group compared with the CON group. This effect was partly reversed by XFZY. Hindlimb unloading increased the expression of glucose-regulated protein 78 (GRP78), cytosine-cytosine-adenosine-adenosine-thymidine/enhancer-binding protein homologous protein (CHOP), C-Jun N-terminal kinase (JNK), and Caspase 12. Treatment with XFZY alleviated this increased protein expression. DISCUSSION: Our results suggest that XFZY could partially reverse the effects of hindlimb unloading on muscle atrophy and perhaps target the sarcoplasmic reticulum stress system, possibly through the GRP78-CHOP-JNK-Caspase 12 pathway.Zhang S, Yuan M, Cheng C, Xia D, Wu S. Chinese herbal medicine effects on muscle atrophy induced by simulated microgravity. Aerosp Med Hum Perform. 2018; 89(10):883-888.
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Medicamentos Herbarios Chinos/farmacología , Suspensión Trasera , Músculo Esquelético/efectos de los fármacos , Atrofia Muscular/patología , Simulación de Ingravidez , Animales , Caspasa 12/efectos de los fármacos , Caspasa 12/metabolismo , Chaperón BiP del Retículo Endoplásmico , Proteínas de Choque Térmico/efectos de los fármacos , Proteínas de Choque Térmico/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/efectos de los fármacos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Sistema de Señalización de MAP Quinasas , Masculino , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Atrofia Muscular/metabolismo , Fosfatidilinositol 3-Quinasa/efectos de los fármacos , Fosfatidilinositol 3-Quinasa/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal , Factor de Transcripción CHOP/efectos de los fármacos , Factor de Transcripción CHOP/metabolismoRESUMEN
Non-alcoholic fatty liver disease (NAFLD) is increasingly prevalent and represents a growing challenge in terms of prevention and treatment. The purpose of this study is to investigate the protective effects of ginsenoside Rg1 (Rg1), an active ingredient of a natural medicine, and further clarify its protective mechanisms, in a mouse model of NAFLD induced by a high-fat diet. Rg1 significantly reduced liver weight, serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), liver free fatty acids (FFAs) and malondialdehyde (MDA) levels, and increased superoxide dismutase (SOD) activity. Rg1 also upregulated the expression of peroxisome proliferator-activated receptor-alpha (PPARα), which stimulated fatty acid beta oxidation and promoted the metabolism of FFAs and TG. It also suppressed the expression of CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP), cysteine-containing aspartate-specific proteases 12 (Caspase 12), and glucose-regulated protein78 (GRP78), which reduced endoplasmic reticulum (ER) stress. Furthermore, Rg1 alleviated liver inflammation by inhibiting the activation of nucleotide binding oligomerization domain (NOD)-like receptor family pyrin domain-containing 3 (NLRP3) and thus reduced the production of inflammatory cytokines, such as interleukin 1-beta (IL-1ß) and interleukin 18 (IL-18). These results suggested that Rg1 may protect against NAFLD, through regulation of lipid peroxidation, ER stress and inflammasome activation.
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Estrés del Retículo Endoplásmico/efectos de los fármacos , Ginsenósidos/farmacología , Inflamasomas/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Hígado/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Panax/química , Animales , Antioxidantes/metabolismo , Antioxidantes/farmacología , Antioxidantes/uso terapéutico , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Caspasa 12/metabolismo , Citocinas/metabolismo , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Chaperón BiP del Retículo Endoplásmico , Femenino , Ginsenósidos/uso terapéutico , Proteínas de Choque Térmico/metabolismo , Inflamación/etiología , Inflamación/metabolismo , Inflamación/prevención & control , Metabolismo de los Lípidos/efectos de los fármacos , Hígado/enzimología , Hígado/metabolismo , Ratones Endogámicos C57BL , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/prevención & control , PPAR alfa/metabolismo , FitoterapiaRESUMEN
Background: Palmitic acid (PA) is the main saturated fatty acid naturally occurring in animal fats and vegetable oils. In recent decades, palm oil, an alternative lipid source containing high amounts of PA, has been widely used to replace fish oil in aquafeed. Objective: We investigated the hepatotoxicity of PA in zebrafish and the underlying mechanism. Methods: One-month-old zebrafish fed a high-fat diet (HFD) containing 16% soybean oil and 3 PA-incorporated HFDs [4%, 8%, and 12% PA (12PA)] for 2 wk (experiment 1) and 4 wk (experiment 2) were used to evaluate PA-induced liver damage and endoplasmic reticulum (ER) stress. Germ-free (GF) zebrafish fed low-fat, high-fat, or 12PA diets for 5 d were used to study the direct effects of PA on liver damage (experiment 3). GF zebrafish colonized with HFD or 12PA microbiota for 48 h were used to elucidate the indirect effects of PA-altered microbiota on liver damage (experiment 4). Last, GF zebrafish colonized with HFD or 12PA microbiota were used to evaluate the effects of different microbiotas on PA absorption (experiment 5). Results: In experiment 1, the proportion of PA in the liver linearly increased as its percentage in dietary lipid increased (r2 = 0.83, P < 0.05). In experiment 2, the expression of glucose-regulated protein 78 (Grp78) and C/EBP-homologous protein (Chop) was higher in the 12PA group than in the HFD group (2.2- and 2.7-fold, respectively; P < 0.05). The activity of caspase-12 was increased by 61.1% in the 12PA group compared with the HFD group (P < 0.05). In experiment 3, caspase-12 activity was higher in the 12PA group than in the HFD group (P < 0.05). In experiment 4, GF zebrafish colonized with PA-altered microbiota had higher caspase-12 activity (P < 0.05) than those colonized by HFD microbiota. In experiment 5, PA-altered microbiota promoted PA absorption (P < 0.05) and aggravated ER stress and liver damage in the context of high-PA feeding. Conclusions: The PA-altered microbiota indirectly induced ER stress and liver damage in zebrafish. Moreover, the PA microbiota promoted the absorption of PA, leading to enhanced PA overflow into the liver and aggravated hepatotoxicity of PA in zebrafish.
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Enfermedad Hepática Inducida por Sustancias y Drogas/microbiología , Grasas de la Dieta/toxicidad , Estrés del Retículo Endoplásmico , Microbioma Gastrointestinal , Hígado/efectos de los fármacos , Ácido Palmítico/toxicidad , Alimentación Animal , Animales , Caspasa 12/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Hígado/metabolismo , Aceite de Palma/química , Aceite de Palma/toxicidad , Pez CebraRESUMEN
Diabetic nephropathy (DN) is by far the most common cause of endstage renal disease (ESRD) in industrial countries, accounting for ~45% of all new ESRD cases in the United States. Grape seed proanthocyanidin extracts (GSPE) are powerful antioxidants, with an antioxidant ability 50fold greater than that of vitamin E and 20fold greater than that of vitamin C. The present study investigated whether GSPE can protect against streptozotocin (STZ)induced DN and aimed to elucidate a possible mechanism. Male Sprague Dawley rats were randomly divided into three groups: Control group (N), diabetes mellitus group (DM) injected with 40 mg/kg STZ, and the GSPE treatment group (intragastric administration of 250 mg/kg/day GSPE for 16 weeks after diabetes was induced in the rats). Blood and kidney samples were collected after treatment. The renal pathological changes were determined with periodic acidSchiff (PAS) staining, while the protein expression levels of glucoseregulated protein 78 (GRP78), phosphorylatedextracellular signalregulated kinase (pERK) and Caspase12 were determined by western blotting and immunohistochemical staining. Apoptosis was determined with a terminal deoxynucleotidyl transferase dUTP nickend labeling (TUNEL) assay. Compared with the DM group, the GSPE group had no significant changes in the blood urea nitrogen (BUN) level and serum creatinine (Scr) level, but showed a significant decline in the renal index (RI) level and 24h urinary albumin level (P<0.05). The histopathology results indicated very little pathological damage in the GSPE group. Compared with the DM group, the GSPE group had a significantly reduced number of TUNELpositive cells (P<0.05), and the GSPE group had an obvious reduction in the protein expression of GRP78, pERK, and Caspase12 (P<0.05). In this study, the results indicated that GSPE can protect renal function and attenuate endoplasmic reticulum stressinduced apoptosis via the Caspase12 pathway in STZinduced DN.
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Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Diabetes Mellitus Experimental/tratamiento farmacológico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Extracto de Semillas de Uva/farmacología , Proantocianidinas/farmacología , Albúminas/genética , Albúminas/metabolismo , Animales , Apoptosis/genética , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Nitrógeno de la Urea Sanguínea , Caspasa 12/genética , Caspasa 12/metabolismo , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/metabolismo , Nefropatías Diabéticas/patología , Esquema de Medicación , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/genética , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Absorción Gástrica , Regulación de la Expresión Génica , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Riñón/efectos de los fármacos , Riñón/metabolismo , Riñón/patología , Masculino , Proteína Quinasa 1 Activada por Mitógenos/genética , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Fosforilación/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , EstreptozocinaRESUMEN
To explore the protective effect of naringin(Nar) on the injury of myocardium tissues induced by streptozotocin(STZ) in diabetic rats and the relationship with oxidative stress and endoplasmic reticulum stress(ERS)ï¼ the male SD rats were intraperitoneally injected with streptozotocin(STZï¼ 60 mg·kg⻹) to establish the diabetic rat model and then randomly divided into the type 1 diabetic rat group(T1DR)ï¼ the low-dose Nar group(Nar25)ï¼ the middle-dose Nar group(Nar50) and the high-dose Nar group(Nar100). The normal rats were designed as control group(Con). Nar25ï¼ Nar50ï¼ Nar100 groups were orally administered with Nar at the doses of 25.0ï¼ 50.0ï¼ 100.0 mg·kg⻹ per dayï¼ respectivelyï¼ while the normal group and the T1DR group were orally administered with saline. At the 8th week after treatmentï¼ fasting plasma glucose and heart mass index were measured. The pathological changes in myocardial tissues were observed by microscope. The cardiac malondialdehyde(MDA) level and superoxide dismutase(SOD) activities were measured. The gene and protein expressions of glucose-regulated protein 78(GRP78)ï¼ C/EBP homologous protein(CHOP)ï¼ cysteinyl aspartate-specific proteinase 12(caspase 12) were detected by qRT-PCR and Western blot. According to the resultsï¼ compared with control groupï¼ the myocardial structure was damagedï¼ the content of MDA was increasedï¼ while the activities of SOD were decreased(P<0.05) in T1DR group. GRP78ï¼ CHOP and caspase 12 mRNA and protein expressions were increased significantly in T1DR group(P<0.05ï¼ P<0.01). Compared with T1DR groupï¼ myocardial structure damage was alleviated in Nar treatment group. The content of MDA was decreasedï¼ while the activities of SOD were increased significantly. The mRNA and protein expressions of GRP78ï¼ CHOP and caspase 12 were increasedï¼ especially in middle and high-dose groups(P<0.05ï¼ P<0.01). After treatment with Nar for 8 weeksï¼ myocardial structure damage was obviously alleviated in Nar treatment groups. The content of MDA was decreasedï¼ while the activities of SOD were increased significantly in myocardial tissues. The mRNA and protein expressions of GRP78ï¼ CHOP and caspase 12 were increasedï¼ especially in middle and high-dose groups(P<0.05ï¼ P<0.01). The findings suggest that Nar may protect myocardium in diabetic rats by reducing mitochondrial oxidative stress injuries and inhibiting the ERS-mediated cell apoptosis pathway.
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Cardiotónicos/farmacología , Cardiomiopatías Diabéticas/tratamiento farmacológico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Flavanonas/farmacología , Estrés Oxidativo/efectos de los fármacos , Animales , Apoptosis , Caspasa 12/metabolismo , Diabetes Mellitus Experimental , Proteínas de Choque Térmico/metabolismo , Masculino , Malondialdehído/metabolismo , Ratas , Ratas Sprague-Dawley , Superóxido Dismutasa/metabolismo , Factor de Transcripción CHOP/metabolismoRESUMEN
SCOPE: We investigated the role of endoplasmic reticulum (ER) stress in the protective effects of EGCG against the neuronal apoptosis in Aß1-42 -induced SH-SY5Y cells and APP/PS1 transgenic mice. METHODS AND RESULTS: Cell viability (CCK8 assay), flow cytometry, Hoechst 33258 staining, immunohistochemistry, transmission electron microscopy (TEM), and western blotting were used. EGCG prevented Aß1-42-induced toxicity in SH-SY5Y cells, increased cell viability, and decreased apoptosis in a dose-dependent manner. In a subsequent mechanism study, it was found that this effect contributed to the down-regulation of GRP78, CHOP, cleaved-caspase-12 and -3. Moreover, EGCG also reduced the cytotoxicity induced by tunicamycin (TM) and thapsigargin (TG), two ER stress activators. Consistent with the in vitro study, EGCG inhibited neuronal apoptosis in the cortex of APP/PS1 transgenic mice, with the mitigation of ER abnormal ultrastructural swelling and the downregulation of ER-stress-associated proteins. CONCLUSION: These results indicate that EGCG attenuates the neurotoxicity in Alzheimer's disease (AD) via a novel mechanism that involves inhibition of ER-stress-associated neuronal apoptosis in vitro and in vivo, suggesting the tremendous potential of EGCG for use in a nutritional preventive strategy against AD.
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Péptidos beta-Amiloides/antagonistas & inhibidores , Apoptosis , Catequina/análogos & derivados , Suplementos Dietéticos , Estrés del Retículo Endoplásmico , Neuronas/metabolismo , Fármacos Neuroprotectores/metabolismo , Fragmentos de Péptidos/antagonistas & inhibidores , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/prevención & control , Péptidos beta-Amiloides/metabolismo , Animales , Caspasa 12/química , Caspasa 12/genética , Caspasa 12/metabolismo , Caspasa 3/química , Caspasa 3/genética , Caspasa 3/metabolismo , Catequina/metabolismo , Catequina/uso terapéutico , Línea Celular Tumoral , Supervivencia Celular , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Corteza Cerebral/ultraestructura , Chaperón BiP del Retículo Endoplásmico , Proteínas de Choque Térmico/agonistas , Proteínas de Choque Térmico/antagonistas & inhibidores , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Ratones Transgénicos , Microscopía Electrónica de Transmisión , Proteínas del Tejido Nervioso/agonistas , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuronas/patología , Neuronas/ultraestructura , Fármacos Neuroprotectores/uso terapéutico , Nootrópicos/metabolismo , Nootrópicos/uso terapéutico , Fragmentos de Péptidos/metabolismo , Distribución Aleatoria , Factor de Transcripción CHOP/agonistas , Factor de Transcripción CHOP/antagonistas & inhibidores , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismoRESUMEN
Palmitate (PA) has been identified to induce cell apoptosis in osteoblasts. The cJun NH2teminal kinase (JNK) signaling pathway and endoplasmic reticulum stress (ERS) were found to be important contributors. Therefore, natural or synthetic agents may antagonize PAinduced apoptosis in osteoblasts, and demonstrate the potential application to reverse osteoporosis. The present study demonstrated that the Lycium barbarum polysaccharide (LBP) is as a major active ingredient of Lycium barbarum and that it can reduce the fatty acid toxicity of PA. Furthermore, this study attempted to elucidate the underlying molecular mechanisms of LBP. Firstly, it was demonstrated via a Cell Counting Kit8 assay, that LBP could significantly increase the viability of MC3T3E1 cells in a dosedependent manner. Flow cytometric analysis indicated that LBP inhibits PAinduced apoptosis in osteoblastic cells. Reverse transcriptionquantitative polymerase chain reaction and western blotting results showed that the expression levels of glucoseregulated protein 78, C/EBP homologous protein and cysteinyl asparate specific proteinase3/9/12, were increased in MC3T3E1 cells following PA treatment. The treatment of the cells with PA resulted in an activation of the ERS and the JNK signaling pathway. These pathways were effectively suppressed by coincubation with LBP. Taken together, PA may cause ERS, in cell apoptosis, and it may further activate the JNK signaling pathway. LBP reversed PAinduced apoptosis in MC3T3E1 cells through inhibition of the activation of the ERSmediated JNK signaling pathway.
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Apoptosis/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Palmitatos/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Biomarcadores , Caspasa 12/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , Chaperón BiP del Retículo Endoplásmico , Proteínas de Choque Térmico/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Factor de Transcripción CHOP/metabolismoRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: As a classical prescription of traditional Chinese medicine (TCM), Jia-Jian-Di-Huang-Yin-Zi decoction (JJDHYZ) has been used to treat the symptoms of neurological disorders with a long history. AIM OF THE STUDY: To evaluate the effects and possible mechanisms of JJDHYZ on a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced subacute mouse model of Parkinson's disease. MATERIALS AND METHODS: Adult male C57BL/6 mice were randomly divided into five groups: control, MPTP, JJDHYZ low dosage (JJDHYZ-L, 8.5g/kg/day), medium dosage (JJDHYZ-M, 17g/kg/day) and high dosage (JJDHYZ-H, 34g/kg/day). Behavioral tests, immunohistochemistry, immunofluorescence, and high-performance liquid chromatography (HPLC) were conducted to evaluate the neuroprotective effects of JJDHYZ. The mechanism was further explored using TdT-mediated dUTP nick end labeling staining and transmission electron microscopy. The protein expression of Bax, Bcl-2, cytochrome c, full-length caspase9, cleaved caspase9, cleaved caspase3, caspase12 and C/EBP homologous protein was assessed. The toxicity on hepatocytes and renal cells was detected using the enzyme-linked immunosorbent assay kits. RESULTS: JJDHYZ-H restored the behavior performance impaired by MPTP, and reduced the loss of tyrosine hydroxylase. Additionally, it blocked the apoptosis, activated cleaved caspase3 and protected the ultrastructural integrity of mitochondria by regulating the expression of proteins in both mitochondrial and endoplasmic reticulum (ER) caspase12 pathways. CONCLUSIONS: JJDHYZ-H showed behavior recovery and dopamine neuron protection by inhibiting the apoptotic activities associated with mitochondrial and ER caspase12 pathways.
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Medicamentos Herbarios Chinos/farmacología , Mitocondrias/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Trastornos Parkinsonianos/tratamiento farmacológico , Animales , Apoptosis/efectos de los fármacos , Caspasa 12/metabolismo , Neuronas Dopaminérgicas/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Medicamentos Herbarios Chinos/administración & dosificación , Retículo Endoplásmico/efectos de los fármacos , Ensayo de Inmunoadsorción Enzimática , Masculino , Medicina Tradicional China , Ratones , Ratones Endogámicos C57BL , Fármacos Neuroprotectores/administración & dosificación , Trastornos Parkinsonianos/fisiopatologíaRESUMEN
The currently available data suggest that natural products may exert significant cytotoxic and immunomodulatory effects. Plant-derived chemotherapeutic agents such as taxol, etoposide or vincristine, currently used in cancer therapy, are prominent examples in this regard. However, there is a need for new and nat- ural anticancer compounds with low or without toxicity to normal cells. One of the active compounds responsible for the immune effects is ß-glucan derived from cereals, fungi, seaweeds, yeasts and bacteria. The recent data suggest that ß-glucans are potent immunomodulators with anticancer properties. Antitumor properties of fungi and yeast derived ß-glucans have been widely recognized, but those polysaccharides are mostly insoluble, creating several problems especially in topical formulation. To overcome the issue of low water solubility, in the current study a more soluble ß-glucan type from oats was chosen for the investigation of its antitumor activities. Cytotoxic effects were studied using a human melanoma cell line (Me45). The effect of electroporation on the antitumor activity of oat ß-glucan was investigated as well. Cellular viability assessment, immuno-cytochemistry and immunofluochemistry were employed to evaluate biologic effects. Our results indicate strong anticancer properties of oat ß-glucan, enhanced by electroporation.
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Antineoplásicos Fitogénicos/farmacología , Electroquimioterapia , Melanoma/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , beta-Glucanos/farmacología , Adulto , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/aislamiento & purificación , Apoptosis/efectos de los fármacos , Avena/química , Caspasa 12/metabolismo , Caspasa 3/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citocromos c/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Melanoma/metabolismo , Melanoma/patología , Fitoterapia , Plantas Medicinales , Neoplasias Cutáneas/metabolismo , Neoplasias Cutáneas/patología , Solubilidad , beta-Glucanos/química , beta-Glucanos/aislamiento & purificaciónRESUMEN
Muscadine grape skin extract (MSKE) is derived from muscadine grape (Vitis rotundifolia), a common red grape used to produce red wine. Endoplasmic reticulum (ER) stress activates the unfolded protein response (UPR) that serves as a survival mechanism to relieve ER stress and restore ER homeostasis. However, when persistent, ER stress can alter the cytoprotective functions of the UPR to promote autophagy and cell death. Although MSKE has been documented to induce apoptosis, it has not been linked to ER stress/UPR/autophagy. We hypothesized that MSKE may induce a severe ER stress response-mediated autophagy leading to apoptosis. As a model, we treated C4-2 prostate cancer cells with MSKE and performed a quantitative Tandem Mass Tag Isobaric Labeling proteomic analysis. ER stress response, autophagy and apoptosis were analyzed by western blot, acridine orange and TUNEL/Annexin V staining, respectively. Quantitative proteomics analysis indicated that ER stress response proteins, such as GRP78 were greatly elevated following treatment with MSKE. The up-regulation of pro-apoptotic markers PARP, caspase-12, cleaved caspase-3, -7, BAX and down-regulation of anti-apoptotic marker BCL2 was confirmed by Western blot analysis and apoptosis was visualized by increased TUNEL/Annexin V staining upon MSKE treatment. Moreover, increased acridine orange, and LC3B staining was detected in MSKE-treated cells, suggesting an ER stress/autophagy response. Finally, MSKE-mediated autophagy and apoptosis was antagonized by co-treatment with chloroquine, an autophagy inhibitor. Our results indicate that MSKE can elicit an UPR that can eventually lead to apoptosis in prostate cancer cells.
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Autofagia/efectos de los fármacos , Extractos Vegetales/farmacología , Vitis/química , Apoptosis/efectos de los fármacos , Caspasa 12/metabolismo , Caspasa 3/metabolismo , Línea Celular Tumoral , Cloroquina/farmacología , Regulación hacia Abajo/efectos de los fármacos , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Proteínas de Choque Térmico/metabolismo , Humanos , Masculino , Microscopía Fluorescente , Extractos Vegetales/química , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Proteómica , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Espectrometría de Masas en Tándem , Regulación hacia Arriba/efectos de los fármacos , Vitis/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
OBJECTIVE: To observe the protective effects of Tongxinluo (TXL) on apoptosis of rat cardiac microvascular endothelial cells (RCMECs) resulting from homocysteine (Hcy) induced endoplasmic reticulum stress (ERS), and to determine the signaling pathway behind its protection. METHODS: Primary cultured RCMECs were isolated from neonatal rats using tissue explant method. The morphology of RCMECs was observed using inverted microscope, identified and differentiated by CD31 immunofluorescence method. Selected were well growing 2nd-4th generations of RCMECs. The optimal action time was determined by detecting the expression of glucose regulated protein 78 (GRP78) using immunofluorescence method. In the next experiment RCMECs were divided into 5 groups, i.e., the blank control group, the Hcy induced group (Hcy 10 mmol/L, 10 h), the Hcy + TXL group (Hcy 10 mmol/L + TXL 400 µg/mL), the Hcy +LY294002 group (Hcy 10 mmol/L + LY294002 5 µmol/L, LY294002 as the inhibitor of PI3K), the Hcy + LY294002 + TXL group (Hcy 10 mmol/L + LY294002 5 µmol/L + TXL 400 µg/mL). The apoptosis rate of RCMECs was detected by flow cytometry. mRNA and protein expressions of GRP78, C/ EBP homologous protein (CHOP), and cysteinyl aspartate specific proteinase-12 (caspase12) were detected by real-time reverse transcription PCR (RT-PCR) and Western blot respectively. Expression levels of phosphorylation of phosphatidylinositol 3-kinase (P-PI3K), total phosphatidylinositol 3-kinase (T- P13K) , phosphorylation of kinase B (P-Akt) , and total kinase B (T-Akt) were detected by Western blot. RESULTS: Ten hours Hcy action time was determined. Compared with the blank control group, the apoptosis rate was increased (22.77%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were increased, protein expressions of P-PI3K and P-Akt,ratios of P-PI3K/T-PI3K and P-Akt/T-Akt were decreased in the Hcy induced group (P < 0.05, P < 0.01). Compared with the Hcy induced group, the apoptosis rate was decreased (10.17%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were decreased, and expression levels of P-PI3K, P-Akt, P-PI3K/T-PI3K, and P-Akt/T-Akt were increased in the Hcy + TXL group (P < 0.05, P < 0.01). Compared with the Hcy + TXL group, the apoptosis rate was increased (17.9%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were increased, expression levels of P-PI3K and P-Akt, ratios of P-PI3K/T-PI3K and P-Akt/T-Akt were decreased in the Hcy + TXL + LY294002 group (P < 0.05, P < 0.01). CONCLUSION: TXL could inhibit the apoptosis of RCMECs resulting from Hcy-induced ERS and its mechanism might be associated with activating PI3K/Akt signaling pathway.
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Apoptosis/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Células Endoteliales/efectos de los fármacos , Transducción de Señal , Animales , Caspasa 12/metabolismo , Células Cultivadas , Cromonas/farmacología , Estrés del Retículo Endoplásmico , Morfolinas/farmacología , Miocardio/citología , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Factor de Transcripción CHOP/metabolismoRESUMEN
In the current study, we examined the potential effect of Ginsenoside Rg3 against gallbladder cancer cells, the underlying signaling mechanisms were also studied. We demonstrated that Rg3 exerted potent cytotoxic and pro-apoptotic activity against established and primary human gallbladder cancer cells. Yet it was safe to non-cancerous gallbladder epithelial cells. At the molecular level, we showed that Rg3 induced endoplasmic reticulum (ER) stress activation, the latter was evidenced by C/EBP homologous protein (CHOP) upregulation, inositol-requiring enzyme 1 (IRE1)/PKR-like endoplasmic reticulum kinase (PERK) phosphorylations, and caspase-12 activation in gallbladder cancer cells. Reversely, the ER stress inhibitor salubrinal, the caspase-12 inhibitor z-ATAD-fmk as well as CHOP shRNA knockdown significantly attenuated Rg3-induced cytotoxicity against gallbladder cancer cells. In vivo, we showed that Rg3 oral administration significantly inhibited GBC-SD gallbladder cancer xenograft growth in nude mice, its activity was, however, compromised with co-administration of the ER stress inhibitor salubrinal. Thus, we suggest that ER stress activation mediates Ginseng Rg3-induced anti-gallbladder cancer cell activity in vitro and in vivo.