Protein Expression Levels Of p-JNK Research Articles

Anti-Amnesia-like Effect of Pinus densiflora Extract by Improving Apoptosis and Neuroinflammation on Trimethyltin-Induced ICR Mice.

This study was conducted to investigate the anti-amnestic property of Korean red pine bark extract (KRPBE) on TMT-induced cognitive decline in ICR mice. As a result of looking at behavioral function, the consumption of KRPBE improved the spatial work ability, short-term learning, and memory ability by Y-maze, passive avoidance, and Morris water maze tests. KRPBE suppressed antioxidant system damage by assessing the SOD activity, reduced GSH content, and MDA levels in brain tissue. In addition, it had a protective effect on cholinergic and synaptic systems by regulating ACh levels, AChE activity, and protein expression levels of ChAT, AChE, SYP, and PSD-95. Also, the KRPBE ameliorated TMT-induced mitochondrial damage by regulating the ROS content, MMP, and ATP levels. Treatment with KRPBE suppressed Aβ accumulation and phosphorylation of tau and reduced the expression level of BAX/BCl-2 ratio and caspase 3, improving oxidative stress-induced apoptosis. Moreover, treatment with KRPBE improved cognitive dysfunction by regulating the neuro-inflammatory protein expression levels of p-JNK, p-Akt, p-IκB-α, COX-2, and IL-1β. Based on these results, the extract of Korean red pine bark, which is discarded as a byproduct of forestry, might be used as an eco-friendly material for functional foods or pharmaceuticals by having an anti-amnesia effect on cognitive impairment.

MG132 protects against lung injury following brain death in rats.

Brain death (BD) results in injury to organs and induces lung donor dysfunction. Since the 20S proteasome abnormality is associated with a variety of diseases, the present study investigated whether it was involved in lung injury following BD in rats, and the effects of the proteasome inhibitor MG132 on lung injury was also assessed. Rats were assigned to a BD group or a control sham group. The BD group of rats were sacrificed at different time points after BD. Administration of MG132 was performed intraperitoneally 30 min before BD. Arterial blood was drawn to measure the oxygenation index [partial artery pressure of oxygen (PaO2)/fractional concentration of inspired oxygen (FiO2)]. The right lung was used for staining with hematoxylin and eosin, immunohistochemistry, immunofluorescence, western blotting and RT-qPCR analysis. The left lung was used to measure the wet and dry weights. Rat alveolar macrophages (NR8383) were treated with MG132 and hypoxia/reoxygenation (H/R) and used for western blotting and flow cytometry. The PaO2/FiO2 ratio decreased after BD; the wet/dry weight ratio, histological lung injury score and protein expression of 20S proteasome β1 and inducible nitric oxide synthase (iNOS) gradually increased in rats after BD. Colocalization in the immunofluorescence between 20S proteasome β1 and iNOS was observed. MG132 treatment increased the PaO2/FiO2 ratio and decreased the wet/dry weight ratio, histological lung injury score and protein expression of 20S proteasome β1 and iNOS in rats after BD. MG132 was revealed to increase NR8383 apoptosis after H/R and to upregulate the protein expression levels of p-JNK and cleaved-caspase 3. Overall, the proteasome inhibitor MG132 could effectively reduce lung injury, which may be associated with its ability to inhibit the expression of the proteasome and promote the apoptosis of alveolar macrophages.

Butylphthalide inhibits nerve cell apoptosis in cerebral infarction rats via the JNK/p38 MAPK signaling pathway.

The aim of the present study was to investigate the influence of butylphthalide on nerve cell apoptosis in rats with cerebral infarction through the c-Jun N-terminal kinase (JNK)/p38 mitogen-activated protein kinase (MAPK) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into sham-operation group (n=12), model group (n=12) and butylphthalide group (n=12). Additionally, qPCR was performed to measure the mRNA expression of Bax and Bcl-2, and a TUNEL assay was conducted to investigate the cell apoptosis. Compared with the sham-operation group, the model group and the butylphthalide group had notably increased Zea-Longa scores (P<0.05), while the butylphthalide group exhibited a markedly decreased Zea-Longa score, compared with the model group (P<0.05). The positive expression of Bax was markedly higher (P<0.05), while that of Bcl-2 was notably lower in the model group and the butylphthalide group (P<0.05), compared with those in the sham-operation group. Furthermore, the positive expression of Bax was notably decreased (P<0.05), while that of Bcl-2 was markedly increased in the butylphthalide group in comparison with those in model group (P<0.05). The model group and the butylphthalide group had markedly higher relative protein expression levels of p-JNK and p-p38 MAPK than the sham-operation group (P<0.05), and the butylphthalide group displayed notably lower relative protein expression levels of p-JNK and p-p38 MAPK than the model group (P<0.05). The relative mRNA expression level of Bax was markedly increased (P<0.05), while that of Bcl-2 was notably decreased in the model group and the butylphthalide group (P<0.05), compared with those in the sham-operation group. Compared with those in the model group, the relative mRNA expression level of Bax decreased markedly (P<0.05), and that of Bcl-2 increased notably in the butylphthalide group (P<0.05). The apoptotic rate was markedly higher in the model group and the butylphthalide group than that in the sham-operation group (P<0.05), but it was notably lower in the butylphthalide group than that in the model group (P<0.05). In conclusion, butylphthalide may inhibit nerve cell apoptosis in rats with cerebral infarction to exert a protective effect, which may be associated with the JNK/p38 MAPK signaling pathway.

Efficacy of Cigu Xiaozhi pill on non-alcoholic steatohepatitis-associated lipoapoptosis through stress-activated c-Jun N-terminal kinase signalling pathway.

To investigate the efficacy of Cigu Xiaozhi pill (, CGXZ) on non-alcoholic steatohepatitis (NASH)-associated lipoapoptosis through the stress-activated c-Jun N-terminal kinase (JNK)/ stress-activated protein kinase signalling pathway. Sixty male Sprague-Dawley rats were randomly divided into the following groups (10rats each): blank control, model, low-dose CGXZ, medium-dose CGXZ, high-dose CGXZ, and positive control (treated with SP600125, a JNK inhibitor). The NASH model was established and the histomorphological characteristics of haematoxylin and eosin-stained liver tissues were examined under a light microscope. Cell apoptosis in liver tissues was assessed via terminal deoxynucleotidyl transferase dUTP nick-end labelling assay. In addition, the mRNA and protein expression levels of p-JNK, p-c-Jun, caspase-8, Fas, and Fas-L were determined via fluorescence-based quantitative real-time PCR, immunohistochemical and Western blot assays. Histopathological examination of the liver showed that the model rats had moderate-to-severe steatosis with infiltration of inflammatory cells as well as significantly higher expression levels of the p-JNK, p-c-Jun, caspase-8, Fas, and Fas-L proteins, compared with those in the blank control group (P < 0.01). Hepatic lobules of the rats in the treatment groups showed significantly reduced vacuolar degeneration and steatosis as well as alleviated inflammatory cell infiltration. The high and medium-dose CGXZ groups exhibited significantly lower mRNA and protein expression levels of p-JNK, p-c-Jun, caspase-8, Fas, and Fas-L, compared with those in the model group (P < 0.05 or P < 0.01). CGXZ pill inhibited the onset of hepatocyte apoptosis by regulating the expression of p-JNK, p-c-Jun, caspase-8, Fas, and Fas-L, thereby exerting therapeutic effects against NASH.

Inhibition effect of disulfiram combined with Cu on the growth of human Burkitt lymphoma cell xenografts in nude mice

Objective To investigate the effects of disulfiram(DS) combined with Cu on the human Burkitt lymphoma cell xenografts in nude mice. Methods Burkitt lymphoma xenograft was established by subcutaneous injection of Raji cell into nude mice after 2 Gy whole body X-irradiation(1×107 Raji cells were resuspended in 200 μl saline). 18 bearing tumor mice were randomly divided into control group, DS group and DS/Cu group. During the experiment, the effects of DS/Cu on the nude mice with tumors were examined, including the tumor volumes, weights and the growth curves of xenograft tumor. Histopathological examination of tumor tissue was observed with optical microscape. The protein expression levels of p-JNK and c-jun were also detected by Western blot. Results Subsequent tumor size and weight in DS or DS/Cu-treated animals were (67.71±2.15) mm3, (33.35±7.74) mm3 and (43.35±4.22) mg, (18.05±2.88) mg. One-way ANOVA analysis indicated that the tumor size and weight in DS or DS/Cu-treated animals were reduced significantly relative to tumors in vehicle-treated animals(F=27.579, P=0.000; F=16.369, P=0.000). Furthermore, multiple comparisons revealed that the DS or DS/Cu-treated animals had significantly reduced tumor size and weight compared with control animals (all P<0.05). There were significant differences in tumor size and weight between DS or DS/Cu-treated animals (both P<0.05). Tumor inhibition rates in DS or DS/Cu group were 63.48 % and 80.24 %, respectively. An increase of apoptosis changes in the xenograft tumor cells in DS or DS/Cu treated mice were more significant. Western blot showed that the p-JNK and c-jun protein expressions in the tumors were improved after the DS or DS/Cu treatment, more obvious in DS/Cu treatment. Conclusion DS/Cu can inhibit the growth of xenografts, and one possible mechanism may involve the regulation of JNK signal pathway. Key words: Disulfiram; Copper; Burkitt lymphoma; JNK signal pathway

Supercritical Fluid Extract of Spent Coffee Grounds Attenuates Melanogenesis through Downregulation of the PKA, PI3K/Akt, and MAPK Signaling Pathways.

The mode of action of spent coffee grounds supercritical fluid CO2 extract (SFE) in melanogenesis has never been reported. In the study, the spent coffee grounds were extracted by the supercritical fluid CO2 extraction method; the chemical constituents of the SFE were investigated by gas chromatography-mass spectrometry (GC-MS). The effects of the SFE and its major fatty acid components on melanogenesis were evaluated by mushroom tyrosinase activity assay and determination of intracellular tyrosinase activity and melanin content. The expression level of melanogenesis-related proteins was analyzed by western blotting assay. The results revealed that the SFE of spent coffee grounds (1–10 mg/mL) and its major fatty acids such as linoleic acid and oleic acid (6.25–50 μM) effectively suppressed melanogenesis in the B16F10 murine melanoma cells. Furthermore, the SFE decreased the expression of melanocortin 1 receptor (MC1R), microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2). The SFE also decreased the protein expression levels of p-JNK, p-p38, p-ERK, and p-CREB. Our results revealed that the SFE of spent coffee grounds attenuated melanogenesis in B16F10 cells by downregulation of protein kinase A (PKA), phosphatidylinositol-3-kinase (PI3K/Akt), and mitogen-activated protein kinases (MAPK) signaling pathways, which may be due to linoleic acid and oleic acid.

Danshensu protects against ischemia/reperfusion injury and inhibits the apoptosis of H9c2 cells by reducing the calcium overload through the p-JNK-NF-κB-TRPC6 pathway.

Ischemia-reperfusion(I/R) plays an important role in myocardial injury. In the present study, we aimed to examine the protective effects of Danshensu(DSS) against I/R injury and to elucidate the underlying mechanisms. For this purpose, H9c2 cells were cultured in hypoxic solution in a hypoxic incubator for 2h, and then cultured in a high oxygen incubator for various periods of time and pre-treated with or without DSS, ammonium pyrrolidine dithiocarbamate(PDTC) or SP600125[a c-Jun N-terminal kinase(JNK) inhibitor]. Cell apoptosis and cytosolic free Ca2+([Ca2+]i) levels were analyzed by flow cytometry. The protein expression levels of JNK, phosphorylated(p-)JNK, nuclear factor-κB(NF-κB) and transient receptor potential cation channel, subfamilyC, member6(TRPC6) were measured by western blot analysis. The mRNA expression levels of JNK were measured by RT-qPCR. The results revealed that TRPC6 protein expression, the cell apoptotic rate and the [Ca2+]i levels increased in a time-dependent manner in the H9c2 cells following the induction of I/R injury. The apoptotic rate and TRPC6 protein expression decreased when the cells were treated with DSS prior to the induction of I/R injury. The knockdown of JNK expression by siRNA decreased the p-JNK and TRPC6 protein expression levels in the H9c2 cells subjected to I/R injury. The protein expression levels of p-JNK and NF-κB in the nucleus increased significantly when the H9c2 cells were subjected to I/R injury, whereas NF-κB expression in the cytoplasm decreased in a time‑dependent manner. However, p-JNK, NF-κB and TRPC6 protein expression, the [Ca2+]i level and cell apoptosis decreased when the H9c2 cells were pre-treated with DSS or SP600125. Therefore, our data suggest that DSS prevents myocardial I/R injury by inhibiting p-JNK activation and NF-κB translocation, which potentially upregulate TRPC6 expression, increase the [Ca2+]i level, and result in the apoptosis of H9c2 cells.

용담화 에탄올 추출물의 항노화 작용 연구

Objective : The goal of this study is to identify the effects of extract of Gentianae sino-ornata (GSO) on the anti-oxidative activity of skin. For this purpose, several functions of GSO were analyzed in terms of skin-lightening activity and wrinkle improvement. Methods : Cell viability was measured by neutral red (NR) assay, and GSO showed highly efficacy in DPPH radical scavenging activity. The level of tyrosinase and matrix metalloproteinase-1 (MMP-1) in media was analyzed by ELISA kit, and the expressions of p-JNK and p-ERK was measured by Western blot. To elucidate inhibitory effects of GSO on melanin synthesis, I determined the tyrosinase activity and melanin production in B16F10 cells. Results : MMP-1 production in UVB-stimulated HDF cells was inhibited by G SO treatments, and also GSO inhibited protein expression levels of p-JNK and p-ERK. GSO significantly reduced tyrosinase activity and melanin synthesis in B16F10 cells. Conclusions : From these results, GSO appears to be effective on skin elasti city increase, wrinkle improvement, whitening as anti-aging activity.

Study on the mechanism of DDP-resistance mediated by phosphate JNK in gastric cancer

To investigate the mechanism of DDP-resistance in gastric cancer cell line SGC7901/DDP mediated by phosphate(p)-JNK. The p-JNK expression was blocked by the JNK inhibitor SP600125. The drug sensitivity was detected by MTT. Cell apoptosis rate was measured by flow cytometry. The expression of p-JNK and P-glycoprotein (P-gp) was examined by Western blot. The expression of both proteins were detected in a tissue microarray containing 168 spots of cancer tissue and 27 spots of normal gastric tissue by SP immunohistochemistry. Pearson method was used to analyze the correlation between p-JNK and P-gp. The drug sensitivity and cell apoptosis rate significantly increased (P<0.01), and the protein expression levels of p-JNK and P-glycoprotein were down-regulated after the inhibition of p-JNK by SP600125 in both SGC7901(p-JNK: 1.17+/-0.03 vs 0.38+/-0.071, P-gp: 0.21+/-0.01 vs 0.06+/-0.01) and SGC7901/DDP (p-JNK: 2.56+/-0.14 vs 1.02+/-0.12, P-gp: 0.77+/-0.05 vs 0.52+/-0.06 )cells(all P<0.01). The protein expression rates of p-JNK and P-glycoprotein were 45.8% and 51.8% respectively in gastric cancer tissue, which were significantly higher than those in normal gastric tissue 7.4% and 18.5% (P<0.01). The correlation of protein expression of p-JNK and P-gp was positive (P<0.01). JNK anti-apoptosis pathway with the regulation of P-gp expression plays an important role in the DDP-resistance of gastric cancer, which may be a novel target for reversing multidrug resistance.