Objective To investigate the protective effects of hydrogen peroxide preconditioning （HPP） on the pheochromocytoma （PC12） cells treated with 1-methyl-4-phenylpyridinium （MPP^＋） and to explore the potential mech...Objective To investigate the protective effects of hydrogen peroxide preconditioning （HPP） on the pheochromocytoma （PC12） cells treated with 1-methyl-4-phenylpyridinium （MPP^＋） and to explore the potential mechanisms. Methods The viability and apoptosis of PC 12 cells were determinded by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay and 4′,6′-diamidino-2-phenylindole （DAPI） staining, respectively. The expressions of 14-3-3 protein and phospholylated p38 mitogen-activated protein kinase （MAPK） were determined by Western blot. Enzyme-linked immunosorbent assay （ELISA） was used to measure the activity of extracellular signal-regulated protein kinase 1/2 （ERK1/2）. Results The cell viability decreased and the number of apoptotic cells increased dramatically in MPP^＋ group compared with that in Control group. HPP induced a significant increase in cell viability and a marked decrease in population of apoptotic cells of the MPP^＋- treated PC 12 cells, accompanied with up-regulation of 14-3-3 protein and increase of ERK 1/2 and p38 MAPK activities. The 14-3-3 protein expression was positively correlated with the phosphorylation of ERK1/2. Furthermore, inhibition of the ERK1/2 with PD98059 abolished the 14-3-3 protein up-regulation in PC 12 cells induced by HPP. Conclusion HPP protects PC 12 cells against MPP＋ toxicity by up-regulating 14-3-3 protein expression through the ERK1/2 and p38 MAPK signaling pathways.展开更多
In the present study, we constructed a lentivirus, FIV-CMV-GFP-miR-7-3, containing the microRNA-7-3 gene and the green fluorescent protein gene, and used it to transfect human glioma U251 cells. Fluorescence microscop...In the present study, we constructed a lentivirus, FIV-CMV-GFP-miR-7-3, containing the microRNA-7-3 gene and the green fluorescent protein gene, and used it to transfect human glioma U251 cells. Fluorescence microscopy showed that 80% of U251 cells expressed green fluorescence. Real-time reverse transcription PCR showed that microRNA-7-3 RNA expression in U251 cells was significantly increased. Proliferation was slowed in transfected U251 cells, and most cells were in the G1 phase of the cell cycle. In addition, the expression of the serine/threonine protein kinase 2 was decreased. Results suggested that transfection with a lentivirus carrying microRNA-7-3 can effectively suppress epidermal growth factor receptor pathway activity in U251 cells, arrest cell cycle transition from GI phase to S phase and inhibit glioma cell growth.展开更多
OBJECTIVE MicroR NA(miR NA)holds promise as a novel therapeutic tool for cancer treatment.However,the transfection efficiency of current delivery systems represents a bottleneck for clinical applications.Here,we demon...OBJECTIVE MicroR NA(miR NA)holds promise as a novel therapeutic tool for cancer treatment.However,the transfection efficiency of current delivery systems represents a bottleneck for clinical applications.Here,we demonstrate that gap junctions mediate an augmentative effect on the antiproliferation mediated by mi R-124-3p in U87 and C6 glioblastoma cells.METHODS The functional inhibition of gap junctions using either si RNA or pharmacological inhibition eliminated the mi R-124-3p-mediated antiproliferation,whereas the enhancement of gap junctions with retinoic acid treatment augmented this mi R-124-3p-mediated antiproliferation.A similar effect was observed in glioblastoma xenograft models.RESULTS More importantly,patch clamp and co-culture assays demonstrated the transmission of mi R-124-3p through gap junction channels into adjacent cells.In further exploring the impact of gap junction-mediated transport of mi R-124-3p on mi R-124-3p target pathways,we found that mi R-124-3p inhibited glioblastoma cell growth in part by decreasing the protein expression of cyclindependent kinase 6,leading to cel cycle arrest at the G0/G1phase;moreover,pharmacological regulation of gap junctions affected this cell cycle arrest.CONCLUSION Our results indicate that the″bystander″effects of functional gap junctions composed of connexin 43 enhance the antitumor effect of mi R-124-3p in glioblastoma cells by transferring mi R-124-3p to adjacent cells,thereby enhancing G0/G1cell cycle arrest.These observations provide a new guiding strategy for the clinical application of mi RNA therapy in tumor treatment.展开更多
Background:Wound management of diabetic foot ulcers(DFUs)is a complex and challenging task,and existing strategies fail to meet clinical needs.Therefore,it is important to develop novel drug candidates and discover ne...Background:Wound management of diabetic foot ulcers(DFUs)is a complex and challenging task,and existing strategies fail to meet clinical needs.Therefore,it is important to develop novel drug candidates and discover new therapeutic targets.However,reports on peptides as molecular probes for resolving issues related to DFUs remain rare.This study utilized peptide RL-QN15 as an exogenous molecular probe to investigate the underlying mechanism of endogenous non-coding RNA in DFU wound healing.The aim was to generate novel insights for the clinical management of DFUs and identify potential drug targets.Methods:We investigated the wound-healing efficiency of peptide RL-QN15 under diabetic con-ditions using in vitro and in vivo experimental models.RNA sequencing,in vitro transfection,quantitative real-time polymerase chain reaction,western blotting,dual luciferase reporter gene detection,in vitro cell scratches,and cell proliferation and migration assays were performed to explore the potential mechanism underlying the promoting effects of RL-QN15 on DFU repair.Results:Peptide RL-QN15 enhanced the migration and proliferation of human immortalized keratinocytes(HaCaT cells)in a high-glucose environment and accelerated wound healing in a DFU rat model.Based on results from RNA sequencing,we defined a new microRNA(miR-4482-3p)related to the promotion of wound healing.The bioactivity of miR-4482-3p was verified by inhibiting and overexpressing miR-4482-3p.Inhibition of miR-4482-3p enhanced the migration and proliferation ability of HaCaT cells as well as the expression of vascular endothelial growth factor B(VEGFB).RLQN15 also promoted the migration and proliferation ability of HaCaT cells,and VEGFB expression was mediated via inhibition of miR-4482-3p expression by the p38 mitogen-activated protein kinase(p38MAPK)and smad3 signaling pathways.Conclusions:RL-QN15 is an effective molecule for the treatment of DFUs,with the underlying mechanism related to the inhibition of miR-4482-3p expression via the p38MAPK and smad3 signaling pathways,ultimately promoting re-epithelialization,angiogenesis and wound healing.This study provides a theoretical basis for the clinical application of RL-QN15 as a molecular probe in promoting DFU wound healing.展开更多
After spinal cord injury, dysregulated miRNAs appear and can participate in inflammatory responses, as well as the inhibition of apoptosis and axon regeneration through multiple pathways. However, the functions of miR...After spinal cord injury, dysregulated miRNAs appear and can participate in inflammatory responses, as well as the inhibition of apoptosis and axon regeneration through multiple pathways. However, the functions of miRNAs in spinal cord ischemia-reperfusion injury progression remain unclear. miRCURY LNATM Arrays were used to analyze miRNA expression profiles of rats after 90 minutes of ischemia followed by reperfusion for 24 and 48 hours. Furthermore, subsequent construction of aberrantly expressed miRNA regulatory patterns involved cell survival, proliferation, and apoptosis. Remarkably, the mitogen-activated protein kinase(MAPK) signaling pathway was the most significantly enriched pathway among 24-and 48-hour groups. Bioinformatics analysis and quantitative reverse transcription polymerase chain reaction confirmed the persistent overexpression of miR-22-3 p in both groups. These results suggest that the aberrant miRNA regulatory network is possibly regulated MAPK signaling and continuously affects the physiological and biochemical status of cells, thus participating in the regulation of spinal cord ischemia-reperfusion injury. As such, miR-22-3 p may play sustained regulatory roles in spinal cord ischemia-reperfusion injury. All experimental procedures were approved by the Animal Ethics Committee of Jilin University, China [approval No. 2020(Research) 01].展开更多
AIM： To address the possibility that insulin-like growth factor （IGF）-Ⅱ is a growth factor and its signaling pathway so as to develop a molecular therapy for hepatoblastoma. METHODS： Huh-6 and HepG2, human hepato...AIM： To address the possibility that insulin-like growth factor （IGF）-Ⅱ is a growth factor and its signaling pathway so as to develop a molecular therapy for hepatoblastoma. METHODS： Huh-6 and HepG2, human hepatoblastoma cell lines, were used. IGF-Ⅱ was added to the medium deprived of serum. Western blot analysis was performed to clarify the expression of IGF-Ⅰ receptor （IGF-IR）. Inhibitors of IGF-IR （piclopodophyllin, PPP）, phosphatidyl-inositol （PI） 3-kinase （LY294002 and Wortmannin）, or mitogen-activated protein （MAP） kinase （PD98059） were added to unveil the signaling pathway of IGF-Ⅱ. Cells were analyzed morphologically with hematoxylin-eosin staining to reveal the mechanism of suppression of cell proliferation. RESULTS： IGF-Ⅱ stimulated cells proliferated to 2.7 （269% ± 76%） （mean ± SD） （Huh-6） and 2.1 （211% ± 85%） times （HepG2）. IGF-IR was expressed in Huh-6 and HepG2. PPP suppressed the cell number to 44% ± 11% （Huh-6） and 39% ± 5% （HepG2）. LY294002 and Wortmannin suppressed the cell number to 30% ± 5% （Huh-6）, 44% ± 0.4% （HepG2）, 49% ± 1.0% （Huh-6） and 46% ± 1.1% （HepG2）, respectively. PD98059 suppressed the cell number to 33% ± 11% for HepG2 but not for Huh-6. When cell proliferation was prohibited, many Huh-6 and HepG2 cells were dead with pyknotic or fragmented nuclei, suggesting apoptosis. CONCLUSION： IGF- Ⅱ was shown to be a growth factor of hepatoblastoma via IGF-Ⅰ receptor and PI3 kinase which were good candidates for target of molecular therapy.展开更多
Tongluojiunao （TLJN） is an herbal medicine consisting of two main components, geniposide and ginsenoside Rg1. TLJN has been shown to protect primary cultured hippocampal neurons. How-ever, its mechanism of action re...Tongluojiunao （TLJN） is an herbal medicine consisting of two main components, geniposide and ginsenoside Rg1. TLJN has been shown to protect primary cultured hippocampal neurons. How-ever, its mechanism of action remains unclear. In the present study, primary cultured hippocampal neurons treated with Aβ1-42 （10 μmol/L） signiifcantly increased the release of lactate dehydroge-nase, which was markedly reduced by TLJN （2 μL/mL）, speciifcally by the component geniposide （26 μmol/L）, but not ginsenoside Rg1 （2.5 μmol/L）. hTe estrogen receptor inhibitor, ICI182780 （1 μmol/L）, did not block TLJN-or geniposide-mediated decrease of lactate dehydrogenase under Aβ1-42-exposed conditions. However, the phosphatidyl inositol 3-kinase or mitogen-activated protein kinase pathway inhibitor, LY294002 （50 μmol/L） or U0126 （10 μmol/L）, respectively blo cked the decrease of lactate dehydrogenase mediated by TLJN or geniposide. hTerefore, these results suggest that the non-classical estrogen pathway （i.e., phosphatidyl inositol 3-kinase or mitogen-activated protein kinase） is involved in the neuroprotective effect of TLJN, speciifcally its component, geniposide, against Aβ1-42-mediated cell death in primary cultured hippocampal neurons.展开更多
We aim to investigate the effect of transforming growth factor （TGF）-β1 on the expression of enhancer of split- and hairy-related protein-2 （SHARP-2） messenger RNA （mRNA） and its signaling pathway. In this stud...We aim to investigate the effect of transforming growth factor （TGF）-β1 on the expression of enhancer of split- and hairy-related protein-2 （SHARP-2） messenger RNA （mRNA） and its signaling pathway. In this study, several cell lines including LLC-PK1 （a porcine kidney tubular epithelial cell line）, MDCK （Madin-Darby canine kidney） and CTLL-2 （cytotoxic T-lymphocyte line） were treated with recombinant human TGF-131, and a series of experiments were carried out, involving Northern blot analysis of total RNA from these cells. Further, several specific chemical inhibitors were applied before TGF-β1 treatment to probe the signaling pathway. The results showed that TGF-β1 can significadtly up-regulate SHARP-2 mRNA expression in the LLC-PK1 cell line. The peak level of induction was found 2 h after TGF-β1 stimulation. While one phospho- inositide 3-kinases （PI-3） kinase inhibitor, LY294002, completely blocked the effect of TGF-131 on SHARP-2 mRNA expression in LLC-PK1 cells at a low concentration, other inhibitors, including PD98059, staurosporine, AG490, wortmannin, okadaic acid and rapamycin, had no effect. The effect of LY294002 was dose-dependent. We conclude that, in LLC-PK1 cells at least, TGF-β1 can effectively induce the SHARP-2 mRNA expression and that the PI-3 kinase pathway can mediate this effect.展开更多
14-3-3 proteins play an important role in the regulation of many cellular processes. The Arabidopsis vacuolar two-pore K＋ channel 1 （TPK1） interacts with the 14-3-3 protein GRF6 （GF14-λ）. Upon phosphorylation of...14-3-3 proteins play an important role in the regulation of many cellular processes. The Arabidopsis vacuolar two-pore K＋ channel 1 （TPK1） interacts with the 14-3-3 protein GRF6 （GF14-λ）. Upon phosphorylation of the putative binding motif in the N-terminus of TPK1, GRF6 binds to TPK1 and activates the potassium channel. In order to gain a deeper understanding of this 14-3-3-mediated signal transduction, we set out to identify the respective kinases, which regulate the phosphorylation status of the 14-3-3 binding motif in TPK1. Here, we report that the calcium-dependent protein kinases （CDPKs） can phosphorylate and thereby activate the 14-3-3 binding motif in TPK1. Focusing on the stress-activated kinase CPK3, we visualized direct and specific interaction of TPK1 with the kinase at the tonoplast in vivo. In line with its proposed role in K＋ homeostasis, TPK1 phosphorylation was found to be induced by salt stress in planta, and both cpk3 and tpkl mutants displayed salt-sensitive phenotypes. Molecular modeling of the TPK1-CPK3 interaction domain provided mechanistic insights into TPK1 stress-regulated phosphorylation responses and pinpointed two arginine residues in the N-terminal 14-3-3 binding motif in TPK1 critical for kinase interaction. Taken together, our studies provide evidence for an essential role of the vacuolar potassium channel TPK1 in salt-stress adaptation as a target of calcium-regulated stress signaling pathways involving Ca2＋, Ca2＋-dependent kinases, and 14-3-3 proteins.展开更多
Background ^11C-4-N-（3-bromoanilino）-6,7-dimethoxyquinazoline （1^11C-PD153035） has been reported as a tracer for imaging human tumors that overexpress epidermal growth factor receptor （EGFR）. However it is still...Background ^11C-4-N-（3-bromoanilino）-6,7-dimethoxyquinazoline （1^11C-PD153035） has been reported as a tracer for imaging human tumors that overexpress epidermal growth factor receptor （EGFR）. However it is still unclear whether ^11C-PD153035 uptake correlates with EGFR expression levels. The objective of this study was to investigate the relationship between ^11C-PD153035 accumulation and EGFR expression levels. Methods Synthesis of ^11C-PD153035 was performed in the Tracerlab FXc system. Accumulation of ^11C-PD153035 by MDA-MB-468, A549 and MDA-MB-231 cells was measured in vitro. There were six tumor-bearing mice in each group. ^11C-PD153035 uptake in tumors was determined by positron emission tomography/computed tomography （PET/CT）. Tumor/normal muscle tissue （TINT） analysis in PET images was applied to quantify the PET data. Sixty minutes after PET/CT scanning, the nude mice were sacrificed and the tumors were excised. The ^11C-PD153035 accumulation in different tumors was determined by a gamma counter. Results Close correlation existed between the uptake and the level of EGFR expression both in vitro and ex vivo （r^2=0.72, P〈0.001; r^2=0.63, P=0.003）. When the static TINT analysis method was applied to analyze the PET data, the observed correlation was again excellent （^2=0.70, P=-0.001）. Conclusions The uptake of PET tracer ^11C-PD153035 closely correlates with the EGFR expression levels in tumor cells. ^11C-PD153035 has the potential to yield useful information for both cancer diagnosis and therapy.展开更多
BACKGROUND： Phototropism is the response a plant exhibits when it is faced with a directional blue light stimulus. Though a seemingly simple differential cell elongation response within a responding tissue that resul...BACKGROUND： Phototropism is the response a plant exhibits when it is faced with a directional blue light stimulus. Though a seemingly simple differential cell elongation response within a responding tissue that results in organ curvature, phototropism is regulated through a complex set of signal perception and transduction events that move from the plasma membrane to the nucleus. In nature phototropism is one of several plant responses that have evolved to optimize photosynthesis and growth. OBJECTIVE： In the present work we will review the state of the field with respect to the molecules and mechanisms associated with phototropism in land plants. METHODS： A systematic literature search was done to identify relevant advances in the field. Though we tried to focus on literature within the past decade （1998-present）, we have discussed and cited older literature where appropriate because of context or its significant impact to the present field. Several previous review articles are also cited where appropriate and readers should seek those out. RESULTS： A total of 199 articles are cited that fulfill the criteria listed above. CONCLUSIONS： Though important numerous and significant advances have been made in our understanding of the molecular, biochemical, cell biological and physiologic mechanisms underlying phototropism in land plants over the past decade, there are many remaining unanswered questions. The future is indeed bright for researchers in the field and we look forward to the next decade worth of discoveries.展开更多
基金the National Natural Science Foundation of China (No. 30570627)
文摘Objective To investigate the protective effects of hydrogen peroxide preconditioning （HPP） on the pheochromocytoma （PC12） cells treated with 1-methyl-4-phenylpyridinium （MPP^＋） and to explore the potential mechanisms. Methods The viability and apoptosis of PC 12 cells were determinded by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay and 4′,6′-diamidino-2-phenylindole （DAPI） staining, respectively. The expressions of 14-3-3 protein and phospholylated p38 mitogen-activated protein kinase （MAPK） were determined by Western blot. Enzyme-linked immunosorbent assay （ELISA） was used to measure the activity of extracellular signal-regulated protein kinase 1/2 （ERK1/2）. Results The cell viability decreased and the number of apoptotic cells increased dramatically in MPP^＋ group compared with that in Control group. HPP induced a significant increase in cell viability and a marked decrease in population of apoptotic cells of the MPP^＋- treated PC 12 cells, accompanied with up-regulation of 14-3-3 protein and increase of ERK 1/2 and p38 MAPK activities. The 14-3-3 protein expression was positively correlated with the phosphorylation of ERK1/2. Furthermore, inhibition of the ERK1/2 with PD98059 abolished the 14-3-3 protein up-regulation in PC 12 cells induced by HPP. Conclusion HPP protects PC 12 cells against MPP＋ toxicity by up-regulating 14-3-3 protein expression through the ERK1/2 and p38 MAPK signaling pathways.
基金supported by the Science and Technology Foundation Program of Jiangsu Province(Tumorigenic nucleostemin genes and adenovirus-based RNA interference targeting to brain tumor stem cell the rapy),No.BK2007072
文摘In the present study, we constructed a lentivirus, FIV-CMV-GFP-miR-7-3, containing the microRNA-7-3 gene and the green fluorescent protein gene, and used it to transfect human glioma U251 cells. Fluorescence microscopy showed that 80% of U251 cells expressed green fluorescence. Real-time reverse transcription PCR showed that microRNA-7-3 RNA expression in U251 cells was significantly increased. Proliferation was slowed in transfected U251 cells, and most cells were in the G1 phase of the cell cycle. In addition, the expression of the serine/threonine protein kinase 2 was decreased. Results suggested that transfection with a lentivirus carrying microRNA-7-3 can effectively suppress epidermal growth factor receptor pathway activity in U251 cells, arrest cell cycle transition from GI phase to S phase and inhibit glioma cell growth.
基金The project supported by National Natural Science Foundation of China(81473234,U1303221)
文摘OBJECTIVE MicroR NA(miR NA)holds promise as a novel therapeutic tool for cancer treatment.However,the transfection efficiency of current delivery systems represents a bottleneck for clinical applications.Here,we demonstrate that gap junctions mediate an augmentative effect on the antiproliferation mediated by mi R-124-3p in U87 and C6 glioblastoma cells.METHODS The functional inhibition of gap junctions using either si RNA or pharmacological inhibition eliminated the mi R-124-3p-mediated antiproliferation,whereas the enhancement of gap junctions with retinoic acid treatment augmented this mi R-124-3p-mediated antiproliferation.A similar effect was observed in glioblastoma xenograft models.RESULTS More importantly,patch clamp and co-culture assays demonstrated the transmission of mi R-124-3p through gap junction channels into adjacent cells.In further exploring the impact of gap junction-mediated transport of mi R-124-3p on mi R-124-3p target pathways,we found that mi R-124-3p inhibited glioblastoma cell growth in part by decreasing the protein expression of cyclindependent kinase 6,leading to cel cycle arrest at the G0/G1phase;moreover,pharmacological regulation of gap junctions affected this cell cycle arrest.CONCLUSION Our results indicate that the″bystander″effects of functional gap junctions composed of connexin 43 enhance the antitumor effect of mi R-124-3p in glioblastoma cells by transferring mi R-124-3p to adjacent cells,thereby enhancing G0/G1cell cycle arrest.These observations provide a new guiding strategy for the clinical application of mi RNA therapy in tumor treatment.
基金supported by grants from the National Natural Science Foundation of China(32360138,32060212,32301054 and 81760648)Key Program of Yunnan Fundamental Research Project(202301AS070036)+1 种基金Outstanding Youth Program of Yunnan Applied Basic Research Project-Kunming Medical University Union Foundation(202301AY070001-301)Project of Yunnan Applied Basic Research Project-Kunming Medical University Union Foundation(202101AY070001-006 and 202101AY070001-036).
文摘Background:Wound management of diabetic foot ulcers(DFUs)is a complex and challenging task,and existing strategies fail to meet clinical needs.Therefore,it is important to develop novel drug candidates and discover new therapeutic targets.However,reports on peptides as molecular probes for resolving issues related to DFUs remain rare.This study utilized peptide RL-QN15 as an exogenous molecular probe to investigate the underlying mechanism of endogenous non-coding RNA in DFU wound healing.The aim was to generate novel insights for the clinical management of DFUs and identify potential drug targets.Methods:We investigated the wound-healing efficiency of peptide RL-QN15 under diabetic con-ditions using in vitro and in vivo experimental models.RNA sequencing,in vitro transfection,quantitative real-time polymerase chain reaction,western blotting,dual luciferase reporter gene detection,in vitro cell scratches,and cell proliferation and migration assays were performed to explore the potential mechanism underlying the promoting effects of RL-QN15 on DFU repair.Results:Peptide RL-QN15 enhanced the migration and proliferation of human immortalized keratinocytes(HaCaT cells)in a high-glucose environment and accelerated wound healing in a DFU rat model.Based on results from RNA sequencing,we defined a new microRNA(miR-4482-3p)related to the promotion of wound healing.The bioactivity of miR-4482-3p was verified by inhibiting and overexpressing miR-4482-3p.Inhibition of miR-4482-3p enhanced the migration and proliferation ability of HaCaT cells as well as the expression of vascular endothelial growth factor B(VEGFB).RLQN15 also promoted the migration and proliferation ability of HaCaT cells,and VEGFB expression was mediated via inhibition of miR-4482-3p expression by the p38 mitogen-activated protein kinase(p38MAPK)and smad3 signaling pathways.Conclusions:RL-QN15 is an effective molecule for the treatment of DFUs,with the underlying mechanism related to the inhibition of miR-4482-3p expression via the p38MAPK and smad3 signaling pathways,ultimately promoting re-epithelialization,angiogenesis and wound healing.This study provides a theoretical basis for the clinical application of RL-QN15 as a molecular probe in promoting DFU wound healing.
基金supported by the National Natural Science Foundation of China,No.81350013(to XYY)。
文摘After spinal cord injury, dysregulated miRNAs appear and can participate in inflammatory responses, as well as the inhibition of apoptosis and axon regeneration through multiple pathways. However, the functions of miRNAs in spinal cord ischemia-reperfusion injury progression remain unclear. miRCURY LNATM Arrays were used to analyze miRNA expression profiles of rats after 90 minutes of ischemia followed by reperfusion for 24 and 48 hours. Furthermore, subsequent construction of aberrantly expressed miRNA regulatory patterns involved cell survival, proliferation, and apoptosis. Remarkably, the mitogen-activated protein kinase(MAPK) signaling pathway was the most significantly enriched pathway among 24-and 48-hour groups. Bioinformatics analysis and quantitative reverse transcription polymerase chain reaction confirmed the persistent overexpression of miR-22-3 p in both groups. These results suggest that the aberrant miRNA regulatory network is possibly regulated MAPK signaling and continuously affects the physiological and biochemical status of cells, thus participating in the regulation of spinal cord ischemia-reperfusion injury. As such, miR-22-3 p may play sustained regulatory roles in spinal cord ischemia-reperfusion injury. All experimental procedures were approved by the Animal Ethics Committee of Jilin University, China [approval No. 2020(Research) 01].
基金Supported by the Japan Society for the Promotion of Science (JSPS) (16590577) and the Kawano Masanori Memorial Foundation for Promotion of Pediatrics (15-6)
文摘AIM： To address the possibility that insulin-like growth factor （IGF）-Ⅱ is a growth factor and its signaling pathway so as to develop a molecular therapy for hepatoblastoma. METHODS： Huh-6 and HepG2, human hepatoblastoma cell lines, were used. IGF-Ⅱ was added to the medium deprived of serum. Western blot analysis was performed to clarify the expression of IGF-Ⅰ receptor （IGF-IR）. Inhibitors of IGF-IR （piclopodophyllin, PPP）, phosphatidyl-inositol （PI） 3-kinase （LY294002 and Wortmannin）, or mitogen-activated protein （MAP） kinase （PD98059） were added to unveil the signaling pathway of IGF-Ⅱ. Cells were analyzed morphologically with hematoxylin-eosin staining to reveal the mechanism of suppression of cell proliferation. RESULTS： IGF-Ⅱ stimulated cells proliferated to 2.7 （269% ± 76%） （mean ± SD） （Huh-6） and 2.1 （211% ± 85%） times （HepG2）. IGF-IR was expressed in Huh-6 and HepG2. PPP suppressed the cell number to 44% ± 11% （Huh-6） and 39% ± 5% （HepG2）. LY294002 and Wortmannin suppressed the cell number to 30% ± 5% （Huh-6）, 44% ± 0.4% （HepG2）, 49% ± 1.0% （Huh-6） and 46% ± 1.1% （HepG2）, respectively. PD98059 suppressed the cell number to 33% ± 11% for HepG2 but not for Huh-6. When cell proliferation was prohibited, many Huh-6 and HepG2 cells were dead with pyknotic or fragmented nuclei, suggesting apoptosis. CONCLUSION： IGF- Ⅱ was shown to be a growth factor of hepatoblastoma via IGF-Ⅰ receptor and PI3 kinase which were good candidates for target of molecular therapy.
基金supported by the National Natural Science Foundation of China No.81072901the New Teacher Fund for Doctor Station,Ministry of Education,No.20120013110013+1 种基金grants from the Nautical Traditional Chinese Medicine Discipline,No.522/0100604054grants from the Nautical Traditional Chinese Medicine Collaborative Innovation Center,No.522/0100604299
文摘Tongluojiunao （TLJN） is an herbal medicine consisting of two main components, geniposide and ginsenoside Rg1. TLJN has been shown to protect primary cultured hippocampal neurons. How-ever, its mechanism of action remains unclear. In the present study, primary cultured hippocampal neurons treated with Aβ1-42 （10 μmol/L） signiifcantly increased the release of lactate dehydroge-nase, which was markedly reduced by TLJN （2 μL/mL）, speciifcally by the component geniposide （26 μmol/L）, but not ginsenoside Rg1 （2.5 μmol/L）. hTe estrogen receptor inhibitor, ICI182780 （1 μmol/L）, did not block TLJN-or geniposide-mediated decrease of lactate dehydrogenase under Aβ1-42-exposed conditions. However, the phosphatidyl inositol 3-kinase or mitogen-activated protein kinase pathway inhibitor, LY294002 （50 μmol/L） or U0126 （10 μmol/L）, respectively blo cked the decrease of lactate dehydrogenase mediated by TLJN or geniposide. hTerefore, these results suggest that the non-classical estrogen pathway （i.e., phosphatidyl inositol 3-kinase or mitogen-activated protein kinase） is involved in the neuroprotective effect of TLJN, speciifcally its component, geniposide, against Aβ1-42-mediated cell death in primary cultured hippocampal neurons.
基金supported by the National Natural Science Foundation of China (Nos. 30471641 and 30872389)the Natural Science Foundation of Zhejiang Province, China (No. Y207088)
文摘We aim to investigate the effect of transforming growth factor （TGF）-β1 on the expression of enhancer of split- and hairy-related protein-2 （SHARP-2） messenger RNA （mRNA） and its signaling pathway. In this study, several cell lines including LLC-PK1 （a porcine kidney tubular epithelial cell line）, MDCK （Madin-Darby canine kidney） and CTLL-2 （cytotoxic T-lymphocyte line） were treated with recombinant human TGF-131, and a series of experiments were carried out, involving Northern blot analysis of total RNA from these cells. Further, several specific chemical inhibitors were applied before TGF-β1 treatment to probe the signaling pathway. The results showed that TGF-β1 can significadtly up-regulate SHARP-2 mRNA expression in the LLC-PK1 cell line. The peak level of induction was found 2 h after TGF-β1 stimulation. While one phospho- inositide 3-kinases （PI-3） kinase inhibitor, LY294002, completely blocked the effect of TGF-131 on SHARP-2 mRNA expression in LLC-PK1 cells at a low concentration, other inhibitors, including PD98059, staurosporine, AG490, wortmannin, okadaic acid and rapamycin, had no effect. The effect of LY294002 was dose-dependent. We conclude that, in LLC-PK1 cells at least, TGF-β1 can effectively induce the SHARP-2 mRNA expression and that the PI-3 kinase pathway can mediate this effect.
基金the Austrian Science Foundation (FWF) to M.T.，grants of the DFG to R.H.，T.D.M.，D.B.
文摘14-3-3 proteins play an important role in the regulation of many cellular processes. The Arabidopsis vacuolar two-pore K＋ channel 1 （TPK1） interacts with the 14-3-3 protein GRF6 （GF14-λ）. Upon phosphorylation of the putative binding motif in the N-terminus of TPK1, GRF6 binds to TPK1 and activates the potassium channel. In order to gain a deeper understanding of this 14-3-3-mediated signal transduction, we set out to identify the respective kinases, which regulate the phosphorylation status of the 14-3-3 binding motif in TPK1. Here, we report that the calcium-dependent protein kinases （CDPKs） can phosphorylate and thereby activate the 14-3-3 binding motif in TPK1. Focusing on the stress-activated kinase CPK3, we visualized direct and specific interaction of TPK1 with the kinase at the tonoplast in vivo. In line with its proposed role in K＋ homeostasis, TPK1 phosphorylation was found to be induced by salt stress in planta, and both cpk3 and tpkl mutants displayed salt-sensitive phenotypes. Molecular modeling of the TPK1-CPK3 interaction domain provided mechanistic insights into TPK1 stress-regulated phosphorylation responses and pinpointed two arginine residues in the N-terminal 14-3-3 binding motif in TPK1 critical for kinase interaction. Taken together, our studies provide evidence for an essential role of the vacuolar potassium channel TPK1 in salt-stress adaptation as a target of calcium-regulated stress signaling pathways involving Ca2＋, Ca2＋-dependent kinases, and 14-3-3 proteins.
文摘Background ^11C-4-N-（3-bromoanilino）-6,7-dimethoxyquinazoline （1^11C-PD153035） has been reported as a tracer for imaging human tumors that overexpress epidermal growth factor receptor （EGFR）. However it is still unclear whether ^11C-PD153035 uptake correlates with EGFR expression levels. The objective of this study was to investigate the relationship between ^11C-PD153035 accumulation and EGFR expression levels. Methods Synthesis of ^11C-PD153035 was performed in the Tracerlab FXc system. Accumulation of ^11C-PD153035 by MDA-MB-468, A549 and MDA-MB-231 cells was measured in vitro. There were six tumor-bearing mice in each group. ^11C-PD153035 uptake in tumors was determined by positron emission tomography/computed tomography （PET/CT）. Tumor/normal muscle tissue （TINT） analysis in PET images was applied to quantify the PET data. Sixty minutes after PET/CT scanning, the nude mice were sacrificed and the tumors were excised. The ^11C-PD153035 accumulation in different tumors was determined by a gamma counter. Results Close correlation existed between the uptake and the level of EGFR expression both in vitro and ex vivo （r^2=0.72, P〈0.001; r^2=0.63, P=0.003）. When the static TINT analysis method was applied to analyze the PET data, the observed correlation was again excellent （^2=0.70, P=-0.001）. Conclusions The uptake of PET tracer ^11C-PD153035 closely correlates with the EGFR expression levels in tumor cells. ^11C-PD153035 has the potential to yield useful information for both cancer diagnosis and therapy.
文摘BACKGROUND： Phototropism is the response a plant exhibits when it is faced with a directional blue light stimulus. Though a seemingly simple differential cell elongation response within a responding tissue that results in organ curvature, phototropism is regulated through a complex set of signal perception and transduction events that move from the plasma membrane to the nucleus. In nature phototropism is one of several plant responses that have evolved to optimize photosynthesis and growth. OBJECTIVE： In the present work we will review the state of the field with respect to the molecules and mechanisms associated with phototropism in land plants. METHODS： A systematic literature search was done to identify relevant advances in the field. Though we tried to focus on literature within the past decade （1998-present）, we have discussed and cited older literature where appropriate because of context or its significant impact to the present field. Several previous review articles are also cited where appropriate and readers should seek those out. RESULTS： A total of 199 articles are cited that fulfill the criteria listed above. CONCLUSIONS： Though important numerous and significant advances have been made in our understanding of the molecular, biochemical, cell biological and physiologic mechanisms underlying phototropism in land plants over the past decade, there are many remaining unanswered questions. The future is indeed bright for researchers in the field and we look forward to the next decade worth of discoveries.
