Objective： Based on a partialsubtilisin-like protease, Prl genomic sequence ofPythium guiyangense which has been cloned before, Panhandle PCR strategy was used to amplify the upstream flanking sequence adjacent to th...Objective： Based on a partialsubtilisin-like protease, Prl genomic sequence ofPythium guiyangense which has been cloned before, Panhandle PCR strategy was used to amplify the upstream flanking sequence adjacent to the known sequence of the Prl gene. Methods： The genomic DNA was firstly digested with BamH I and then treated with calf intestinal alkaline phosphatase（CIAP）. Next, a 5＇ phosphorylated oligonucleotide was ligated to the 5＇ ends of BamH I -digested DNA. After denaturation, intmstrand annealing and polymemse extension, a pan with a handle was formed,and lastly the nested PCR was performed. Results： A 864 bp product was amplified,which was adjacent to the known sequence of Prl gene.The gene has been accessed by GenBank （Accession：JQ975036）. Conclusion： Panhandle PCR is a quick and convenient approach for amplifying and identifying unknown partner genes,which facilitates cloning full-length Prl gene展开更多
The regenerative capacity of peripheral nerves is limited after nerve injury.A number of growth factors modulate many cellular behaviors,such as proliferation and migration,and may contribute to nerve repair and regen...The regenerative capacity of peripheral nerves is limited after nerve injury.A number of growth factors modulate many cellular behaviors,such as proliferation and migration,and may contribute to nerve repair and regeneration.Our previous study observed the dynamic changes of genes in L4–6 dorsal root ganglion after rat sciatic nerve crush using transcriptome sequencing.Our current study focused on upstream growth factors and found that a total of 19 upstream growth factors were dysregulated in dorsal root ganglions at 3,9 hours,1,4,or 7 days after nerve crush,compared with the 0 hour control.Thirty-six rat models of sciatic nerve crush injury were prepared as described previously.Then,they were divided into six groups to measure the expression changes of representative genes at 0,3,9 hours,1,4 or 7 days post crush.Our current study measured the expression levels of representative upstream growth factors,including nerve growth factor,brain-derived neurotrophic factor,fibroblast growth factor 2 and amphiregulin genes,and explored critical signaling pathways and biological process through bioinformatic analysis.Our data revealed that many of these dysregulated upstream growth factors,including nerve growth factor,brain-derived neurotrophic factor,fibroblast growth factor 2 and amphiregulin,participated in tissue remodeling and axon growth-related biological processes Therefore,the experiment described the expression pattern of upstream growth factors in the dorsal root ganglia after peripheral nerve injury.Bioinformatic analysis revealed growth factors that may promote repair and regeneration of damaged peripheral nerves.All animal surgery procedures were performed in accordance with Institutional Animal Care Guidelines of Nantong University and ethically approved by the Administration Committee of Experimental Animals,China(approval No.20170302-017)on March 2,2017.展开更多
Age-related macular degeneration(AMD)is a complicated disease that causes irreversible visual impairment.Increasing evidences pointed retinal pigment epithelia(RPE)cells as the decisive cell involved in the progress o...Age-related macular degeneration(AMD)is a complicated disease that causes irreversible visual impairment.Increasing evidences pointed retinal pigment epithelia(RPE)cells as the decisive cell involved in the progress of AMD,and the function of anti-oxidant capacity of PRE plays a fundamental physiological role.Nuclear factor erythroid 2 related factor 2(Nrf2)is a significant transcription factor in the cellular anti-oxidant system as it regulates the expression of multiple anti-oxidative genes.Its functions of protecting RPE cells against oxidative stress(OS)and ensuing physiological changes,including inflammation,mitochondrial damage and autophagy dysregulation,have already been elucidated.Understanding the roles of upstream regulators of Nrf2 could provide further insight to the OS-mediated AMD pathogenesis.For the first time,this review summarized the reported upstream regulators of Nrf2 in AMD pathogenesis,including proteins and miRNAs,and their underlying molecular mechanisms,which may help to find potential targets via regulating the Nrf2 pathway in the future research and further discuss the existing Nrf2 regulators proved to be beneficial in preventing AMD.展开更多
Hepatocellular carcinoma(HCC)is an aggressive malignancy that is often refractory to chemotherapy and immune checkpoint inhibitors.This therapeutic resistance is driven in part by the persistence of cancer stem-like c...Hepatocellular carcinoma(HCC)is an aggressive malignancy that is often refractory to chemotherapy and immune checkpoint inhibitors.This therapeutic resistance is driven in part by the persistence of cancer stem-like cells(CSCs)and the development of an immune-cold tumor microenvironment.However,the upstream regulators that coordinate these malignant features remain poorly defined.In this study,we identified dysadherin as a novel upstream activator of YAP that promotes both CSC plasticity and immune evasion through the FAK/YAP/TEAD2 signaling axis.Using single-cell transcriptomic analysis,in vitro assays,and multiple in vivo models including a humanized immune mouse system,we showed that dysadherin enhances the expression of pluripotency genes,such as OCT4 and upregulates PD-L1.These changes support stem-like tumor behavior and contribute to T-cell exclusion,fostering an immunosuppressive niche.Notably,genetic knockdown or peptide-based pharmacologic inhibition of dysadherin effectively restored antitumor immune activation,suppressed metastasis and improved therapeutic responsiveness.Our findings reveal a mechanistic link between dysadherin-mediated cell adhesion signaling and the transcriptional regulation of both stemness and immune escape.Collectively,these findings establish the dysadherin/YAP axis as a key driver of HCC progression and resistance,and highlight it as a compelling therapeutic target that could overcome treatment failure in advanced liver cancer.展开更多
基金Supported by the Guangxi Department of education scientific research funds,China(No.200103YB154)
文摘Objective： Based on a partialsubtilisin-like protease, Prl genomic sequence ofPythium guiyangense which has been cloned before, Panhandle PCR strategy was used to amplify the upstream flanking sequence adjacent to the known sequence of the Prl gene. Methods： The genomic DNA was firstly digested with BamH I and then treated with calf intestinal alkaline phosphatase（CIAP）. Next, a 5＇ phosphorylated oligonucleotide was ligated to the 5＇ ends of BamH I -digested DNA. After denaturation, intmstrand annealing and polymemse extension, a pan with a handle was formed,and lastly the nested PCR was performed. Results： A 864 bp product was amplified,which was adjacent to the known sequence of Prl gene.The gene has been accessed by GenBank （Accession：JQ975036）. Conclusion： Panhandle PCR is a quick and convenient approach for amplifying and identifying unknown partner genes,which facilitates cloning full-length Prl gene
基金supported by the Natural Science Foundation of Jiangsu Higher Education Institutions of China(Major Program),No.16KJA310005(to SYL)the Natural Science Foundation of Nantong City of China,No.JC2018058(to TMQ)the Priority Academic Program Development of Jiangsu Higher Education Institutions of China
文摘The regenerative capacity of peripheral nerves is limited after nerve injury.A number of growth factors modulate many cellular behaviors,such as proliferation and migration,and may contribute to nerve repair and regeneration.Our previous study observed the dynamic changes of genes in L4–6 dorsal root ganglion after rat sciatic nerve crush using transcriptome sequencing.Our current study focused on upstream growth factors and found that a total of 19 upstream growth factors were dysregulated in dorsal root ganglions at 3,9 hours,1,4,or 7 days after nerve crush,compared with the 0 hour control.Thirty-six rat models of sciatic nerve crush injury were prepared as described previously.Then,they were divided into six groups to measure the expression changes of representative genes at 0,3,9 hours,1,4 or 7 days post crush.Our current study measured the expression levels of representative upstream growth factors,including nerve growth factor,brain-derived neurotrophic factor,fibroblast growth factor 2 and amphiregulin genes,and explored critical signaling pathways and biological process through bioinformatic analysis.Our data revealed that many of these dysregulated upstream growth factors,including nerve growth factor,brain-derived neurotrophic factor,fibroblast growth factor 2 and amphiregulin,participated in tissue remodeling and axon growth-related biological processes Therefore,the experiment described the expression pattern of upstream growth factors in the dorsal root ganglia after peripheral nerve injury.Bioinformatic analysis revealed growth factors that may promote repair and regeneration of damaged peripheral nerves.All animal surgery procedures were performed in accordance with Institutional Animal Care Guidelines of Nantong University and ethically approved by the Administration Committee of Experimental Animals,China(approval No.20170302-017)on March 2,2017.
基金Supported by Capital Medical University Scientific Research Grant for Undergraduate Students(No.XSKY2023026).
文摘Age-related macular degeneration(AMD)is a complicated disease that causes irreversible visual impairment.Increasing evidences pointed retinal pigment epithelia(RPE)cells as the decisive cell involved in the progress of AMD,and the function of anti-oxidant capacity of PRE plays a fundamental physiological role.Nuclear factor erythroid 2 related factor 2(Nrf2)is a significant transcription factor in the cellular anti-oxidant system as it regulates the expression of multiple anti-oxidative genes.Its functions of protecting RPE cells against oxidative stress(OS)and ensuing physiological changes,including inflammation,mitochondrial damage and autophagy dysregulation,have already been elucidated.Understanding the roles of upstream regulators of Nrf2 could provide further insight to the OS-mediated AMD pathogenesis.For the first time,this review summarized the reported upstream regulators of Nrf2 in AMD pathogenesis,including proteins and miRNAs,and their underlying molecular mechanisms,which may help to find potential targets via regulating the Nrf2 pathway in the future research and further discuss the existing Nrf2 regulators proved to be beneficial in preventing AMD.
基金supported by the National Research Foundation of Korea(NRF)grants funded by the Korean government(MSIT)(RS-2023-NR077237,RS-2024-00411137,RS-2024-00441157,and RS-2023-00223591)by a CNUH-GIST Research Collaboration Grant,funded by the Gwangju Institute of Science and Technology(GIST)in 2025The biospecimens and data used for this study were provided by the Biobank of Chonnam National University Hwasun Hospital,a member of the Korea Biobank Network。
文摘Hepatocellular carcinoma(HCC)is an aggressive malignancy that is often refractory to chemotherapy and immune checkpoint inhibitors.This therapeutic resistance is driven in part by the persistence of cancer stem-like cells(CSCs)and the development of an immune-cold tumor microenvironment.However,the upstream regulators that coordinate these malignant features remain poorly defined.In this study,we identified dysadherin as a novel upstream activator of YAP that promotes both CSC plasticity and immune evasion through the FAK/YAP/TEAD2 signaling axis.Using single-cell transcriptomic analysis,in vitro assays,and multiple in vivo models including a humanized immune mouse system,we showed that dysadherin enhances the expression of pluripotency genes,such as OCT4 and upregulates PD-L1.These changes support stem-like tumor behavior and contribute to T-cell exclusion,fostering an immunosuppressive niche.Notably,genetic knockdown or peptide-based pharmacologic inhibition of dysadherin effectively restored antitumor immune activation,suppressed metastasis and improved therapeutic responsiveness.Our findings reveal a mechanistic link between dysadherin-mediated cell adhesion signaling and the transcriptional regulation of both stemness and immune escape.Collectively,these findings establish the dysadherin/YAP axis as a key driver of HCC progression and resistance,and highlight it as a compelling therapeutic target that could overcome treatment failure in advanced liver cancer.
