Background： Ovarian serous adenocarcinoma can be divided into low- and high-grade tumors, which exhibit substantial differences in pathogenesis, clinicopathology, and prognosis. This study aimed to investigate the d...Background： Ovarian serous adenocarcinoma can be divided into low- and high-grade tumors, which exhibit substantial differences in pathogenesis, clinicopathology, and prognosis. This study aimed to investigate the difl＇erences in the PH domain leucine-rich repeat protein phosphatase （PHLPP）, tbrkhead llomeobox type O3a （FoxO3a）, and RAD51 protein expressions, and their associations with prognosis in patients with low- and high-grade ovarian serous adenocarcinomas. Methods： The PH LPP, FoxO3a, and RA D51 protein expressions were examined in 94 high- and 26 low-grade ovarian serous adenocarcinomas by immunohistochemistry. The differences in expression and their relationships with pathological features and prognosis were analyzed. Results： In high-grade serous adenocarcinomas, the positive rates of PHLPP and goxO3a were 24.5% and 26.6%, while in low-grade tumors, they were 23.1% and 26.9%, respectively （P 〈 0.05 vs. the control specimens; low- vs. high-grade： P 〉 0.（15）. The positive rates of RAD51 were 70.2% and 65.4% in high- and low-grade serous adenocarcinomas, respectively （P 〈 0.（15 vs. the control specimens; low- vs. high-grade： P 〉 0.05）. Meanwhile, in high-grade tumors, Stage Ⅲ/Ⅳ tumors and lymph node and omental metastases were significantly associated with lower PHLPP and FoxO3a and higher RAD51 expression. The 5-year survival rates of patients with PHLPP- and FoxO3a-positive high-grade tumors （43.5% and 36.0%） were significantly higher than in patients with PHLPP-negative tumors （5.6% and 7.2%, respectively; P 〈 0.05）. Similarly, the 5-year survival rate of RAD5 l-positive patients （3.0%） was significantly lower than in negative patients （42.9%： P〈 0.05）. In low-grade tumors, the PHLPP, FoxO3a, and RAD51 expressions were not significantly correlated with lymph node metastasis, omental metastasis, Federation of Gynecology and Obstetrics stage, or prognosis. Conclusions： Abnormal PHLPP, FoxO3a, and RAD51 protein expressions may be involved in the development of high- and low-grade ovarian serous adenocarcinomas, suggesting conlmon molecular pathways. Decreased PH LPP and FoxO3a and increased RAD51 protein expression may be important molecular markers for poor prognosis, and RAD51 may be an independent prognosis factor, of high-grade, but not low-grade, ovarian serous adenocarcinomas.展开更多
NF-kappaB plays a critical role in cell survival,apoptosis,and inflammatory responses.Serine/threoninespecific phosphatases(PPs)represent the second major class of enzymes that catalyze the dephosphorylation of protei...NF-kappaB plays a critical role in cell survival,apoptosis,and inflammatory responses.Serine/threoninespecific phosphatases(PPs)represent the second major class of enzymes that catalyze the dephosphorylation of proteins.The roles of PPs regulating NF-kappaB activities are poorly understood.Here we describe an RNAi-based screen to identify the PPs that involve in regulating NFkappaB signaling.Thirty-four candidate PPs siRNAs were synthesized and primarily screened by NF-kappaB reporter gene assay in HeLa cells.PHLPP,one of the protein phosphatase type 2C family members(PP2C),was identified as a positive regulator of NF-kappaB signaling.Knock-down of PHLPP dramatically attenuated TNFα-stimulated NF-kappaB transcriptional activation.Knockdown of PHLPP led to enhancement of NF-kappaB/p65 nuclear import and retention,but decreased TNFα-induced phosphorylation at Ser276 on p65.This critical phosphorylation was also drastically reduced by knock-down of PKCalpha and Akt1,two important serine/threonine kinases dephosphorylated by PHLPP.The results together suggest that PHLPP-Akt-PKC may represent an important signaling loop that activates NF-kappaB/p65 signaling through critical serine phosphorylation.展开更多
Axonal growth inhibitors are released during traumatic injuries to the adult mammalian central nervous system, including after spinal cord injury. These molecules accumulate at the injury site and form a highly inhibi...Axonal growth inhibitors are released during traumatic injuries to the adult mammalian central nervous system, including after spinal cord injury. These molecules accumulate at the injury site and form a highly inhibitory environment for axonal regeneration. Among these inhibitory molecules, myelinassociated inhibitors, including neurite outgrowth inhibitor A, oligodendrocyte myelin glycoprotein, myelin-associated glycoprotein, chondroitin sulfate proteoglycans and repulsive guidance molecule A are of particular importance. Due to their inhibitory nature, they represent exciting molecular targets to study axonal inhibition and regeneration after central injuries. These molecules are mainly produced by neurons, oligodendrocytes, and astrocytes within the scar and in its immediate vicinity. They exert their effects by binding to specific receptors, localized in the membranes of neurons. Receptors for these inhibitory cues include Nogo receptor 1, leucine-rich repeat, and Ig domain containing 1 and p75 neurotrophin receptor/tumor necrosis factor receptor superfamily member 19(that form a receptor complex that binds all myelin-associated inhibitors), and also paired immunoglobulin-like receptor B. Chondroitin sulfate proteoglycans and repulsive guidance molecule A bind to Nogo receptor 1, Nogo receptor 3, receptor protein tyrosine phosphatase σ and leucocyte common antigen related phosphatase, and neogenin, respectively. Once activated, these receptors initiate downstream signaling pathways, the most common amongst them being the Rho A/ROCK signaling pathway. These signaling cascades result in actin depolymerization, neurite outgrowth inhibition, and failure to regenerate after spinal cord injury. Currently, there are no approved pharmacological treatments to overcome spinal cord injuries other than physical rehabilitation and management of the array of symptoms brought on by spinal cord injuries. However, several novel therapies aiming to modulate these inhibitory proteins and/or their receptors are under investigation in ongoing clinical trials. Investigation has also been demonstrating that combinatorial therapies of growth inhibitors with other therapies, such as growth factors or stem-cell therapies, produce stronger results and their potential application in the clinics opens new venues in spinal cord injury treatment.展开更多
目的探讨磷酸酶PHLPP1在人脐静脉内皮细胞(HUVEC)中的基础表达及转基因对其增殖的影响。方法体外培养的HUVEC分3组处理,分别为未转染组、转染pcDNA3-GFP组和转染pcDNA3HA-PHLPP组。通过构建pcDNA3HA-PHLPP1质粒并瞬时转染HUVEC。以细胞...目的探讨磷酸酶PHLPP1在人脐静脉内皮细胞(HUVEC)中的基础表达及转基因对其增殖的影响。方法体外培养的HUVEC分3组处理,分别为未转染组、转染pcDNA3-GFP组和转染pcDNA3HA-PHLPP组。通过构建pcDNA3HA-PHLPP1质粒并瞬时转染HUVEC。以细胞计数及噻唑盐比色法测定细胞增殖能力,Western blot ting定量磷酸酶PHLPP1蛋白表达水平。结果基础状态下HUVEC不表达PHLPP1。转染pcDNA3HA-PHLPP1组明显增加PHLPP1表达,与正常对照组、pcDNA3-GFP组比较差异显著(均P<0.01)。3组的细胞增殖指标无明显差异(P>0.05),其中MTT吸收度A值分别是0.134±0.015,0.133±0.014,0.137±0.016,细胞计数为(8.293±0.962)×105,(7.937±0.101)×105,8.127±0.112)×105。结论 PHLPP可能不是调节HUVEC增殖的最重要信号蛋白。展开更多
Obstructive sleep apnea can worsen the prognosis of subarachnoid hemorrhage.Howeve r,the underlying mechanism remains unclear.In this study,we established a mouse model of subarachnoid hemorrhage using the endovascula...Obstructive sleep apnea can worsen the prognosis of subarachnoid hemorrhage.Howeve r,the underlying mechanism remains unclear.In this study,we established a mouse model of subarachnoid hemorrhage using the endovascular perforation method and exposed the mice to intermittent hypoxia for 8 hours daily for 2 consecutive days to simulate sleep apnea.We found that sleep apnea aggravated brain edema,increased hippocampal neuron apoptosis,and worsened neurological function in this mouse model of subarachnoid hemorrhage.Then,we established an in vitro HT-22 cell model of hemin-induced subarachnoid hemorrhage/intermittent hypoxia and found that the cells died,and lactate dehydrogenase release increased,after 48 hours.We further investigated the underlying mechanism and found that sleep apnea increased the expression of hippocampal neuroinflammatory factors interleukin-1β,interleukin-18,inte rleukin-6,nuclear factorκB,pyro ptosis-related protein caspase-1,pro-caspase-1,and NLRP3,promoted the prolife ration of astrocytes,and increased the expression of hypoxia-inducible factor 1αand apoptosis-associated speck-like protein containing a CARD,which are the key proteins in the hypoxia-inducible factor 1α/apoptosis-associated speck-like protein containing a CARD signaling pathway.We also found that knockdown of hypoxia-inducible factor 1αexpression in vitro greatly reduced the damage to HY22 cells.These findings suggest that sleep apnea aggravates early brain injury after subarachnoid hemorrhage by aggravating neuroinflammation and pyroptosis,at least in part through the hypoxia-inducible factor 1α/apoptosis-associated speck-like protein containing a CARD signaling pathway.展开更多
Leucine-rich repeat （LRR） receptor-like kinases （RLKs）, evolutionarily related LRR receptor-like proteins （RLPs） and receptor-like cytoplasmic kinases （RLCKs） have important roles in plant signaling, and their...Leucine-rich repeat （LRR） receptor-like kinases （RLKs）, evolutionarily related LRR receptor-like proteins （RLPs） and receptor-like cytoplasmic kinases （RLCKs） have important roles in plant signaling, and their gene subfamilies are large with a complicated history of gene duplication and loss. In three pairs of closely related lineages, including Arabidopsis thaliana and A. lyrata （Arabidopsis）, Lotus japonicus, and Medicago truncatula （Legumes）, Oryza sativa ssp. japonica, and O. sativa ssp. indica （Rice）, we find that LRR RLKs comprise the largest group of these LRR-related subfamilies, while the related RLCKs represent the smal est group. In addition, comparison of orthologs indicates a high frequency of reciprocal gene loss of the LRR RLK/LRR RLP/RLCK subfamilies. Furthermore, pairwise comparisons show that reciprocal gene loss is often associated with lineage-specific duplication（s） in the alternative lineage. Last, analysis of genes in A. thaliana involved in development revealed that most are highly conserved orthologs without species-specific duplication in the two Arabidopsis species and originated from older Arabidopsis-specific or rosid-specific duplications. We discuss＆amp;nbsp;potential pitfal s related to functional prediction for genes that have undergone frequent turnover （duplications, losses, and domain architecture changes）, and conclude that prediction based on phylogenetic relationships wil likely outperform that based on sequence similarity alone.展开更多
miR-190-5p是小分子非编码RNA,参与调控肺腺癌细胞的增殖和转移。对miR-190-5p调控肺腺癌细胞的增殖和转移的机制深入研究会帮助了解到肺腺癌的发生机制和探究潜在的药物干预靶点。本文研究了miR-190-5p对肺腺癌A549细胞的增殖和转移的...miR-190-5p是小分子非编码RNA,参与调控肺腺癌细胞的增殖和转移。对miR-190-5p调控肺腺癌细胞的增殖和转移的机制深入研究会帮助了解到肺腺癌的发生机制和探究潜在的药物干预靶点。本文研究了miR-190-5p对肺腺癌A549细胞的增殖和转移的影响,解释了miR-190-5p促进肺腺癌生长的机制;通过感染miR-190-5p的慢病毒表达系统,克隆了稳定过表达miR-190-5p的肺腺癌A549细胞系;通过生物信息学方法和荧光素酶报告实验确认了miR-190-5p的靶基因。结果发现,稳定过度表达的miR-190-5p能促进A549细胞的增殖和转移;确定了miR-190-5p的靶点是PH域富含亮氨酸重复的蛋白磷酸酶1(Leucine-rich repeat-rich protein phosphatase 1 in the PH domain,PHLPP1),miR-190-5p降低PHLPP1蛋白水平;稳定表达的miR-190-5p增加了A549细胞中E盒结合锌指蛋白1(E box-binding zinc finger protein1,TCF8/ZEB1)、锌指转录因子(Zinc finger transcription factor,Snail)、具有凝血酶原基序的解体蛋白和金属蛋白酶1(a disintegrin and metalloproteinase with thrombospondin motifs 1,ADAMTS1)的表达和蛋白激酶B(protein kinase B,AKT)在Ser473的磷酸化,而过表达PHLPP1削弱了miR-190-5p的功能,降低了miR-190-5p促进A549细胞的增殖、迁移和侵袭能力;miR-190-5p通过抑制PHLPP1的表达而增强了A549细胞的增殖和转移。这表明,miR-190-5p可能可以作为肺腺癌治疗的潜在靶点。展开更多
文摘Background： Ovarian serous adenocarcinoma can be divided into low- and high-grade tumors, which exhibit substantial differences in pathogenesis, clinicopathology, and prognosis. This study aimed to investigate the difl＇erences in the PH domain leucine-rich repeat protein phosphatase （PHLPP）, tbrkhead llomeobox type O3a （FoxO3a）, and RAD51 protein expressions, and their associations with prognosis in patients with low- and high-grade ovarian serous adenocarcinomas. Methods： The PH LPP, FoxO3a, and RA D51 protein expressions were examined in 94 high- and 26 low-grade ovarian serous adenocarcinomas by immunohistochemistry. The differences in expression and their relationships with pathological features and prognosis were analyzed. Results： In high-grade serous adenocarcinomas, the positive rates of PHLPP and goxO3a were 24.5% and 26.6%, while in low-grade tumors, they were 23.1% and 26.9%, respectively （P 〈 0.05 vs. the control specimens; low- vs. high-grade： P 〉 0.（15）. The positive rates of RAD51 were 70.2% and 65.4% in high- and low-grade serous adenocarcinomas, respectively （P 〈 0.（15 vs. the control specimens; low- vs. high-grade： P 〉 0.05）. Meanwhile, in high-grade tumors, Stage Ⅲ/Ⅳ tumors and lymph node and omental metastases were significantly associated with lower PHLPP and FoxO3a and higher RAD51 expression. The 5-year survival rates of patients with PHLPP- and FoxO3a-positive high-grade tumors （43.5% and 36.0%） were significantly higher than in patients with PHLPP-negative tumors （5.6% and 7.2%, respectively; P 〈 0.05）. Similarly, the 5-year survival rate of RAD5 l-positive patients （3.0%） was significantly lower than in negative patients （42.9%： P〈 0.05）. In low-grade tumors, the PHLPP, FoxO3a, and RAD51 expressions were not significantly correlated with lymph node metastasis, omental metastasis, Federation of Gynecology and Obstetrics stage, or prognosis. Conclusions： Abnormal PHLPP, FoxO3a, and RAD51 protein expressions may be involved in the development of high- and low-grade ovarian serous adenocarcinomas, suggesting conlmon molecular pathways. Decreased PH LPP and FoxO3a and increased RAD51 protein expression may be important molecular markers for poor prognosis, and RAD51 may be an independent prognosis factor, of high-grade, but not low-grade, ovarian serous adenocarcinomas.
基金This research was supported by the National High Technology Research and Development Program of China(863 Program)(No.2006AA02Z191),the Bureau of Science and Technology of Guangzhou,China(No.2007Z1-E4041)Guangzhou Economic&Technological Development District(GETDD S&T Project)(2007G-P029).
文摘NF-kappaB plays a critical role in cell survival,apoptosis,and inflammatory responses.Serine/threoninespecific phosphatases(PPs)represent the second major class of enzymes that catalyze the dephosphorylation of proteins.The roles of PPs regulating NF-kappaB activities are poorly understood.Here we describe an RNAi-based screen to identify the PPs that involve in regulating NFkappaB signaling.Thirty-four candidate PPs siRNAs were synthesized and primarily screened by NF-kappaB reporter gene assay in HeLa cells.PHLPP,one of the protein phosphatase type 2C family members(PP2C),was identified as a positive regulator of NF-kappaB signaling.Knock-down of PHLPP dramatically attenuated TNFα-stimulated NF-kappaB transcriptional activation.Knockdown of PHLPP led to enhancement of NF-kappaB/p65 nuclear import and retention,but decreased TNFα-induced phosphorylation at Ser276 on p65.This critical phosphorylation was also drastically reduced by knock-down of PKCalpha and Akt1,two important serine/threonine kinases dephosphorylated by PHLPP.The results together suggest that PHLPP-Akt-PKC may represent an important signaling loop that activates NF-kappaB/p65 signaling through critical serine phosphorylation.
基金a Ph D fellowship by FCT-Fundacao para a Ciência Tecnologia (SFRH/BD/135868/2018)(to SSC)。
文摘Axonal growth inhibitors are released during traumatic injuries to the adult mammalian central nervous system, including after spinal cord injury. These molecules accumulate at the injury site and form a highly inhibitory environment for axonal regeneration. Among these inhibitory molecules, myelinassociated inhibitors, including neurite outgrowth inhibitor A, oligodendrocyte myelin glycoprotein, myelin-associated glycoprotein, chondroitin sulfate proteoglycans and repulsive guidance molecule A are of particular importance. Due to their inhibitory nature, they represent exciting molecular targets to study axonal inhibition and regeneration after central injuries. These molecules are mainly produced by neurons, oligodendrocytes, and astrocytes within the scar and in its immediate vicinity. They exert their effects by binding to specific receptors, localized in the membranes of neurons. Receptors for these inhibitory cues include Nogo receptor 1, leucine-rich repeat, and Ig domain containing 1 and p75 neurotrophin receptor/tumor necrosis factor receptor superfamily member 19(that form a receptor complex that binds all myelin-associated inhibitors), and also paired immunoglobulin-like receptor B. Chondroitin sulfate proteoglycans and repulsive guidance molecule A bind to Nogo receptor 1, Nogo receptor 3, receptor protein tyrosine phosphatase σ and leucocyte common antigen related phosphatase, and neogenin, respectively. Once activated, these receptors initiate downstream signaling pathways, the most common amongst them being the Rho A/ROCK signaling pathway. These signaling cascades result in actin depolymerization, neurite outgrowth inhibition, and failure to regenerate after spinal cord injury. Currently, there are no approved pharmacological treatments to overcome spinal cord injuries other than physical rehabilitation and management of the array of symptoms brought on by spinal cord injuries. However, several novel therapies aiming to modulate these inhibitory proteins and/or their receptors are under investigation in ongoing clinical trials. Investigation has also been demonstrating that combinatorial therapies of growth inhibitors with other therapies, such as growth factors or stem-cell therapies, produce stronger results and their potential application in the clinics opens new venues in spinal cord injury treatment.
文摘目的探讨磷酸酶PHLPP1在人脐静脉内皮细胞(HUVEC)中的基础表达及转基因对其增殖的影响。方法体外培养的HUVEC分3组处理,分别为未转染组、转染pcDNA3-GFP组和转染pcDNA3HA-PHLPP组。通过构建pcDNA3HA-PHLPP1质粒并瞬时转染HUVEC。以细胞计数及噻唑盐比色法测定细胞增殖能力,Western blot ting定量磷酸酶PHLPP1蛋白表达水平。结果基础状态下HUVEC不表达PHLPP1。转染pcDNA3HA-PHLPP1组明显增加PHLPP1表达,与正常对照组、pcDNA3-GFP组比较差异显著(均P<0.01)。3组的细胞增殖指标无明显差异(P>0.05),其中MTT吸收度A值分别是0.134±0.015,0.133±0.014,0.137±0.016,细胞计数为(8.293±0.962)×105,(7.937±0.101)×105,8.127±0.112)×105。结论 PHLPP可能不是调节HUVEC增殖的最重要信号蛋白。
基金the Natural Science Foundation of Jiangsu Province(Youth Program),No.BK20190129National Scientific Program of Jiangsu Colleges and Universities of China,No.19KJB320012(both to LY)。
文摘Obstructive sleep apnea can worsen the prognosis of subarachnoid hemorrhage.Howeve r,the underlying mechanism remains unclear.In this study,we established a mouse model of subarachnoid hemorrhage using the endovascular perforation method and exposed the mice to intermittent hypoxia for 8 hours daily for 2 consecutive days to simulate sleep apnea.We found that sleep apnea aggravated brain edema,increased hippocampal neuron apoptosis,and worsened neurological function in this mouse model of subarachnoid hemorrhage.Then,we established an in vitro HT-22 cell model of hemin-induced subarachnoid hemorrhage/intermittent hypoxia and found that the cells died,and lactate dehydrogenase release increased,after 48 hours.We further investigated the underlying mechanism and found that sleep apnea increased the expression of hippocampal neuroinflammatory factors interleukin-1β,interleukin-18,inte rleukin-6,nuclear factorκB,pyro ptosis-related protein caspase-1,pro-caspase-1,and NLRP3,promoted the prolife ration of astrocytes,and increased the expression of hypoxia-inducible factor 1αand apoptosis-associated speck-like protein containing a CARD,which are the key proteins in the hypoxia-inducible factor 1α/apoptosis-associated speck-like protein containing a CARD signaling pathway.We also found that knockdown of hypoxia-inducible factor 1αexpression in vitro greatly reduced the damage to HY22 cells.These findings suggest that sleep apnea aggravates early brain injury after subarachnoid hemorrhage by aggravating neuroinflammation and pyroptosis,at least in part through the hypoxia-inducible factor 1α/apoptosis-associated speck-like protein containing a CARD signaling pathway.
基金supported by the United States National Science Foundation(DEB 1026200 to M.J.S.and IOS 1257316 to F.E.T.)
文摘Leucine-rich repeat （LRR） receptor-like kinases （RLKs）, evolutionarily related LRR receptor-like proteins （RLPs） and receptor-like cytoplasmic kinases （RLCKs） have important roles in plant signaling, and their gene subfamilies are large with a complicated history of gene duplication and loss. In three pairs of closely related lineages, including Arabidopsis thaliana and A. lyrata （Arabidopsis）, Lotus japonicus, and Medicago truncatula （Legumes）, Oryza sativa ssp. japonica, and O. sativa ssp. indica （Rice）, we find that LRR RLKs comprise the largest group of these LRR-related subfamilies, while the related RLCKs represent the smal est group. In addition, comparison of orthologs indicates a high frequency of reciprocal gene loss of the LRR RLK/LRR RLP/RLCK subfamilies. Furthermore, pairwise comparisons show that reciprocal gene loss is often associated with lineage-specific duplication（s） in the alternative lineage. Last, analysis of genes in A. thaliana involved in development revealed that most are highly conserved orthologs without species-specific duplication in the two Arabidopsis species and originated from older Arabidopsis-specific or rosid-specific duplications. We discuss＆amp;nbsp;potential pitfal s related to functional prediction for genes that have undergone frequent turnover （duplications, losses, and domain architecture changes）, and conclude that prediction based on phylogenetic relationships wil likely outperform that based on sequence similarity alone.
文摘miR-190-5p是小分子非编码RNA,参与调控肺腺癌细胞的增殖和转移。对miR-190-5p调控肺腺癌细胞的增殖和转移的机制深入研究会帮助了解到肺腺癌的发生机制和探究潜在的药物干预靶点。本文研究了miR-190-5p对肺腺癌A549细胞的增殖和转移的影响,解释了miR-190-5p促进肺腺癌生长的机制;通过感染miR-190-5p的慢病毒表达系统,克隆了稳定过表达miR-190-5p的肺腺癌A549细胞系;通过生物信息学方法和荧光素酶报告实验确认了miR-190-5p的靶基因。结果发现,稳定过度表达的miR-190-5p能促进A549细胞的增殖和转移;确定了miR-190-5p的靶点是PH域富含亮氨酸重复的蛋白磷酸酶1(Leucine-rich repeat-rich protein phosphatase 1 in the PH domain,PHLPP1),miR-190-5p降低PHLPP1蛋白水平;稳定表达的miR-190-5p增加了A549细胞中E盒结合锌指蛋白1(E box-binding zinc finger protein1,TCF8/ZEB1)、锌指转录因子(Zinc finger transcription factor,Snail)、具有凝血酶原基序的解体蛋白和金属蛋白酶1(a disintegrin and metalloproteinase with thrombospondin motifs 1,ADAMTS1)的表达和蛋白激酶B(protein kinase B,AKT)在Ser473的磷酸化,而过表达PHLPP1削弱了miR-190-5p的功能,降低了miR-190-5p促进A549细胞的增殖、迁移和侵袭能力;miR-190-5p通过抑制PHLPP1的表达而增强了A549细胞的增殖和转移。这表明,miR-190-5p可能可以作为肺腺癌治疗的潜在靶点。
