为了获得高纯度的重组人长效促卵泡激素FSH-CTP,收集表达FSH-CTP的CHO细胞培养上清,经过染料亲和层析、DEAE阴离子交换层析以及疏水层析三步层析制备,所得样品进行分子量、等电点测定和Western Blot鉴定,雌性大鼠卵巢增重法测定体内生...为了获得高纯度的重组人长效促卵泡激素FSH-CTP,收集表达FSH-CTP的CHO细胞培养上清,经过染料亲和层析、DEAE阴离子交换层析以及疏水层析三步层析制备,所得样品进行分子量、等电点测定和Western Blot鉴定,雌性大鼠卵巢增重法测定体内生物活性。结果显示：经3步纯化所得样品纯度可达98%以上,表观分子量约为47 k Da,MALDI-TOF-MS测得结果为35.2 k Da,等电点4.0-2.8,Western Blot结果呈阳性,测活结果表明,FSH-CTP的活性比重组人卵泡刺激素明显偏高。纯化所得样品纯度良好,为该药物的进一步研究打下了基础。展开更多
Animal populations,with a known history of introduction events,provide opportunities to study the dynamics of how rapid shi s in ecological context a ect behavioral(e.g.,responses to brood parasitism) and life-history...Animal populations,with a known history of introduction events,provide opportunities to study the dynamics of how rapid shi s in ecological context a ect behavioral(e.g.,responses to brood parasitism) and life-history(e.g.,clutch and egg parameters) traits.We studied the European Green nch(Carduelis chloris) introduced to New Zealand,regarding foreign-egg rejection behaviors and also compared their clutch characteristics with data from the source populations in the United Kingdom.Although previously this species had been considered an unsuitable host for the Common Cuckoo(Cuculus canorus),and not impacted by selection pressure associated with brood parasitism,we found that Green nches in our study population were able to eject experimental eggs at low frequencies.In contrast,nest desertion rates were similar in experimentally parasitized and control unmanipulated nests,implying that nest desertion is not an antiparasite adaptation in this species.Contrary to previous studies,we did not nd signi cant di erences in clutch and egg sizes between introduced and source populations.is study emphasizes(1) the importance of using control treatments in studies of host responses to experimental parasitism,(2) including apparently unsuitable hosts of brood parasites,and(3) meta-replicating prior studies to further the process of gaining and validating scienti c knowledge.展开更多
To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukem...To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukemic cells, the cDNA encoding the signal and extracellular domains of murine CD80 was generated by PCR amplification from plasmid pcDNMB7 containing the full length cDNA of murine CD80 and those of murine IgG1, in which the Fc fragment was obtained through RT-PCR amplification from murine spleen cells. These two cDNAs were then cloned in tandem into eukaryotic expression vector pcDNA3.0 and the resultant recombinant plasmid pcDNA/CD80-IgG was then transfected to Chinese hamster ovary cells with liposome transfection reagent. The cell clones constitutively expressing CD80-IgG fusion protein were obtained by G418 screening. Western blotting and dot ELISA assay were used to detect the expression of the fusion protein in the supernatants of these cells. Meanwhile, the fusion protein expressed was then purified with affinity chromatography, and its biological activity was demonstrated by flow cytometry, MTr colorimetry and ELISA assay. The experimental resuits showed that these two inserts were successfully cloned into plasmid pcDNA3.0, and the highly purified fusion protein was obtained. This fusion protein was proved to be able to upregulate the density of CD80 on leukemic cells, deliberately promote the proliferative reactions of mouse allogenic lymphocytes and increase the killing activity against WEHI-3 cells from 49.7 % up to 84.6 %. In addition, this fusion protein could also enhance the IL-2 secretion from allogenic lymphocytes activated by tumorspecific antigens. It is concluded that the recombinant vector constructed can be functionally expressed in the mammalian cells, thus providing a solid foundation for the further investigation on the mechanism to eliminate the immune escape of leukemic cells in vivo.展开更多
Understanding the cell-of-origin of ovarian high grade serous cancer(HGSC)is the prerequisite for efficient prevention and early diagnosis of this most lethal gynecological cancer.Recently,a mesenchymal type of ovaria...Understanding the cell-of-origin of ovarian high grade serous cancer(HGSC)is the prerequisite for efficient prevention and early diagnosis of this most lethal gynecological cancer.Recently,a mesenchymal type of ovarian HGSC with the poorest prognosis among ovarian cancers was identified by both TCGA and AOCS studies.The cell-of-origin of this subtype of ovarian cancer is unknown.While pursuing studies to understand the role of the Hippo pathway in ovarian granulosa cell physiology and pathology,we unexpectedly found that the Yes-associated protein 1(YAP1),the major effector of the Hippo signaling pathway,induced dedifferentiation and reprogramming of the ovarian granulosa cells,a unique type of ovarian follicular cells with mesenchymal lineage and high plasticity,leading to the development of high grade ovarian cancer with serous features.Our research results unveil a potential cell-of-origin for a subtype of HGSC with mesenchymal features.展开更多
文摘为了获得高纯度的重组人长效促卵泡激素FSH-CTP,收集表达FSH-CTP的CHO细胞培养上清,经过染料亲和层析、DEAE阴离子交换层析以及疏水层析三步层析制备,所得样品进行分子量、等电点测定和Western Blot鉴定,雌性大鼠卵巢增重法测定体内生物活性。结果显示：经3步纯化所得样品纯度可达98%以上,表观分子量约为47 k Da,MALDI-TOF-MS测得结果为35.2 k Da,等电点4.0-2.8,Western Blot结果呈阳性,测活结果表明,FSH-CTP的活性比重组人卵泡刺激素明显偏高。纯化所得样品纯度良好,为该药物的进一步研究打下了基础。
基金supported by Human Frontier Science Program awards (RGY69/07 to TG,PC,and MEH and RGY86/12 to TG and MEH)
文摘Animal populations,with a known history of introduction events,provide opportunities to study the dynamics of how rapid shi s in ecological context a ect behavioral(e.g.,responses to brood parasitism) and life-history(e.g.,clutch and egg parameters) traits.We studied the European Green nch(Carduelis chloris) introduced to New Zealand,regarding foreign-egg rejection behaviors and also compared their clutch characteristics with data from the source populations in the United Kingdom.Although previously this species had been considered an unsuitable host for the Common Cuckoo(Cuculus canorus),and not impacted by selection pressure associated with brood parasitism,we found that Green nches in our study population were able to eject experimental eggs at low frequencies.In contrast,nest desertion rates were similar in experimentally parasitized and control unmanipulated nests,implying that nest desertion is not an antiparasite adaptation in this species.Contrary to previous studies,we did not nd signi cant di erences in clutch and egg sizes between introduced and source populations.is study emphasizes(1) the importance of using control treatments in studies of host responses to experimental parasitism,(2) including apparently unsuitable hosts of brood parasites,and(3) meta-replicating prior studies to further the process of gaining and validating scienti c knowledge.
文摘To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukemic cells, the cDNA encoding the signal and extracellular domains of murine CD80 was generated by PCR amplification from plasmid pcDNMB7 containing the full length cDNA of murine CD80 and those of murine IgG1, in which the Fc fragment was obtained through RT-PCR amplification from murine spleen cells. These two cDNAs were then cloned in tandem into eukaryotic expression vector pcDNA3.0 and the resultant recombinant plasmid pcDNA/CD80-IgG was then transfected to Chinese hamster ovary cells with liposome transfection reagent. The cell clones constitutively expressing CD80-IgG fusion protein were obtained by G418 screening. Western blotting and dot ELISA assay were used to detect the expression of the fusion protein in the supernatants of these cells. Meanwhile, the fusion protein expressed was then purified with affinity chromatography, and its biological activity was demonstrated by flow cytometry, MTr colorimetry and ELISA assay. The experimental resuits showed that these two inserts were successfully cloned into plasmid pcDNA3.0, and the highly purified fusion protein was obtained. This fusion protein was proved to be able to upregulate the density of CD80 on leukemic cells, deliberately promote the proliferative reactions of mouse allogenic lymphocytes and increase the killing activity against WEHI-3 cells from 49.7 % up to 84.6 %. In addition, this fusion protein could also enhance the IL-2 secretion from allogenic lymphocytes activated by tumorspecific antigens. It is concluded that the recombinant vector constructed can be functionally expressed in the mammalian cells, thus providing a solid foundation for the further investigation on the mechanism to eliminate the immune escape of leukemic cells in vivo.
基金supported by the National Cancer Institute/the National Institute of Health(1R01CA197976,1R01CA201500)Vincent Memorial Hospital Foundation+6 种基金the Vincent Center for Reproductive Biologythe Olson Center for Women’s HealthUniversity of Nebraska Medical Center Graduate Studies Fellowshipthe Fred&Pamela Buffett Cancer Center(LB595)Colleen’s Dream FoundationMarsha Rivkin Center for Ovarian Cancer Research(the Barbara Learned Bridge Funding Award)the Co BRE grant from the Nebraska Center for Cellular Signaling/the National Institute of General Medical Science/the National Institute of Health(5P30GM106397)。
文摘Understanding the cell-of-origin of ovarian high grade serous cancer(HGSC)is the prerequisite for efficient prevention and early diagnosis of this most lethal gynecological cancer.Recently,a mesenchymal type of ovarian HGSC with the poorest prognosis among ovarian cancers was identified by both TCGA and AOCS studies.The cell-of-origin of this subtype of ovarian cancer is unknown.While pursuing studies to understand the role of the Hippo pathway in ovarian granulosa cell physiology and pathology,we unexpectedly found that the Yes-associated protein 1(YAP1),the major effector of the Hippo signaling pathway,induced dedifferentiation and reprogramming of the ovarian granulosa cells,a unique type of ovarian follicular cells with mesenchymal lineage and high plasticity,leading to the development of high grade ovarian cancer with serous features.Our research results unveil a potential cell-of-origin for a subtype of HGSC with mesenchymal features.
