PTH融合蛋白研究进展

甲状旁腺激素(Parathyroid Hormone)是由甲状旁腺主细胞合成和分泌的一种单链多肽激素,是人体内维持血钙恒定的主要激素.PTH在人体内的生理功能非常广泛,PTH类似物已被开发成为治疗骨质疏松症的一系列新型药物.为了克服人PTH分子量小、在人体内代谢快、临床应用不方便的缺点,长效融合蛋白体的研究逐渐成为一种趋势,并且取得了一定的成就.主要依据国内外对PTH重组体和PTH融合蛋白体的研究现状,对PTH的构效关系与生理活性、PTH类似物的研究成果及PTH的应用前景进行综述.     

PTHrP受体胞外区基因的克隆及其在大肠杆菌中的表达

PTHrP(甲状旁腺激素相关蛋白,Parathyroid hormone related protein)与多种肿瘤的生物学行为有关,尤其对肿瘤的骨转移起重要作用。应用其可溶性受体阻断它的病理作用是一个非常值得研究的课题。从肾癌组织中提取RNA,采用逆转录PCR (RT-PCR)方法获得PTHrP受体N端胞外区543碱基对的带信号肽的cDNA片段。将其核苷酸序列与国外发表的资料相比,基因中第262位核苷酸由T变为C,相应密码子由TAC变为CAC,导致氨基酸变异,即由酪氨酸变为组氨酸。将获得的受体基因克隆到原核表达载体pET-3a,在大肠杆菌得到高效表达,经ELISA证实表达产物可与PTHrP特异性结合。体外生物学活性检测表明表达产物可阻断PTHrP的生物学活性。     

人甲状旁腺激素的研究

人甲状旁腺激素(hPTH)是甲状旁腺分泌的多肽激素。它能与骨基质和肾细胞膜上专一性的受体相结合,将调节细胞中钙磷浓度的信号传导到膜内。hPTH活性片段在N端,其N端氨基酸序列与牛、猪PTH高度同源。hPTH及其活性片段在治疗骨及肌肉疾病方面有重要作用,重组hPTH已获成功。     

DLC1在肿瘤进程中的新角色

原发肿瘤向重要器官,如骨、肺、脑、肝等的转移是恶性肿瘤患者死亡的主要原因。对于女性而言,乳腺癌是发病率较高的一类恶性肿瘤。早期乳腺癌通过合理治疗可实现根治,但晚期会发生原位癌向多器官的转移。其中,大约70%的晚期乳腺癌会发生骨转移。在乳腺癌骨转移过程中,转化生长因子β(TGFβ)-甲状旁腺激素样激素(PTHLH)通路发挥着关键性的调控作用,但对这一通路活性本身受哪些因子的影响方面现在知之甚少。因此,中国科学院上海生命科学研究院健康科学研究所胡国宏课题组利用多种肿瘤转移模型,探索影响TGFβ-PTHLH通路活性的因素,最终发现DLC1在乳腺癌骨转移过程中起关键性的负调控作用。DLC1功能的行使依赖于它失活RHO-ROCK通路,继而抑制TGFβ诱导的SMAD3连接区磷酸化。DLC1对后者的抑制降低了TGFβ诱导的PTHLH的转录和分泌,从而减少了骨转移微环境中的成熟破骨细胞,最终阻碍了乳腺癌的破骨性转移。该项研究首次证明DLC1-RHO信号通路在调控乳腺癌骨转移微环境中具有重要作用,也为临床上治疗乳腺癌骨转移提供了新思路。     

超强激素

超强激素董为人李进李福山（第一军医大学细胞生物学实验室,广州５１０５１５）关键词超强激素糖蛋白激素糖蛋白激素为一组进化保守的激素,参与生殖和代谢调节,广泛存在于种属间,如鳗鱼和人类。该类激素的家族包括垂体腺分泌的促性腺激素,如促卵泡素（ＦＳＨ）、黄体...     

重组人甲状旁腺激素肽（1-34）在大肠杆菌中的高效融合表达及纯化

人工合成人甲状旁腺激素1-34肽段 (PTH1-34)的cDNA序列,克隆到大肠杆菌蛋白表达载体pThioHis中,获得了高表达菌株。经发酵、破菌、金属鳌合层析、反相层析和凝胶层析后获得了纯度大于95%的hPTH1-34。hPTH1-34肽N端测序和质谱分子量测定结果与天然PTH1-34一致。生物学活性研究表明,hPTH1-34在体外具有刺激腺苷酸环化酶的作用。     

人组织激肽释放酶4在激素依赖性肿瘤中的研究进展

人组织激肽释放酶基因家族由KLK1-KLK15构成,编码一组丝氨酸蛋白酶。研究发现KLK基因家族涉及癌细胞的多种生物学功能,且其表达受类固醇激素的调节。人组织激肽释放酶4是丝氨酸蛋白酶家族的一个成员,在多种激素依赖性肿瘤如卵巢癌、前列腺癌、乳腺癌、子宫内膜癌中高表达,且表达量受雌激素、孕激素、雄激素不同程度的调节。近年来很多文献报道人组织激肽释放酶4涉及癌细胞的增殖、上皮间质转化及细胞外基质的降解等过程,可能促进了肿瘤的发生、发展,且与激素依赖性肿瘤的预后不良有关。这些研究显示人组织激肽释放酶4与激素依赖性肿瘤关系密切,是其潜在的肿瘤标记物和治疗靶点,随着研究的进一步深入,有望应用于激素依赖性肿瘤的早期诊断、病程监测和治疗。     

甲状旁腺激素的基因表达调控

甲状旁腺激素(parathyroid hormone,PTH)是人体骨转换的主要决定因子,它在骨质疏松的发生、预防和治疗中发挥着重要作用,也是目前应用于临床的主要的促骨形成药物之一。PTH的表达和分泌受到Ca2+浓度、钙敏感受体(calcium-sensing recepter,Ca SR)、维生素D、成纤维细胞生长因子(fibroblast growth factor-23,FGF-23)、雌激素等因素影响。本文就近年来PTH基因表达调控的研究进展进行了简要综述,以期为该领域的研究和应用提供有益参考。     

类固醇激素作用的非基因组效应

类固醇激素作用的非基因组效应崔肇春（大连医科大学生化教研室,大连１１６０２７）关键词类固醇激素非基因组效应六十年代发现了类固醇激素的胞内受体,促进了类固醇激素作用机制的研究。众所周知,类固醇激素作用的途径是它直接进入细胞后,与胞内受体结合成激素－受体...     

超数排卵常用的激素

人教版高中生物学选修3中提到超数排卵要用促性腺激素处理。简要介绍超数排卵常用的几种激素。     

Gene for parathyroid hormone-like peptide is on mouse chromosome 6

The single-copy parathyroid hormone-like peptide gene (Pthlh) was assigned to mouse chromosome 6 using a rat PTHLH cDNA as hybridization probe in the Southern blot analysis of DNAs isolated from a panel of mouse x Chinese hamster cell hybrids. The mouse parathyroid hormone gene (Pth) has previously been assigned to mouse chromosome 7 and the PTHLH and PTH genes have also been shown to be on different chromosomes in human and rat. Therefore, despite significant amino-terminal sequence homology between the PTHLH and PTH peptides, as well as similarities in the structural organization of the human PTHLH and PTH genes, the genes encoding these peptides have discrete chromosomal locations in the mouse, rat, and man.     

The roles of parathyroid hormone-like hormone during mouse preimplantation embryonic development

Parathyroid hormone-like hormone (PTHLH) was first identified as a parathyroid hormone (PTH)-like factor responsible for humoral hypercalcemia in malignancies in the 1980s. Previous studies demonstrated that PTHLH is expressed in multiple tissues and is an important regulator of cellular and organ growth, development, migration, differentiation, and survival. However, there is a lack of data on the expression and function of PTHLH during preimplantation embryonic development. In this study, we investigated the expression characteristics and functions of PTHLH during mouse preimplantation embryonic development. The results show that Pthlh is expressed in mouse oocytes and preimplantation embryos at all developmental stages, with the highest expression at the MII stage of the oocytes and the lowest expression at the blastocyst stage of the preimplantation embryos. The siRNA-mediated depletion of Pthlh at the MII stage oocytes or the 1-cell stage embryos significantly decreased the blastocyst formation rate, while this effect could be corrected by culturing the Pthlh depleted embryos in the medium containing PTHLH protein. Moreover, expression of the pluripotency-related genes Nanog and Pou5f1 was significantly reduced in Pthlh-depleted embryos at the morula stage. Additionally, histone acetylation patterns were altered by Pthlh depletion. These results suggest that PTHLH plays important roles during mouse preimplantation embryonic development.     

Precise chromosomal locations of the genes for dentatorubral-pallidoluysian atrophy (DRPLA), von Willebrand factor (F8vWF) and parathyroid hormone-like hormone (PTHLH) in human chromosome 12p by deletion mapping

The precise chromosomal localization of the gene for dentatorubral-pallidoluysian atrophy (DRPLA) was detected by deletion mapping. Segregation patterns of genotypes of polymerase chain reaction products of DRPLA, von Willebrand factor (F8vWF), antigen CD4(p55) (CD4) and parathyroid hormone-like hormone (PTHLH) loci were studied in patients with del(12)(p13.3p13.3), del(12)(p12.3-p11.2), del(12)(p12.1-p11.2), del(12) (p11.2p11.2) and their parents. The gene for DRPLA was assigned to p13.1-p12.3 of chromosome 12. In addition, genes for F8vWF and PTHLH were mapped to p13.2 and p11.2 of chromosome 12, respectively.     

Effects of parathyroid hormone like hormone (PTHLH) antagonist, PTHLH(7-34), on fetoplacental development and growth during midgestation in rats

Parathyroid hormone-like hormone (PTHLH) secretion has been reported in human amnion, chorion, decidual cytotrophoblast, syncytiotrophoblast, endometrium, and myometrium; however, the functions of PTHLH during pregnancy, particularly during placenta formation and fetal development, are not well understood. We examined whether neutralization of PTHLH action using PTHLH antagonist, PTHLH(7-34), in rats during early gestation affects fetal and placental growth. Rats received s.c. a daily dose of either 0, 4, 12, or 36 microg of PTHLH(7-34) infused continuously through mini-osmotic pumps from Day 8 through Day 15 of pregnancy. Fetal weights measured on Day 15 were significantly decreased in rats treated with all the doses of PTHLH(7-34) compared to controls, and decreases in placental weights were significant at the 12-microg dose. TUNEL assay demonstrated an increased number of apoptotic cells in placenta of treated rats, including rats treated with the 4-microg dose. Cleaved caspase 3 (CASP3), caspase 9 (CASP9) (P < 0.05) and poly-ADP-ribose polymerase (PARP1) (P < 0.01) expression was increased and BCL2 (P < 0.01) expression was decreased in rats treated with 4 microg PTHLH(7-34) compared to that in control. Placental cytochrome c expression was increased (P < 0.01) in cytosolic and decreased (P < 0.01) in mitochondrial fraction in PTHLH(7-34)-treated rats. Caspase 8 expression was not affected by the treatment. Immunohistochemical analysis of platelet endothelial cell adhesion molecule (PECAM1) showed higher staining intensity in control than in treated rats. In conclusion, these results suggests that PTHLH plays a role in early pregnancy, and that antagonization of PTHLH action causes fetoplacental growth restriction through activation of mitochondrial pathway of apoptosis in the placenta and through decreased expression of PECAM1.     

Localization of mouse parathyroid hormone-like peptide gene (Pthlh) to distal chromosome 6 using interspecific backcross mice and in situ hybridization.

The single copy parathyroid hormone-like peptide gene (Pthlh) was mapped to distal mouse chromosome 6 using genetic linkage analysis with a panel of DNA samples from interspecific backcross mice. In all 114 meiotic events examined, the Pthlh locus cosegregated with the locus for the Kirsten ras-2 gene (Kras-2) which was previously localized to distal mouse chromosome 6. In addition, Pthlh was localized to chromosome 6 band F-G and the mouse parathyroid hormone Pth was localized to chromosome 7 band F, by in situ hybridization. These studies confirm the previous localization of Pthlh to mouse chromosome 6 using somatic cell hybrids and show that the Pthlh/PTHLH locus is a part of a conserved linkage group between distal mouse chromosome 6 and the proximal segment of the short arm of human chromosome 12.     

Assignment of the rat parathyroid hormone-like peptide gene (PTHLH) to chromosome 4: evidence for conserved synteny between human chromosome 12, mouse chromosome 6, and rat chromosome 4

The gene coding for rat parathyroid hormone-like peptide (PTHLH) was previously assigned to rat chromosome 2 (Hendy et al., 1988). We reexamined this assignment. According to our results, the gene is on rat chromosome 4. Taking into account the known localizations of the KRAS2 (Kras-2) oncogene and the PTHLH gene, this assignment strongly suggests that a synteny group is conserved on rat chromosome 4, mouse chromosome 6, and human chromosome 12.     

microRNA expression profiles and the potential competing endogenous RNA networks in NELL-1-induced human adipose-derived stem cell osteogenic differentiation

Studies have indicated that Nel-like molecule-1 (NELL-1) was an osteoblast-specific cytokine and some specific microRNAs (miRNAs) could serve as competing endogenous RNA (ceRNA) to partake in osteogenic differentiation of human adipose-derived stem cells (hASCs). The aim of this study was to explore the potential functional mechanisms of recombinant human NELL-1 protein (rhNELL-1) during hASCs osteogenic differentiation. rhNELL-1 was added to osteogenic medium to activate osteogenic differentiation of hASCs. High-throughput RNA sequencing (RNA-Seq) was performed and validated by real-time quantitative polymerase chain reaction. Gene ontology functional annotation and Kyoto Encyclopedia of Genes and Genomes pathway analysis were performed to detect the functions of differentially expressed miRNAs and genes. Coding-noncoding gene co-expression network and ceRNA networks were constructed to predict the potential regulatory role of miRNAs. A total of 1010 differentially expressed miRNAs and 1762 differentially expressed messenger RNAs (mRNAs) were detected. miRNA-370-3p, bone morphogenetic protein 2 (BMP2), and parathyroid hormone like hormone (PTHLH) were differentially expressed during NELL-1-induced osteogenesis. Bioinformatic analyses demonstrated that these differentially expressed miRNAs and mRNAs enriched in Rap1 signaling pathway, PI3K-Akt signaling pathway, p53 signaling pathway, Glucagon signaling pathway, and hypoxia-inducible factor-1 signaling pathway, which were important pathways related to osteogenic differentiation. In addition, miRNA-370-3p and has-miR-485-5p were predicted to interact with circ0001543, circ0002405, and ENST00000570267 in ceRNA networks. Based on the gain or loss of functional experiments by transfection, the results showed that miR-370-3p was a key regulator in osteogenic differentiation by targeting BMP2 and disturbing the expression of PTHLH, and participated in NELL-1-stimulated osteogenesis. The present study provided the primary data and evidence for further exploration on the roles of miRNAs and ceRNAs during NELL-1-induced ossification of hASCs.     

Demonstration that cyclic adenosine 3',5'-monophosphate mediates the lipolytic action of parathyroid hormone.

Studies were carried out with rat epididymal fat pads first to compare the effects of the synthetic N-terminal 1-34 peptide of bovine parathyroid hormone and of the native hormone to determine whether this portion of the molecule is responsible for the lipolytic action of the hormone and second to determine whether this biologic action of parathyroid hormone is mediated by cyclic adenosine 3',5'-monophosphate. The N-terminal polypeptide was as effective as the native hormone in stimulating lipolysis in the concentration range between 10(-8) M and 10(-6) M. Parathyroid hormone stimulated lipolysis by isolated fat cells. The concentration of cyclic adenosine 3',5'-monophosphate in the fat pads was significantly increased by the hormone (10(-6)M). Lipolytic stimulation by parathyroid hormone (10(-6)M) was diminished by insulin (100 muU/ml) and prostaglandin E1 (1 mug/ml), both of which are known inhibitors of lipolysis. The findings indicate that the amino-terminal 1-34 peptide portion of parathyroid hormone is responsible for the lipolytic action and that this effect is mediated through cyclic adenosine 3',5'-monophosphate.     

The structure of human parathyroid hormone-related protein(1-34) in near-physiological solution

Parathyroid hormone-related protein plays a major role in the pathogenesis of humoral hypercalcemia of malignancy. Under normal physiological conditions, parathyroid hormone-related protein is produced in a wide variety of tissues and acts in an autocrine or paracrine fashion. Parathyroid hormone-related protein and parathyroid hormone bind to and activate the same G-protein-coupled receptor. Here we present the structure of the biologically active NH2-terminal domain of human parathyroid hormone-related protein(1-34) in near-physiological solution in the absence of crowding reagents as determined by two-dimensional proton magnetic resonance spectroscopy. An improved strategy for structure calculation revealed the presence of two helices, His-5-Leu-8 and Gln-16-Leu-27, connected by a flexible linker. The parathyroid hormone-related protein(1-34) structure and the structure of human parathyroid hormone(1-37) as well as human parathyroid hormone(1-34) are highly similar, except for the well defined turn, His-14-Ser-17, present in parathyroid hormone. Thus, the similarity of the binding affinities of parathyroid hormone and parathyroid hormone-related protein to their common receptor may be based on their structural similarity.