Amyloid beta protein precursor is possibly a heparan sulfate proteoglycan core protein

The amyloid beta protein peptide is a major constituent of amyloid plaque cores in Alzheimer's disease and is apparently derived from a higher molecular weight precursor. It is now shown that the core protein of a heparan sulfate proteoglycan secreted from a nerve cell line (PC12) has an amino acid sequence and a size very similar to those of the amyloid beta protein precursor and that these molecules are antigenically related. This amyloid beta protein precursor-related protein is not found in the conditioned medium of a variant cell line (F3 PC12) that does not secrete heparan sulfate proteoglycan. The synaptic localization and metabolism of this class of proteoglycans are consistent with its potential involvement in central nervous system dysfunction.     

Cleavage of amyloid beta peptide during constitutive processing of its precursor

The amyloid beta peptide (A beta P) is a small fragment of the much larger, broadly distributed amyloid precursor protein (APP). Abundant A beta P deposition in the brains of patients with Alzheimer's disease suggests that altered APP processing may represent a key pathogenic event. Direct protein structural analyses showed that constitutive processing in human embryonic kidney 293 cells cleaves APP in the interior of the A beta P, thus preventing A beta P deposition. A deficiency of this processing event may ultimately prove to be the etiological event in Alzheimer's disease that gives rise to senile plaque formation.     

Deposits of amyloid beta protein in the central nervous system of transgenic mice.

Alzheimer's disease is characterized by widespread deposition of amyloid in the central nervous system. The 4-kilodalton amyloid beta protein is derived from a larger amyloid precursor protein and forms amyloid deposits in the brain by an unknown pathological mechanism. Except for aged nonhuman primates, there is no animal model for Alzheimer's disease. Transgenic mice expressing amyloid beta protein in the brain could provide such a model. To investigate this possibility, the 4-kilodalton human amyloid beta protein was expressed under the control of the promoter of the human amyloid precursor protein in two lines of transgenic mice. Amyloid beta protein accumulated in the dendrites of some but not all hippocampal neurons in 1-year-old transgenic mice. Aggregates of the amyloid beta protein formed amyloid-like fibrils that are similar in appearance to those in the brains of patients with Alzheimer's disease.     

Platelet coagulation factor XIa-inhibitor, a form of Alzheimer amyloid precursor protein

An inhibitor of coagulation factor XIa was purified from serum-free conditioned medium of HepG2 liver cells. Platelets stimulated with thrombin or calcium ionophore (A23187) secrete a protein functionally and immunologically identical to the inhibitor, implying a role for this inhibitor in hemostasis. Analysis of the amino-terminal amino acid sequence and immunologic reactivity showed the inhibitor to be a truncated form of the Alzheimer's amyloid precursor protein that contains a Kunitz-type serine protease inhibitor domain and at least a portion of the amyloid beta protein. It inhibits factor XIa and trypsin with a Ki of 450 +/- 50 pM and 20 +/- 10 pM, respectively. Heparin (1 unit/ml) did not significantly effect inhibition of trypsin, but inhibition of XIa was 15 times greater (Ki = 25 +/- 15 pM) in the presence of heparin.     

高良姜提取物对PC12细胞的神经保护作用

通过建立H2O2介导的PC12细胞损伤模型;采用MTT法检测细胞存活率,利用荧光显微镜观察细胞形态;检测细胞中乳酸脱氢酶漏出率,丙二醛(MDA)含量,超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GGSH-Px)活性,评价高良姜提取物对H2O2介导的PC12细胞损伤的保护作用。结果表明,高良姜提取物能明显降低细胞内乳酸脱氢酶漏出率,降低细胞内MDA含量,提高SOD和GGSH-Px的活性,且具有浓度依赖性,由此推断,高良姜提取物对H2O2介导的PC12细胞损伤有明显保护作用。     

Amyloid beta protein precursor gene and hereditary cerebral hemorrhage with amyloidosis (Dutch)

Human hereditary cerebral hemorrhage with amyloidosis of the Dutch type (HCHWA-D), an autosomal dominant form of cerebral amyloid angiopathy (CAA), is characterized by extensive amyloid deposition in the small leptomeningeal arteries and cortical arterioles, which lead to an early death of those afflicted in their fifth or sixth decade. Immunohistochemical and biochemical studies have indicated that the amyloid subunit in HCHWA-D is antigenically related to and homologous in sequence with the amyloid beta protein isolated from brains of patients with Alzheimer's disease and Down syndrome. The amyloid beta protein is encoded by the amyloid beta protein precursor (APP) gene located on chromosome 21. Restriction fragment length polymorphisms detected by the APP gene were used to examine whether this gene is a candidate for the genetic defect in HCHWA-D. The data indicate that the APP gene is tightly linked to HCHWA-D and therefore, in contrast to familial Alzheimer's disease, cannot be excluded as the site of mutation in HCHWA-D.     

A mutation in the amyloid precursor protein associated with hereditary Alzheimer's disease

Alzheimer's disease is a form of localized amyloidosis characterized by cerebral cortical amyloid plaques, neurofibrillary tangles, and amyloid deposits within the walls of leptomeningeal vessels. Although most cases of Alzheimer's disease are sporadic, kindreds with autosomal-dominant inheritance of the syndrome suggest that a single mutation may be important in pathogenesis. Direct sequencing of DNA from a family with autopsy-proven Alzheimer's disease revealed a single amino acid substitution (Phe for Val) in the transmembrane domain of the amyloid precursor protein. This mutation correlates with the presence of Alzheimer's disease in all patients in this study, and may be the inherited factor causing both amyloid fibril formation and dementia.     

多巴胺对PC12细胞活性的影响及三七皂苷-Rg1的保护作用

采用细胞化学及生物化学方法研究了三七皂苷-Rg1时多巴胺引起的PC12细胞损伤的保护作用。结果表明：三七皂苷-Rg1对PCl2细胞损伤有明显的改善作用，可显著提高PC12细胞的增殖活性。同时降低培养上清中乳酸脱氢酶的释放量夏细胞内凋亡蛋白Bax的表达，这提示三七皂苷-Rg1对神经系统由于多巴胺氧化引起的损伤有明显的治疗和保护作用。     

Neurotrophic and neurotoxic effects of amyloid beta protein: reversal by tachykinin neuropeptides

The amyloid beta protein is deposited in the brains of patients with Alzheimer's disease but its pathogenic role is unknown. In culture, the amyloid beta protein was neurotrophic to undifferentiated hippocampal neurons at low concentrations and neurotoxic to mature neurons at higher concentrations. In differentiated neurons, amyloid beta protein caused dendritic and axonal retraction followed by neuronal death. A portion of the amyloid beta protein (amino acids 25 to 35) mediated both the trophic and toxic effects and was homologous to the tachykinin neuropeptide family. The effects of the amyloid beta protein were mimicked by tachykinin antagonists and completely reversed by specific tachykinin agonists. Thus, the amyloid beta protein could function as a neurotrophic factor for differentiating neurons, but at high concentrations in mature neurons, as in Alzheimer's disease, could cause neuronal degeneration.     

Atomic view of a toxic amyloid small oligomer

Amyloid diseases, including Alzheimer's, Parkinson's, and the prion conditions, are each associated with a particular protein in fibrillar form. These amyloid fibrils were long suspected to be the disease agents, but evidence suggests that smaller, often transient and polymorphic oligomers are the toxic entities. Here, we identify a segment of the amyloid-forming protein αB crystallin, which forms an oligomeric complex exhibiting properties of other amyloid oligomers: β-sheet-rich structure, cytotoxicity, and recognition by an oligomer-specific antibody. The x-ray-derived atomic structure of the oligomer reveals a cylindrical barrel, formed from six antiparallel protein strands, that we term a cylindrin. The cylindrin structure is compatible with a sequence segment from the β-amyloid protein of Alzheimer's disease. Cylindrins offer models for the hitherto elusive structures of amyloid oligomers.     

抗氧化中药复方浸提液对PC12细胞铅中毒的防护作用

研究了由茯苓、熟地黄、山茱萸、枸杞及麦饭石组成的复方(PRMFM)的浸提液对PC12细胞铅中毒的防护作用。结果表明:PC12细胞暴露在500 mmol/L的醋酸铅环境中时,细胞的膜脂过氧化程度增大,活性氧(O-2,H2O2)含量增多,抗氧化酶(Superoxide dismutase, SOD; Peroxidase, POD; Catalase, CAT)活性降低。若在细胞遭受铅中毒之前用PRMFM浸提液预处理,可抑制由铅引起的细胞膜脂过氧化程度的增大、活性氧(O-2,H2O2)含量的增多及抗氧化酶(SOD, POD, CAT)活性的降低。表明该中药复方对铅中毒的细胞有明显的防护作用。     

乙醇等有机溶剂对肿瘤细胞PC12的毒性剂量分析

分析一定浓度的甲醇、乙醇、丙酮和甘油作用下肿瘤细胞 PC12形态和生存率的变化,确定上述有机溶剂对细胞的毒性剂量。研究发现,PC12细胞对上述溶剂表现出不同的耐受性,其中乙醇的毒性剂量最低,20 ml/L浓度作用24 h细胞生存率就下降近40%;该浓度的丙酮和甲醇干预 PC12细胞24 h对应的细胞生存率则分别下降约20%和15%;甘油的作用相对缓和,但随着干预浓度逐渐增大时,也表现明显的细胞毒性。该研究可为后续药理研究提供参考资料。     

Tracking SNARE complex formation in live endocrine cells

Syntaxin, synaptosome-associated protein of 25 kD (SNAP25), and vesicle-associated membrane protein/synaptobrevin are collectively called SNAP receptor (SNARE) proteins, and they catalyze neuronal exocytosis by forming a "core complex." The steps in core complex formation are unknown. Here, we monitored SNARE complex formation in vivo with the use of a fluorescent version of SNAP25. In PC12 cells, we found evidence for a syntaxin-SNAP25 complex that formed with high affinity, required only the amino-terminal SNARE motif of SNAP25, tolerated a mutation that blocks formation of other syntaxin-SNAP25 complexes, and assembled reversibly when Ca2+ entered cells during depolarization. The complex may represent a precursor to the core complex formed during a Ca2+-dependent priming step of exocytosis.     

Lipoprotein uptake by neuronal growth cones in vitro

Macrophages that rapidly enter injured peripheral nerve synthesize and secrete large quantities of apolipoprotein E. This protein may be involved in the redistribution of lipid, including cholesterol released during degeneration, to the regenerating axons. To test this postulate, apolipoprotein E-associated lipid particles released from segments of injured rat sciatic nerve and apolipoprotein E-containing lipoproteins from plasma were used to determine whether sprouting neurites, specifically their growth cones, possessed lipoprotein receptors. Pheochromocytoma (PC12) cells, which can be stimulated to produce neurites in vitro, were used as a model system. Apolipoprotein E-containing lipid particles and lipoproteins, which had been labeled with fluorescent dye, were internalized by the neurites and their growth cones; the unmetabolized dye appeared to be localized to the lysosomes. The rapid rate of accumulation in the growth cones precludes the possibility of orthograde transport of the fluorescent particles from the PC12 cell bodies. Thus, receptor-mediated lipoprotein uptake is performed by the apolipoprotein B,E(LDL) (low density lipoprotein) receptors, and in the regenerating peripheral nerve apolipoprotein E may deliver lipids to the neurites and their growth cones for membrane biosynthesis.     

原花青素影响Aβ25-35诱导PC12细胞周期变化与细胞凋亡的可能机制

目的探讨原花青素（Proanthocyanidins,PAC）对β-淀粉样肽（25—35）[β amyloid peptide-（25—35）,Aβ25-35]诱导体外血清饥饿培养的PCI2细胞周期异常与凋亡保护作用的可能机制。方法种人培养瓶或板的PC12细胞贴壁后用常用血清饥饿培养使细胞同步于岛期,30mg／L的PAC预处理血清饥饿培养的PC12细胞,加入终浓度为25μmol／L Aβ25-35处理0～20h,通过RT-PCR和Western blot从mRNA及蛋白水平检测细胞周期蛋白依赖性激酶4（Cyclin—dependent kinase-4,CDK4）、磷酸化的视网膜纤维母细胞瘤蛋白（phosphorylated Retinoblastoma protein,pRb）、腺病毒E2启动子结合因子1（Adenovirus E2 factor-1,E2 F1）、B细胞淋巴瘤／白血病关联X蛋白（B-cell lymphoma／leukemia-2 Associated X protein,bax）基因表达的变化。结果与Aβ25—35诱导组比较,CDK4、E2F1、bax mRNA表达和CDK4、pR6、Bax蛋白表达降低。结论PAC可能通过下调CDK4、pRb、E2F1的表达,从而降低bax的活化,对Aβ25-35诱导的PC12细胞周期异常与凋亡起保护作用的。     

A plant peptide encoded by CLV3 identified by in situ MALDI-TOF MS analysis

The Arabidopsis CLAVATA3 (CLV3) gene encodes a stem cell-specific protein presumed to be a precursor of a secreted peptide hormone. Matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) applied to in situ Arabidopsis tissues determined the structure of a modified 12-amino acid peptide (MCLV3), which was derived from a conserved motif in the CLV3 sequence. Synthetic MCLV3 induced shoot and root meristem consumption as cells differentiated into other organs, displaying the typical phenotype of transgenic plants overexpressing CLV3. These results suggest that the functional peptide of CLV3 is MCLV3.     

Diverse gene expression for isotypes of murine serum amyloid A protein during acute phase reaction

Serum amyloid A protein (SAA) is a precursor for a major component of amyloid fibrils, which, upon deposition, cause secondary amyloidosis in diseases such as rheumatoid arthritis. In mice, SAA is encoded by at least three genes, which show diverse expression during inflammation. Furthermore, in amyloidosis-resistant SJL mice, the gene expression for one SAA isotype, SAA2, is defective, although SAA2 gene expression is normal in amyloidosis-susceptible BALB/c mice. Because only SAA2-derived products deposit in mouse amyloid tissues, the resistance of SJL mice to amyloidosis seems to be due to defective SAA2 gene expression. Thus, the study emphasizes the importance of SAA gene structure in determining susceptibility to amyloidosis.     

玫瑰花对β-淀粉样蛋白诱导PC12神经细胞毒性的抑制作用

[目的]研究玫瑰花提取物对β-淀粉样蛋白（Aβ25-35）诱导的损伤PC12细胞的保护作用。[方法]PC12细胞加入Aβ25-35共同孵育后,采用四甲基偶氮唑盐（MTT）法检测PC12细胞活力,比色法检测细胞内丙二醛（MDA）含量,酶联免疫法检测细胞内Nf-κb水平。[结果]玫瑰花提取物对PC12无显著细胞毒性,可以显著抑制Aβ25-35诱导的PC12细胞毒性。与模型组相比,添加玫瑰花乙酸乙酯萃取物的样品组能降低PC12细胞内MDA浓度和Nf-κb表达。[结论]玫瑰花提取物对β-淀粉样蛋白致PC12细胞损伤具有一定的抑制作用,其作用机理可能与降低Aβ25-35所致的PC12细胞内的氧化应激有关。     

Signaling pathways for PC12 cell differentiation: making the right connections

A key issue in signal transduction is how signaling pathways common to many systems-so-called canonical signaling cassettes-integrate signals from molecules having a wide spectrum of activities, such as hormones and neurotrophins, to deliver distinct biological outcomes. The neuroendocrine cell line PC12, derived from rat pheochromocytoma, provides an example of how one canonical signaling cassette-the Raf --> mitogen-activated protein kinase kinase (MEK) --> extracellular signal-regulated kinase (ERK) pathway-can promote distinct outcomes, which in this case include neuritogenesis, gene induction, and proliferation. Two growth hormones, epidermal growth factor (EGF) and nerve growth factor (NGF), use the same pathway to cause PC12 proliferation and differentiation, respectively. In addition, pituitary adenylate cyclase-activating polypeptide (PACAP), a neurotransmitter that also causes differentiation, uses the same canonical cassette as NGF but in a different way. The Connections Map for PC12 Cell Differentiation brings into focus the complex array of specific cellular responses that rely on canonical signal transduction systems.