胆固醇对金属离子磷脂酰甘油单分子膜成膜的影响

目的：本文主要研究不同条件胆固醇（Cholesterol,简称Chol）和金属离子（钾、镁离子）对磷脂酰甘油（Phosphatidylglycerols,简称PG）相互作用后形成的单分子膜的影响。方法：首先以金属离子作为亚相,研究胆固醇的量对磷脂酰甘油单分子膜的影响;其次加入同等量的胆固醇量,亚相为不同金属离子时,对磷脂酰甘油单分子膜的影响,最后分析磷脂酰甘油LB膜的π-A曲线,即曲线外扩、相变点、膜压等等变化特征。结果：随着胆固醇的增多,金属离子磷脂酰甘油单分子膜形成π-A曲线变化逐渐明显;当加入同等量的胆固醇时,随着金属离子价态的逐渐增高,磷脂酰甘油单分子膜形成的π-A曲线的变化也逐渐明显。结论：其一：胆固醇对金属离子磷脂酰甘油单分子膜成膜质量是有影响的。其二,金属离子对胆固醇与磷脂酰甘油混合形成的单分子膜同样也是有的影响。     

磷脂酰肌醇—4—激酶的免疫分析

在化猪肝微粒体磷脂酰肌醇－４－激酶（ＰＩ－４－Ｋ）的基础上,制备该酶的免疫血清,提纯ＩｇＧ,并建立了ＰＩ４－Ｋ的ＥＬＩＳＡ,进行不同组织中ＰＩ４－Ｋ的免疫沉淀分析和免疫定量。结果证明：猪肝微粒体ＰＩ４－Ｋ的抗血清或抗体ＩｇＧ可沉淀猪肾和猪肺的微粒膜,猪肝细胞膜和人中性粒细胞细胞膜ＴｒｉｔｏｎＸ－１００增溶液（ＴＸＳＭ）中的ＰＩ４－Ｋ表明不同组织,不同亚细胞和不同种属的ＰＩ４－Ｋ有相似的抗原性。猪肝     

磷脂酰肌醇－4－激酶的的免疫分析

在纯化猪肝微粒体磷脂酰肌醇－４－激酶（ＰＩ－４－Ｋ）的基础上,制备该酶的免疫血清、提纯ＩｇＧ,并建立了ＰＩ４－Ｋ的ＥＬＩＳＡ,进行不同组织中ＰＩ４－Ｋ的免疫沉淀分析和免疫定量。结果证明：猪肝微粒体ＰＩ４－Ｋ的抗血清或抗体ＩｇＧ可沉淀猪肾和猪肺的微粒体膜、猪肝细胞膜和人中性粒细胞细胞膜ＴｒｉｔｏｎＸ－１００增溶液（ＴＸＳＭ）中的ＰＩ４－Ｋ,表明不同组织、不同亚细胞器和不同种属的ＰＩ４－Ｋ有相似的抗原性。猪肝、肾、肺、胃、小肠和大肠ＴＸＳＭ中ＰＩ４－Ｋ的活力和酶蛋白含量相仿,急性中性粒细胞白血病患者的中性粒细胞膜上ＰＩ４－Ｋ的含量和活力都增加３．５倍左右．加上ＰＩ４－Ｋ抗体沉淀各组织ＰＩ４－Ｋ活力和酶蛋白的百分率基本相同,支持文献报道组织中微粒体或细胞膜中的ＰＩＫ主要是ＰＩ４－Ｋ,而免疫性不同的ＰＩ３－Ｋ含量极少的观点．     

磷脂酰胆碱特异性磷脂酶C的研究

随着人们对信号转导认识的逐步加深,各种磷脂酶在信号通路中的作用也日渐受到重视,并日趋明了。其中磷脂酶Ａ２（ＰＬＡ２）、磷脂酰胆碱特异性磷脂酶Ｄ（ＰＣ－ＰＬＤ）的基因已克隆,对磷脂酰肌醇特异性磷脂酶Ｃ（ＰＩ－ＰＬＣ）也有较深了解,而对磷脂酰胆碱特异性磷...     

苯丙氨酸稀土配合物与磷脂脂质体相互作用的研究

用ＤＳＣ、ＦＴ－ＩＲ和Ｒａｍａｎ光谱法研究了苯丙氨酸（Ｐｈｅ）及其稀土配合物Ｌｎ（Ｐｈｅ）Ｃｌ３．６Ｈ２Ｏ（Ｌｎ＝Ｓｍ．ｌａ．Ｇａ．Ｙｂ）对ＤＰＰＣ热致性的影响,并从分子水平上探讨了它们与ＤＰＰＣ相互作用的机理。结果表明：Ｐｈｅ对ＤＰＰＣ从凝胶态到液晶态的相变温度Ｔｍ影响不大,但磷脂酰链的构象有序度降低;Ｌｎ（Ｐｈｅ）Ｃｌ３．６Ｈ２Ｏ像Ｌｎ３＋一样,使ＤＰＰＣ的Ｔｍ显著升高,但前者的升高程度小于后者,同时磷脂链的构象有序度升高。上述ＤＰＰＣ脂质体二元体系的铸膜ＦＴＩＲ结果与ＤＰＰＣ脂质体基本上相同,配合物Ｌｎ（Ｐｈｅ）Ｃｌ３．６Ｈ２Ｏ的配合键是离子型的．     

地塞米松和肺成纤维细胞对大鼠胎肺表面活性物质合成的影响

为提高ＤＥＸ对新生儿呼吸窘迫综合征（ＮＲＤＳ）的防治,本文研究了地塞米松（ＤＥＸ）促进肺表面活性物质（ＰＳ）合成的作用机制。用分离培养的２０天大鼠胚胎肺泡Ⅱ型细胞为材料,观察了单独用ＤＥＸ、用与不用ＤＥＸ刺激后的成纤维细胞条件培养液（ＤＥＸＦＣＭ、ＦＣＭ）对肺泡Ⅱ型细胞ＰＳ中磷脂合成及特异性肺表面活性物质蛋白质ＳＰＢ、ＳＰＣｍＲＮＡ表达的影响。结果表明,ＤＥＸ本身及未用ＤＥＸ刺激的ＦＣＭ对肺泡Ⅱ型细胞的上述三种指标均没有影响,而ＤＥＸ刺激后的ＤＥＸＦＣＭ使这三个指标有不同程度的增加效应。提示糖皮质激素刺激肺泡Ⅱ型细胞ＰＳ合成增加是由成纤维细胞介导的     

猪骨羟基磷灰石的结构稳定性

在不同酸碱条件下对猪骨处理,利用转靶Ｘ射线衍射（ＸＲＤ）、红外吸收（ＩＲ）、差热分析（ＤＴＡ）分别对猪骨的物相和微结构进行了分析,并与纯的羟基磷灰石（ＨＡ）进行了比较,研究了猪骨中纳米羟基磷灰石的结构稳定性。实验表明猪骨在弱酸弱碱溶液中羟基磷灰石相是稳定的,但晶粒尺寸、结晶度和非晶含量有所变化。６００°Ｃ的高温处理,可改善其结晶度,这对开发新的骨移植材料有重要意义     

乳腺癌转移中的磷脂酰胆碱和溶血磷脂酰胆碱分析

我们以前曾报道花生四烯酸（arachidonic acid,AA)代谢产物可以促进乳腺癌细胞增殖和迁移。为了进一步寻找维持高转移乳腺癌细胞中AA高水平代谢的内源机制,深入探求AA代谢促进乳腺癌细胞转移的分子机理,我们应用HPLC/ESI/MSⁿ技术检测和分析了乳腺癌MCF-7和高转移乳腺癌LM-MCF-7细胞中溶血磷脂酰胆碱（lysophosphatidylcholines,LysoPCs）和磷脂酰胆碱（phosphatidylcholines,PCs）的成分和含量。我们发现了10种LysoPC的含量在LM-MCF-7细胞中显著高于MCF-7细胞,有6种PC可水解产生AA,它们在LM-MCF-7细胞中的含量显著低于MCF-7细胞,提示这些溶血磷脂含量的升高和磷脂含量的降低可能与乳腺癌转移相关。在LM-MCF-7细胞中,COX-2抑制剂吲哚美辛（indomethacin,Indo）和LOX抑制剂（nordihydroguaiaretic acid,NDGA）共同作用可明显下调cPLA2的活性,应用HPLC-ESI-MSⁿ技术比较cPLA2活性下调前后LM-MCF-7细胞中LysoPC和PC含量的变化,发现其中4种PC可被cPLA2水解产生AA。我们还发现,细胞内LysoPC与PC的比值可以反映cPLA2的活性。通过以上研究我们进一步证实了由cPLA2活性调节的AA释放及代谢对乳腺癌转移具有重要作用。     

云南高原水稻幼苗的抗冷性与其活性氧清除系统的关系

经低温（２℃或５℃）暗处理（１－５ｄ）的耐寒性不同的云南高原水稻幼苗，超氧物歧化酶（ＳＯＤ），过氧化物酶（ＰＯＸ）的活性均有不同程度的下降。胁迫后光下（２８℃，２５００ｌｘ）恢复，耐寒性较强的品种ＳＯＤ，ＰＯＸ活性均比对照值明显提高，耐寒性弱的品种仍低于对照值。随着低温时间的延长，低温胁迫程度的加深和光下恢复，抗坏血酸（ＡＳＡ）和谷胱甘肽（ＧＳＨ）含量逐渐减少，丙二醛（ＭＤＡ）含量则逐渐增加。耐寒性强的品种ＡＳＡ和ＧＳＨ含量减少较小，ＭＤＡ含量增加也较小。     

荧光光谱法研究铈离子与二棕榈酰磷脂酰胆碱（DPPC）脂质体的相互作用

通过荧光光谱法研究了稀土元素Ｃｅ３＋与二棕榈酰磷脂酰胆碱（ＤＰＰＣ）脂质体的相互作用,结果表明,Ｃｅ３＋－ＤＰＰＣ体系的激发波长在２４７ｎｍ,２９４ｎｍ,发射波长在３４４ｎｍ,与水合Ｃｅ３＋的荧光光谱完全不同。这表明Ｃｅ３＋与ＤＰＰＣ形成了复合物,该复合物的荧光光谱同Ｃｅ３＋－ＤＨＰ复合物的荧光光谱一致。可以认为Ｃｅ３＋与ＤＰＰＣ分子上的磷酸基团相作用。     

山莨菪碱与膜相互作用的小角X射线衍射研究

应用小角X射线衍射和差示扫描量热法研究了中药提取物山莨菪碱与DPPC多层膜的相互作用.结果表明,山莨菪碱不仅能降低DPPC多层膜的相变温度,而且能使多层膜发生分相现象,根据衍射结果并采用推广傅里叶合成去卷积法计算了脂双分子层膜的电子密度剖面图.从中我们估计山莨菪碱分子可能分布于脂双分子层极性头部的内侧     

Interaction of cholesterol with galactocerebroside and galactocerebroside-phosphatidylcholine bilayer membranes

The interaction of the galactocerebroside, N-palmitoylgalactosylsphingosine (NPGS), with cholesterol has been studied by differential scanning calorimetry (DSC) and x-ray diffraction. Thermal and structural studies demonstrate complex behavior characterized by two endothermic transitions: transition I (TI approximately equal to 50-60 degrees C) corresponding to an NPGS-cholesterol bilayer gel----bilayer liquid crystal transition II (TII where TI less than TII less than TNPGS) corresponding to an NPGS bilayer crystal (stable E form)----bilayer liquid crystal transition. For mixtures containing from 6 to 80 mol % cholesterol, x-ray diffraction studies at 22 degrees C (T less than TI) indicate two separate lamellar phases; an NPGS crystal bilayer phase and a cholesterol monohydrate phase. For cholesterol concentrations less than 50 mol % at TI less than T less than TII, NPGS-cholesterol liquid crystal bilayer and excess NPGS crystal bilayer phases are observed. For greater than 50 mol % cholesterol concentrations at these temperatures, an excess cholesterol monohydrate phase coexists with the NPGS-cholesterol liquid crystal bilayers. At T greater than TII, complete NPGS-cholesterol miscibility is only observed for less than 50 mol % cholesterol concentrations, whereas at greater than 50 mol % cholesterol an excess cholesterol phase is present. The solid phase immiscibility of cerebroside and cholesterol at low temperatures is suggested to result from preferential NPGS-NPGS associations via hydrogen bonding. The unique thermal and structural behavior of NPGS-cholesterol dispersions is contrasted with the behavior of cholesterol-phosphatidycholine and cholesterol-sphingomyelin bilayers. Thermal and structural studies of NPGS in dipalmitoylphosphatidylcholine (DPPC)/cholesterol (1:1, molar ratio) bilayers have been performed. For dispersions containing less than 20 mol % NPGS at 22 degrees C there are no observable calorimetric transitions and x-ray diffraction studies indicate complete lipid miscibility. At greater than 20 mol % NPGS, a high temperature transition is observed that is shown by x-ray diffraction studies to be due to an excess NPGS crystal bilayer----liquid crystal bilayer transition. Complete miscibility of NPGS in DPPC/cholesterol bilayers is observed at T greater than TNPGS. The properties of NPGS/DPPC/cholesterol bilayers are discussed in terms of the lipid composition of the myelin sheath.     

Effect of cholesterol on the formation of an interdigitated gel phase in lysophosphatidylcholine and phosphatidylcholine binary mixtures

We previously reported that 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) forms an interdigitated gel phase in the presence of 1-palmitoyl-sn-glycero-3-phosphocholine (16:0LPC) at concentrations below 30 mol%. In the present investigation, fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH), X-ray diffraction, and differential scanning calorimetry (DSC) were used to investigate the effect of cholesterol on the phase behavior of 16:0LPC/DPPC binary mixtures. At 25 degrees C, 30 mol% 16:0LPC significantly decreases the DPH fluorescence intensity during the transition of DPPC from the L(beta') phase to the L(betaI) phase. However, the addition of cholesterol to 16:0LPC/DPPC mixtures results in a substantial increase in fluorescence intensity. The changes in DPH fluorescence intensity reflect the probe's redistribution from an orientation parallel to the acyl chain to the center of the bilayer, suggesting a bilayer structure transition from interdigitation to noninterdigitation. The normal repeat period of small angle X-ray diffraction patterns can be restored and a reflection appears at 0.42 nm with a broad shoulder around 0.41 nm in wide angle X-ray diffraction patterns when 10 mol% cholesterol is incorporated into 30 mol% 16:0LPC/DPPC vesicles, indicating that the mixtures are in the gel phase (L(beta')). Moreover, DSC results demonstrate that 10 mol% cholesterol is sufficient to significantly decrease the main enthalpy, cooperativity and lipid chain melting of 30 mol% 16:0LPC/DPPC binary mixtures, which are L(betaI), indicating that the transition of the interdigitated phase is more sensitive to cholesterol than that of the noninterdigitated phase. Our data imply that the interdigitated gel phase induced by 16:0LPC is prevented in the presence of 10 mol% cholesterol, but unlike ethanol, an increasing concentration of 16:0LPC is not able to restore the interdigitation structure of the lipid mixtures.     

Galactocerebroside-phospholipid interactions in bilayer membranes.

Differential scanning calorimetry (DSC) and x-ray diffraction have been used to study the interaction of hydrated N-palmitoylgalactosylsphingosine (NPGS) and dipalmitoylphosphatidylcholine (DPPC). For mixtures containing less than 23 mol% NPGS, complete miscibility of NPGS into hydrated DPPC bilayers is observed in both the bilayer gel and liquid-crystal phases. X-ray diffraction data demonstrate insignificant differences in the DPPC-bilayer gel-phase parameters on incorporation of up to 23 mol% NPGS. At greater than 23 mol% NPGS, additional high-temperature transitions occur, indicating phase separation of cerebroside. For these cerebroside concentrations, at 20 degrees C, x-ray diffraction shows two lamellar phases, hydrated DPPC-NPGS gel bilayers (d = 64 A) containing 23 mol% NPGS, and NPGS "crystal" bilayers (d = 55 A). On heating to temperatures greater than 45 degrees C, the mixed DPPC-NPGS bilayer phase undergoes chain melting, and on further increasing the temperature progressively more NPGS is incorporated into the liquid-crystal DPPC-NPGS bilayer phase. At temperatures greater than 82 degrees C (the transition temperature of hydrated NPGS), complete lipid miscibility is observed at all DPPC/NPGS molar ratios.     

胆固醇对脂双层结构影响的SAXS和STM研究

用小角X射线散射（SAXS)和扫描隧道显微镜（STM）技术分别研究了模拟生物膜脂质体的结构以及胆固醇对生物膜双层结构的影响。结果表明,在扫描隧道显微镜照片中,磷脂分子在石墨表面形成规则的二维点状排列图像;磷脂胆固醇脂质体在石墨表面形成规则的二维波纹状排列图像。用小角X射线散射研究结果表明,DPPC脂质体是片层相结构,DPPC＋Chol脂质体是复相片层结构,DPPE＋Chol脂质体是片层立方相结构,DPPC＋DPPE＋Chol脂质体是立方六角形相结构。     

Properties of ganglioside GM1 in phosphatidylcholine bilayer membranes.

Gangliosides have been shown to function as cell surface receptors, as well as participating in cell growth, differentiation, and transformation. In spite of their multiple biological functions, relatively little is known about their structure and physical properties in membrane systems. The thermotropic and structural properties of ganglioside GM1 alone and in a binary system with 1,2-dipalmitoyl phosphatidylcholine (DPPC) have been investigated by differential scanning calorimetry (DSC) and x-ray diffraction. By DSC hydrated GM1 undergoes a broad endothermic transition TM = 26 degrees C (delta H = 1.7 kcal/mol GM1). X-ray diffraction below (-2 degrees C) and above (51 degrees C) this transition indicates a micellar structure with changes occurring only in the wide angle region of the diffraction pattern (relatively sharp reflection at 1/4.12 A-1 at -2 degrees C; more diffuse reflection at 1/4.41 A-1 at 51 degrees C). In hydrated binary mixtures with DPPC, incorporation of GM1 (0-30 mol%; zone 1) decreases the enthalpy of the DPPC pretransition at low molar compositions while increasing the TM of both the pre- and main transitions (limiting values, 39 and 44 degrees C, respectively). X-ray diffraction studies indicate the presence of a single bilayer gel phase in zone 1 that can undergo chain melting to an L alpha bilayer phase. A detailed hydration study of GM1 (5.7 mol %)/DPPC indicated a conversion of the DPPC bilayer gel phase to an infinite swelling system in zone 1 due to the presence of the negatively charged sialic acid moiety of GM1. At 30-61 mol % GM1 (zone 2), two calorimetric transitions are observed at 44 and 47 degrees C, suggesting the presence of two phases. The lower transition reflects the bilayer gel --> L alpha transition (zone 1), whereas the upper transition appears to be a consequence of the formation of a nonbilayer, micellar or hexagonal phase, although the structure of this phase has not been defined by x-ray diffraction. At > 61 mol % GM1 (zone 3) the calorimetric and phase behavior is dominated by the micelle-forming properties of GM1; the presence of mixed GM1/DPPC micellar phases is predicted.     

Effects of cholesterol on phospholipid membranes: inhibition of the interdigitated gel phase of F-DPPC and F-DPPC/DPPC

Unlike the parent phospholipid, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), the monofluorinated analog, 1-palmitoyl-2-(16-fluoropalmitoyl)sn-glycero-3-phosphocholine (F-DPPC), spontaneously forms an interdigitated gel phase (L(β)I) below the main transition temperature (T(m)). We have examined the effects of introducing cholesterol to F-DPPC and 1:1 F-DPPC/DPPC membranes using a combination of DSC, optical density, fluorescence intensity and polarization, (31)P NMR, and X-ray diffraction techniques. Cholesterol increases the fluidity of the gel phase, broadens the main transition, and decreases the main transition enthalpy. However, these results also reveal that there is an unusually large degree of phase coexistence between the L(β)I and non-interdigitated gel phases when cholesterol is added. Cholesterol encourages this phase segregation by partitioning into the thicker non-interdigitated domains. At higher cholesterol concentrations, the majority or all of the L(β)I phase of F-DPPC and 1:1 F-DPPC/DPPC is eliminated and is replaced by a non-interdigitated liquid-ordered (l(o)) phase with properties similar to DPPC/cholesterol. Consequently, cholesterol mitigates the influence the CF moiety has on the thermodynamic phase behavior of F-DPPC. Our findings demonstrate that there are multiple characteristics of cholesterol-rich membranes that disfavor interdigitation.     

Phase diagram of stigmasterol-dipalmitoylphosphatidylcholine mixtures dispersed in excess water

As a simple model of rafts in plant cells, the effect of stigmasterol, one of the predominant sterols in plant plasma membranes, on the phase behavior of dipalmitoylphosphatidylcholine (DPPC) multilayers has been studied by X-ray diffraction (XRD), differential scanning calorimetry (DSC), and freeze-fracture electron microscopy (FFEM) techniques. A partial phase diagram of the binary system has been constructed. Particularly, the stigmasterol concentrations of the "left endpoint" and "right endpoint" of the three-phase line have been determined using the newly developed linear and nonlinear fitting method. They are 6.2 and 23.7 mol%, respectively. Furthermore, the resemblance and difference of phase diagrams of DPPC/stigmasterol, DPPC/cholesterol, and DPPC/ergosterol have been compared and the efficiency of these sterols in promoting the formation of the liquid-ordered domains (rafts) have also been discussed.     

Biophysical perturbations induced by ethylazinphos in lipid membranes

Perturbations induced by ethylazinphos on the physical organization of dipalmitoylphosphatidylcholine (DPPC) and DPPC/cholesterol membranes were studied by differential scanning calorimetry (DSC) and fluorescence polarization of 2-, 6-, 12-(9-anthroyloxy) stearic acids and 16-(9-anthroyloxy) palmitic acid. Ethylazinphos (50 and 100 microM) increases the fluorescence polarization of the probes, either in the gel or in the fluid phase of DPPC bilayers, and this concentration dependent effect decreases from the surface to the bilayer core. Additionally, the insecticide displaces the phase transition to a lower temperature range and broadens the transition profile of DPPC. A shifting and broadening of the phase transition is also observed by DSC. Furthermore at insecticide/lipid molar ratios higher than 1/7, DSC thermograms, in addition to the normal transition centered at 41 degrees C, also display a new phase transition centered at 45.5 degrees C. The enthalpy of this new transition increases with insecticide concentration, with a corresponding decrease of the main transition enthalpy. Ethylazinphos in DPPC bilayers with low cholesterol (< or = 20 mol%) perturbs the membrane organization as described above for pure DPPC. However, cholesterol concentrations higher than 20 mol% prevent insecticide interaction, as revealed by fluorescence polarization and DSC data. Apparently, cholesterol significantly modulates insecticide interaction by competition for similar distribution domains in the membrane. The present results strongly support our previous hypothesis that ethylazinphos locates in the cooperativity region, i.e. the region of C1-C9 atoms of the acyl chains, and extends to the lipid-water interface, where it increases lipid packing order sensed across all the thickness of the bilayer. Additionally, and, on the basis of DSC data, a lateral regionalization of ethylazinphos is here tentatively suggested.     

Phase diagram of stigmasterol-dipalmitoylphosphatidylcholine mixtures dispersed in excess water

As a simple model of rafts in plant cells, the effect of stigmasterol, one of the predominant sterols in plant plasma membranes, on the phase behavior of dipalmitoylphosphatidylcholine (DPPC) multilayers has been studied by X-ray diffraction (XRD), differential scanning calorimetry (DSC), and freeze-fracture electron microscopy (FFEM) techniques. A partial phase diagram of the binary system has been constructed. Particularly, the stigmasterol concentrations of the “left endpoint” and “right endpoint” of the three-phase line have been determined using the newly developed linear and nonlinear fitting method. They are 6.2 and 23.7 mol%, respectively. Furthermore, the resemblance and difference of phase diagrams of DPPC/stigmasterol, DPPC/cholesterol, and DPPC/ergosterol have been compared and the efficiency of these sterols in promoting the formation of the liquid-ordered domains (rafts) have also been discussed.