2型糖尿病患者糖化血红蛋白与血糖相关性分析

目的:探讨2型糖尿病患者糖化血红蛋白(Hb A1c)与空腹血糖(FPG)检测结果的相关性。方法:选取2015年1月至2015年6月在门诊确诊的152例2型糖尿病患者作为研究对象,分别用葡萄糖氧化酶法检测FPG,用高效液相色谱法检测Hb A1c。取Hb A1c7%为A组;Hb A1c7-10%为B组;Hb A1c10%为C组,将与其相对应的FPG值进行相关性分析。结果:152例总体FPG(8.96±3.54)mmol/L;Hb A1c(7.87±2.16)%,r(总)=0.777684;其中152例2型糖尿病患者中,A组66例,FPG(6.82±1.06)mmol/L、Hb A1c(6.17±0.50)%,r(A)=0.33668(P0.05);B组62例,FPG(9.16±2.35)mmol/L、Hb A1c(8.15±0.95)%,r(B)=0.40925(P0.05);C组24例,FPG(14.34±4.56)mmol/L、Hb A1c(11.83±1.67)%,r(C)=0.43877(P0.05)。其Hb A1c与FPG的结果分析显示,随着FPG水平增高,Hb A1c也相应升高,二者呈正相关。结论:联合检测血糖和Hb A1c对糖尿病的治疗、预后判定具有显著意义,是评价血糖控制方案的金标准,建议定期进行检测。     

妊娠糖尿病患者血清HbA1c与CRP水平的相关性分析

目的:为明确妊娠期糖尿病(gestational diabetes mellitus,GDM)患者血清糖化血红蛋白(glycosylated hemoglobin,Hb A1c)与C反应蛋白(C-reactive protein,CRP)的关系,本研究检测了GDM患者血糖、血清Hb A1c与CRP水平,并对Hb A1c与CRP的相关性进行了分析。方法:以68例2015年4月至2017年4月于我院诊治的GDM孕妇为研究对象,所有患者均符合《妇产科学》中关于GDM的诊断标准,另选取68例正常孕妇为对照组。采用葡萄糖氧化酶法检测血糖水平(空腹血糖及餐后2 h血糖水平),采用免疫凝集法检测和比较两组血清糖化血红蛋白(HbA1c)水平,采用免疫透射比浊法检测血清C反应蛋白(CRP)水平,并分析HbA1c与CRP的相关性。结果:GDM组患者空腹血糖(fasting plasma glucose,FPG)、2 h血糖(plasma glucose,PG)、血清HbA1c和CRP水平均显著高于正常组(P0.05),GDM患者血清Hb A1c和CRP水平呈显著正相关关系(r=0.654,P0.05)。结论:GDM患者血清HbAlC和CRP水平相较于正常孕妇有显著提高,且二者呈显著的正相关关系,二者联合检测可能作为GDM早期诊断的筛查的重要参考指标。     

空腹血葡萄糖及糖化血红蛋白在慢性肝病患者中诊断糖尿病的对比研究

目的:比较空腹血葡萄糖(FPG)和糖化血红蛋白(HbA1c)在肝病人群中糖尿病(DM)诊断中的意义.方法:对142例未诊断为DM的对象同时检测FPG和HbA1c.分别按1999年世界卫生组织(WHO)诊断糖尿病标准(FPG-≥ 7.0 mmol/L)和美国糖尿病协会(ADA)新标准(HbA1c≥ 6.5％),对研究对象进行分组(DM和非DM组),比较2个检测指标在诊断DM中的差异.以FPG为诊断DM的金标准,计算HbA1c诊断DM的灵敏度、特异度、预测值、似然比等诊断性能.结果:FPG均值在2种标准诊断的DM患者之间差异有统计学意义(P＜0.05),在2种标准诊断的非DM患者之间差异则无统计学意义(P＞0.05).HbA1c均值在2种标准诊断的DM患者间差异无统计学意义(P＞0.05),在2种标准诊断的非DM患者间差异无统计学意义(P＞0.05).HbA1c≥6.5％诊断糖尿病的灵敏度(S)为100％.特异度(Sp)为93％,阴性似然比(-LR)为0.00和阳性似然比(+LR)为14.29.结论:HbA1c在肝病人群中诊断糖尿病有较高的诊断灵敏度和特异度.     

胰岛素联合六味地黄丸治疗老年早期糖尿病肾病患者的临床疗效研究

目的:探讨胰岛素联合六味地黄丸治疗老年早期糖尿病肾病患者的临床疗效。方法:收集我院收治的老年早期糖尿病肾病患者100例,随机分为对照组和实验组,每组各50例,对照组患者血糖情况给予胰岛素1次~2次/d,实验组患者在此基础上给予六味地黄丸1丸/次,2次/d,口服。连续4周。治疗结束后,对患者血清糖基化血红蛋白(Hb A1c)、同型半胱氨酸(HCY)、胱抑素C(Cys-C)水平的变化及临床疗效进行检测并比较。结果:与治疗前相比,两组患者的Hb A1c、HCY、Cys-C均下降(P0.05);与对照组相比,实验组患者Hb A1c、HCY、Cys-C水平较低(P0.05),实验组患者治疗总有效率较高(P0.05)。结论:胰岛素联合六味地黄丸能够有效改善老年早期糖尿病肾病患者的糖稳态及肾功能,临床疗效较好。     

2型糖尿病患者糖化血红蛋白水平与颈动脉内-中膜厚度的相关性

目的:分析2型糖尿病(T2DM)患者糖化血红蛋白(Hb A1c)水平与颈动脉内-中膜厚度(CIMT)的相关性。方法:选择在我院内分泌科住院的T2DM患者328名,对入组患者进行Hb A1c、血生化指标检测以及CIMT测量等。根据CIMT值分为CIMT正常组(0.9 mm)和CIMT增厚组(0.9 mm),并对CIMT的相关危险因素进行多因素Logistic回归分析。结果:(1)328名T2DM患者中,CIMT正常154例,CIMT增厚174例;(2)Pearson相关分析显示,总胆固醇(TC)、Hb A1c水平与IMT值呈正相关(P0.05)。(3)单因素分析示,CIMT正常组和CIMT增厚组两组间年龄(t=4.132,P=0.041)、收缩压(t=8.456,P0.01)、Hb A1c≥9.0%(x~2=9.912,P0.01)、总胆固醇(t=5.549,P=0.018)、甘油三酯(t=6.592,P=0.008)、尿酸(t=9.618,P0.01)、空腹血糖(t=4.592,P=0.037)间差异有统计学意义;(4)多因素Logistic回归分析示,年龄、Hb A1c≥9%、收缩压、总胆固醇是T2DM患者CIMT增厚的独立危险因素(P0.05)。结论:Hb A1c与CIMT增厚明显相关;且Hb A1c≥9%是CIMT增厚的独立危险因素。     

检测指标对糖尿病慢性肾脏疾病诊断率的影响

为研究常见临床检测指标糖尿病慢性肾脏疾病诊断率的影响,本次研究选取了143例Ⅱ型糖尿病患者,按照尿白蛋白排泄率(UAER)将患者则会分为A组(单纯糖尿病组)、B组(合并早期慢性肾病组,UAER大于20μg/min,且小于200μg/min)、C组(合并临床期慢性肾病组,UAER大于200μg/min)。检测患者糖化血红蛋白(Hb A1c)、血尿素氮(BUN)、血肌酐(Scr)、血清尿酸(SUA)、血清甘油三酯(TG)、血清总胆固醇(TC)、肾小球过滤率(GFR)、尿转铁蛋白(TF)以及身体质量指数(BMI),并分析上述指标与糖尿病慢性肾脏疾病的关系。研究显示,TC、TF是与糖尿病慢性肾病关系最为密切的危险因素(p0.05),且与UAER具有正相关性(p0.05);TC、TF联合诊断曲线下面积(AUC)为0.96。这提示TC、TF两种指标联合检测可能会提高糖尿病慢性肾病的诊断率。     

糖化血红蛋白与糖尿病及其并发症的关系

目的:探讨糖化血红蛋白(HbAlc)与糖尿病诊断、疗效评价及并发症的关系。方法:选择2型糖尿病患者250例和健康体检者150例,分别测定空腹血糖(FPG)、2h血糖(2hPG)及糖化血红蛋白(HbA1c),统计学分析HbA1c与FPG、2hPG的相关性;分析HbA1c与糖尿病并发症发生的关系。结果:糖尿病组FPG、2hPG及HbA1c水平均显著高于对照组(P<0.01);糖尿病伴有并发症患者的HbAlc明显高于无并发症者(P<0.05),HbA1c水平与糖尿病并发症的发生率存在高度相关性(P<0.01)。结论:检测外周血中HbA1c水平对2型糖尿病诊断、疗效评价具有重要临床价值,控制糖化血红蛋白对预防糖尿病并发症的发生具有重要意义。     

2 型糖尿病患者凝血功能水平与血管病变发生的相关性

目的:探讨2型糖尿病患者凝血功能及血清肿瘤坏死因子α(TNF-α)及白细胞介素18(IL-18)水平与糖尿病血管病变的关系。方法:选择2014年6月-2015年10月在我院接受治疗的2型糖尿病患者83例作为研究对象,根据患者疾病进展情况将其分为血管病变组(45例)和无血管病变组(38例)。测定两组患者凝血酶时间(TT)、活化部分凝血酶原时间(APTT)、凝血酶原时间(PT)、纤维蛋白原(Fg)、血糖(Glu)、糖化血红蛋白(Hb A1c)、肿瘤坏死因子α(TNF-α)及白细胞介素18(IL-18)水平;采用logistics回归分析2型糖尿病患者发生血管病变的危险因素。结果:血管病变组患者Glu,Hb A1c及Fg水平均显著高于无血管病变组,而APTT及PT水平均显著低于无血管病变组,差异均具有统计学意义(P0.05);两组间TT水平比较,差异无统计学意义(P0.05)。血管病变组患者血清中TNF-α及IL-18水平均高于无血管病变组患者,差异具有统计学意义(P0.05)。Hb A1c,Fg,TNF-α以及IL-18水平异常是2型糖尿病患者发生血管病变的独立危险因素(OR=1.23,1.45,2.632,3.884,P0.05)。结论:凝血功能紊乱及炎症反应是2型糖尿病患者发生血管病变的重要因素,临床应给予重视。     

2 型糖尿病与干眼症的相关性研究及其危险因素分析

目的:探讨2型糖尿病和干眼症的关系并分析2型糖尿病发生干眼症的危险因素。方法:纳入2型糖尿病患者220例为观察组和健康人群50例为对照组,采集所有研究对象眼表失衡指数(OSDI)、泪膜破裂时间(TBUT)、泪液分泌试验(SIt),以及观察组性别、年龄、糖尿病病程、血糖、Hb A1c、HOMA-IR、血清CRP,对比分析两组患者干眼症发病率及干眼症症状,采用多因素Logistic回归分析影响2型糖尿病发生干眼症的危险因素。结果:对照组干眼症发病率(9/100,9.00%)明显低于观察组(108/440眼,24.52%)(P0.05)。观察组OSDI评分明显高于对照组(P0.05)。观察组TBUT、SIt明显小于对照组,两组间差异有统计学意义(P0.05)。性别、年龄、糖尿病病程、OSDI、TBUT、SIt、Hb A1c和2型糖尿病患者发生干眼症具有一定的相关性(P0.05)。年龄、糖尿病病程、TBUT、Hb A1c是2型糖尿病患者发生干眼症的危险因素(B0,OR1)。结论:2型糖尿病和干眼症具有一定的相关性,糖尿病患者年龄、糖尿病病程、血糖控制水平是2型糖尿病发生干眼症的独立危险因素。     

芪卫颗粒对2型糖尿病大鼠肾脏氧化应激和病理的影响

目的:研究芪卫颗粒对2型糖尿病大鼠肾脏氧化应激和病理的影响。方法:先诱导2型糖尿病大鼠模型50只,按照随机数字表法将大鼠分为A组和B组,每组25只,另选取25只正常鼠为对照组;A组给予芪卫颗粒,B组给予a-硫辛酸,均治疗3个月,检测3组血糖(BG)、糖化血红蛋白(Hb A1c)、血脂、肾功能指标、超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽氧化物酶(GSH-Px),并观察肾脏的病理变化。结果:A组BG、Hb A1c和血脂水平均显著优于B组,差异有统计学意义(P0.05);A组肾功能指标与对照组比较无统计学意义(P0.05),A组肾功能指标显著优于B组,差异有统计学意义(P0.05);A组SOD、MDA和GSH-Px均优于B组,差异有统计学意义(P0.05);A组肾小球病理变化显著优于B组,差异有统计学意义(P0.05)。结论:芪卫颗粒对2型糖尿病大鼠肾脏氧化应激有一定抑制作用,能改善肾功能和病理。     

Post-translational Regulation of the Type III Inositol 1,4,5-Trisphosphate Receptor by miRNA-506

The type III isoform of the inositol 1,4,5-trisphosphate receptor (InsP₃R3) is apically localized and triggers Ca²⁺ waves and secretion in a number of polarized epithelia. However, nothing is known about epigenetic regulation of this InsP3R isoform. We investigated miRNA regulation of InsP₃R3 in primary bile duct epithelia (cholangiocytes) and in the H69 cholangiocyte cell line, because the role of InsP₃R3 in cholangiocyte Ca²⁺ signaling and secretion is well established and because loss of InsP₃R3 from cholangiocytes is responsible for the impairment in bile secretion that occurs in a number of liver diseases. Analysis of the 3′-UTR of human InsP₃R3 mRNA revealed two highly conserved binding sites for miR-506. Transfection of miR-506 mimics into cell lines expressing InsP₃R3–3′UTR-luciferase led to decreased reporter activity, whereas co-transfection with miR-506 inhibitors led to enhanced activity. Reporter activity was abrogated in isolated mutant proximal or distal miR-506 constructs in miR-506-transfected HEK293 cells. InsP₃R3 protein levels were decreased by miR-506 mimics and increased by inhibitors, and InsP₃R3 expression was markedly decreased in H69 cells stably transfected with miR-506 relative to control cells. miR-506-H69 cells exhibited a fibrotic signature. In situ hybridization revealed elevated miR-506 expression in vivo in human-diseased cholangiocytes. Histamine-induced, InsP₃-mediated Ca²⁺ signals were decreased by 50% in stable miR-506 cells compared with controls. Finally, InsP₃R3-mediated fluid secretion was significantly decreased in isolated bile duct units transfected with miR-506, relative to control IBDU. Together, these data identify miR-506 as a regulator of InsP₃R3 expression and InsP₃R3-mediated Ca²⁺ signaling and secretion.     

Identification of Key Residues Determining Species Differences in Inhibitor Binding of Microsomal Prostaglandin E Synthase-1

Microsomal prostaglandin E synthase-1 (MPGES1) is induced during an inflammatory reaction from low basal levels by pro-inflammatory cytokines and subsequently involved in the production of the important mediator of inflammation, prostaglandin E₂. Nonsteroidal anti-inflammatory drugs prevent prostaglandin E₂ production by inhibiting the upstream enzymes cyclooxygenases 1 and 2. In contrast to these conventional drugs, a new generation of NSAIDs targets the terminal enzyme MPGES1. Some of these compounds potently inhibit human MPGES1 but do not have an effect on the rat orthologue. We investigated this interspecies difference in a rat/human chimeric form of the enzyme as well as in several mutants and identified key residues Thr-131, Leu-135, and Ala-138 in human MPGES1, which play a crucial role as gate keepers for the active site of MPGES1. These residues are situated in transmembrane helix 4, lining the entrance to the cleft between two subunits in the protein trimer, and regulate access of the inhibitor in the rat enzyme. Exchange toward the human residues in rat MPGES1 was accompanied with a gain of inhibitor activity, whereas exchange in human MPGES1 toward the residues found in rat abrogated inhibitor activity. Our data give evidence for the location of the active site at the interface between subunits in the homotrimeric enzyme and suggest a model of how the natural substrate PGH₂, or competitive inhibitors of MPGES1, enter the active site via the phospholipid bilayer of the membrane.     

Measurement of Metabolic Rate in Drosophila using Respirometry

Metabolic disorders are a frequent problem affecting human health. Therefore, understanding the mechanisms that regulate metabolism is a crucial scientific task. Many disease causing genes in humans have a fly homologue, making Drosophila a good model to study signaling pathways involved in the development of different disorders. Additionally, the tractability of Drosophila simplifies genetic screens to aid in identifying novel therapeutic targets that may regulate metabolism. In order to perform such a screen a simple and fast method to identify changes in the metabolic state of flies is necessary. In general, carbon dioxide production is a good indicator of substrate oxidation and energy expenditure providing information about metabolic state. In this protocol we introduce a simple method to measure CO₂ output from flies. This technique can potentially aid in the identification of genetic perturbations affecting metabolic rate.     

Structural basis of pre-mRNA recognition by the human cleavage factor Im complex

The cleavage factor I(m) (CF I(m)), consists of a 25 kDa subunit (CF I(m)25) and one of three larger subunits (CF I(m)59, CF I(m)68, CF I(m)72), and is an essential protein complex for pre-mRNA 3'-end cleavage and polyadenylation. It recognizes the upstream sequence of the poly(A) site in a sequence-dependent manner. Here we report the crystal structure of human CF I(m), comprising CF I(m)25 and the RNA recognition motif domain of CF I(m)68 (CF I(m)68RRM), and the crystal structure of the CF I(m)-RNA complex. These structures show that two CF I(m)68RRM molecules bind to the CF I(m)25 dimer via a novel RRM-protein interaction mode forming a heterotetramer. The RNA-bound structure shows that two UGUAA RNA sequences, with anti-parallel orientation, bind to one CF I(m)25-CF I(m)68RRM heterotetramer, providing structural basis for the mechanism by which CF I(m) binds two UGUAA elements within one molecule of pre-mRNA simultaneously. Point mutation and kinetic analyses demonstrate that CF I(m)68RRM can bind the immediately flanking upstream region of the UGUAA element, and CF I(m)68RRM binding significantly increases the RNA-binding affinity of the complex, suggesting that CF I(m)68 makes an essential contribution to pre-mRNA binding.     

Comparison of gas chromatography-mass spectrometry,radioimmunoassay and bioassay for the quantification of gibberellin A9 in norway spruce (Picea abies (L.) Karst.)

Quantitative estimates of gibberellin A₉ in Norway spruce extracts obtained by gas chromatography-mass spectrometry, radioimmunoassay (RIA_ and bioassay were compared after successive purifications of the extracts. The extracts were assayed in several dilutions with and without the addition of standard gibberellin A₉, thus showing the effect of extract components on the response of the assays. Radioimmunoassay produced estimates comparable to gas chromatography-mass spectrometry after one purification step by high-performance liquid chromatography. Extracts purified by polyvinylpyrrolidone-column chromatography and solvent partitioning but not high-performance liquid chromatography resulted in inaccurate RIA estimates. The performance of the RIA could be monitored by logit-log transformations of the standard curve and extract dilution curve and by calculating the slope of the standard addition curve. It was, however, not possible to correct for the interference caused by extract components by the standard addition procedure. Quantifications by Tan-ginbozu dwarf-rice bioassay were accurate, but a large and unpredictable variation makes it unsuitable for quantitative determinations.Abbreviations FW fresh weight - GA₉ gibberellin A₉ - GA₉–Me methylated GA₉ - GC-MS gas chromatography-mass spectrometry - HPLC high performance liquid chromatography - MID multiple-ion detection - RIA radioimmunoassay     

Differential Regulation of Multiple Steps in Inositol 1,4,5-Trisphosphate Signaling by Protein Kinase C Shapes Hormone-stimulated Ca2+ Oscillations

How Ca²⁺ oscillations are generated and fine-tuned to yield versatile downstream responses remains to be elucidated. In hepatocytes, G protein-coupled receptor-linked Ca²⁺ oscillations report signal strength via frequency, whereas Ca²⁺ spike amplitude and wave velocity remain constant. IP₃ uncaging also triggers oscillatory Ca²⁺ release, but, in contrast to hormones, Ca²⁺ spike amplitude, width, and wave velocity were dependent on [IP₃] and were not perturbed by phospholipase C (PLC) inhibition. These data indicate that oscillations elicited by IP₃ uncaging are driven by the biphasic regulation of the IP₃ receptor by Ca²⁺, and, unlike hormone-dependent responses, do not require PLC. Removal of extracellular Ca²⁺ did not perturb Ca²⁺ oscillations elicited by IP₃ uncaging, indicating that reloading of endoplasmic reticulum stores via plasma membrane Ca²⁺ influx does not entrain the signal. Activation and inhibition of PKC attenuated hormone-induced Ca²⁺ oscillations but had no effect on Ca²⁺ increases induced by uncaging IP₃. Importantly, PKC activation and inhibition differentially affected Ca²⁺ spike frequencies and kinetics. PKC activation amplifies negative feedback loops at the level of G protein-coupled receptor PLC activity and/or IP₃ metabolism to attenuate IP₃ levels and suppress the generation of Ca²⁺ oscillations. Inhibition of PKC relieves negative feedback regulation of IP₃ accumulation and, thereby, shifts Ca²⁺ oscillations toward sustained responses or dramatically prolonged spikes. PKC down-regulation attenuates phenylephrine-induced Ca²⁺ wave velocity, whereas responses to IP₃ uncaging are enhanced. The ability to assess Ca²⁺ responses in the absence of PLC activity indicates that IP₃ receptor modulation by PKC regulates Ca²⁺ release and wave velocity.     

Light-induced pH changes in leaves of C4 plants

Light-induced changes in the fluorescence of the pH-indicating dyes pyranine or 5-(and 6-)carboxy-2, 7-dichlorofluorescein (CDCF) which had been fed to leaves were examined to monitor cellular pH changes. After short-term feeding of pyranine (pK 7.3) to leaves of Amaranthus caudatus L., a NAD-malic-enzyme-type C₄ plant, vascular bundles and surrounding cells became fluorescent. Fluorescence emission from mesophyll cells required longer feeding times. In CO₂-free air, pyranine fluorescence increased much more on illumination after mesophyll cells had become fluorescent than when only the vascular bundles and the bundle sheath of Amaranthus leaves had been stained. After short feeding times and in the absence of actinic illumination, CO₂ decreased pyranine fluorescence very slowly in Amaranthus and rapidly in C₃ leaves. After prolonged feeding times, the extent of the light-dependent increase in pyranine fluorescence was several times greater in different C₄ plants than in C₃ species. The kinetics of the fluorescence changes were also remarkably different in C₃ and C₄ plants. Carbon dioxide (500 l · l^–1) suppressed the light-induced increase in pyranine fluorescence more in C₄ than in C₃ leaves. Light-dependent changes in light scattering, which are indicative of chloroplast energization, and in 410-nm transmission, which indicate chloroplast movement, differed kinetically from those of the changes in pyranine fluorescence. Available evidence indicated that light-dependent changes in pyranine fluorescence did not originate from the apoplast of leaf cells. Microscopic observation led to the conclusion that, after prolonged feeding times or prolonged incubation, changes in pyranine fluorescence emitted from C₄ leaves reflect pH changes mainly in the cytosol of mesophyll cells. A transient acidification reaction indicated by quenching of pyranine fluorescence in the dark-light transient and not observed in C₃ species is attributed to the carboxylation of phosphoenolpyruvate. After short feeding times and in the absence of actinic illumination, CO₂ (250 l l^–1) decreased pyranine fluorescence very slowly in Amaranthus and more rapidly in C₃ leaves. After prolonged feeding times, both the rate and the extent of CO₂-dependent quenching of pyranine fluorescence increased, but the increase was insufficient to indicate the presence of highly active carbonic anhydrase in the compartment from which pyranine fluorescence was emitted. In contrast to pyranine, CDCF (pK 4.8) did not increase but rather decreased its fluorescence on illumination of an Amaranthus leaf, indicating acidification of an acidic compartment, most probably the vacuole of green leaf cells. The pattern of the acidification reaction was similar in C₄ and C₃ leaves. The remarkably large extent of the light-dependent increase in pyranine fluorescence from leaves of C₄ species and its slow kinetics are proposed to be caused by an alkalization of the cytosol which in the absence of CO₂ is larger in the mesophyll than in the bundle sheath. It gives rise to deprotonation of dye originally located in the mesophyll and, in addition, of dye which diffuses from the bundle sheath into the mesophyll following a pH gradient. Implications of slow diffusional transport of pyranine and CO₂ between mesophyll and bundle-sheath cells and the fast metabolite transport required in C₄ photosynthesis are discussed.Abbreviations CDCF 5-(and 6-)carboxy-2,7-dichlorofluorescein - DHAP dihydroxyacetone phosphate - PGA 3-phosphoglycerate This work was supported by the Sonderforschungsbereiche 176 and 251 of the University of Würzburg and by the Gottfried-Wilhelm-Leibniz Program of the Deutsche Forschungsgemeinschaft. A.S.R. was the recipient of a fellowship of the Alexander-von-Humboldt Foundation. We are grateful to Mrs. S. Neimanis for cooperation.     

Single particle analysis of thylakoid proteins from Thermosynechococcus elongatus and Synechocystis 6803: localization of the CupA subunit of NDH-1

The larger protein complexes of the cyanobacterial photosynthetic membrane of Thermosynechoccus elongatus and Synechocystis 6803 were studied by single particle electron microscopy after detergent solubilization, without any purification steps. Besides the "standard" L-shaped NDH-1L complex, related to complex I, large numbers of a U-shaped NDH-1MS complex were found in both cyanobacteria. In membranes from Synechocystis DeltacupA and DeltacupA/cupB mutants the U-shaped complexes were absent, indicating that CupA is responsible for the U-shape by binding at the tip of the membrane-bound arm of NDH-1MS. Comparison of membranes grown under air levels of CO(2) or 3% CO(2) indicates that the number of NDH-1MS particles is 30-fold higher under low-CO(2).     

Targeted deletion of LPA5 identifies novel roles for lysophosphatidic acid signaling in development of neuropathic pain

Lysophosphatidic acid (LPA) is a bioactive lipid that serves as an extracellular signaling molecule acting through cognate G protein-coupled receptors designated LPA(1-6) that mediate a wide range of both normal and pathological effects. Previously, LPA(1), a G(αi)-coupled receptor (which also couples to other G(α) proteins) to reduce cAMP, was shown to be essential for the initiation of neuropathic pain in the partial sciatic nerve ligation (PSNL) mouse model. Subsequent gene expression studies identified LPA(5), a G(α12/13)- and G(q)-coupled receptor that increases cAMP, in a subset of dorsal root ganglion neurons and also within neurons of the spinal cord dorsal horn in a pattern complementing, yet distinct from LPA(1), suggesting its possible involvement in neuropathic pain. We therefore generated an Lpar5 null mutant by targeted deletion followed by PSNL challenge. Homozygous null mutants did not show obvious base-line phenotypic defects. However, following PSNL, LPA(5)-deficient mice were protected from developing neuropathic pain. They also showed reduced phosphorylated cAMP response element-binding protein expression within neurons of the dorsal horn despite continued up-regulation of the characteristic pain-related markers Caα(2)δ(1) and glial fibrillary acidic protein, results that were distinct from those previously observed for LPA(1) deletion. These data expand the influences of LPA signaling in neuropathic pain through a second LPA receptor subtype, LPA(5), involving a mechanistically distinct downstream signaling pathway compared with LPA(1).