胰高血糖素样肽-2对小鼠小肠缺血/再灌注损伤的保护作用

目的:观察胰高血糖素样肽-2(GLP-2)对缺血/再灌注损伤小鼠小肠的保护效应.方法:采用肠缺血/再灌注(I/R)模型,将32只小鼠随机分为4组(n=8)假手术(Sham)组、I/R组、I/R GLP-2保护组和I/R 谷氨酰胺(GLN)阳性对照组.光镜观察小肠黏膜形态学改变.检测小肠绒毛高度和隐窝深度;小肠组织二胺氧化酶(DAO)活性;肠系膜淋巴结(MLN)细菌易位率.结果:与假手术组相比,I/R组部分小肠绒毛坏死脱落,绒毛高度下降,隐窝变浅(P＜0 01);小肠组织DAO活性降低(P＜0.01);MLN细菌易位率增加(P＜0.05).与I/R组比,GLP-2组肠绒毛损害明显减轻,DAO活性回升(P＜0.01),细菌易位率回降(P＜0.05).结论:GLP-2对缺血/再灌注损伤小鼠小肠的形态结构及肠屏障功能具有保护作用.     

胰高血糖素样肽-2研究的新进展

胰高血糖素样肽-2(glucagon-like peptide-2,GLP-2)是胰高血糖素原基因转录、翻译后处理加工的33氨基酸的多肽,GLP-2经二酰肽酶Ⅳ水解后,则失去生物学活性。GLP-2作为一种肠上皮特异性生长因子,能促进正常肠黏膜的生长及损伤肠上皮的修复。GLP-2通过作用于GLP-2受体(GLP-2R)来发挥生物学作用。GLP-2R在肠道的广泛分布(肠上皮细胞、肠内在神经元、肠内分泌细胞、肠黏膜下的肌纤维母细胞),提示GLP-2可能通过直接、间接等多条途径发挥生物学作用。本文概括介绍GLP-2的特性、生理作用及机制等方面的研究进展。     

下丘脑室旁核注射GLP-1 对糖尿病大鼠中枢GLP-1 受体表达

目的:探讨下丘脑室旁核(hypothalamic paraventricular nucleus,PVN)注射GLP-1(胰高血糖素样肽-1)对糖尿病大鼠胃排空的影响及机制。方法:30只Wistar大鼠随机分为正常对照组(NC组)、糖尿病组(DM组)和GLP-1干预组(GLP-1组),每组各10只。DM组和GLP-1组腹腔注射链脲佐菌素,三组大鼠均PVN区埋置套管,恢复7d,GLP-1组微量注射0.5μg/0.5μl的GLP-1,NC组和DM组大鼠PVN区微量注射等体积生理盐水。甲基纤维素-酚红灌胃法检测胃排空;半定量RT-PCR检测大鼠下丘脑GLP-1RmRNA的表达。结果:DM组胃排空率较NC组明显升高(P<0.05),GLP-1组胃排空明显低于DM组(P<0.05),GLP-1组和NC组差异无统计学意义(P>0.05)。GLP-1组下丘脑GLP-1RmRNA的表达明显高于DM组和NC组(P<0.05),并与胃排空率成负相关(P<0.05)。DM组和NC组差异无统计学意义(P>0.05)。结论:PVN区注射GLP-1可以抑制糖尿病大鼠早期胃排空加速,作用机制可能和促进下丘脑GLP-1受体表达有关。     

室旁核注射胰高血糖素样肽-1对不同病程糖尿病大鼠胃排空的影响（英文）

目的：研究下丘脑室旁核（paraventricular nucleus,PVN）注射胰高血糖素样肽-1（GLP-1）对糖尿病早期大鼠胃排空的影响,并探讨其相关作用机制。方法：60只清洁级雄性Wistar大鼠随机分为正常对照组（NC组）,糖尿病组（DM组）,GLP-1干预组（GLP-1组）,每组各20只,后两组腹腔注射链脲佐菌素（STZ）制备糖尿病模型,分别于注射STZ2周、6周后每组随机取半数进行实验,实验前于无菌条件下大鼠一侧下丘脑PVN区埋置套管,GLP-1组经套管注入GLP-1,NC组及DM组注入等体积生理盐水。酚红灌胃法检测胃排空率,酶联免疫吸附法（ELISA）测定血浆GLP-1浓度,半定量RT-PCR法测定胃窦、胃底GLP-1RmRNA表达。结果：注射STZ 2周后,DM组较NC组胃排空率显著升高（P〈0.01）。GLP-1组胃排空率低于DM组（P〈0.01）,血浆GLP-1浓度高于DM组及NC组（P均〈0.05）,胃窦GLP-1RmRNA表达明显高于DM组、NC组（P均〈0.01）。注射STZ 6周后,DM组胃排空率高于NC组（P〈0.01）。GLP-1组较DM组胃排空率显著降低（P〈0.01）,血浆GLP-1浓度、胃窦GLP-1RmRNA表达显著高于DM组、NC组（P均〈0.01）。结论：下丘脑PVN区注射GLP-1后,可减慢糖尿病大鼠初期加速的胃排空,原因可能与血浆GLP-1浓度及胃窦GLP-1RmRNA表达增加有关。     

室旁核注射胰高血糖素样肽-1对不同病程糖尿病大鼠胃排空的影响

目的:研究下丘脑室旁核(paraventricular nucleus,PVN)注射胰高血糖素样肽-1(GLP-1)对糖尿病早期大鼠胃排空的影响,并探讨其相关作用机制.方法:60只清洁级雄性Wistar大鼠随机分为正常对照组(NC组),糖尿病组(DM组),GLP-1干预组(GLP-1组),每组各20只,后两组腹腔注射链脲佐菌素(STZ)制备糖尿病模型,分别于注射STZ2周、6周后每组随机取半数进行实验,实验前于无菌条件下大鼠一侧下丘脑PVN区埋置套管,GLP-1组经套管注入GLP-1,NC组及DM组注入等体积生理盐水.酚红灌胃法检测胃排空率,酶联免疫吸附法(ELISA)测定血浆GLP-1浓度,半定量RT-PCR法测定胃窦、胃底GLP-1RmRNA表达.结果:注射STZ2周后,DM组较NC组胃排空率显著升高(P＜0.01).GLP-1组胃排空率低于DM组(P＜0.01),血浆GLP-1浓度高于DM组及NC组(P均＜0.05),胃窦GLP-1RmRNA表达明显高于DM组、NC组(P均＜0.01).注射STZ 6周后,DM组胃排空率高于NC组(P＜0.01).GLP-1组较DM组胃排空率显著降低(P＜0.01),血浆GLP-1浓度、胃窦GLP-1RmRNA表达显著高于DM组、NC组(P均＜0.01).结论:下丘脑PVN区注射GLP-1后,可减慢糖尿病大鼠初期加速的胃排空,原因可能与血浆GLP-1浓度及胃窦GLP-1RmRNA表达增加有关.     

一种GLP-1过表达肠类器官构建的方法

胰高血糖素样肽1(glucagon-like peptide 1,GLP-1)作为一种肠促胰岛素,主要由肠道L细胞分泌,由于其能够有效促进胰岛素的释放从而降低血糖,因此GLP-1及其类似物在2型糖尿病的治疗上具有良好的应用前景.本研究优化了慢病毒感染类器官的方法,利用该方法成功构建了GLP-1过表达的小鼠小肠类器官(o...     

室旁核注射GLP-1 及拮抗剂对大鼠胃组织nesfatin-1 表达的影响

目的:探讨下丘脑室旁核(pareventricular,PVN)注射胰高血糖素样肽-1(GLP-1)及其受体拮抗剂Exendin(9-39)后胃组织核组蛋白2(NUCB2)/nesfatin-1表达的影响。方法:选取48只雄性Wistar大鼠,随机分为6组,生理盐水组,四种不同剂量GLP-1组(0.003 nmol/10μL,0.03 nmol/10μL,0.3 nmol/10μL,3 nmol/10μL),30 nmol Exendin(9-39)+3 nmol GLP-1(E+G)组,每组8只。PVN区埋置套管并按每组要求分别经套管给予GLP-1及Exendin(9-39)等药物。给药2小时后处死大鼠并取胃组织,实时荧光定量RT-PCR法检测各组胃组织NUCB2 m RNA表达。另外生理盐水组,3 nmo L GLP-1组及E+G组每组分别随机取6只大鼠的部分胃组织,用免疫组织化学法测胃粘膜NUCB2/nesfatin-1蛋白的表达情况。结果:实时荧光定量RT-PCR法发现3 nmo L GLP-1组大鼠胃组织NUCB2 m RNA表达量高于生理盐水组,差异有统计学意义(P0.05),而其余各组大鼠胃组织NUCB2 m RNA表达与生理盐水组比较无统计学差异(P0.05)。免疫组化结果显示3 nmo L GLP-1组胃粘膜NUCB2/nesfatin-1蛋白表达与生理盐水组、E+G组比较有统计学差异(P0.05),生理盐水组大鼠胃粘膜NUCB2/nesfatin-1蛋白表达与E+G组比较无明显差异(P0.05)。结论:PVN注射GLP-1能够促进胃组织NUCB2/nesfatin-1的表达,这一作用可能是通过激活GLP-1受体来完成的。     

CFTR基因在小鼠肠道组织中的差异表达

目的研究肠道组织CFTR基因表达与分泌性腹泻发生的关系。方法选取KM小鼠24只,雌雄各半,随机分为3组（每组8只）：对照组经小鼠腹腔注射0.2 mL生理盐水,实验组小鼠经腹腔注射LPS[6 mg/（kg·bw）]分别作用1 h、8 h,于注射后通过小鼠精神状态、肠道组织形态学判定分泌性腹泻模型的建立,利用荧光定量PCR法检测各段肠道组织CFTR基因的表达。结果 LPS成功诱导小鼠发生了分泌性腹泻;CFTR基因在小鼠十二指肠、空肠、回肠和结肠组织中均有不同的表达丰度,以结肠最高,但各段肠道间差异不显著;与对照组相比,LPS上调了十二指肠、空肠和回肠CFTR基因的转录,下调了结肠CFTR基因的转录。结论提示肠道组织CFTR基因转录水平的上调与LPS诱导分泌性腹泻的发生密切相关,且在各肠段发挥的作用不同,其中空肠在氯离子（Cl-）分泌中发挥主要作用,结肠的作用最弱。     

菌群失调腹泻抗生素造模对小鼠肠黏膜的影响

目的研究抗生素造模对菌群失调腹泻小鼠肠黏膜的影响。方法正常组给予无菌生理盐水0.35m L/(只·次)灌胃,其余各组采用头孢拉定胶囊和硫酸庆大霉素注射液用无菌生理盐水配成浓度为62.5g/L抗生素混合液0.35 m L/(只·次)灌胃。每天2次,连续5 d。造模成功后,采集小鼠空肠、回肠和结肠,分别测量绒毛高度、隐窝深度、淋巴细胞数和肠黏膜厚度,观察肠黏膜情况。结果菌群失调腹泻抗生素造模后,空肠、回肠和结肠的隐窝深度及肠黏膜厚度与正常组相比,差异无统计学意义(P0.05);造模后的结肠淋巴细胞数与正常组相比较,高于正常组,差异有统计学意义(P0.05),造模后的空肠绒毛高度与正常组相比,明显低于正常组,差异有统计学意义(P0.01)。结论菌群失调腹泻抗生素造模使结肠淋巴细胞数增高,空肠绒毛高度降低。     

GFP荧光标记GLP-1受体细胞系的建立

为建立胰高血糖素样肽1受体(glucagon-like peptide 1 receptor,GLP-1R)药物筛选模型,真核表达载体pCMV6/GFP/GLP-1R构建好后,转染至U2OS细胞,转染后的细胞经G418筛选获得单克隆细胞株。该细胞株经Western-blot和GLP-1类似物检测,结果表明GLP-1R在该细胞株高表达并对GLP-1类似物有着高灵敏度与特异性的反应,可以用于GLP-1受体激动剂的筛选。     

Cell Wall,Cytoskeleton, and Cell Expansion in Higher Plants

To accommodate two seemingly contradictory biological roles in plant physiology, providing both the rigid structural support of plant cells and the adjustable elasticity needed for cell expansion, the composition of the plant cell wall has evolved to become an intricate network of cellulosic, hemicellulosic, and pectic polysaccharides and protein. Due to its complexity, many aspects of the cell wall influence plant cell expansion, and many new and insightful observations and technologies are forthcoming. The biosynthesis of cell wall polymers and the roles of the variety of proteins involved in polysaccharide synthesis continue to be characterized. The interactions within the cell wall polymer network and the modification of these interactions provide insight into how the plant cell wall provides its dual function. The complex cell wall architecture is controlled and organized in part by the dynamic intracellular cytoskeleton and by diverse trafficking pathways of the cell wall polymers and cell wall-related machinery. Meanwhile, the cell wall is continually influenced by hormonal and integrity sensing stimuli that are perceived by the cell. These many processes cooperate to construct, maintain, and manipulate the intricate plant cell wall--an essential structure for the sustaining of the plant stature, growth, and life.     

Dissecting the Heat Stress Response in Chlamydomonas by Pharmaceutical and RNAi Approaches Reveals Conserved and Novel Aspects

To study how conserved fundamental concepts of the heat stress response （HSR） are in photosynthetic eukaryotes, we applied pharmaceutical and antisense/amiRNA approaches to the unicellular green alga Chlamydomonas reinhardtii. The Chlamydomonas HSR appears to be triggered by the accumulation of unfolded proteins, as it was induced at ambient temperatures by feeding cells with the arginine analog canavanine. The protein kinase inhibitor staurosporine strongly retarded the HSR, demonstrating the importance of phosphorylation during activation of the HSR also in Chlamydomonas. While the removal of extracellular calcium by the application of EGTA and BAPTA inhibited the HSR in moss and higher plants, only the addition of BAPTA, but not of EGTA, retarded the HSR and impaired thermotoler- ance in Chlamydomonas. The addition of cycloheximide, an inhibitor of cytosolic protein synthesis, abolished the attenu- ation of the HSR, indicating that protein synthesis is necessary to restore proteostasis. HSP90 inhibitors induced a stress response when added at ambient conditions and retarded attenuation of the HSR at elevated temperatures. In addition, we detected a direct physical interaction between cytosolic HSP90A/HSP70A and heat shock factor 1, but surprisingly this interaction persisted after the onset of stress. Finally, the expression of antisense constructs targeting chloroplast HSP70B resulted in a delay of the cell＇s entire HSR, thus suggesting the existence of a retrograde stress signaling cascade that is desensitized in HSP7OB-antisense strains.     

Strigolactone-Regulated Hypocotyl Elongation Is Dependent on Cryptochrome and Phytochrome Signaling Pathways in Arabidopsis

Seedling development including hypocotyl elongation is a critical phase in the plant life cycle. Light regula- tion of hypocotyl elongation is primarily mediated through the blue light photoreceptor cryptochrome and red/far-red light photoreceptor phytochrome signaling pathways, comprising regulators including COP1, HY5, and phytochrome- interacting factors （PIFs）. The novel phytohormones, strigolactones, also participate in regulating hypocotyl growth. However, how strigolactone coordinates with light and photoreceptors in the regulation of hypocotyl elongation is largely unclear. Here, we demonstrate that strigolactone inhibition of hypocotyl elongation is dependent on cryp- tochrome and phytochrome signaling pathways. The photoreceptor mutants cry1 cry2, phyA, and phyB are hyposensi- tive to strigolactone analog GR24 under the respective monochromatic light conditions, while cop1 and pifl pif3 pif4 pif5 （pifq） quadruple mutants are hypersensitive to GR24 in darkness. Genetic studies indicate that the enhanced respon- siveness of cop1 to GR24 is dependent on HY5 and MAX2, while that of pifq is independent of HY5. Further studies demonstrate that GR24 constitutively up-regulates HY5 expression in the dark and light, whereas GR24-promoted HY5 protein accumulation is light- and cryptochrome and phytochrome photoreceptor-dependent. These results suggest that the light dependency of strigolactone regulation of hypocotyl elongation is likely mediated through MAX2-dependent promotion of HY5 expression, light-dependent accumulation of HY5, and PIF-regulated components.     

Redox Regulation of Arabidopsis Mitochondrial Citrate Synthase

Citrate synthase has a key role in the tricarboxylic （TCA） cycle of mitochondria of all organisms, as it cata- lyzes the first committed step which is the fusion of a carbon-carbon bond between oxaloacetate and acetyl CoA. The regulation of TCA cycle function is especially important in plants, since mitochondrial activities have to be coordinated with photosynthesis. The posttranslational regulation of TCA cycle activity in plants is thus far almost entirely unexplored. Although several TCA cycle enzymes have been identified as thioredoxin targets in vitro, the existence of any thioredoxin-dependent regulation as known for the Calvin cycle, yet remains to be demonstrated. Here we have investigated the redox regulation of the Arabidopsis citrate synthase enzyme by site-directed mutagenesis of its six cysteine residues. Our results indicate that oxidation inhibits the enzyme activity by the formation of mixed disulfides, as the partially oxidized citrate synthase enzyme forms large redox-dependent aggregates. Furthermore, we were able to demonstrate that thioredoxin can cleave diverse intraas well as intermolecular disulfide bridges, which strongly enhances the activity of the enzyme. Activity measurements with the cysteine variants of the enzyme revealed important cysteine residues affecting total enzyme activity as well as the redox sensitivity of the enzyme.     

Organelle pH in the Arabidopsis Endomembrane System

The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by combining the improved-solubility feature of solubility-modified green fluorescent protein （GFP） （smGFP） with the pH-sensing capabil- ity of the pHluorins and codon optimized for expression in Arabidopsis. PEpHluorin （plant-solubility-modified ecliptic pHluorin） gradually loses fluorescence as pH is lowered with fluorescence vanishing at pH 6.2 and PRpHluorin （plant- solubility-modified ratiomatric pHluorin）, a dual-excitation sensor, allowing for precise measurements. Compartment- specific sensors were generated by further fusing specific sorting signals to PEpHluorin and PRpHluorin. Our results show that the pH of cytosol and nucleus is similar （pH 7.3 and 7.2）, while peroxisomes, mitochondrial matrix, and plastidial stroma have alkaline pH. Compartments of the secretory pathway reveal a gradual acidification, spanning from pH 7.1 in the endoplasmic reticulum （ER） to pH 5.2 in the vacuole. Surprisingly, pH in the trans-Golgi network （TGN） and mul- tivesicular body （MVB） is, with pH 6.3 and 6.2, quite similar. The inhibition of vacuolar-type H＋-ATPase （V-ATPase） with concanamycin A （ConcA） caused drastic increase in pH in TGN and vacuole. Overall, the PEpHluorin and PRpHluorin are excellent pH sensors for visualization and quantification of pH in vivo, respectively.     

How does the host-specialized aphid deal with food deficiency？

Aphis gossypii Glover shows obvious host specialization, with cucurbit- and cotton-specialized biotypes or host races in many regions. Because its annual natal hostcrops senesce earlier the cucurbit-specialized biotype may suffer food deficiency. The method this biotype uses to overcome this challenge is still poorly understood. In orderto understand the potential of the cucurbit-specialized biotype aphids in host shift and usage, the performance of this biotype on cotton （Gossypium hirsutum）, a common butpoor quality host plant, was explored in this study. The cucurbit-specialized aphids could establish populations on cotton only when these plants had at least nine leaves, and subsequent populations developed rather slowly. The presence of whitefly populations on cotton improved the success rate of cucurbit-specialized aphids. The cucurbit-specialized aphidswere mainly distributed on the older leaves of cotton, with only a few settling on the upper leaves. The cucurbit-specialized aphids reared on cotton for 40, 54 and 61 days stillmaintained strong preference for their natal host plant, cucumber （Cucumis sativus）, rather than cotton, and their net reproductive rates and intrinsic rates of natural increase weredramatically lower when they were transferred onto new six-leaf cotton plants or detached leaves. Therefore, we concluded that the cucurbit-specialized aphids have the potentialto utilize mature or whitefly-stressed cotton plants, but that this feeding experience on cotton did not alter their specialization for cucurbits. Some cotton plants could act as atemporary host for the cucurbit-specialized aphids to overcome food deficiency arising from senescing cucurbits.     

Plastid Signals and the Bundle Sheath： Mesophyll Development in Reticulate Mutants

The development of a plant leaf is a meticulously orchestrated sequence of events producing a complex organ comprising diverse cell types. The reticulate class of leaf variegation mutants displays contrasting pigmentation between veins and interveinal regions due to specific aberrations in the development of mesophyll cells. Thus, the reticulate mutants offer a potent tool to investigate cell-type-specific developmental processes. The discovery that most mutants are affected in plastid-localized, metabolic pathways that are strongly expressed in vasculature-associated tis- sues implicates a crucial role for the bundle sheath and their chloroplasts in proper development of the mesophyll cells. Here, we review the reticulate mutants and their phenotypic characteristics, with a focus on those in Arabidopsis thali- ana. Two alternative models have been put forward to explain the relationship between plastid metabolism and meso- phyll cell development, which we call here the supply and the signaling hypotheses. We critically assess these proposed models and discuss their implications for leaf development and bundle sheath function in C3 species. The characteriza- tion of the reticulate mutants supports the significance of plastid retrograde signaling in cell development and highlights the significance of the bundle sheath in C3 photosynthesis.     

Knights in Action： Lectin Receptor-Like Kinases in Plant Development and Stress Responses

The Receptor-Like Kinase （RLK） is a vast protein family with over 600 genes in Arabidopsis and 1100 in rice. The Lectin RLK （LecRLK） family is believed to play crucial roles in saccharide signaling as well as stress perception. All the LecRLKs possess three domains： an N-terminal lectin domain, an intermediate transmembrane domain, and a C-terminal kinase domain. On the basis of lectin domain variability, LecRLKs have been subgrouped into three subclasses： L-, G-, and C-type LecRLKs. While the previous studies on LecRLKs were dedicated to classification, comparative structural analysis and expression analysis by promoter-based studies, most of the recent studies on LecRLKs have laid special emphasis on the potential of this gene family in regulating biotic/abiotic stress and developmental pathways in plants, thus mak- ing the prospects of studying the LecRLK-mediated regulatory mechanism exceptionally promising. In this review, we have described in detail the LecRLK gene family with respect to a historical, evolutionary, and structural point of view. Furthermore, we have laid emphasis on the LecRLKs roles in development, stress conditions, and hormonal response. We have also discussed the exciting research prospects offered by the current knowledge on the LecRLK gene family. The multitude of the LecRLK gene family members and their functional diversity mark these genes as both interesting and worthy candidates for further analysis, especially in the field of crop improvement.