美洲大蠊(Periplaneta americana L.)抗菌肽杀菌作用初步观察

报道了美洲大蠊(PeriplanetaamericanaL.)抗菌肽对大肠杆菌(EscherichiacoliK88)的作用过程.电镜观察表明,美洲大蠊抗菌肽首先使细菌的外层及细胞质膜损伤,形成开口,导致内容物外泄而死亡,最后菌体崩解成碎片.

Periplaneta Americana Extract Ameliorates LPS-induced Acute Lung Injury Via Reducing Inflammation and Oxidative Stress

Objective Acute lung injury(ALI)is an acute clinical syndrome characterized by uncontrolled inflammation response,which causes high mortality and poor prognosis.The present study determined the protective effect and underlying mechanism of Periplaneta americana extract(PAE)against lipopolysaccharide(LPS)-induced ALI.Methods The viability of MH-S cells was measured by MTT.ALI was induced in BALB/c mice by intranasal administration of LPS(5 mg/kg),and the pathological changes,oxidative stress,myeloperoxidase activity,lactate dehydrogenase activity,inflammatory cytokine expression,edema formation,and signal pathway activation in lung tissues and bronchoalveolar lavage fluid(BALF)were examined by H&E staining,MDA,SOD and CAT assays,MPO assay,ELISA,wet/dry analysis,immunofluorescence staining and Western blotting,respectively.Results The results revealed that PAE obviously inhibited the release of proinflammatory TNF-α,IL-6 and IL-1βby suppressing the activation of MAPK/Akt/NF-κB signaling pathways in LPS-treated MH-S cells.Furthermore,PAE suppressed the neutrophil infiltration,permeability increase,pathological changes,cellular damage and death,pro-inflammatory cytokines expression,and oxidative stress upregulation,which was associated with its blockage of the MAPK/Akt/NF-κB pathway in lung tissues of ALI mice.Conclusion PAE may serve as a potential agent for ALI treatment due to its anti-inflammatory and anti-oxidative properties,which correlate to the blockage of the MAPK/NF-κB and AKT signaling pathways.

Potential Risk of Transmission of Escherichia coli and Salmonella spp. Infections by the Musca domestica Fly and the Periplaneta americana Cockroach in the City of Cotonou (South Benin)

Background: The occurrence of bacterial infections sometimes involves synanthropic flies and cockroaches, as mechanical vectors of pathogenic microorganisms. The aim of this study was to identify the different species of flies and cockroaches that cohabit with humans in the city of Cotonou and determine the contribution of Musca domestica and Periplaneta americana in the transmission of bacteria responsible for infections in humans. Methods: Capture sessions during the day for flies and at night for cockroaches were carried out in 4 arrondissements of the city of Cotonou. The insects collected were transported to the Center of Research Entomological of Cotonou for identification, then sent to the Section Hygiene of Water and Food for microbiological analysis, enumeration and testing for human pathogenic bacteria. Results: A total of 351 flies and 26 cockroaches were collected at the sites, including two synanthropic species belonging to two families each. Musca domestica and Periplaneta americana were the most abundant species found after identification in the city of Cotonou. Thermo-tolerant coliforms and Escherichia coli were found on all specimens at varying concentrations. Salmonella spp. bacteria were identified in the fly population from the Dantokpa dump. Conclusion: Flies and cockroaches can transmit microorganisms to humans. The presence of Salmonella spp. among the specimens revealed that infections frequently caused by contaminated food or water are also transmitted to humans by flies. The presence of these germs on flies and cockroaches represents a potential risk of mechanical transmission to humans. It is, therefore, essential to continue investigations in order to assess the species of bacteria propagated, improve control strategies against these troublesome insects and adopt better hygiene conditions for better living.

Periplaneta americana extract used in patients with systemic inflammatory response syndrome

BACKGROUND:Periplaneta americana extract is recognized to have a positive effect on gastrointestinal mucosa.This study aimed to investigate the effects of periplaneta americana extract on immune function,nutrition status and gastrointestinal complications of early enteral nutrition patients with systemic inflammatory response syndrome(SIRS).METHODS:Patients with SIRS were randomly divided into two groups:treatment and control groups.All patients in the two groups received conventional therapy including enteral nutrition,but periplaneta americana extract,an additional Chinese medicine,was given to the patients in the treatment group.At the beginning of treatment(0 day)and 1,3,and 7 days after treatment,the levels of immunoglobulin(Ig A),total lymphocyte count(TLC),total protein(TP)and prealbumin(PA)were respectively tested in patients'venous blood.The incidences of bloating,diarrhea,aspiration pneumonia and high blood sugar at 7 days after treatment were recorded.The mortality of the patients in 28 days was recorded.RESULTS:At 3 and 7 days after treatment,the levels of Ig A and TLC in the treatment group were higher than those in the control group(P<0.05).At 7 days after treatment,the levels of TP and PA in the treatment group were higher than those in the control group(P<0.05).The incidences of bloating and diarrhea in the treatment group were lower than those in the control group,the differences were significant(P<0.05).The mortality of treatment group was lower than that of the control group(P>0.05).CONCLUSION:Periplaneta americana extract could reduce gastrointestinal complications and improve immune function and nutritional status in patients with systemic inflammatory response syndrome.

美洲大蠊研粉干预脊髓半横断大鼠运动功能和自噬蛋白Beclin-1的表达

背景:脊髓损伤有氧化应激、炎症、凋亡、自噬等机制参与其中,激活自噬可改善脊髓损伤后的神经运动功能,对脊髓起保护作用。目的:探讨美洲大蠊研粉对脊髓半横断损伤大鼠后肢运动功能和损伤区域自噬蛋白Beclin-1的影响。方法:取30只6-8周龄SD大鼠,采用随机数字表法分为3组,每组10只:假手术组仅打开椎板,暴露脊髓后缝合;生理盐水组与美洲大蠊组均建立左侧脊髓半横断损伤模型,术后第1天开始分别灌胃给予生理盐水与美洲大蠊研粉,连续给药14 d。术后6 h及1,3,7,14 d行BBB评分,观察后肢运动功能的情况;给药14 d后处死大鼠,采用免疫组化染色、Western Blot检测、免疫荧光染色检测脊髓半横断损伤区域自噬蛋白Beclin-1的表达情况。结果与结论:①随着术后时间的延长,生理盐水组、美洲大蠊组大鼠BBB评分逐渐升高;与假手术组比较,生理盐水组、美洲大蠊组术后6 h及1,3,7,14 d的BBB评分均降低(P<0.05);美洲大蠊组术后7,14 d的BBB评分高于生理盐水组(P<0.05);②免疫组化染色显示,假手术组Beclin-1呈弱阳性表达,主要位于胞浆;生理盐水组Beclin-1主要位于胞浆,可见部分核膜表达;美洲大蠊组Beclin-1主要位于胞浆,可见部分核膜表达;生理盐水组、美洲大蠊组Beclin-1阳性细胞比例多于假手术组(P<0.05),美洲大蠊组Beclin-1阳性细胞比例多于生理盐水组(P<0.05);③Western Blot检测与免疫荧光染色显示,生理盐水组、美洲大蠊组Beclin-1蛋白表达高于假手术组(P<0.05),美洲大蠊组Beclin-1蛋白表达高于生理盐水组(P<0.05);④结果表明,美洲大蠊研粉能够改善脊髓半横断损伤大鼠的后肢运动功能,机制可能是美洲大蠊研粉中的多糖类物质增加了自噬蛋白Beclin-1的表达。

Exploration on Separation of Periplaneta Americana Small Peptides with Tricine-SDS-PAGE System

[ Objective] This study aimed to explore a Tricine-SDS-PAGE method for separation and purification of Periplaneta Americana small peptides. [ Method] By comparing the separation effects of three types of gels with different ratios on P. Americana small peptides, an appropriate Tricine-SDS-PAGE method for separation of small peptides was established. [ Result] Standard low molecular weight Marker was preliminarily separated to a certain extent with polyacrylamide gel electraphoresis, while the sample mixture was not separated due to the different levels of aggregation in separating gel, spacer gel and stacking gel. [Condusion] More appropriate polyacrylamide gel electrephoresis separation method or other better separation methods were to be further explored for separating peptide mixture with smaller molecular weight.

基于TLR4/MyD88/NF-κB通路探讨美洲大蠊虫粉对脊髓损伤大鼠的保护作用

目的探讨美洲大蠊虫粉对脊髓损伤大鼠的保护作用和可能的机制。方法将48只SD大鼠随机分为假手术组、盐水组、美洲大蠊虫粉组、TOLL样受体4(toll-like receptors,TLR4)抑制剂组。除假手术组外其余3组均构建大鼠脊髓半横断损伤模型。术后假手术组不做治疗,盐水组与美洲大蠊虫粉组分别给予生理盐水和美洲大蠊虫粉(630 mg/kg)灌胃处理,TLR4抑制剂组给予TLR4抑制剂(3 mg/kg)腹腔注射处理。术后1、3、7、14 d运用BBB评分评估大鼠后肢运动功能,苏木素-伊红(HE)染色观察脊髓组织病理改变,免疫组化观察神经元数目变化,酶联免疫法检测炎性因子IL-1、IL-6、IL-10和TNF-α表达,Western Blot检测TLR4、髓样分化因子(myeloid differentiation factor 88,MyD88)和NF-κB p65表达。结果与假手术组相比,盐水组BBB评分、神经元数目显著下降,而病理损伤程度、IL-1、IL-6、TNF-α、TLR4、MyD88、NF-κB p65水平显著增加(P<0.05),与盐水组相比,美洲大蠊虫粉组和TLR4抑制剂组增加了BBB评分、神经元数目、降低了病理损伤程度、IL-1、IL-6、TNF-α、TLR4、MyD88、NF-κB p65水平(P<0.05),与TLR4抑制剂组相比,美洲大蠊虫粉组更好地增加了BBB评分、神经元数目,降低病理损伤程度、IL-1、TNF-α表达。结论美洲大蠊虫粉可以通过抑制TLR4/MyD88/NF-κB通路减少脊髓损伤后炎性因子的产生,起到保护神经和促进运动恢复的作用。

Separation of Antitumor Active Plypeptides from Periplaneta Americana by Ultra Filtration Technology

[Objective] The paper was to establish ultrafiltration membrane technology for separation of antitumor active plypeptides from Periplaneta Americana. [Method] Hollow fiber ultrafiltration membrane（3 kDa）was used to separate antitumor active plypeptides from P. Americana. Through single factor test, the impact of rotation speed, concentration ratio, pH and operation time on separation effort was studied, and the separation technology conditions were further optimized through orthogonal test. [Result] The best technology conditions for separation of antitumor active plypeptides from P. Americana were as follows： rotation speed 4 500 r/min, concentration ratio 30%, pH 11, and the rotation speed was the major influencing factor. [Conclusion] The study laid foundation for industrialization of ultrafiltration membrane separation of antitumor active plypeptides from P. Americana.

肝龙胶囊对Balb/c裸鼠腋下移植瘤的影响

目的探讨肝龙胶囊对Balb/c裸鼠腋下移植瘤的影响。方法将人肝癌敏感株BEL-7402接种于裸鼠腋下造模,造模成功后将其分为模型组、索拉非尼组、CII-3组、脱脂膏组、肝龙胶囊高、中、低剂量组,正常裸鼠为空白组,连续给药14d。比较各组的抑瘤率、脏器指数、生化指标、缺氧微环境相关蛋白及其mRNA表达水平。结果各药物治疗组均能抑制肿瘤生长,其中肝龙胶囊中剂量组的平均抑瘤率最高;与模型组相比,各药物治疗组裸鼠的心脏指数、肾脏指数、肝脏指数、肺脏指数均有所升高,脾脏指数均有所下降,但多数变化不大;各项生化指标显示,肝龙胶囊安全性良好,并未出现明显的组织损伤;索拉非尼组、肝龙胶囊中剂量组裸鼠的低氧诱导因子1α(hypoxia-inducible factor-1α,HIF-1α)、血管内皮生长因子(vascular endothelial growth factor,VEGF)和血管内皮生长因子受体2(vascular endothelial growth factor receptor 2,VEGFR2)的mRNA表达均显著低于模型组(P<0.05);免疫组织化学结果显示,各药物治疗组的VEGFR2、HIF-1α、VEGF、Ki-67和CD31蛋白表达均表现出不同程度的抑制,其中以索拉非尼组、肝龙胶囊中剂量组及CII-3组的抑制作用较强。结论肝龙胶囊可有效抑制Balb/c裸鼠腋下移植瘤的生长,并下调移植瘤中缺氧微环境相关蛋白及其mRNA表达水平。

锰超富集植物垂序商陆(Phytolacca americana L．)的认定

针对目前国内锰超富集植物商陆属(Phytolacca)植物名称混乱的状态,在重新审查锰超富集植物——商陆(Phytolacca acinosa Roxb.)的野外标本和温室培养植物的基础上,比对中国科学院昆明植物所标本馆的腊叶标本,并查阅相关文献资料,以期正确认定锰超富集累植物的学名。结果表明,湘潭锰矿尾矿废弃地原生的锰超富集植物实为垂序商陆(Phytolacca americana L.),国内外相关研究论文中出现的商陆和美洲商陆实为垂序商陆的同物异名。这一植物名称的认定,将对避免锰超富集植物研究重复进行、保证相关研究正常有序开展具有重要意义。

美洲商陆(Phytolacca americana L.)——一种新的Mn积累植物

通过野外调查和水培试验,研究了锰在美州商陆体内积累情况及对其生长发育的影响,发现并鉴定出美洲商陆(Phytolacca am ericana L.)是一种新的M n积累植物。调查结果发现,美洲商陆对土壤中高含量的锰有很强的忍耐、吸收和积累能力,叶片中M n含量为5160￣8000m g·kg-1,平均为6490m g·kg-1。营养液培养试验证明,美洲商陆对生长介质中的M n有很强的忍耐能力,当生长介质中M n浓度高达10m m ol·L-1,叶片中M n含量达到11.76g·kg-1,植株仍生长正常。地上部M n含量及其积累量均随生长介质中M n浓度的增加而增加,当生长介质中M n浓度为50m m ol·L-1时,叶片中M n含量达到47.06g·kg-1。结果表明,美洲商陆是我国发现的一种新的M n积累植物,为今后探明植物积累M n的机理和M n污染土壤的植物修复提供了一种新的种质资源。

不同生境美洲商陆(Phytolacca americana L.)根系差异表达蛋白筛选

某铀尾矿区生长着大量美洲商陆（Phytolacca americana L.）,目前还未见美洲商陆耐受及富集铀相关蛋白质组学研究报道。为了找到铀耐受及富集表达相关蛋白,建立了铀尾矿区和对照区2种不同生境美洲商陆根系蛋白质组研究技术。通过全蛋白质制备条件优化、2DE电泳条件摸索以及染色方法选择,得到了重现性好、分离效果和清晰度都很高的美洲商陆根系双向电泳差异凝胶图谱。经软件分析,筛选出了铀耐受及富集相关蛋白24个,上调表达11个、下调表达13个。

Two New Steroidal Glycosides from Fermented Leaves of Agave americana

Two new spirostanol glycosides named agamenoside A and B, were isolated from the fermented leaves of Agave americana. Their structures were elucidated as (23S, 25R)-5a -spirostan-3b, 6a, 23-triol 3-O-a-L-rhamnopyranosyl-(13)- b-D-glucopyranosyl-(12)-[b-D- xylopyranosyl-(13)]-b-D-glucopyranosyl-(14)-b-D-galactopyranoside (1) and (25R)-5a-spiro- stan-3b, 6a-diol 3-O-b-D-glucopyranosyl-(12)-[ b-D-xylopyranosyl-(13)]- b-D-glucopyra- nosyl-(14)- b-D-galactopyranoside (2) by a combination of chemical and spectral methods.

A New Pregnane Glycoside from Fermented Leaves of Agave americana

A new minor pregnane glycoside was isolated from the fermented leaves of Agave americana. Its structure was elucidated as (20S)-5a-pregnane-3? 20-diol 20-O--D-glucopyrano- side (1) by spectral methods.

Two New Spirostanol Steroidal Sapogenins from Fermented Leaves of Agave americana

Two new spirostanol sapogenins named agavegenin A and B were isolated from the fermented leaves of Agave americana L. Their structures were elucidated as (23S, 25R)-5a- spirostan-3b, 6a, 11a, 23-tetraol (1) and (23S, 25S)-5a-spirostan-3b, 23, 27-triol (2) by spectral methods.

美洲大蠊提取物体外对肝癌耐药细胞BEL-7402/5-FU自噬和侵袭转移的影响

目的:探讨美洲大蠊提取物CⅡ-3和脱脂膏对肝癌耐5-氟尿嘧啶细胞(BEL-7402/5-FU)自噬和转移侵袭能力的影响。方法:以肝癌敏感细胞BEL-7402和肝癌耐药细胞BEL-7402/5-FU作为实验对象,给予索拉菲尼、CⅡ-3和脱脂膏处理,检测自噬标志蛋白的生成,同时检测细胞中自噬相关因子和侵袭转移相关因子的水平以及细胞侵袭转移能力的变化。结果:耐药细胞组中p62蛋白低表达,给予阳性药索拉菲尼、CⅡ-3和脱脂膏后,细胞中p62蛋白表达量明显升高。和敏感细胞相比,耐药细胞中LC3B生成增多,给予CⅡ-3后LC3B生成减少。CⅡ-3和脱脂膏给药组均能明显抑制自噬和侵袭转移的部分相关因子的表达,包括降低ATG5、PIK3C3、BECLIN、LC3B、MMP-9、Akt、4EBP1mRNA表达。划痕试验结果显示,相比于敏感细胞,耐药细胞具有更强的侵袭转移能力,给予索拉菲尼和CⅡ-3作用后均可显著抑制细胞的侵袭转移,而脱脂膏则无显著的抑制效果。结论:肝癌耐药细胞具有很强的自噬和侵袭转移能力,而美洲大蠊提取物CⅡ-3和脱脂膏体外均能明显抑制耐药细胞的自噬和侵袭转移,但仅能影响部分相关因子。

HPLC法同时测定美洲大蠊标准汤剂中6种成分含量

目的:建立HPLC法同时测定美洲大蠊标准汤剂中尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷、原儿茶酸的含有量。方法:采用WATERS Symmetry Shield C 18色谱柱(250 mm×4.6 mm,5μm);甲醇-0.1%磷酸水作为流动相,梯度洗脱;体积流量0.6 mL/min;检测波长254 nm;柱温20℃。结果:测定显示尿嘧啶、次黄嘌呤、尿苷、肌苷、鸟苷、原儿茶酸分别在0.041~0.341μg(r=0.9994)、0.097~0.810μg(r=0.9994)、0.020~0.168μg(r=0.9999)、0.277~2.308μg(r=0.9999)、0.016~0.135μg(r=0.9997)、0.131~1.096μg(r=0.9998)范围内线性关系良好,平均加样回收率分别为99.93%(RSD=1.18%)、98.03%(RSD=1.05%)、97.17%(RSD=1.29%)、100.19%(RSD=1.91%)、98.94%(RSD=1.11%)、100.70%(RSD=1.05%)。十批美洲大蠊标准汤剂各成分含量范围分别为:尿嘧啶0.14~0.21 mg/g、次黄嘌呤0.22~0.36 mg/g、尿苷0.06~0.08 mg/g、肌苷2.17~3.15 mg/g、鸟苷0.03~0.07 mg/g、原儿茶酸0.35~0.56 mg/g,处于均值的±30%范围内。结论:美洲大蠊标准汤剂含量测定方法准确、可靠,可用于美洲大蠊配方颗粒的质量参考。

载美洲大蠊有效部位CII-3 PDLLA微球的制备及制剂学评价

采用乳化溶剂挥发法,以PEG修饰的PDLLA为原材料,制备载美洲大蠊有效部位CII-3 PDLLA微球。通过单因素试验以及正交试验优化制备工艺,并对微球进行表征,通过对微球的体外释放规律和稳定性进行初步研究。结果显示,所制备的微球载药量为1.22%,包封率为43.03%,平均粒径为97μm。微球形态圆整,表面存在密集小孔,具有良好的释药特性。初步稳定性考察表明,微球能够在一定程度上保护美洲大蠊有效部位CII-3不受到胃酸以及各种酶的影响从而失活,可为CII-3的进一步开发应用提供理论依据。

美洲大蠊提取物对小鼠3LL肺癌的抑制作用及其机制探讨

背景与目的美洲大蠊系列昆虫药品康复新在体外能诱导多种肿瘤细胞系发生细胞凋亡。本研究旨在探讨美洲大蠊提取物对C57BL/6J小鼠3LL肺癌的抑制效应及其作用机制。方法荷3LL肺癌的C57BL/6J小鼠随机分为生理盐水组、美洲大蠊提取物高、低剂量组,通过检测各组小鼠的体重变化和抑瘤率,应用流式细胞术进行DNA倍体分析、TUNEL分析和凋亡相关基因表达改变的检测。结果与生理盐水组比较,美洲大蠊提取物高、低剂量组的抑瘤率分别为41.24%及81.08%。流式细胞术检测美洲大蠊提取物能诱导肿瘤细胞凋亡,随着药物剂量的增高,其凋亡率也升高,S期及G2/M期细胞数大量减少,细胞被阻滞在G0/G1期;流式细胞术TUNEL检测结果显示,细胞凋亡和坏死同时存在。流式细胞法分析3LL肺癌细胞凋亡相关基因显示,无论是低剂量组还是高剂量组,美洲大蠊提取物均能上调Fas、FasR、p53等凋亡基因表达,下调Bcl-2抑制凋亡基因表达。结论美洲大蠊提取物对3LL肺癌生长具有一定的抑制作用,能促进肿瘤细胞凋亡,其机制可能与调控细胞凋亡相关基因的表达有关。

Fe_(3)O_(4)@ZIF-8-表面辅助激光解吸/电离质谱法检测美洲大蠊多肽

本实验在室温下合成了以Zn作为金属中心,2-甲基咪唑作为连接剂的磁性金属有机框架材料Fe_(3)O_(4)@ZIF-8,以Fe_(3)O_(4)@ZIF-8作为固体基质,通过建立Fe_(3)O_(4)@ZIF-8-表面辅助激光解吸/电离质谱(SALDI-MS)法,实现了美洲大蠊小分子多肽IPMTAPGL-NH_(2)、LTAPGL-NH_(2)、SLMTGPGL-NH_(2)、SLHTAPGL-NH_(2)的快速检测。实验优化了基质浓度、检测限和点样方法等条件,考察了该基质对美洲大蠊多肽IPMTAPGL-NH_(2)的质谱信号影响。与传统基质辅助激光解析电离质谱法(MALDI-MS)相比,Fe_(3)O_(4)@ZIF-8-SALDI-MS法检测背景更干净,目标多肽响应更强,检测灵敏度更高。该法可应用于血样中美洲大蠊多肽IPMTAPGL-NH_(2)的快速检测。