Activity of dihydroartemisinin against Leishmania donovani both in vitro and vivo

Visceral leishmaniasis (VL), also known as kala-azar, is a vector-borne disease caused by protozoa of the Leishmania donovani complex. It is usually fatal if untreated. Pentavalent antimonials have been the mainstay of treatment for more than 60 years, but these drugs have serious side effects and progressive antimonial resistance.1 Other proven therapeutics such as amphotericin B, pentamidine, and paromomycin are costly without oral formulation and unpleasant side effects.2 Thus, new chemotherapeutic drugs are required to supplement or replace currently available drugs.     

Activity of dihydroartemisinin against Leishmania donovani both in vitro and vivo

Visceral leishmaniasis (VL), also known as kala-azar, is a vector-borne disease caused by protozoa of the Leishmania donovani complex. It is usually fatal if untreated. Pentavalent antimonials have been the mainstay of treatment for more than 60 years, but thesedrugs have serious side effects and progressive antimonialresistance. Other proven therapeutics such as amphotericin B, pentamidine, and paromomycin arecostly without oral formulation and unpleasant side effects. Thus, new chemotherapeutic drugs are required to supplement or replace currently availabledrugs.     

骨髓发现杜氏利什曼原虫小儿黑热病2例

黑热病病原体杜氏利什曼原虫的无鞭毛体，主要寄生在肝、脾、骨髓、淋巴结等器官的巨噬细胞内，常引起全身症状，如发热、肝脾肿大（脾进行性肿大）贫血等。我院于2005年5月-8月收治2例黑热病患儿，骨髓检查发现巨噬细胞胞浆内及细胞外有杜氏利什曼原虫，现报告如下。     

Epidemiology of leishmaniases in Kenya: V. Wider search for breeding habitats of phlebotomine sandflies in three kala-azar endemic foci

Intensive investigations on the breeding sites of phebotomine sandflies were undertaken in Kitui, Machakos and Baringo foci of leishmaniases in Kenya. A total of 473 soil samples weighing approximately 4,244 kg were collected from termite hills, animal burrows, tree hole, human dwellings, animal enclosures, under tree canopy, open ground, chicken coop and rock crevices habitats and incubated in the field laboratories. 267 samples weighing approximately 3002 kg were positive, producing 6419 sandflies comprising 17 different species. This study resulted in the identification of both perennial and seasonal breeding sites of most of the phlebotomine sandfly species found in these three leishmaniases foci.     

Difference in DNA sequences in SSU rDNA variable regions among pathogens isolated from different epidemic foci of visceral leishmaniasis in China

Objective To confirm the existence of point mutations in the SSU rDNA variable regions of 5 Leishmania donovani (L. d. ) isolates from different epidemic foci in China. Methods Specific SSU rDNA fragments from nuclear DNA of 7 Leishmania species/isolates were amplified by PCR and then cloned into pGEM[R- T Easy Vectors.After that, the specific fragments were sequenced by an automated DNA sequencer. Results Sequence analysis showed that the amplified DNA fragments of 7 Leishmania species/isolates were all 392 bp in length.All 5 point mutations were located in two unique sequence blocks (UQ- Ⅰ and UQ- Ⅱ), and no insertions or deletions were found.The identities of comparison of Leishmania in GeneBank were more than 98%. Conclusion Five point mutations exist in the SSU rDNA variable region of 5 L. d. isolates from different epidemic foci of visceral leishmaniasis (VL) in China. Sequence differences of the SSU rDNA variable region exist among L. d.isolates from different foci.     

华北地区阴道毛滴虫同工酶分析

目的：研究我国华北地区（北京，河北唐山、河北承德）阴道毛滴虫4个分离株的同工酶类型。（2）方法：采用聚丙烯酰胺凝胶电泳（PAGE）与同工酶染色法进行分析。（3）结果：检测MDH，LDH，G6PD，PGM5种 同工酶。在G6PD，PGI酶谱中，4株完全相同，在MDH，LDH，PGM酶谱中，北京1株，北京1株与承德株，唐山株不同。（4）结论：承德株，唐山株与北京1株，北京2株大多数同工酶谱存在显性差异，说明华北地区阴道毛滴虫虫株可能至少存在2种不同的酶谱类型。     

不同地理株间日疟原虫SSUrRNA基因序列比较

目的测定我国间日疟原虫不同地理株红内期SSUrRNA基因序列,比较分析其分子特征。方法收集深圳、海南、湖北和河南四地间日疟患者血样5份（其中海南2份）,并提取核酸DNA,采用PCR从核酸提取物中扩增出间日疟原虫SSUrRNA基因片段,纯化后分别与pGEM—Teasy质粒连接构建重组子并转化大肠杆菌JM109;阳性克隆以双酶切鉴定后,双脱氧末端终止法测定序列,采用BLAsT和MEGA4软件分析序列相互关系特征。结果5株间日疟原虫SSUrRNA基因扩增片段大小一致,约为998bp;核酸序列测定结果显示,所克隆的SSUrRNA基因片段均含有998个核苷酸,不同地理株间SSUrRNA基因序列有9个位点存在变异的可能,两两比对的同源性均高于99．5％,其中河南与湖北株序列的同源性为100．0％;与GenBank中报道的国外6株间日疟原虫相同序列作分子系统进化分析。国内5株间日疟原虫SSUrRNA基因序列与Sa11株遗传距离小,亲缘关系接近。结论测定的国内间日疟原虫SSUrRNA基因序列在不同地理株间相对保守．其间存在的单核苷酸多态性与地理变化有一定程度的关系。     

Sequence analysis and genotypes of glutamate rich protein of Plasmodium falciparum isolates from different malaria endemic areas in China

To sequence the gene encoding glutamate rich protein (GLURP) and identify the genotypes of geographically different Plasmodium falciparum (P. f ) isolates from China. Methods The gene of R2 repeat region of GLURP was amplified by nested polymerase chain reaction and cloned into T-vector. The nucleotide sequence of GLURP gene was determined by automatic sequencer (Dideoxy termination method) and analyzed by DNA Star software. Results At least 7different GLURPgenotypesranging from 600 bp to 1 500 bp were found in Yunnan and Hainan provinces. R2 region of GLURP gene consisted of several repeat units. Each repeat unit was composed of 19-20 residues which were shown to be highly conserved. GLURP gene was also size polymorphic due to differences in the number of repeat units, whereas the repeat sequence was conserved. Sequence analysis showed that DNA sequences and deduced amino acid sequences were highly homologous among the geographically dispersed isolates or various isolates from the same geographical region. No obvious differences were found in the GLURP gene sequences among geographically different isolates. Conclusion GLURP gene is highly structure conserved and size polymorphic, and so is useful in searching for malaria vaccine candidate antigen and developing a genotyping method for malaria research.     

原位杂交技术测定放烧复合伤创面aFGF mRNA的表达

目的 :探讨放烧复合伤创面愈合过程中aFGF的作用。方法 :用SD大鼠制备深Ⅱ°烧伤、放烧复合伤模型 ,分别于模型后 12h和 1、3、7d取背部创面皮肤组织 ,采用原位杂交技术测aFGFmRNA的表达。结果 :烧伤组大鼠在上述 4个时相点aFGFmRNA表达明显增加 ,而放烧复合伤组仅在第 7天有少量表达 ;aFGFmRNA的表达部位位于真皮及皮下。结论 :提示aFGF有助于创伤的愈合 ;放烧复合伤创面愈合延迟与aFGFmRNA表达低下及延迟有关     

不同来源国产血竭的薄层色谱和紫外光谱鉴别

目的：比较6种不同来源国产血竭的质量差异。方法：以剑叶龙血树含皮及木质部为原料，采用回流提取、索氏提取，超声提取，低温膨化提取，并与市售的通过传统工艺提取的云南血竭和海南血竭为对照，对它们进行薄层层析和紫外光谱鉴别。结果：薄层对应斑点基本相同，紫外光谱基本一致。结论：6种不同来源的国产血竭的成分基本一致，而以龙血素C作为评价指标，其质量存在一定差异。     

Identification of differentially expressed genes in anagen dermal papilla by suppression subtractive hybridization

Background We constructed a cDNA subtractive library of dermal papilla cells (DPCs) in anagen with suppression subtractive hybridization (SSH) technique and clone differentially expressed genes related to DPCs in anagen. Methods Total mRNA was isolated from DPCs of anagen and telogen follicles. Moreover, singlestrand (ss) and double-strand (ds) cDNAs were synthesized in turn using SMART PCR cDNA synthesis technology, ds cDNAs then were digested with Rsa I and divided into two groups, and ligated to the specific adaptor 1 and adaptor 2R, respectively. After cDNAs were hybridized with each other twice and underwent two rounds of nested PCR. PCR products were ligated with arms of T/A plasmid vectors to set up the subtractive library. Selected clones were demonstrated by reverse Northern blot and sequenced. The acquired sequence data were aligned against the Genbank nucleotide database. Results cDNA subtractive library of DPCs in anagen follicles was set up successfully with high subtractive efficiency, Thirty-five genes were identified in this study with 22 known functional genesand 13 unknown functional genes. Conclusions All results confirm the effectiveness and sensitivity of SSH in detecting differentially expressed genes from a small amount of clinical samples. Information about such alterations in gene expression could be useful for elucidating the genetic events in hair follicle growth regulation.     

Raji细胞中EB病毒染色体整合的荧光原位杂交定位

目的:对Raji细胞中EBV整合位点进行染色体定位.方法:用Southern免疫印迹检测Raji细胞中EBV DNA,应用G显带和荧光原位杂交(fluorescence in situ hybridization,FISH)对Raji细胞中EBV整合位点进行染色体定位.结果:Raji细胞gDNA经BamHI酶切消化后,与同位素标记的Probe-1(EBV基因组13 232～16 189)杂交,阳性片段大小为10 kb和4 kb,与Probe-2(EBV基因组5～3271)杂交,阳性片段大小为23 kb.Raji细胞中EBV整合位点有1 P,1 q,2 q,3 P,3 q,4 q,5 q,6 q,7 P,7 q,9 q,11 P,14 q,15 q等,其中4 q,2 q,1 q,7 q为EBV DNA整合的高频位点.计数33个整合信号,分别有7,4,4,4个信号位于染色体4 q,2 q,1 q,7 q,这4个位点的信号占所有信号的构成比为64%,15号以后的染色体及2条性染色体均未见EBV整合.结论:本研究用G显带和FISH方法首次对Raji细胞中EBV整合位点进行了定位,Raji细胞中EBV整合具有一些高频位点,是一种非随机性整合.     

用点杂交法检测人促红细胞生成素(hEPO)的整合

目的为制作生产人促红细胞生成素的转基因鸡，把人促红细胞生成素（hEPO）基因以质粒介导用脂质体法导入鸡胚内，然后检测人促红细胞生成素基因的整合。方法以PCR方法进行筛选后，再用点杂交法进行鉴定。即把人促红细胞生成素基因PCR扩增阳性产物用地高辛标记，作为探针和鸡整个基因组进行点杂交。结果点杂交阳性结果有4个。结论得到了整合人促红细胞生成素基因的鸡胎。