火木层孔菌发酵菌粉及其组分对小鼠移植性肝癌H22的体内生长作用

采用肝癌H22荷瘤小鼠的肿瘤模型,对火木层孔菌(桑黄)Phellinus igniarius发酵菌粉及其各提取物组分的体内抗肿瘤活性进行评价。结果表明,火木层孔菌发酵菌粉及其各提取物组分对荷瘤小鼠肝癌H22都具有一定的抗肿瘤作用并能延长荷瘤小鼠生存期,其中,火木层孔菌发酵菌粉(1,000mg/kg/d)及其组分I(I多糖组分)(360mg/kg/d)具有较为显著的抗肿瘤作用,抑瘤率分别为33.5%和40.3%。组织病理学研究结果表明在菌粉及其多糖组分作用后,肿瘤细胞坏死细胞明显增多,免疫组化检测表明火木层孔菌发酵菌粉及其多糖组分能明显的降低瘤组织中Bcl-2基因蛋白的表达,提高小鼠瘤组织中的Bax基因蛋白的表达。火木层孔菌发酵菌粉及其多糖组分具有较好的体内抗肿瘤活性。     

香菇C91-3菌LP1蛋白在小鼠体内的 抗肿瘤免疫作用

目的通过观察注射C91-3菌LP1蛋白对H22荷瘤小鼠的影响,探讨LP1蛋白在小鼠体内的抗肿瘤免疫作用。方法使用鼠肝癌H22细胞接种于BALB/C小鼠右腋下,建立小鼠H22实体瘤模型。取上述建立成功的H22实体瘤模型小鼠64只,体重20~25g;分为A、B两组,每组32只。A组再分为LP1实验Ⅰ组(300μg/只)、LP1实验Ⅱ组(100μg/只)、PBS对照组和顺铂对照组(4 mg/kg),每组8只,各组隔日给药1次,共给药5次。A组用于检测LP1蛋白作用后在小鼠体内对H22肿瘤的抑瘤作用、血清中IL-2含量以及脾中NK细胞活性等生理指标。B组按同样的方法分组,隔日给药1次,直至荷瘤小鼠死亡,记录各组小鼠的生存期,计算生命延长率。结果 LP1蛋白可以延长H22荷瘤小鼠的生存期,LP1实验组生存期达16.6d,较PBS对照组13.2d有明显提高。LP1蛋白在体内对H22实体瘤具有一定的抑制作用,对H22实体瘤进行病理切片、HE染色观察后发现,LP1实验组中H22肿瘤组织较PBS对照组肿瘤组织内出现炎性细胞浸润,局部可见坏死现象。使用ELISA法和LDH法分别检测H22荷瘤小鼠血清中IL-2含量以及NK细胞活性,发现LP1实验组IL-2水平和NK细胞活性较PBS对照组和顺铂对照组显著提高。结论 LP1蛋白可延长H22荷瘤小鼠的生存期限,提高小鼠的生存质量,具有一定的肿瘤抑制作用。其抑制作用主要是由增强H22荷瘤小鼠自身免疫力,提高NK细胞活性,发挥机体自身肿瘤免疫功能造成的。     

膜型Tim-3分子促进荷瘤小鼠抗肿瘤免疫应答的研究

目的 研究膜型Tim-3分子对H22肝癌细胞生长的抑制效应,探讨膜型Tim-3分子对荷瘤小鼠免疫系统的影响.方法 以H22肝癌细胞接种于BALB/c小鼠大腿肌肉建立小鼠实体瘤模型,采用原位注射裸DNA的方法在小鼠体内表达膜型Tim-3进行基因治疗,观察Tim-3对肿瘤生长的抑制作用;采用RTPCR技术在肿瘤生长不同时期检测Tim-3对4-1BB、IFN-γ、galectin-9等免疫相关基因表达的影响;流式细胞术检测膜型Tim-3对脾细胞增殖活性及细胞毒活性的影响;观察Tim-3 ＋4-1 BBL协同抗肿瘤作用.结果 体内转染表达Tim-3对肿瘤的生长有明显的抑制作用.流式细胞术结果显示,在小鼠荷瘤早期,Tim-3可提高脾细胞在特异性抗原刺激下的增殖反应和对H22肿瘤细胞的细胞毒作用;Tim-3与4-1BBL协同作用时抗肿瘤作用更加明显.结论 膜型Tim-3可在免疫启动阶段作为正向免疫调节因子增强抗肿瘤免疫应答,并可与4-1 BBL协同产生更强的抑瘤效应.     

滇姜花二萜成分的抗肿瘤活性研究

对滇姜花粗提物、滇姜花素A(1)和姜花酮(2)进行了动物体内抗肿瘤活性测试,结果表明它们均能显著性抑制小鼠体内移植性肿瘤H22的生长,其中滇姜花素A对小鼠H22肿瘤生长抑制率达54.27%,作用最强。体外抗肿瘤实验发现,滇姜花素C(3)对体外培养的人类乳腺癌细胞株MDA-MB-231具有较强的细胞毒作用,并具有明显的剂量效应关系。     

参芝发酵产物的抗肿瘤活性及其对小鼠免疫功能的影响

为了探索红参水煎液的灵芝发酵产物对H22荷瘤小鼠的抗肿瘤活性及其对小鼠免疫功能的影响,通过体内抗肿瘤实验和增强免疫功能实验从抑瘤率、对免疫器官的影响指数、对非特异性免疫、体液免疫及细胞免疫的影响5个方面对该产物做了功能性评价。结果表明,在抗肿瘤实验中,参芝发酵产物高剂量组的抑瘤率达到51.65%,脾指数和胸腺指数均高于对照组和环磷酰胺(CTX)组;增强免疫功能实验中,3个实验的给药组小鼠和对照组小鼠相比都有显著性差异(P<0.01)。由此可见,将灵芝与人参在发酵层次上配伍具有显著的抑制荷瘤小鼠肿瘤生长及增强小鼠免疫功能的作用。     

木蹄层孔菌子实体提取物对H22荷瘤小鼠体内抗肿瘤活性的影响

研究了木蹄层孔菌子实体的石油醚提取物、氯仿提取物、甲醇提取物对体内抑制肿瘤活性及对免疫功能的影响。建立H22荷瘤小鼠模型,观察木蹄层孔菌不同提取物对荷瘤小鼠的抑瘤效果,通过抑瘤率、免疫器官指数及生存时间的影响来评价不同提取物的活性。结果表明：木蹄层孔菌子实体的石油醚提取物、氯仿提取物、甲醇提取物均有一定的抑制肿瘤作用,其中石油醚提取物下层沉淀的抑制作用最显著,当质量分数为100 mg/kg时抑瘤率高达56.29%,接近阳性药的抑瘤率58.78%,可使小鼠的体质量、脾指数和胸腺指数增加,延长H22荷瘤小鼠的生存时间。木蹄层孔菌石油醚提取物下层沉淀在一定的剂量范围内,能较好地抑制小鼠肿瘤的生长,提高机体的免疫功能。     

酯苷胶囊对小鼠移植瘤的抑制作用

目的:探讨酯苷胶囊对小鼠移植性肿瘤的抑制作用。方法:采用小鼠移植性肿瘤模型,以5-FU为阳性对照组,观测2、4、8mg.kg-1酯苷胶囊对小鼠H22、S180肉瘤和HCA肝癌模型动物的抗肿瘤作用。结果:酯苷胶囊对H22、S180和HCA移植瘤的抑制率分别为36.8%～65.3%,19.0%～41.4%,46.8%～52.3%。结论:酯苷胶囊具有较强的抗肿瘤作用,显著延长荷瘤小鼠的生命。     

CRM197修饰的肿瘤细胞裂解物疫苗抗肝癌作用研究

[目的]探讨CRM197能否增强H22肝癌细胞裂解物疫苗的抗肿瘤活性。[方法]反复冻融H22细胞制备裂解物,与CRM197偶联,制备H22-CRM197疫苗,以小鼠皮下移植瘤模型考察疫苗抗肿瘤活性,并对免疫学机制进行探讨。[结果]与PBS组相比,H22-CRM197免疫显著降低了荷瘤小鼠肿瘤重量(0.53±0.20 g VS 2.04±0.43 g,p0.01);与PBS组比较,H22-CRM197组小鼠免疫血清中检测到高滴度的抗-H22抗体(p0.01);H22-CRM197免疫能够有效地刺激脾淋巴细胞的增殖并诱导产生了明显靶向H22细胞的细胞毒性T淋巴细胞杀伤作用。[结论]CRM197可以显著增强H22肝癌细胞裂解物疫苗的抗肿瘤活性。     

桦木酸对H22荷瘤小鼠肿瘤细胞凋亡及细胞周期的影响

研究桦木酸对H22荷瘤小鼠生命延长率、肿瘤细胞凋亡及细胞周期的影响。结果表明桦木酸能够明显延长H22荷瘤小鼠生生存时间,其中低、中剂量效果显著（P〈0．05）;利用DNA结合性荧光探针直接对细胞DNA染色后FCM分析,桦木酸可能是通过影响H22肿瘤细胞S期而诱导肿瘤细胞凋亡。     

双歧杆菌及其发酵液的抗肿瘤机制的研究

目的 通过比较双歧杆菌及其发酵液的抗肿瘤作用,对其抑瘤作用和相关机制进行初步探讨。方法 取BALB／c小鼠,分别予H22细胞和S180细胞腹腔荷瘤,荷瘤后给予不同的治疗。分别观察2种肿瘤细胞荷瘤小鼠的生存期。右腋下荷瘤后,进行病理切片观察。MTT法计算给予不同方法后的抑瘤率。并计算淋巴细胞转化率。制备电镜标本。观察其超微结构。结果 双歧杆菌死菌液、活菌液能够延长荷瘤小鼠的生存期,而双歧杆菌发酵液不能延长其生存期,病理切片可见作用后的肿瘤组织内部和间质有大量炎性细胞浸润。双歧杆菌死菌液、活菌液对肿瘤细胞株有抑杀作用。电镜观察死菌液作用后的H22细胞可见典型的凋亡表现。结论 双歧杆菌死菌液、活菌液在体内外均有较好的抑瘤作用,其发酵液并未显示出抑瘤作用。说明双歧杆菌的抗肿瘤作用主要来源于菌体的作用。     

牛樟芝固态发酵产物的降胆固醇作用

本实验主要探究牛樟芝(S-29)固态发酵产物对高脂饮食小鼠胆固醇调节的影响。小鼠随机分为正常组、高脂模型组、护肝片阳性对照组、固态发酵组及液态发酵组;小鼠经高脂饲料喂养6周,相应物质灌胃4周。检测小鼠血清及肝脏相关指标; q-PCR检测胆固醇代谢相关基因的mRNA表达量。结果表明,与模型组比较,固态发酵组小鼠血清游离脂肪酸(NEFA)及谷丙转氨酶(ALT)浓度显著降低,分别降低了38. 5%和40. 7%;肝脏总胆固醇(TC)浓度显著降低,降低了23. 5%;低密度脂蛋白受体(LDL-R)的mRNA表达量显著增加,增加了3. 6倍。结果证明,牛樟芝固态发酵产物具有较好的降胆固醇作用,其主要机制可能是通过上调LDL-R基因的表达,以促进胆固醇的分解代谢,进而降低小鼠体内胆固醇浓度。     

不同培养方式对牛樟芝活性组分的影响

牛樟芝因含有多种活性成分且在抗癌、保肝等方面有着良好的功效而备受关注。本文考察了固态发酵以及不同液态发酵方式对牛樟芝活性产物的影响。结果显示,固态发酵不同谷物对活性产物的合成有明显的影响,青稞作为发酵谷物基质,其活性产物种类多且含量高,Antrodin C、Antroquinonol产量分别为5 901.27 mg/kg、3 715.76 mg/kg。常规液态发酵菌丝体中主要含有Antrodins类化合物,以Antrodin C为主,但不能合成Antroquinonol类化合物。通过添加前体物辅酶Q_0可以诱导牛樟芝菌丝体在液态发酵中合成Antroquinonol;在此基础上,添加植物油进行同步萃取发酵,能够显著提高牛樟芝菌丝体活性成分种类和产量,除了诱导合成Antroquinonol之外,还有大量Antroquinonol B合成;同时,大多数活性成分在发酵过程中不断被富集萃取,Antrodin C、Antroquinonol产量分别为289.80 mg/L、95.39 mg/L。Antrodin C产量比常规液态发酵提高了7.06倍,而Antroquinonol产量比诱导发酵提高了203.89%。     

基于人工神经网络-遗传算法的樟芝发酵培养基优化

采用优化模型对药用丝状真菌樟芝的复杂发酵过程进行建模,并获得最优发酵培养基组成.对樟芝发酵过程中的形态变化过程进行了观察,并分别采用人工神经网络(ANN)和响应面法(RSM)对樟芝发酵过程进行建模,同时采用遗传算法(GA)优化了发酵培养基组成.结果表明,ANN模型比RSM模型具有更好的实验数据拟合能力和预测能力,GA计算得到樟芝生物量理论最大值为6.2 g/L,并获得发酵最佳接种量及培养基组成:孢子浓度1.76× 105个/mL,葡萄糖29.1 g/L,蛋白胨9.4 g/L,黄豆粉2.8 g/L.在最佳培养条件下,樟芝生物量为(6.1±0.2)g/L.基于ANN-GA的优化方法可用于优化其他丝状真菌的复杂发酵过程,从而获得生物量或活性代谢产物.     

响应面法优化深层发酵樟芝总三萜的提取工艺(英文)

采用响应面法对樟芝深层培养中总三萜的提取工艺进行优化。根据Box-Benhnken的中心组合实验设计原理,在单因素实验的基础上,选取溶剂浓度、提取温度和液固比为变量,应用响应面法进行三因素三水平的实验设计。以总三萜得率作为响应值,建立了樟芝总三萜提取的回归模型,对其提取条件进行进一步优化。结果表明,优化的总三萜提取条件为乙醇浓度86%,提取温度75℃,液固比37。进一步的实验也验证了该模型的有效性。     

Enhancement of benzene clastogenicity by praziquantel in mice

Praziquantel (PQ) is a commonly used drug to treat patients with schistosomiasis. Previous studies using cells in vitro have shown that PQ can enhance the mutagenic activities of known mutagens. We have conducted a cytogenetic - urine metabolite study to determine the in vivo clastogenic and co-clastogenic potential of PQ with a ubiquitous environmental contaminant, benzene (BZ). 16 groups of adult male ICR mice (5 animals per group) were used. They were negative control, solvent controls (cremophore E1 3%, olive oil and combined), positive control (BZ 440 mg/kg b.w.) and 11 exposed groups. To test for clastogenicity of PQ, mice were treated orally with 100, 400, 800 and 1200 mg/kg b.w. PQ and sacrificed 30 h later for determination of micronuclei (MN) frequency in bone-marrow polychromatic erythrocytes (PCE). None of these PQ does induced an increase of MN frequency. On the other hand, BZ induced, as expected, a high frequency of MN (46.4 +/- 6.34/1000 PCE). The enhancement effect of PQ was tested in 7 groups of mice using 3 different protocols. Mice were treated with 440 mg/kg b.w. BZ and 1 h later with 0, 100, 200, 400, 800 and 1200 mg/kg b.w. PZ. In another group, 800 mg/kg PQ was administered at 3 h after BZ exposure. In the last group, PQ (800 mg/kg) was administered at 1 h prior to BZ exposure. Results from the first combined exposure group showed a significant PQ dose-dependent increase in the frequency of MN in PCE (p less than 0.05). The increase with the two high doses of praziquantel is significantly higher (p less than 0.05) than the MN frequencies in the benzene control and the expected value based on the additive effects of the two agents. Studies with other combined treatment groups showed that the induction of MN was highest when PQ was administered at 1 h before BZ exposure. Moreover, the presence of BZ metabolites (muconic acid, phenol, catechol and hydroquinone) in urine was studied in 6 of the combined treatment groups. This metabolite study revealed that PQ enhanced the metabolism of BZ towards the pathway to form muconaldehyde which is converted to muconic acid in urine. In conclusion, our study showed that PQ is not a clastogen but can enhance the clastogenic activity of BZ in vivo by shifting the metabolic pathways of BZ towards formation of muconaldehyde which may be responsible for the enhancement effect.     

Phylogenetic analysis of <Emphasis Type="Italic">Antrodia</Emphasis> species and <Emphasis Type="Italic">Antrodia camphorata</Emphasis> inferred from internal transcribed spacer region

The species of Antrodia are one of the difficult-to-classify and obscure groups of poroid Aphyllophorales based on morphological appearance. However, it is becoming increasingly important to reliably identify the entire suite of Antrodia camphorata strains and Antrodia species due to the potential pharmaceutical value of their biologically active ingredients. In this study, the internal transcribed spacer (ITS) region of the ribosomal RNA gene (rDNA) was sequenced and phylogenetically analyzed in a number of Antrodia fungal species and strains. ITS amplicons from the Antrodia species tested ranged in size from 543 to 610 bp; the size of the ITS of A. camphorata strains ranged from 592 to 596 bp. The overall sizes of ITS2 and 5.8S ribosomal RNA gene of all A. camphorata strains tested in this study were shown to be 217 and 158 bp, respectively. A phylogenetic analysis of ITS data generated, which included sequences of 11 A. camphorata strains and nine other Antrodia species, showed three clearly distinct groups. Group 1 includes A. camphorata, Antrodia salmonea, and Antrodia carbinca strains. Within Group 2, Antrodia sinuosa and Antrodia xantha were clustered together. Group 3 contained Antrodia albida, A. heteromorpha, A. serialis, and A. malicola. The observed sequence diversity among ITS alleles provided an effective tool for differentiating strains of A. camphorata, A. salmonea, A. xantha, A. sinuosa, or A. serialis. Polymorphisms arising within the ITS1-5.8S-ITS2 region can provide practical markers for establishing a foundation for the further expansion of an ITS sequence database of medically important fungi.     

用正交试验法筛选樟芝菌适宜发酵培养基

用正交试验法对樟芝菌 (AntrodiacamphorataZang&Su)的发酵培养基进行了研究。选取马铃薯淀粉、葡萄糖、麦芽糖、酵母膏、蛋白胨、麸皮、KH2 PO4 、MgSO4 ·7H2 O等作为试验因素 ,采用L2 7(31. 3)正交表设计五因素、三水平的正交试验 ,对所选各因素的添加量分别作了试验 ,对菌丝干重和发酵液多糖含量结果作了方差分析 ,确定了合适的发酵培养基。试验结果表明 :樟芝菌的深层发酵适宜培养基是 :马铃薯淀粉 1 .0 0 % ,葡萄糖2. 0 0 % ,酵母膏 0 . 10 % ,蛋白胨 0 . 10 % ,麸皮 1 .0 0 % (或 0. 5 0 % ) ,KH2 PO4 0 . 10 % ,MgSO4 ·7H2 O 0 . 0 5 %。     

重组别藻蓝蛋(rAPC)抑瘤活性及其对免疫作用的研究

目的:观察重组别藻蓝蛋白(rAPC)对接种H22肝癌细胞小鼠的抑瘤活性及其免疫作用.方法:昆明小鼠随机分为5组(10只/组),即模型组、rAPC低、中、高剂量组(25,50,100mg/kg·d)、环磷酰胺组(CY对照组).建立小鼠肝癌1422移植瘤模型,次日除模型组外,其余4组分别以不同剂量的rAPe、CY灌胃,于15d后处死,完整剥离出肿瘤、胸腺、脾脏,准确称重,计算抑瘤率、胸腺指数和脾指数,并采用放免法(PIA)测定血清中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)的水平.结果:rAPC低剂量组小鼠H22肿瘤质量增长较模型组缓慢(P＜0.05),中、高剂量组较模型组显著缓慢(P＜0.01),抑瘤率分别为25.2%、36.7%、43.1%;rAPC各剂量组能升高胸腺指数、脾指数和血清中细胞因子IL-6、TNF-α的水平.结论:rAPC可有效抑制H22肝癌的生长,促进小鼠胸腺和脾脏的生长发育,提高小鼠的免疫功能,从而抑制肿瘤的生长.     

Anti-genotoxic effect of Aloysia triphylla infusion against acrylamide-induced DNA damage as shown by the comet assay technique

Aloysia triphylla a perennial, bushy plant originally from South America has long been used in traditional medicine. Its aqueous extract contains considerable amounts of polyphenolic compounds, namely flavonoids and phenolic acids. In view of the interest in natural phenolic compounds as antioxidant in preventive medicine, this study was undertaken to investigate the chemoprotective effects of cedron leaves infusion against the genetic damage induced by acrylamide (AA) by using the alkaline version of the comet assay technique. Mice were separated in nine groups (eight animals each): (I) untreated, (II) negative control, (III) treated with infusion of cedron leaves 5%, 20 days twice a day, (IV) treated with AA (5 mg/kg b.w.), (V) treated with AA (20 mg/kg b.w.), (VI) treated with AA (30 mg/kg b.w.), (VII) treated with AA (50 mg/kg b.w.), (VIII) pretreated with infusion and treated with AA (50 mg/kg b.w.) and (IX) positive control (cyclophosphamide, 20 mg/kg b.w.). Three hundred blast cells were digitally evaluated per animal from three different slides (100 each). Media of tail moment (TM) values were analyzed by ANOVA test. No statistical differences (p>0.05) were found between untreated animals, negative control and infusion-treated mice. A single dose of AA-induced genetic damage as revealed by a statistically significant increase in TM values (p<0.01). Pretreatment with infusion prior to AA injection significantly reduces the capacity of AA to induce genetic damage. In these conditions, tail moments values did not differ from data obtained in negative control (p>0.05) and exhibit statistical differences from animals treated only with AA (p<0.01). Cell viability was at least 90% in all cases as measured by the trypan blue exclusion method. The ferric reducing ability of plasma (FRAP) method reveals that the plasma of infusion-treated mice has a significantly higher antioxidant capacity than plasma from controls (p<0.01). The results suggest that the infusion could exerts an in vivo chemo protective action, probably due to its scavenging potency towards free radicals.