Chlorella vulgaris triggers apoptosis in hepatocarcinogenesis-induced rats

Chlorella vulgaris (CV) has been reported to have antioxidant and anticancer properties. We evaluated the effect of CV on apoptotic regulator protein expression in liver cancer-induced rats. Male Wistar rats (200∼250 g) were divided into eight groups: control group (normal diet), CDE group (choline deficient diet supplemented with ethionine in drinking water to induce hepatocarcinogenesis), CV groups with three different doses of CV (50, 150, and 300 mg/kg body weight), and CDE groups treated with different doses of CV (50, 150, and 300 mg/kg body weight). Rats were sacrificed at various weeks and liver tissues were embedded in paraffin blocks for immunohistochemistry studies. CV, at increasing doses, decreased the expression of anti-apoptotic protein, Bcl-2, but increased the expression of pro-apoptotic protein, caspase 8, in CDE rats, which was correlated with decreased hepatoctyes proliferation and increased apoptosis as determined by bromodeoxy-uridine (BrdU) labeling and terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling (TUNEL) assay, respectively. Our study shows that CV has definite chemopreventive effect by inducing apoptosis via decreasing the expression of Bcl-2 and increasing the expression of caspase 8 in hepatocarcinogenesis-induced rats. Project supported by Department of Biochemistry, Faculty of Medicine, UKM Medical Center, Universiti Kebangsaan Malaysia, and the Malaysian Ministry of Science and Technological Innovation (MOSTI)     

Expression and significance of cyclin D1, p27kip1 protein in bronchioloalveolar carcinoma

INTRODUCTION Bronchioloalveolar carcinoma (BAC) is a par-ticular subtype of pulmonary adenocarcinoma de-rived from clara cell and type II pneumocyte. BAC cells grow along and within alveolar spaces while the alveolar framework of the lung is preserved. The incidence of BAC appears to be rising recently. The etiology and pathogenesis of this unique neoplastic disease are still unclear; many studies of oncogene and tumor suppressor gene expression include BAC with all adenocarcinoma o…     

Ki-67在肝细胞肝癌中的表达及其临床意义

[目的]探讨抗增殖核蛋白的单克隆抗体(Ki-67)在肝细胞肝癌(HCC)中的表达情况及临床意义.[方法]采用免疫组织化学方法检测51例HCC病人术后存档的石蜡标本中Ki-67的表达情况.[结果]HCC中Ki-67阳性表达率58.8％.Ki-67表达阳性率与肿瘤大小、门脉癌栓和肿瘤分级相关,与肝硬化、病灶数目、包膜、AFP及HBsAg不相关,Ki-67阳性表达组HCC病人术后无瘤生存期均显著低于阴性表达组.[结论]Ki-67对HCC的浸润转移均起促进作用,是HCC预后的重要标志物.     

Expression and significance of cyclin D1, p27kip1 protein in bronchioloalveolar carcinoma

Purpose: To investigate the relationship between expression of cell cycle-related protein cyclin D1, p27kipl and the pathogenesis of bronchioloalveolar carcinoma (BAC) and the value of prediction of prognosis. Methods: Cyclin D 1 and p27kip 1 protein were detected by immunohistochemical En Vision method in 43 BACs. Results: The positivity of cyclin D 1 in BAC was 65.1% (28/43), which was significantly higher than that in normal pulmonary tissue (0/13), P＜0.01. No statistically significant association was found between cyclin D1 expression data and sex, age, tobacco-use history, histologic subtype (mucinous vs nonmucinous), stromal fibrosis, lymph node metastasis, clinical stage or postoperative survival period (P＞0.05), while cyclin D1 expression was found to be negatively correlated with tumor size (P＜0.05). The positivity of p27kipl in BACs was 51.2% (22/43), significantly lower than that in normal pulmonary tissue (12/13), P＜0.01. p27kipl expression level was not associated with sex, age, tobacco-use history, tumor size or histologic subtype (P＞0.05), but was negatively correlated with stromal fibrosis, lymph node metastasis and clinical stage (P＜0.05); and positively associated with postoperative survival period (P＜0.01). The survival rate of p27kipl positive group was significantly higher than that of p27kipl negative group (P＜0.01). No statistically significant correlation was found between cyclin D 1 and p27kipl expression. Conclusions: Increased cyclin D1 expression and decreased p27kip 1 expression are related to the pathogenesis of BAC;decreased p27kipl expression is associated with metastasis progression; immunodetection ofp27kip 1 is useful for assessment of prognosis.     

HDAC1、8在肺泡Ⅱ型上皮细胞间质转化中的表达及意义

目的：观察TGF-β1作用下,肺泡Ⅱ型上皮细胞RLE-6TN在EMT过程中HDAC1、8的表达情况及TSA对TGF-β1诱导细胞EMT的影响。方法：将TGF-β1加入到体外培养的细胞中,于不同时间收取细胞,采用WB及Real-time RT-PCR检测HDAC1、8及E-cad、α-SMA的表达情况。然后将TGF-β1加入经过TSA预处理的细胞中,于不同时间收取细胞,重新检测上述基因。结果：加入TGF-β1后,E-cad蛋白表达下调;α-SMA蛋白表达上调;HDAC1蛋白表达上调;HDAC8蛋白表达下调。E-cad mRNA表达下调;α-SMA及HDAC8的mRNA均于12 h表达上调,6 h、24 h表达下调;HDAC1 mRNA表达于6 h下调,24 h上调。加入TSA后,E-cad蛋白表达上调;α-SMA、HDAC1及HDAC8蛋白的表达均下调。E-cad mRNA表达上调;α-SMA mRNA表达于6 h、12 h上调,24 h下调;HDAC1 mRNA表达下调;HDAC8 mRNA表达上调。结论：TGF-β1体外诱导的RLE-6TN细胞EMT,可以通过应用TSA抑制其获得α-SMA表型、失去E-cad表型,从而部分逆转TGF-β1诱导的肺泡Ⅱ型上皮EMT。     

Screening the active constituents of Chinese medicinal herbs as potent inhibitors of Cdc25 tyrosine phosphatase, an activator of the mitosis-inducing p34cdc2 kinase

Objective: To screen and evaluate the active constituents of Chinese medicinal herbs as potent inhibitors of Cdc25 phosphatase. Methods: The affinity chromatography purified glutashione-S-transferase/Cdc25A phosphatase fusion protein and Cdc2/cyclin B from the extracts of starfish M phase oocytes are used as the cell cycle-specific targets for screening the antimitotic constituents. We tested 9 extracts isolated from the Chinese medicinal herbs and vegetables including the agents currently used in cancer treatment by measuring the inhibition of Cdc25A phosphatase and Cdc2 kinase activity. The antitumor activity of the extracts was also evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and flow cytometry. Results: Cdc25A inhibitory activity and antitumor activity are detected in the extracts isolated from three Chinese medicinal herbs Agrimona pilosa; Herba solani lyrati; Galla chinesis. Conclusion: We found three extracts isolated from Chinese medicinal herbs have potential inhibitory activity of Cdc25 phosphatase using a highly specific mechanism-based screen assay for antimitotic drug discovery.     

Influence of gonadotropin-releasing hormone agonist on the effect of chemotherapy upon ovarian cancer and the prevention of chemotherapy-induced ovarian damage: an experimental study with nu/nu athymic mice

Background and objective: Gonadotropin-releasing hormone (GnRH) plays an important role in the regulation of ovarian function and ovarian cancer cell growth. In this study, we determined whether administration of the GnRH agonist (GnRHa), triporelin, prior to cisplatin treatment affects cisplatin and/or prevents cisplatin-induced ovarian damage. Methods: nu/nu mice were injected with ovarian cancer OVCAR-3 cells intraperitoneally. After two weeks, the mice were treated with saline (control), cisplatin, GnRHa, or cisplatin plus GnRHa for four weeks. At the end of the experimental protocol, blood, tumor, ovary, and uterine tissues were resected for hematoxylin and eosin (H&E) staining, immunohistochemical analyses of Ki67, nuclear factor-κB (NF-κB), and caspase-3, transmission electron microscopy of apoptosis, or enzyme-linked immunosorbent assay (ELISA) analyses of anti-Mullerian hormone (AMH). Results: Cisplatin treatment effectively inhibited tumor growth in mice treated with human ovarian cancer cells; however the treatment also induced considerable toxicity. Immunohistochemical analyses showed that Ki67 expression was reduced in cisplatin-treated mice compared to control (P<0.05), but there was no statistically significant differences between cisplatin-treated mice and cisplatin plus GnRHa-treated mice (P>0.05), while expressions of NF-κB and caspase-3 were reduced and induced, respectively, in cisplatin-treated mice and cisplatin plus GnRHa-treated mice. Apoptosis occurred in the GnRHa, cisplatin, and cisplatin plus GnRHa-treated mice, but not in control mice. Ovaries exposed to GnRHa in both GnRHa mice and cisplatin-treated mice (combination group) had significantly more primordial and growth follicles and serum levels of AMH than those in the control mice and cisplatin-treated mice (P<0.05). Conclusions: Administration of GnRHa to mice significantly decreased the extent of ovarian damage induced by cisplatin, but did not affect the anti-tumor activity of cisplatin.     

Immunomodulatory function of orally administered thymosin α1

INTRODUCTION The thymus is a vital immune organ and plays a very important role in the development and mainte- nance of the lymph system. The extract of animal thymus has been used in clinical practice as an ad- junct treatment in patients with carcinoma, viral in- fection and immunodeficiency, but the extract con- taining miscellaneous animal proteins may easily lead to allergy in patients. Thymosin α1 (Tα1) is the most potent ingredient in thymosin and the biological ac- tivity of pur…     

Chlorella vulgaris triggers apoptosis in hepatocarcinogenesis-induced rats

Chlorella vulgaris (CV) has been reported to have antioxidant and anticancer properties. We evaluated the effect of CV on apoptotic regulator protein expression in liver cancer-induced rats. Male Wistar rats (200-250 g) were divided into eight groups: control group (normal diet), CDE group (choline deficient diet supplemented with ethionine in drinking water to induce hepatocarcinogenesis), CV groups with three different doses of CV (50, 150, and 300 mg/kg body weight), and CDE groups treated with different doses of CV (50, 150, and 300 mg/kg body weight). Rats were sacrificed at various weeks and liver tissues were embedded in paraffin blocks for immunohistochemistry studies. CV, at increasing doses, decreased the expression of anti-apoptotic protein, Bcl-2, but increased the expression of pro-apoptotic protein, caspase 8, in CDE rats, which was correlated with decreased hepatoctyes proliferation and increased apoptosis as determined by bromodeoxy-uridine (BrdU) labeling and terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling (TUNEL) assay, respectively. Our study shows that CV has definite chemopreventive effect by inducing apoptosis via decreasing the expression of Bcl-2 and increasing the expression of caspase 8 in hepatocarcinogenesis-induced rats.     

Expression of MMP-9 in hepatic sinusoidal obstruction syndrome induced by Gynura segetum

Background and objective:Hepatic sinusoidal obstruction syndrome(HSOS) is characterized by painful hepatomegaly,ascites,increased body weight,and jaundice.Gynura segetum(Compositae),a plant widely used in Chinese traditional medicine,often leads to the development of HSOS.However,the mechanism is unclear.The aim was to study the role of matrix metalloproteinase-9(MMP-9) in the onset of HSOS induced by Gynura segetum.Methods:Twenty-five male Sprague-Dawley rats were randomly divided into two groups.Twenty were exposed to 600 mg/kg daily Gynura segetum extract solution for three weeks;five control rats were exposed to tap water alone.Liver sections were evaluated by light microscopy with a modified scoring system.Routine transmission electron microscopy(TEM) methods were used to evaluate the ultrastructual features of fixed liver tissue,and blood samples were collected to determine liver enzyme concentrations.MMP-9 expression was assessed by both immunohistochemical staining and enzyme-linked immunosorbent assay(ELISA) methods.Results:A stable and reproducible rat model of HSOS was achieved by long-term exposure to Gynura segetum extract.The treated rats presented clinical symptoms and the histopathological manifestation of HSOS,including abnormal liver enzyme concentrations(alanine aminotransferase(ALT):(84.8±13.62) vs.(167.0±72.63) U/L,P<0.05;aspartate aminotransferase(AST):(27.6±6.31) vs.(232.8±108.58) U/L,P<0.05).Hematoxylin and eosin(H&E) staining and TEM together revealed deposition of red blood cells,the damage and destruction of hepatic sinusoidal endothelial cells,collapse of hepatic sinusoids,hemorrhage of subendothelial cells,atrophy and destruction of hepatocytes,etc.Compared with controls,the expression of MMP-9 in the blood sample,the lung and liver tissues of HSOS rats was increased.Conclusions:MMP-9 may have an important role in early pathological changes of HSOS,and thus the onset of the disease.     

莫诺苷对局灶性脑缺血-再灌注大鼠海马CyclinD1及CDK6的影响

目的：观察莫诺苷对局灶性脑缺血-再灌注大鼠细胞周期蛋白Cyclin D1及CDK6的影响。方法：用线栓法制备大鼠局灶性脑缺血-再灌注模型。将15只Sprague-Dawley（SD）大鼠随机分为假手术组,模型组,莫诺苷组（270、90、30 mg·kg-1）。利用免疫组化方法检测大鼠患侧海马Cyclin D1、CDK6表达。结果：与假手术组相比,模型组Cyclin D1及CDK6表达显著增加;给予莫诺苷治疗后,与模型组相比,莫诺苷（270、90 mg·kg-1）剂量组能显著降低大鼠Cyclin D1及CDK6的表达。结论：莫诺苷能通过降低脑缺血-再灌注后Cyclin D1及CDK6的表达而发挥神经保护作用。     

天麻素对模型大鼠海马Caspase-3表达的影响

目的：观察天麻素对癫痫大鼠海马Caspase-3表达的影响及其脑保护作用。方法：120只健康成年雄性Wistar大鼠随机分为对照组、PTZ（戊四氮）组、VPA（丙戊酸钠）组、Gb（天麻素大剂量）组和Gs（天麻素小剂量）组（n=24）。对照组和PTZ组分别以生理盐水（4mL/kg·d）灌胃;VPA组给予丙戊酸20mg．kg^-1,Gb组和Gs组分别给予天麻素200mg．kg^-1和100mg．kg^-1;每天1次,连续7天。造模第一天,除对照组大鼠外,余者均腹腔注射戊四氮75mg／kg,记录动物行为学变化。于致痫后12h、2d、5d和7d相应时间点取材,制备脑标本,免疫细胞化学术检测caspase-3表达。结果：致痫后12h,P11Z组Caspase-3有微量表达,其余各组几乎无表达;2—7d。PTZ组Caspase-3表达增加。与PTZ组比较,Gb组、Gs组及VPA组Caspase-3阳性表达降低,差异显著（P〈0．05）;与VPA组比较,Gb组和Gs组Caspase-3表达无显著差异（P〉0．05）。结论：天麻素能降低致痫大鼠海马神经元Caspase-3表达;可能通过抑制神经元凋亡发挥脑保护作用。     

肝大部切除术后PGE1对肝功能保护机制研究进展

综述了近年手术对肝功能损伤的机制和前列腺素1（PGE1）对肝功能保护机制的研究进展。肝大部切除术可通过缺血再灌注损伤和/或急性期应答反应,导致肝功能损害,PGE1能通过细胞上相应受体和信号转导系统,调控各种炎症反应因子产生,对肝细胞保护和再生有重要作用。     

Expression of bone morphogenetic protein 2, 4, and related components of the BMP signaling pathway in the mouse uterus during the estrous cycle

The objective was to investigate the expression of bone morphogenetic protein (BMP) family members in the mouse uterus during the estrous cycle by real-time polymerase chain reaction (PCR) and immunohistochemistry. Uterine samples from Swiss ICR mice were collected and dissected free of surrounding tissue. One uterine horn was snap frozen in liquid nitrogen immediately after collection and stored at −80 °C for RNA extraction, and the other was fixed in 40 mg/ml paraformaldehyde at room temperature for immunolocalization of BMP2 protein. Real-time PCR analysis showed that the expression level of Bmp2 was significantly higher at proestrus than at estrus and metestrus (P<0.05). The relative abundance of Bmp4 exhibited significant fluctuations, but there were no statistically significant differences between the expression levels of Bmp2 and Bmp4 (P>0.05). The expression levels of Bmpr1a and Bmpr2 remained unchanged during estrous cycles. However, the level of Bmpr1b mRNA decreased significantly at estrus (P<0.05), increasing subsequently at metestrus. Furthermore, the level of Bmpr1b mRNA was significantly lower than those of Bmpr1a and Bmpr2 mRNA at the corresponding stages (P<0.05). All three receptor-regulated Smads (R-Smads) detected were differentially expressed in the mouse uterus and the expression levels of Smad1 and Smad5 were significantly higher than that of Smad8 (P<0.05). In addition, the expression level of Smad4 did not change substantially throughout the estrous cycle. Immunohistochemical experiments revealed that BMP2 protein was differentially expressed and localized mainly in the uterine luminal and glandular epithelial cells throughout the estrous cycle. In conclusion, our results provide information about the variation in the mRNA levels of Bmp2 and Bmp4 and related components of the BMP signaling pathway. The data provide quantitative and useful information about the roles of endometrial BMP proposed and demonstrated by others, such as the degradation and remodeling of the endometrium.     

大肠癌组织中CD105和Ki-67的表达及其临床病理学意义

目的探讨大肠癌组织中CD105和Ki-67的表达及其意义。方法采用免疫组织化学PV-9000法检测50例大肠癌组织和10例正常大肠黏膜组织中CD105和Ki-67的表达,检测肿瘤内微血管密度（MVD）以及计算增殖指数（PI）。结果Ki-67和CD105在大肠癌组织中呈高表达（51．34±24．44,16．44±6．33）,与正常黏膜组织中的表达（21．10±8.66,4．00±1．25）相比较,差异有极显著的统计学意义（P〈0．01）。Ki-67的表达与大肠癌的分化程度、临床分期和淋巴结的转移有关,有显著统计学意义（P〈0．05）。MVD与大肠癌的分化程度、临床分期有关（P〈0．05）,但与淋巴结转移无关（P〉0．05）。Ki-67在大肠癌高微血管密度组的表达（68.28±16．91）明显高于低微血管密度组（30．59±13．90）,差异非常显著（P〈0．01）,Ki钾的表达与MYD呈正相关（r=0．28,P〈0.01））。结论CD105和Ki-67在大肠腺癌的发生发展中起着重要作用,并且同时检测两者的表达对大肠腺癌的抗血管治疗和判断其预后有一定应用价值。     

束缚应激小鼠对溶血空斑的影响

目的：研究束缚应激对小鼠溶血空斑形成试验的影响．方法：４０只小鼠,随机等分为对照组和应激组,用直接空斑形成试验（Ｐｌａｑｕｅ－ｆｏｒｍｉｎｇｃｅｌｌａｓｓｎｙ,ＰＦＣ）玻片法,测其溶血空斑形成数量．结果：应激组溶血空斑形成数量明显降低（Ｐ＜０．０１）．结论：束缚应激可致小鼠体液免疫功能降低．     

Thyroid dysfunction,either hyper or hypothyroidism,promotes gallstone formation by different mechanisms

We have investigated comprehensively the effects of thyroid function on gallstone formation in a mouse model. Gonadectomized gallstone-susceptible male C57BL/6 mice were randomly distributed into three groups each of which received an intervention to induce hyperthyroidism, hypothyroidism, or euthyroidism. After 5 weeks of feeding a lithogenic diet of 15% (w/w) butter fat, 1% (w/w) cholesterol, and 0.5% (w/w) cholic acid, mice were killed for further experiments. The incidence of cholesterol monohydrate crystal formation was 100% in mice with hyperthyroidism, 83% in hypothyroidism, and 33% in euthyroidism, the differences being statistically significant. Among the hepatic lithogenic genes, Trβ was found to be up-regulated and Rxr down-regulated in the mice with hypothyroidism. In contrast, Lxrα, Rxr, and Cyp7α1 were up-regulated and Fxr down-regulated in the mice with hyperthyroidism. In conclusion, thyroid dysfunction, either hyperthyroidism or hypothyroidism, promotes the formation of cholesterol gallstones in C57BL/6 mice. Gene expression differences suggest that thyroid hormone disturbance leads to gallstone formation in different ways. Hyperthyroidism induces cholesterol gallstone formation by regulating expression of the hepatic nuclear receptor genes such as Lxrα and Rxr, which are significant in cholesterol metabolism pathways. However, hypothyroidism induces cholesterol gallstone formation by promoting cholesterol biosynthesis.     

细胞周期蛋白D1和细胞周期蛋白依赖激酶4在宫颈鳞癌中的表达

目的：探讨Cyclin D1和CDK4在正常宫颈上皮、CIN和宫颈鳞癌中表达状况。方法：收集子宫颈活检及手术标本118例，其中正常鳞状上皮14例，CIN25例，宫颈鳞癌79例。采用SP免疫组织化学方法，检测Cyclin D1和CDK4蛋白的表达情况。结果：Cyclin D1和CDK4在正常宫颈上皮、CIN和宫颈鳞癌的中表达水平存在显著性差异（P〈0．05），在不同分化程度的宫颈鳞癌间无显著性差异（P〉0．05）。结论：在宫颈鳞癌发生过程中伴右Cyclin D1和CDK4表达水平的增高．检测其含量改变有助于宫颈鳞癌诊断。