裸细胞型ALK阳性间变性大细胞淋巴瘤1例并文献复习

回顾性分析陕西省人民医院收治的1例胸壁裸细胞型淋巴瘤激酶(anaplastic lymphoma kinase,ALK)阳性间变性大细胞淋巴瘤的临床病理特点并进行文献复习。该患者为51岁女性,因40 d前发现胸壁包块入院。超声示:左侧胸壁肌层内囊实性包块,大小约为8 cm×6 cm×3 cm;腹腔及后腹膜多发异常增大淋巴结,腹腔积液。行胸壁包块穿刺活体组织检查,小条穿刺组织镜下所见:纤维组织内有弥漫性小圆细胞浸润,细胞质嗜酸性,核呈圆形或卵圆形,部分核偏位,呈“马蹄形”。免疫组织化学染色示:细胞角蛋白、CD2、CD3、CD5、CD7、CD20、CD38、CD138、CD56及细胞周期蛋白D1均为阴性,ALK和CD30均为强阳性。T细胞抗原受体基因重排阳性。病理诊断为裸细胞型ALK阳性间变性大细胞淋巴瘤。该肿瘤非常罕见,其诊断需结合临床表现、组织学形态、免疫表型及分子遗传学改变进行综合判断,避免误诊。

从结缕草的裸细胞再生植株

日本草坪(ジャバン·タヘフグラス)的研究组在日本最先确立了从结缕草的原生质体中再生植株的技术.这为使用基因工程方法育成抗病性,抗虫性和绿化期长的品种开辟了道路.也可以说是对不能使用农药的高尔夫球场作出了分子生物学的答案.详情于3月31日在东京召开的日本育种学会上发表了.报告这项工作的是该公司的猪熊千惠、杉浦清之、赵彻、金子诚二等.猪熊等从市售的结缕草种子诱导愈伤组织,并使原生质体的再生获得成功.愈伤诱导效率最高的条件是在LS培养基中添加(5 mg/1)2,4-D,放置在28℃和黑暗条件下.把在这阶段诱导的愈伤组织移植到添加了3 mg/l的2,4-D和氨基酸的N6培养基上,在28℃和黑暗条件下进行液体培养. 进一步分离原生质体,用在“K8培养基”添加2,4-D的培养基培养,形成细胞团后。

海藻酸钙微球与海藻酸钙-聚赖氨酸-海藻酸钙微囊包裹对不同种类细胞生长的影响

目的:观察海藻酸钙微球或海藻酸钙-聚赖氨酸-海藻酸钙微囊包裹对不同特性细胞生长的影响。方法:实验于2004-04/2005-04在解放军总医院老年医学研究所细胞生物学研究室完成。细胞:牛肾上腺嗜铬细胞、Swiss小鼠胚胎成纤维细胞株(3T3)、人早幼粒白血病细胞株。仪器和试剂:加拿大Sun氏静电微囊发生仪;体视显微镜;海藻酸钠,聚赖氨酸、氯化钙、柠檬酸钠。制备海藻酸钙微球和海藻酸钙-聚赖氨酸-海藻酸钙微囊,分别用其包裹3T3细胞、牛肾上腺嗜铬细胞及人早幼粒白血病细胞。按照细胞的不同及包裹材料的不同,分为裸细胞组、海藻酸钙微球组、海藻酸钙-聚赖氨酸-海藻酸钙微囊组。在光镜下观察体外培养的微球、微囊及细胞的形态、存活率、增殖率。细胞增殖率=培养后细胞数-培养前细胞数/培养前细胞数×100%。结果:光镜下见包裹后即刻海藻酸钙微球和海藻酸钙-聚赖氨酸-海藻酸钙微囊呈大小均一的半透明圆球,直径150～300μm,表面光滑。培养24h时,牛肾上腺嗜铬细胞、3T3或人早幼粒白血病细胞均呈单个细胞,均匀分散于微球或微囊内。培养120h时,牛肾上腺嗜铬细胞、3T3和人早幼粒白血病细胞在海藻酸钙-聚赖氨酸-海藻酸钙微囊内聚集成团块,而在海藻酸钙微球内仍呈单个细胞散在分布。①海藻酸钙微球或海藻酸钙-聚赖氨酸-海藻酸钙微囊内高增殖活性的3T3或人早幼粒白血病细胞的存活率显著高于牛肾上腺嗜铬细胞(74.38±7.72,89.08±3.05,87.48±3.43,92.40±7.56,72.67±2.08,89.33±5.13,P<0.01);海藻酸钙-聚赖氨酸-海藻酸钙微囊内细胞存活率均显著高于海藻酸钙微球内同一种细胞的存活率(87.88±3.38,89.08±3.05,89.63±5.55,85.33±4.86,92.40±7.56,95.08±3.81,83.67±5.51,89.33±5.13,85.33±2.08,P<0.01)。②在不同状态下,培养72和120h时,人早幼粒白血病细胞或3T3细胞的增殖率存在差异,海藻酸钙微球或海藻酸钙-聚赖氨酸-海藻酸钙微囊内同种细胞的增殖率显著高于裸细胞组(2.54±0.65,5.20±2.38,2.30±0.70,2.26±0.83,3.89±1.89,13.37±16.14,P<0.05);而在不同状态下牛肾上腺嗜铬细胞的增殖率差异无显著性(P>0.05)。③在相同状态下,培养72和120h时,人早幼粒白血病细胞增殖率明显高于3T3或牛肾上腺嗜铬细胞(18.66±7.76,39.11±16.06,P<0.01);而3T3与牛肾上腺嗜铬细胞的增殖率差异无显著性(P>0.05)。④对同一种细胞,培养24,72和120h时,海藻酸钙微球与海藻酸钙-聚赖氨酸-海藻酸钙微囊相比,其细胞的增殖率差异均无显著性(P>0.05)。结论:在培养条件下,海藻酸钙微球或海藻酸钙-聚赖氨酸-海藻酸钙微囊包裹似更利于该3种细胞的生长;海藻酸钙-聚赖氨酸-海藻酸钙微囊包裹较海藻酸钙微球包裹更利于同种细胞的生长。

急性白血病免疫分型中的少见类型多系表达及“裸型”分析

目的：研究急性白血病（ＡＬ）细胞抗原表达特征及临床意义。方法：选用１４～１６ 种单克隆抗体，采用免疫荧光的方法，对９６例ＡＬ患者进行免疫分型。结果：９ 例为多系表达，即同时表达三系以上的抗原（急性髓细胞白血病－ＡＭＬ７ 例，急性淋巴细胞白血病－ＡＬＬ２ 例），ＡＭＬ中多系表达发生率为１０．８％ ，ＡＬＬ中为６．５％ 。“裸细胞”型的发生率ＡＭＬ中７．７％ ，ＡＬＬ中６．４％ 。结论：呈多系表达的ＡＭＬ多为Ｍ５ 型，ＣＤ１４高表达，Ｐ－１７０ 高表达，ＣＲ率低；

HIF-1α及相关基因与无功能垂体腺瘤侵袭性的相关性研究

目的探讨HIF-1α及其下游相关基因与垂体裸细胞瘤和嗜酸性细胞瘤侵袭性的相关性。方法选择46例裸细胞瘤(侵袭性28例,非侵袭性18例),46例嗜酸细胞瘤(侵袭性11例,非侵袭性35例)和3例正常垂体组织。采用q RT-PCR检测HIF-1α基因及其下游相关基因MMP9、ECAD和NCAD在正常垂体组织和肿瘤组织的m RNA表达情况。结果 HIF-1α基因表达在侵袭性裸细胞瘤高于非侵袭性裸细胞瘤(P=0.026);而HIF-1α基因表达在侵袭性嗜酸性细胞瘤和非侵袭性嗜酸性细胞瘤之间,无明显统计学差异。MMP9基因表达在侵袭性裸细胞瘤高于非侵袭性裸细胞瘤(P=0.023)。NCAD和ECAD基因表达在侵袭性裸细胞瘤和非侵袭性裸细胞瘤之间,无明显统计学差异。结论 HIF-1α基因与裸细胞瘤侵袭性相关,其可能通过上调MMP9参与裸细胞瘤侵袭生长。HIF-1α可作为侵袭性裸细胞瘤的特异性标记物。

Improvement of in vitro Cytoplasmic Maturation of Denuded Porcine Oocytes by Coculture with Follicular Mural Granulosa Cells

[Objective] This study aimed to improve the in vitro maturation quality of denuded porcine oocytes and provide scientific basis for establishing a stable and efficient denuded oocyte culture system. [Method] The first polar body extrusion rate, oocyte glutathione （GSH） content, positive rate of brilliant cresyl blue （BCB） staining and development potential of activated oocytes or fertilized oocytes were employed as main indicators to investigate the effects of follicular mural granulosa cell （MGC） coculture on cytoplasmic maturation of cumulus cell-removal oocytes （Denuded Oocyte, DO）. [Result] According to in vitro maturation results, compared with DO group, the first polar body extrusion rate of porcine oocytes in DO＋MGC group was not significantly different, but the nuclear maturation process was improved and was more similar to that in COC （cumulus-oocyte complex） group. Detection of GSH content in mature oocytes showed that there was no significant difference between DO＋ MGC group （optical density of 1 053.67） and COC group （optical density of 1 426.00） or between DO＋MGC group and COC＋GC group （optical density of 1 541.00）, however, GSH content in mature oocytes of DO group （optical density of 724.67） was significantly lower than that of COC group and COC＋GC group （P0.05）. Detection of glucose-6-phosphate dehydrogenase （G6PDH） activity showed that there was no significant difference in BCB positive oocyte rate between DO ＋MGC group （88.26% ） and COC group （92.75%） or between DO＋MGC group and DO group （82.86% ）, however, BCB positive oocyte rate of DO group was significantly lower than that of COC group （P0.05）. Furthermore, the cleavage rate and blastocyst rate of activated mature oocytes derived from DO ＋MGC group （94.98% and 43.67% , respectively） were significantly higher than those from DO group （52.54% and 8.97%, respectively） （P0.05）, and were not significantly different compared with those from COC group （97.11% and 38.30%, respectively）. In addition, the cleavage rate of fertilized oocytes derived from DO＋MGC group （72.65%） showed no significant difference compared with that from DO group （63.59%）, but the blastocyst rate of DO＋MGC group was significantly higher than that of DO group （9.88%） （P0.05）. [Conclusion] MGC coculture can significantly improve the in vitro cytoplasmic maturation quality of denuded porcine oocytes, thereby enhancing the subsequent developmental potential.

Cytological Mechanism of Cytoplasmic Inheritance in Pinus tabulaeformis: Ⅱ. Transmission of Male and Female Organelles During Fertilization and Proembryo Development

In an earlier report the ultrastructure and nucleoid organelles of male gamete in Pinus tabulaeformis Carr. have been described. Presently, the ultrastructure of the cytoplasm of the egg cell and pollen tube—immediately before fertilization and during cytoplasmic transmission of male gametophyte—has been described for the same species. The fate of parental plastids and mitochondria in the proembryo has also been followed. The mature egg cell contains a large amount of mitochondria, but seems to lack normal plastids. Most plastids have transformed into large inclusions. Apart from the large inclusions, there are abundant small inclusions and other organelles in the egg cell. During fertilization, pollen tube penetrates into the egg cell at the micropylar end and thereafter the contents are released. Plastid and mitochondrion of male origin are lacking near the fusing sperm_egg nuclei. The second sperm nucleus—not involved in karyogamy—remains at a site near the receptive vacuole. This nucleus is surrounded by large amount of male cytoplasm containing mixed organelles from the sperm cell, tube cell, and egg cell. At the free nuclear proembryo stage, organelles of male and female origin are visible in the perinucleus_cytoplasmic zone. Most of the mitochondria have the same morphological features as those in the egg cell. Some of the mitochondria appear to have originated from the sperm and tube cells. Plastids are most likely of male gametophyte origin because they have similar appearance as those of the sperm and tube cell. Large inclusions in the egg cell become vacuole_like. Paternal plastids have been incorporated into the neocytoplasm of the proembryo. In the cellular proembryo, maternal mitochondria are more abundant. Plastids resembling those of the sperm and tube cell are still present. These cytological results clearly show that in P. tabulaeformis, plastids are inherited paternally and mitochondria bipaternally. The cytological mechanism of plastid and mitochondrion inheritance in gymnosperm is discussed.

Change in expression of apoptosis genes after hyperthermia,chemotherapy and radiotherapy in human colon cancer transplanted into nude mice

AIM:To investigate the change in expression of p53 ,Bcl-2 ,and Bax genes in human colon cancer cells transplanted into nude mice after hyperthermia,chemotherapy,radiotherapy,thermochemotherapy,thermoradiotherapy and thermochemoradiotherapy. METHODS:Human colon cancer cell line (HT29) was transplanted into the hind limbs of nude mice. Under laboratory simulated conditions of hyperthermia (43℃,60 min),the actual radiation doses and doses of mitomycin C (MMC) were calculated in reference to the clinical radiotherapy for human rectal cancer and chemotherapy prescription for colon cancer. The mice were divided into 6 groups according to the treatment approaches:hyperthermia,chemotherapy,radiotherapy,thermochemotherapy,thermoradiotherapy,and thermochemoradiotherapy. The mice were sacrificed at different time points and the tumor tissue was taken for further procedures. The morphologic changes in membrane,cytoplasm and nuclei of tumor cells of p53,Bcl-2,and Bax after treatment,were observed by immunohistochemistry staining. RESULTS:All of the six treatment modalities down-regulated the expression of p53,Bcl-2 and up-regulated the expression of Bax at different levels. The combined therapy of hyperthermia,with chemotherapy,and/or irradiation showed a greater effect on down-regulating the expression of p53 (0.208 ± 0.009 vs 0.155 ± 0.0115,P < 0.01) and Bcl-2 (0.086 ± 0.010 vs 0.026 ± 0.0170,P < 0.01) and up-regulating Bax expression (0.091 ± 0.0013 vs 0.207 ± 0.027,P < 0.01) compared with any single therapy.CONCLUSION:Hyperthermia enhances the effect of radio-and chemotherapy on tumors by changing the expression of apoptosis genes,such as p53,Bcl-2 and Bax.

Anticancer activity of genistein on implanted tumor of human SG7901 cells in nude mice

AIM： To investigate genistein-induced apoptosis of implanted tumors of SG7901 cells in nude mice, and the relationship between this apoptosis and expression of Bcl-2 and Bax. METHODS： Establishing a transplanted tumor model by injecting human SG7901 cells into subcutaneous tissue of nude mice. Genistein （0.5, 1 and 1.5 mg/kg） was directly injected adjacent to the tumor, six times at 2-d intervals. Then, changes in tumor volume were measured continuously and tumor inhibition rate of each group was calculated. We observed the morphological alterations by transmission electron microscopy （TEN）, measured the apoptotic rate by the TUNEL staining method, and detected the expression of apoptosisregulated gene Bcl-2 and bax by immunohistochemical staining and RT-PCR. RESULTS： Genistein 0.5, 1 and 1.5 mg/kg significantly inhibited carcinoma growth when it was injected near the tumor by 10.8%, 29.9% and 39.6%, respectively. Genistein induced implanted tumor cells to undergo apoptosis, with apoptotic characteristics seen by TEM. The apoptosis index was increased progressively with increasing genistein dose （28.9% ± 1.2%, 33.8% ±1.6% and 37.7% ±1.2%）. The positive rate of Bcl-2 protein was decreased progressively （11.9%± 0.9%, 5.9%± 0.7% and 4.2% ±0.6%）, and the positive rate of bax protein was increased progressively （0.9% ±1.7%, 24.9% ±0.8% and 29.6% ± 1.7%） by immunohistochemical staining, with increasing dose of genistein. The density of Bcl-2 mRNA decreased progressively and the density of bax mRNA increased progressively with elongation of time by RT-PCR. CONCLUSION： Genistein was able to induce apoptosisof transplanted tumor cells. This apoptosis may be mediated by down-regulation of the apoptosis-regulated gene Bcl-2 and up-regulation of apoptosis-regulated gene bax.

Effect of 5-LOX/COX-2 common inhibitor DHDMBF30 on pancreatic cancer cell Capan2

AIM: To study the effect of 5-lipoxygenase/cyclooxy- genase-2 (5-LOX/COX-2) dual inhibitor 7-tert-butyl-2, 3-dihydro-3, 3-dimethyl substituted dihydrofuran 30 (DHDMBF30) on proliferation and apoptosis of the pancreatic cancer cell line Capan-2 and the effect of DHDMBF30 on human pancreatic cancer in a nude mouse model. METHODS: Investigate the effect of 5-LOX/COX-2 dual inhibitor DHDMBF30 on proliferation and apoptosis of the pancreatic cancer cell line Capan-2 by RT-PCR, MTT assay, FCM and electron microscope. Cell line Capan-2 was inoculated percutaneously on the outer thigh of 12 nude mice. The VEGF mRNA of transplantation tumor was detected by RT-PCR. RESULTS: DHDMBF30 inhibits the proliferation of cell line Capan2, reduces the expression of 5-LOX, COX-2 and VEGF. After Capan2 was treated with DHDMBF30, we found that the apoptosis peak of the experimental group was significantly higher than that of the contrast group (3.08 ± 1.89 vs 27.67 ± 0.52, P < 0.001). The tumor weight of the DHDMBF30 group was significantly lower than PBS control groups (1.35 ± 0.47 vs 2.92 ± 0.73, P < 0.01). Expression of VEGF in the DHDMBF30 group was significantly decreased. CONCLUSION: DHDMBF30 inhibits the proliferation ofthe pancreatic cell line Capan2, and induces apoptosis and inhibits the growth of pancreatic cancer in nude mice.

Angiostatin inhibits pancreatic cancer cell proliferation and growth in nude mice

AIM: To observe the biologic behavior of pancreatic cancer cells in vitro and in vivo, and to explore the potential value of angiostatin gene therapy for pancreatic cancer.METHODS: The recombinant vector pcDNA3.1(+)-angiostatin was transfected into human pancreatic cancer cells PC-3 with Lipofectamine 2000, and paralleled with the vector and mock control. Angiostatin transcription and protein expression were determined by immunofluorescence and Western blot. The stable cell line was selected by G418. The supernatant was collected to treat endothelial cells. Cell proliferation and growth in vitro were observed under microscope. Cell growth curves were plotted.The troms-fected or untroms-fected cells overexpressing angiostatin vector were implanted subcutaneously into nude mice. The size of tumors was measured, and microvessel density count (MVD) in tumor tissues was assessed by immunohistochemistry with primary anti-CD34antibody.RESULTS: After transfected into PC-3 with Lipofectamine 2000 and selected by G418, macroscopic resistant cell clones were formed in the experimental group transfected with pcDNA 3.1(+)-angiostatin and vector control. But untreated cells died in the mock control. Angiostatin protein expression was detected in the experimental group by immunofluorescence and Western-blot. Cell proliferation and growth in vitro in the three groups were observed respectively under microscope. After treatment with supernatant, significant differences were observed in endothelial cell (ECV-304) growth in vitro. The cell proliferation and growth were inhibited. In nude mice model, markedly inhibited tumorigenesis and slowed tumor expansion were observed in the experimental group as compared to controls, which was parallel to the decreased microvessel density in and around tumor tissue.CONCLUSION: Angiostatin does not directly inhibit human pancreatic cancer cell proliferation and growth in vitro,but it inhibits endothelial cell growthin vitro. It exerts the anti-tumor functions through antiangiogenesis in a paracrine way in vivo.

Anticancer activity of resveratrol on implanted human primary gastric carcinoma cells in nude mice

AIM: To investigate the apoptosis of implanted primary gastric cancer cells in nude mice induced by resveratrol and the relation between this apoptosis and expression of bcl-2 and bax. METHODS: A transplanted tumor model was established by injecting human primary gastric cancer cells into subcutaneous tissue of nude mice. Resveratrol (500 mg/kg, 1 000 mg/kg and 1 500 mg/kg) was directly injected beside tumor body 6 times at an interval of 2 d. Then changes of tumor volume were measured continuously and tumor inhibition rate of each group was calculated. We observed the morphologic alterations by electron microscope, measured the apoptotic rate by TUNEL staining method, detected the expression of apoptosis-regulated genes bcl-2 and baxby immunohistoch-emical staining and PT-PCR. RESULTS: Resveratrol could significantly inhibit carcinoma growth when it was injected near the carcinoma. An inhibitory effect was observed in all therapeutic groups and the inhibition rate of resveratrol at the dose of 500 mg/kg, 1 000 mg/kg and 1 500 mg/kg was 10.58%, 29.68% and 39.14%, respectively. Resveratrol induced implanted tumor cells to undergo apoptosis with apoptotic characteristics, including morphological changes of chromatin condensation, chromatin crescent formation, nucleus fragmentation. The inhibition rate of 0.2 mL of normal saline solution, 1 500 mg/kg DMSO, 500 mg/kg resveratrol, 1 000 mg/kg resveratrol, and 1 500 mg/kg resveratrol was 13.68±0.37%, 13.8±0.43%, 48.7±1.07%, 56.44±1.39% and 67±0.96%, respectively. The positive rate of bd-2 protein of each group was 29.48±0.51%, 27.56±1.40%, 11.86±0.97%, 5.7±0.84% and 3.92±0.85%, respectively by immunohistochemical staining. The positive rate of bax protein of each group was 19.34±0.35%, 20.88±0.91%, 40.02±1.20%, 45.72±0.88% and 52.3±1.54%, respectively by immunohistochemical staining. The density of bcl-2 mRNA in 0.2 mL normal saline solution, 1 500 mg/kg DMSO, 500 mg/kg resveratrol, 1 000 mg/kg resveratrol, and 1 500 mg/kg resveratrol decreased progressively and the density of bax mRNA in 0.2 mL normal saline solution, 1 500 mg/kg DMSO, 500 mg/kg resveratrol, 1 000 mg/kg resveratrol, and 1 500 mg/kg increased progressively with elongation of time by RT-PCR. CONCLUSION: Resveratrol is able to induce apoptosis of transplanted tumor cells. This apoptosis may be mediated by down-regulating apoptosis-regulated gene bcl-2 and up-regulating the expression of apoptosis-regulated gene bax.

Inhibition of betulinic acid to growth and angiogenesis of human colorectal cancer cell in nude mice

Objective： Angiogenesis plays a major role in the pathogenesis of many disorders. Vascular endothelial growth factor （VEGF） has been shown to be the key regulator of normal and pathological angiogenesis. Many studies showed that decreased expression of VEGF has been inhibited growth and migration of cancer cells. The aim of this study was to explore the effects of Betulinic acid on the VEGF expression and the growth of colorectal cell SW480 xenografts in nude mice. Methods： The xenografts derived from colorectal cell SW480 were established in BALB/C nude mice. Inoculated mice were randomly divided into negative control （corn oil）, low dose betulinic acid group （20 mg/kg/d） and high dose group （40 mg/kg/d）. After 22 days, the animals were sacrificed; tumor volume and weights were measured. The mRNA level of VEGF was analyzed by quantitative real-time polymerase chain reaction. The expression of VEGF protein was detected by immunohistochemistry. Results： The tumor weight was significantly lower in low and high dose groups than in corn oil group （1.12 ＋ 0.04, 0.43 ＋ 0.02 vs 2.08 ＋ 0.07; P 〈 0.05）. The mRNA levels of VEGF was also significantly lower in betulinic acid treated groups （0.72 ＋ 0.02, 0.38 ＋ 0.01; P 〈 0.05） than in control group （1.08 ＋ 0.04）. H＆E staining showed tumor tissue necrosis was observed in treatment groups. The positive expression of VEGF was lower in low and high dose groups than in corn eil group. Gray scale increased in low dose group and high dose group （121.1 ＋ 2.8, 156.2 ＋ 3.3, P 〈 0.05）. Conclusion： Betulinic acid had significant inhibitory effect on VEGF expression and tumors growth of human colorectal cancer xenografts in vivo, and down-regulation of VEGF expression may account for one of the molecular mechanisms of the anticancer effects of betulinic acid.

The effect of pre-low-dose X-ray radiation on tumor inhibition of HepG2 cells in tumor-bearing nude mice

Objective:The aim of this study was to discuss the effect of pre-low-dose X-ray radiation on P53,Bcl-2 and apoptosis of HepG2 cells in tumor-bearing nude mouse,and further explore the mechanism of low doses radiation.Methods:HepG2 cells were implanted subcutaneously into nude mice.14 days after the implanting,these mice were divided into 6 groups randomly,S group (sham-irradiation 0 cGy),D1 group (7.5 cGy,dosage rate=7.5 cGy/min),D2 group,(200 cGy,dosage rate=100 cGy/min),D1 + 2 h + D2 group,D1 + 6 h + D2 group and D1 + 12 h + D2 group.Tumor-bearing mice in each experimental group were executed at 24 h after the last irradiation.P53 and Bcl-2 were detected by immunohistochemical staining,the tumor tissues apoptosis were detected in site (Tunel).Results:Each combined exposure groups (D1 + 2 h + D2 group,D1 + 6 h + D2 group and D1 + 12 h + D2 group) compared with the D2 group,the percentages of positive P53 and Bcl-2 were decreased obviously,and the apoptotic indexs were increased (P < 0.01).Conclusion:Pre-low-dose radiation combined with the conventional radiation can increase the apoptosis of tumor tissues by decreasing the expression of P53 and Bcl-2,it can enhance the anti-tumor effect of conventional radiation,and it can have actual clinical significance on supporting radiotherapy.

Establishment of nude mice model of human osteosarcoma cell MG63 with different potential of metastasis

Objective: To establish a nude mice model of human osteosarcoma lung metastasis. Methods: The growth of human osteosarcoma cell sublines M8 and M6 was determined by MTT assay. 2 × 107 cells were injected into the tail vein of nude mice. Mice were sacrificed started on week 4 after injection, and lung metastases were evaluated under both mac-roscopic and microscopic observation with HE staining. Results: The growth of low-metastatic subline M6 was lower than high-metastatic sublines M8. Seventeen mice after injected M8 had occurred lung metastases while only one mice had oc-curred in M6 group. Moreover, M8 cells within metastases were arrangement disorder with variable nuclear hyperchromasia. Conclusion: A mouse model for human osteosarcoma cancer lung metastasis can be established by injection different ability of metastasis MG63 cells into tail vein.

Anti-tumor effect of bufalin on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice

Objective: To investigate anti-tumor effect of bufalin on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice. Methods: BEL-7402 cells of human hepatocellular carcinoma were inoculated to form subcutaneous tumors in nude mice by subcutaneous injection. Then the subcutaneous tumors were implanted into the liver of nude mice, and the orthotopic transplantation tumor models of human hepatocellular carcinoma were established. Seventy-five models were randomized into 5 groups ( n = 15) . Bufalin was injected intraperitoneally into the 3 groups at dose of 1.5,1 and 0.5 mg/kg for day 15 - 24, respectively. NS group were injected equal volume saline as above and adriamycin were injected intraperitoneally into ADM group at dose of 8.0 mg/kg for day 15. Ten mice in each group were killed at day 25 and detected on morphological and ultrastructural changes in myocardium, brain, liver, kidney and tumor tissues by pathology and electron microscope. The survival time in each group were observed. Results: The tumor volumes in each group of bufalin were reduced significantly compared with NS group (P < 0.01), the survival time were prolonged in group Bu 1 and Bu 2 compared with NS group ( P < 0.05), and tumor tissues were mainly necrosis in severe or moderate degree in Bu 1, Bu 2 groups, and mild degree or moderate degree in Bu 3 group. No morphological changes were detected in myocardium, brain, liver and kidney tissues, respectively. Apoptotic characteristics could be seen in tumor tissues of group Bu 1 and group Bu 2. Conclusion: Bufalin has significant anti-tumor effects on the orthotopic transplantation tumor model of human hepatocellular carcinoma in nude mice without marked toxicity. To guide cell apoptosis may be one of its anti-tumor mechanism of bufalin.

Mechanism of anti-tumor effect of HIF-1α silencing on cervical cancer in nude mice

Objective： The aim of our study was to observe the anti-tumor effect of silencing the expression of HIF-1α on cervical cancer in nude mice and to explore its mechanism of action. Methods： Human cervical cancer cell line Siha cells were divided into 3 groups： mock control group, control group transfected with scrambled sequence plasmid, and experimental group transfected with pU-HIF-la-shRNA eukaryotic expression plasmid. Cultured cells of the three groups were inoculated in nude mice to establish cervical cancer-bearing nude mice. HIF-la RNAi assay was performed to evaluate the tumor-suppressive effect of HIF-1α silencing on cervical cancer-bearing nude mice. Immunohistochemistry and Western blot were used to observe the distribution and protein expression of HIF-1α and GLUT1, while RT-PCR was adopted to detect the gene expression of HIF-1α, GLUTI and HK I1. The product of glycolysis （tactic acid） and apoptosis in tumor cells were detected by colorimetry and semi-quantitative TUNEL staining, respectively. Results： The tumor growth in experimental group was significantly slower than that in the two control groups （P 〈 0.05）. In the 50th day after transplantation, the tumor weight in the experimental group was （1.90 ± 0.28） g, significantly lower than （2.95 ± 0.77） g in the control group and （2.54 ± 0.56） g in the mock group （P 〈 0.01）. In the experimental group, the gene and protein levels of HIF-1α were 0.45 ± 0.04 and 1.25 ± 0.92, and the levels of GLUT1 were 0.32 ± 0.02 and 1.25 ± 0.48, respectively. Both indicators in HIF-la and GLUT1 were lower than that in the two control groups （P 〈 0.05）. The expression levels of HK Ⅱ gene and lactic acid in the experimental group were lower than that in the two control groups （P 〈 0.05）, but the apoptotic cells were much more numerous in the experimental group than that in matched control groups （P 〈 0.01）. Conclusion： The 9ene therapy by siRNAtargeted silencing of HIF-1α may down-regulate its downstream genes GLUT1 and HK Ⅱ expression, therefore, to reduce the tumor glycolysis activity and promote tumor cell apoptosis, and exert a tumor-suppressing effect in vivo.

Lidamycin Induces Apoptosis of B-Cell Lymphoma Cells and Inhibits Xenograft Growth in Nude Mice

OBJECTIVE To study the cytotoxicity of Lidamycin （LDM） and its induction of apoptosis in Raji and Daudi cells of B-cell lymphoma, and the inhibition of growth of the lymphoma Raji xenograft in nude mice. METHODS MTT assay was used to observe the inhibition by LDM on the proliferation of the Raji and Daudi ceils. Annexin V-FITC/PI double-stain, in combination with flow cytometry （FCM）, was used to determine the induction of apoptosis by LDM in Raji cells. The B-cell lymphoma Raji xenograft model in nude mice was set up to detect the in vivo antitumor activity of LDM. RESULTS LDM markedly inhibited the proliferation of the Raji and Daudi cells in vitro, with IC50 values of 7.13×10^-11 mol/L and 2.91×10^-10 mol/L, respectively. The apoptotic rates of Raji cells were respectively 77.98% and 67.63% at 0.5 nmol/L and 0.25 nmol/L of LDM, indicating an obvious induction of apoptosis in Raji cells. LDM inhibited the formation and growth of human B-cell lymphoma Raji xenograft in nude mice. The inhibition rates of tumor growth were respectively 74.9% and 65.2% in LDM at dosage group of 0.05 mg/kg and 0.025 mg/kg, suggesting an apparent prolongation of survival time in the nude mouse bearing lymphoma. CONCLUSION LDM can effectively induce apoptosis of the B-cell lymphoma cells and inhibit the xenograft growth in nude mice.

Survivin ASODN targeted therapy in XWLC-05 cell transplanted nude mice

Objective:The aim of this study was to study the inhibiting effect of survivin mRNA on transplanted XWLC-05 tumor on nude mice.Methods:We established XWLC-05 transplanted nude mice model.44 mice would be divided randomly into 4 groups:control group (blank),Lip group (simple liposome),survivin SODN group (transfected by sense oligonucleotide) and survivin ASODN group (transfected by antisense oligonucleotide).We would study general activities of nude mice in these 4 groups,measure the size of tumor and calculate the tumor inhabiting rate also.Pathological methods were applied in the analysis of the effect of different treatment on heart,kidney and liver of nude mice in these 4 groups.Results:Tumor grew slowly and size,weight of tumor was lower in survivin ASODN group when compared with that of others.Nude mice of survivin ASODN group showed lower growth index and tumor inhabiting rate was significantly higher than that of other groups (P < 0.05).Transplanted tumor on nude mice in control group (blank),Lip group,and survivin SODN group grew bigger as time passed and there was no significance among them (P > 0.05).We found a great deal of tumor cell necrosis in survivin ASODN group.No death of nude mice was observed in all 4 groups and we did not found obvious lesion in vital organs.Conclusion:Survivin ASDON could be used for the inhibitation of subcutaneously transplanted tumor in nude mice without obvious lesion in vital organs.

Inhibitory effects of transfected Bcl-XL antisense oligodeoxynucleotide to proliferation and growth in esophageal carcinoma cell line and human esophageal carcinoma xenograft of nude mice

Objective： The aim of our study was to investigate the biological effects of Bcl-XL antisense oligodeoxynucleotide （ASODN） transfected into cultured esophageal carcinoma cells and human esophageal carcinoma xenograft in nude mice. Methods： Cationic liposome-mediated ASODN was used to transfect esophageal carcinoma cells. RT-PCR, Western blot, MTT assay, flow cytometry, and in situ apoptosis cells detection （TUNEL detection） were used to systematically study the biological effects of transfected cells both in vitro and in vivo. Results： In this study, the results showed that the proliferation of esophageal carcinoma cells in ASODN group decreased significantly when compared with the control group （P 〈 0.05）, at 57.3% Bcl-XL mRNA inhibitory rate, and a significant decreasing of Bcl-XL protein expression, at the apoptosis rates of （31.1 ＋ 5.8）% and 35.0% by flow cytometry and TUNEL assay respectively （P 〈 0.01, when compared with control groups）. It also showed that the growth of human esophageal carcinoma in nude mice of ASODN group was significantly inhibited （P 〈 0.05）, together with a significant decreased expression level of Bcl-XL mRNA and protein, and an induced tumor cell apoptosis in nude mice. Conclusion： Our result indicates BcI-XL ASODN can effectively inhibit the proliferation of esophageal carcinoma cells in vitro and tumor growth in vivo. The suppression of Bcl-XL expression by ASODN may offer both a therapeutic approach and an important theoretic foundation for gene therapy against esophageal carcinoma.