Effect of the chemoprotectant tempol on anti-tumor activity of cisplatin / 南方医科大学学报

OBJECTIVE@#To investigate the effect of the chemoprotectant tempol on the anti-tumor activity of cisplatin (DDP).@*METHODS@#The cellular toxicity of tempol in human colon cancer SW480 cells and mouse colon cancer CT26 cells were evaluated using MTT and cell counting kit-8 assays. CalcuSyn software analysis was used to determine the interaction between tempol and DDP in inhibition of the cell viability. A subcutaneous homograft mouse model of colon cancer was established. The mice were randomly divided into control group, tempol group, cisplatin group and tempol + DDP treatment group with intraperitoneal injections of the indicated agents. The tumor size, body weight and lifespan of the mice were measured, and HE staining was used to analyze the cytotoxic effect of the agents on the kidney and liver. Immunohistochemistry and Western blotting were performed to detect the expression of Bax and Bcl2 in the tumor tissue, and TUNEL staining was used to analyze the tumor cell apoptosis. The level of reactive oxygen species (ROS) in the tumor tissue was determined using flow cytometry.@*RESULTS@#Tempol showed inhibitory effects on the viability of SW480 and CT26 cells. CalcuSyn software analysis showed that tempol had a synergistic anti-tumor effect with DDP (CI < 1). In the homograft mouse model, tempol treatment alone did not produce obvious anti-tumor effect. HE staining showed that the combined use of tempol and DDP alleviated DDP-induced fibrogenesis in the kidneys, but tempol also reduced the anti-tumor activity of DDP. Compared with the mice treated with DDP alone, the mice treated with both tempol and DDP had a significantly larger tumor size ( < 0.01) and a shorter lifespan ( < 0.05). Tempol significantly reversed DDP-induced expression of Bax and Bcl2 in the tumor tissue and tumor cell apoptosis ( < 0.001), and obviously reduced the elevation of ROS level in the tumor tissue induced by DDP treatment ( < 0.05).@*CONCLUSIONS@#Tempol can attenuate the anti-tumor effect of DDP while reducing the side effects of DDP. Caution must be taken and the risks and benefits should be carefully weighed when considering the use of tempol as an anti-oxidant to reduce the toxicities of DDP.

Establishment of lung metastasis model of human primary malignant melanoma in the small intestine in nude mice / 中华消化外科杂志

Objective To provide an ideal animal model for exploring the pathogenesis and experimental treatment of malignant melanoma in the small intestine.Methods Fresh tissue of lung metastatic lesions from patients with malignant melanoma of the smallintestine were transplanted into mucosa of the small intestine in nude mice.After 4 times of screening.the tissue of the lung metastatic lesions from the nude mice were transplanted into the small intestine of additionat nude mice.Tumorgenecity and metastasis of transplanted tumors were observed,and were analyzed by morphology,karyotype and flow cytometry.Results A lung metastatic model of human primary malignant melanoma of the small intestine in nude mice was successfully constructed and named HSIM-0601.Massive melanin granules and melanin complex were seen in cytoplasm of tumor cells.Immunohistochemical straining of S-100 and HMB-45 were positive.The number of chromosome was between 57 and 59.DNA index was 1.49.HSIM-0601 was passed for 26 generations.A total of 173 nude mice were used for tumor transplantation.The growth rate of the transplanted tumors and resuscitation rate of liquid nitrogen cryopreservation were both 100%.In HSIM-0601.lung metastasis rate was 100%(173/173)and lymph node metastasis rate was 61.3%(106/173).Conclusions The HSIM-0601 successfully mimics the natural clinicopathologic course of patients with primary small intestinal melanoma,and provides an ideal animal model for research on pathogenesis,metastasis and experimentM therapy of malignant lymphoma in the small intestine.

Lentiviral vector for RNA interference of human Snail gene inhibits proliferation and invasion of nasopharyngeal carcinoma 5-8F cell line / 第三军医大学学报

Objective To construct a recombinant lentiviral expression vector for RNA interference (RNAi) of human Snail gene and to study its effects on the proliferation and invasion of nasopharyngeal carcinoma cell line 5-8F. Methods The effective sequence of short hairpin RNAs (shRNA) targeting Snail gene was designed and cloned into the linear pLVTHM vector after enzyme digestion. After confirmation by DNA sequencing, 5-8F cells were infected with the viral supernatants. The cells with stable Snail gene knock-down were separated by fluorescence activated cell sorter (FASC). The expression of Snail mRNA was detected by real time RT-PCR. MTT and cell invasion assay were used to detect the proliferation and invasion of 5-8F cells after plVTHM-siSnail transfection. Results The lentivirus vector plVTHM-siSnail was constructed successfully. The separated 5-8F-plVTHM-siSnail exhibited significant knock-down of Snail mRNA expression. Slower proliferation and decreased cells to permeate through the Matrigel were found after plVTHM-siSnail transfection (P

Establishment and characteristics of orthotopically transplanted model of human primary malignant spleen lymphoma in nude mice / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To establish three orthotopically transplanted model of human primary malignant spleen lymphoma in the nude mice.</p><p><b>METHODS</b>Orthotopic transplantation of histologically intact human primary malignant splenic lymphoma tissue obtained from patients was introduced into the splenic parenchyma of nude mice. Tumorigenicity, invasion, metastasis and morphological characteristics of the transplanted tumor were studied by light microscopy, electron microscopy and immunohistochemical methods.</p><p><b>RESULTS</b>The first kind, a strain of human primary malignant spleen lymphoma (non-Hodgkin's, cleaved B cell, BFNHL-HMN-1) screened from 11 patients which had been passaged in vivo for 41 generations, a second kind, a liver metastasis model of human primary malignant spleen lymphoma (non-Hodgkin's, cleaved B cell, LM-BFNHL-HMN-2) which had been passaged for 47 generations and a third kind of human primary malignant spleen lymphoma (non-Hodgkin's, T-immunoblastic cell, TINHL-HMN-3) having passaged for 37 generations were all successfully transplanted in 611 nude mice. Models of BFNHL-HMN-1 and TINHL-HMN-3 tumor gave nodular growth and lymph node metastasis in the spleen hilum but without any metastasis in the abdominal lymph nodes or organs. In the LM-BFNHL-HMN-2 model, not only did the tumor cells grow in the spleen, but in spleen hilum, lymph nodes and liver also. The orthotopically transplanted tumor cells were similar to the original human tumor in light histopathology, ultrastructure features, DNA content and chromosomal karyotype.</p><p><b>CONCLUSION</b>These three models are able to serve as useful tools for the study of biologic characteristics and experimental treatment of human primary malignant lymphoma.</p>

Establishment of high metastasis models of human primary colorectal lymphoma in nude mice by orthotopic transplantation / 解放军医学杂志

Objective To reproduce high metastasis models of human primary colorectal lymphoma in nude mice by orthotopic transplantation, and to investigate their biologic features. Methods Histologically fresh lymphoma tissues from primary and liver metastatic lesions of human primary colorectal lymphoma obtained during operations were transplanted into colorectal mucosa of nude mice. Tumorgenecity, invasion, metastasis, morphology (light microscopy, electron microscopy and immunohistochemistry), karyotype analysis and DNA content of the orthotopically transplanted tumors were studied. Results According to the new WHO classification of malignant lymphoma, a strain of liver metastasis model of human primary colonic lymphoma (non-Hodgkin's, B cell) in nude mice by orthotopic transplantation (HCBL-0301), and a strain of high metastasis model of human primary rectal lymphoma (non-Hodgkin's, B cell) in nude mice by orthotopic transplantation (HRBL-0302) were successfully screened from four cases of human primary colorectal lymphoma. Histopathology of transplanted tumors showed high grade non-Hodgkin's large B cell lymphoma. The cells were positive for CD19, CD20, CD22 and CD45, but negative for CD3 and CD7. The number of chromosome was between 55 and 69. DNA index (DI) was 1.59~1.71 (i.e. heteroploid). So far, HCBL-0301 and HRBL-0302 had been passaged for 31 and 27 generations in nude mice, respectively, and transplantation was successful in 326 nude mice. Since the third generation, both the growth rate of transplantation and rate of resuscitation after being thawed from liquid nitrogen cryopreservation were 100%. In HCBL-0301, metastasis to the right lobe of liver was most common and metastatic rate was 100%; additionally, rates metastasis to of lymph node and peritoneal seeding were 67.4%. In HRBL-0302, metastasis to the left and right lobes of liver was most common with metastasis rate of 63.7%, and rates of metastasis to lymph node and peritoneal seeding were 56.4%. Transplanted human primary colorectal lymphoma could autonomously and invasively grow in the colorectum of nude mice, with occurrence of hematogenic, lymph node and implantation metastases. Conclusions The study successfully replicated high metastasis models of human primary colorectal lymphoma in nude mice by orthotopic transplantation. HCBL-0301 and HRBL-0302 models can be used in the research on pathogenesis, mechanism of invasion and metastasis and experimental therapy of human primary colorectal lymphoma.

Reproduction of a nude mouse model orthotopical transplantation of huma n primary malignant lymphoma of the liver for the study of its biological characteristics / 解放军医学杂志

Objective To provide an experimental tool for explo ri ng pathogenesis and experimental treatment strategies for primary lymphoma of th e liver. Methods Histologically intact lymphoma tissues from pa tients with primary lymphoma of the liver were transplanted into hepatic parench yma of nude mice. Tumorgenecity, invasion, metastasis, morphological characteris tics(light microscopy, electron microscopy and immunohistochemistry), serologica l test(AFP, HBsAg and LDH), karyotype analysis and DNA content of orthotopically transplanted tumors were studied. Results Orthotopically trans planted model of human primary malignant lymphoma of the liver in nude mice(desi gnated as HLBL-0102) was successfully reproduced. Histopathology of transplante d tumors showed primary lymphoma of the liver(non-Hodgkin's, B cell). The cells were positive for CD19,CD20,CD45RO and CD79a, but negative for CD3 and CD7. Ser ological tests showed that the serum was AFP negative and HBsAg positive, and t he concentration of LDH was 1267.5U/L. The number of chromosome was between 55 and 59. DNA index(DI) was 1.57~1.61(i.e. heteroploid). So far, the strain HLB L-0102 has grown for 3 years and been passaged for 37 generations in nude mice. Altogether 283 nude mice were used for transplantation. The growth rate and res uscitation rate of liquid nitrogen cryopreservation of transplanted tumors were both 100%. The transplanted tumors autonomoasly and invasively grew in the live r of nude mice, damaging adjacent liver tissues, bile ducts, and veins in the po rtal area. There was no involvement of other tissues and organs or distal lymph nodes. Orthotopically transplanted tumors were consistent in histopathological a nd ultrastructural features, DNA content and chromosomal karyotype with the orig inal human tumor. Conclusions The study is the first attempt to successfully reproduce orthotopically transplantation model of human primary ma lignant lymphoma of the liver in nude mice. HLBL-0102 completely replicates th e natural clinical process of primary lymphoma of the liver, and provides an ide al animal model for further research on the biology and experimental therapeutic strategies of primary lymphoma of the liver.

Reproduction of a murine model of liver metastasis of human primary malignant lymphoma of small intestine by continuous screening in vivo / 解放军医学杂志

Objective A liver metastasis model of human primary malignant lymphoma of small intestine was reproduced in nude mice in an attempt to provide an ideal animal model for elucidating the mechanism of liver metastasis of primary small intestinal lymphoma.Methods A piece of surgically obtained liver metastatic tissue of small intestinal lymphoma was implanted into the mucous membrane of small intestine in nude mice to reproduce the model.After metastasis of this tumor to the liver occurred,a portion of the metastais was transplanted into the mucous membrane of small intestine of another nude mouse.This process was repeated 4 times in order to obtain a cell line with the characteristics of high malignant lymphoma metastasis to liver.The survival rate of the experimental animals,regional invasion rate and metastasis rate were observed,and the morphological characteristics(light microscopy,electron microscopy and immunohistochemistry),karyotype analysis and DNA content of the neoplastic cells were also determined.Results A liver metastasis model of human primary malignant lymphoma of small intestine in nude mice(termed HSIL-0402)was successfully reconstructed,by repeated implantation of liver metastatic tumor in vivo.Histopathology showed HSIL-0402 tumor was a high grade non-Hodgkin's lymphoma.Immunohistology showed the cells were CD19,CD20,CD45 and CD79a positive,but negative for CD3 and CD7.The modal number of chromosome was between 56-69.DNA index(DI)was 1.61?0.37,which showed heteroploid.So far,HSIL-0402 had been maintained for 27 passages in nude mice,exhibiting 100% of transplantability,liver metastatic rate,and resuscitation rate after liquid nitrogen cryopreservation.Lymph node metastasis and celiac seeding occurred in 67.4% and 61.7% of a total of 156 observed animals.The HSIL-0402 model displayed various manifestations reminiscent of highly metastatic invasive behavior in nude mice,including invasive growth,hematogenous metastasis,lymph node metastasis and celiac seeding.Conclusion The present study successfully re-established a spontaneous liver metastasis model of human primary malignant lymphoma of small intestine in nude mice,HSIL-0402,which provides an ideal animal model for the researches on biological mechanism of liver metastasis and anti-metastasis therapy of human primary small intestinal lymphoma.

Reblication of high metastasis model of malignant lymphoma of human small intestine in nude mice / 解放军医学杂志

Objective To replicate an ideal animal model for exploring pathogenesis and experimental treatment of malignant lymphoma of small intestine. Methods Fresh lymphoma tissues derived from primary lesion and liver metastasis of malignant lymphoma of human small intestine obtained during operation were respectively transplanted into mucosa of small intestine and subcutaneous space in the interscapular region in nude mice. Tumorigenicity, invasion and metastasis of transplanted tumors were observed. Morphology (light microscopy, electron microscopy and immunohistochemistry), karyotype analysis, DNA quantitative assay (FCM) were also studied. Results From five patients of malignant lymphoma of small intestine, tumor tissue from 3 patients was successfully transplanted. According to the new classification of the World Health Organization, a strain of high metastatic model of orthotopically transplanted malignant lymphoma in nude mice (HSIL-0101) and another of subcutaneously transplanted highly metastatic model were reproduced from the same human neoplasm (non-Hodgkin B cell type); orthotopically Pathological study showed that the tumor was high-grade large B-cell lymphoma. Histochemitry showed that it was CD19, CD20, CD22 and CD45 positive, and CD3 and CD7 negative. The number of chromosome is ranged from 55 to 59; DI (DNA Index) was 1.47～1.61 (ie, heteroploid). HSIL-0101 and HSIL-0102 had been passaged for 32 and 38 generations in nude mice separately. 357 nude mice were transplanted. Rate of growth of neoplasm transplantation and resuscitation rate of liquid nitrogen cryopreservation were both 100%. In HSIL-0101, metastasis rate of liver and lymph node was 100%. In HSIL-0102, metastasis rate of liver was 63.5% and metastasis rate of lymph node was 62.7%. Transplanted tumors invasively grew in small intestine and subcutaneous region of nude mice. Blood metastasis was found (liver and spleen metastases). There were also lymph metastasis and seeding metastasis in the peritoneal cavity. Conclusions The study first successfully established spontaneous high metastasis models of malignant lymphoma of human small intestine in nude mice by orthotopic and subcutaneous transplantation. HSIL-0101 and HSIL-0102 could be used to carry out the research on pathogenesis, invasion, metastasis, and experimental therapy of malignant lymphoma of small intestine.

ORTHOTOPIC TRANSPLANT MODEL OF HUMAN PANCREATIC CANCER IN NUDE MICE AND THEIR BIOLOGICAL PROPERTIES / 解放军医学杂志

Two cell strains from human pancreatic cancer taken surgically were transplanted to the pancreas of pureline BALB/C-nu/nu nude mice and 9 generations of PTNMP-1 and 6 generations of PTNMP-2 were obtained. Biological properties were studied. It yielded a transplant success rate of 95% - 100% and large amount of CEA. Analysis of the karyotype confermed that they retained the karyotype of human cancer cells. Some lymphatic and blood stream metastasis and invasion could be found in the nude mice orthotopically transplanted with tumour, showing that the transplanted tumour had characteristics identical with the donor tumour.