Animals ; Carcinogenicity Tests ; Cytotoxicity Tests, Immunologic ; Humans ; Mutagenicity Tests ; Soot ; toxicity

With the increased use of cell therapy in the veterinary sector, there is a growing demand for the development of cell-based medicinal products and the determination of their safety. Currently, the Korean Animal and Plant Quarantine Agency has established a guideline for evaluating the safety of cell-based medicinal products for animal use. The guideline includes items related to definition, classification, management, manufacturing procedure and quality control (standard and test method), stability testing, toxicity testing, pharmacological testing, and performance of clinical trials. In addition, testing protocols related to safety assessment of animal cell-based products such as chromosome karyotyping, tumorigenicity testing, confirmatory testing of biodistribution and kinetics, and target animal safety testing are described in detail. Moreover, because cell-based medicinal products are novel therapies, deviations from traditional designs may be justified in order to obtain relevant safety information on the treatment. Additionally, this guideline can be amended on the basis of new scientific findings.
Animals ; Carcinogenicity Tests ; Cell- and Tissue-Based Therapy ; Classification ; Karyotyping ; Kinetics ; Plants ; Quality Control ; Quarantine ; Toxicity Tests

Preclinical evaluation of medical devices (prototype products) offers the opportunity to investigate and study the intended use of device materials. Preclinical evaluation programs are designed to determine the efficacy, safety, and biocompatibility of biomaterials, prostheses, and medical devices. The purpose of safety testing is to determine if a material presents potential harm to the human; it evaluates the interaction of the material with the in vivo environment and determines the effect of the host on the implant. Preclinical evaluation is the determination of the ability of the prototype product to perform with appropriate host response in a specific application, considered from the perspective of human clinical use. Therefore, preclinical data should include materials science and engineering, biology, biochemistry, medicine, host reactions and their evaluation, the testing of biomaterials, and the degradation of materials in a biological environment.
Animal ; Carcinogenicity Tests ; Equipment and Supplies*/adverse effects ; Hemolysis ; Human ; Pyrogens/toxicity ; Sterilization

OBJECTIVETo predict the carcinogenicity of polycyclic aromatic hydrocarbons (PAHs) by cell transformation assay using BALB/c 3T3 cells and HPV16-E6E7-transfected BALB/c 3T3 cells (BALB/c-E6E7 cells).

METHODSThe cell transformation assays induced by PAHs using BALB-E6E7 cells and BALB/c 3T3 cells.

RESULTSThe initiating and promoting activities of PAHs examined in a BALB-E6E7 cell transformation assay were similar to in a BALB/c 3T3 cell transformation assay, which was up to the standard of agents classified by the IARC. There were much more transformed foci appeared and much shorter time consumed to accomplish phenotypic alterations in the BALB/c-E6E7 cell transformation assay than in the BALB/c 3T3 cell transformation assay. The BALB/c-E6E7 cell transformation assay was superior to the BALB/c 3T3 cell transformation assay in cost and labor performance, the sensitivity of transformation response.

CONCLUSIONThe BALB/c-E6E7 cell transformation assay, with a satisfied prediction performance of initiating activity and promoting activity, would improve the overall process of safety and risk assessment of carcinogenicity.

Animals ; BALB 3T3 Cells ; Carcinogenicity Tests ; Cell Transformation, Neoplastic ; Mice ; Mice, Inbred BALB C ; Polycyclic Aromatic Hydrocarbons ; toxicity

OBJECTIVETo study the acute toxicity of C25P polypeptide, a CCR5 antagonist, in mice and its carcinogenic effect in vitro.

METHODSThe acute toxicity of C25P polypeptide in mice was assessed by determining the maximum tolerated dose (MTD). The mice were given C25P at the dose of 3.64 g/kg by tail vein injection, and the control mice received saline (40 ml/kg) injection. The mice were continuously observed for 14 days after the administration and sacrificed on day 14 for routine blood test, examination of the blood biochemistry and pathological examination. The carcinogenicity of C25P polypeptide in vitro was evaluated in cultured cell lines by chromosome aberration test, cell transformation test and non-anchorage dependent growth test.

RESULTSNo mice died following administration of the drug, but 3 mice showed mild adverse reactions. The rats in both groups showed an increase in the body weight at a comparable rate. GPT increased and ALP decreased significantly in C25P polypeptide group (P<0.05). Most of the organs of the rats treated with in C25P polypeptide remained normal, but 3 mice showed pathologies in the lung, spleen and liver. Chromosome aberration test, cell transformation test and non-anchorage-dependent growth test all yielded negative results for C25P polypeptide.

CONCLUSIONC25P polypeptide is a low-toxicity drug that produces no apparent acute toxicity in mice or obvious carcinogenicity in vitro.

Animals ; CCR5 Receptor Antagonists ; Carcinogenicity Tests ; Chemokines ; toxicity ; Female ; Male ; Mice ; Mice, Inbred Strains ; Mutagenicity Tests ; Peptides ; toxicity ; Toxicity Tests, Acute

Human fibroblasts were developed for cellular therapy with the aim of correcting of depressed scars, but the safety of that in vivo is unclear. In this study, we assessed the safety of human fibroblasts by investigating the tumorigenicity, 13-week toxicity and through distribution studies. In the tumorigenicity test, nude mice were divided into three dosage level treatment groups with a negative/positive control group. At 6 months after intradermal transplantation, all of the treatment groups showed no development of a nodule on the injection sites and organs. Toxicity studies were performed using ICR and BALB/c mice for 13 weeks. The mice were divided into three dosage level treatment groups with a control and a syngeneic group. There was no treatment-related effect on clinical signs, mortality, body weight, food/water consumption, hematology, serum biochemistry, urine, necropsy findings and histopathological findings in any groups. These results suggest that the no-observed-effect level (NOEL) of the human fibroblasts was greater than 7.5x10(7) cells/kg for mice. In the distribution study, groups were treated with fibroblasts labeled with a fluorescent dye (CM-DiI) at low and high doses with a control and a syngeneic group. At 24 hours, a large percentage of the labeled fibroblasts were observed at the dermal layer. At 3 months, fluorescence of the labeled fibroblasts continued to be observed. Other tissues were not detected the fluorescence at any time. These studies demonstrate that the safety of human fibroblasts is reasonable with no toxic effect, no tumorigenicity and retention in the dermis. Our studies define preclinical safety testing standards relevant to the development of cellular therapeutics.
Animals ; Biochemistry ; Body Weight ; Carcinogenicity Tests ; Cicatrix ; Dermis ; Fibroblasts ; Fluorescence ; Hematology ; Humans ; Mice ; Mice, Nude ; No-Observed-Adverse-Effect Level ; Retention (Psychology) ; Tissue Therapy ; Transplants

OBJECTIVETo discuss the tumorigenicity of immortalized endothelial cells differentiated from embryonic stem cells.

METHODSThe embryoid bodies (EB) formed in vitro from embryonic stem cells, were induced to differentiate into many "round cells" (the precursor of endothelial cells). These "round cells" later formed the vascular tube-like structures. To immortalize these cells, human telomerase reverse transcriptase (hTERT) cDNA was transfected into "round cells" by lipofectin, RT-PCR and immunocytochemistry were used to evaluate the immortalized cells. And the tumorigenicity of these cells were evaluated by being injected into nude mice subcutaneously.

RESULTS95% of these transfected cells expressed Flk-1, CD34 and vWF, and could proliferate in large quantity in vitro (cell number was doubled in 2 days, and increased 12 times in 3 days), and were able to form tubular structures.

CONCLUSIONSThese results suggest that hTERT cDNA transfection can immortalize induced endothelial cells and tumorigenicity is found after immortalized cells are injected into nude mice subcutaneously.

Animals ; Carcinogenicity Tests ; Cell Differentiation ; Cells, Cultured ; Embryonic Stem Cells ; cytology ; Endothelial Cells ; cytology ; Endothelium, Vascular ; cytology ; Mice ; Mice, Inbred Strains ; Mice, Nude ; Telomerase ; genetics ; Transfection

OBJECTIVETo investigate the application of 293 cells to detect suspected carcinogens and provide experimental evidence by using in vitro cell transformation assay and tumorigenicity study.

METHODSThe transformation systems of cells cultured in vitro have been adopted to clarify the tumor promotive activity of Microcystin LR (MC-LR). The malignant transformation of 293 cells induced by MC-LR is tested by several methods including clone forming in soft agarose, serum requirement assay and tumor forming in mice to define the promotive activity of 293 cells.

RESULTS293 cell acted like tumor cells after induced by MC-LR: serum dependence decreased, anchorage independence growth in soft agarose and formed cell clones, malignant tumors appeared in SCID mice.

CONCLUSION293 cells were easy to culture and sensitive to environmental carcinogens so that can be used in detection of suspicious carcinogens.

Animals ; Carcinogenicity Tests ; methods ; Carcinogens ; toxicity ; Cell Line ; drug effects ; Cell Transformation, Neoplastic ; drug effects ; Humans ; Mice ; Mice, SCID ; Microcystins ; toxicity

BACKGROUND/AIMS: It is essential to develop an in vitro culture model of primary hepatocytes for the study of hepatocellular function and the pathogenesis of hepatitis C virus (HCV) infection. In this study, we have established the immortalized primary human hepatocyte (IPHH) and performed in vitro culture of HCV derived from human patient. METHODS: Primary human hepatocytes were isolated from surgically resected liver tissue and then were immortalized by transfection with the SV40 large T antigen. The characterization of the IPHH during culture was analyzed by immunocytochemistry, RT-PCR, Western blot, ELISA, and soft agar assay. Next, sera and/or liver tissue homogenates from surgically resected liver tissues of patients with HCV infection were inoculated for the culture of HCV in IPHH. After HCV RNA extraction from IPHH and culture media, positive or negative stranded HCV RNA was examined by specific nest RT-PCR. RESULTS: IPHH expressed liver-associated proteins but did not express alpha-fetoprotein. Also IPHH showed ammonia removal activity. With regard to its malignant potential, colony formation in soft agar assay was not observed. Next, positive and negative stranded HCV RNAs in IPHH infected with patient's sera plus liver tissue homogenates were clearly detected whereas those in IPHH infected with only patient's sera were not detected. CONCLUSIONS: These results demonstrated the phenotypic characteristics of IPHH and the feasibility in vitro culture system of HCV infected human samples. This system might be useful for study of pathogenesis of HCV infection or hepatocyte-based applications.
Antigens, Viral, Tumor/genetics ; Base Sequence ; Carcinogenicity Tests ; Cell Culture Techniques ; Cells, Cultured ; Cells, Immobilized ; Hepacivirus/isolation & purification/*physiology ; Hepatocytes/metabolism/physiology/*virology ; Humans ; Liver Function Tests ; Models, Biological ; RNA Probes ; RNA, Viral/analysis ; Reverse Transcriptase Polymerase Chain Reaction

Carcinogenesis is a multi-stage process that generally consists of at least three steps; initiation, promotion, and progression. If one of these carcinogenic steps were suppressed or delayed, the cancer could be prevented. Cancer chemoprevention is defined to be inhibition or reversal of the carcinogenic process by the specific chemical agents and is a novel approach to cancer management alternative to conventional chemotherapy. Chlorophylln(CHL), a water-soluble derivative of chlorophyll, containing sodium and copper, has been known to be strong antimutagen in several test systems, but its mechanism of antimutagenic action is unknown. In the present experiment, the possibility of CHL as chemopreventive drugs on 7,12-dimethylbenz[a]anthracene(DMBA)-induced hamster buccal pouch carcinogenesis was investigated by mutagenicity test, carcinogenicity test, and frequency or spectrum of H-ras mutations in the both of DMBA-induced and chlorophylln-pretreated-DMBA induced tumor by polymerase chain reaction and non-isotopic restriction fragment length polymorphism. The treatment of CHL reduced the yields and multiplicity of the 0.5% DMBA-induced tumor, 86% to 62.5% and 3.7+/-0.6 to 1.4+/-0.3, respectively. The occurrence of histidine revertant by 20 micromole DMBA was inhibited 25.6 to 81.7% by 1 to 5 microM CHL in a dose-dependent manner. The mutation rates of H-ras gene in DMBA-induced and CHL-pretreated-DMBA induced tumor were 96%, 94% of which the most mutations were in codon 12/13. These results suggest that CHL inhibits the carcinogenic action of DMBA by the formation of complex between CHL and DMBA or the inhibition of the activation of DMBA in vivo. But CHL did not affect the mutation rates or its spectrum in already formed tumor.
9,10-Dimethyl-1,2-benzanthracene ; Animals ; Carcinogenesis ; Carcinogenicity Tests ; Cheek* ; Chemoprevention ; Chlorophyll ; Codon ; Copper ; Cricetinae* ; Drug Therapy ; Genes, ras ; Histidine ; Mutagenicity Tests ; Mutation Rate ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Sodium