Study on the Low Dose X Ray-induced Frequency of Mcie Marrow and Human Micronucleated Reticulocytes by Flow Cytometric Automation Detection
|School||Third Military Medical University|
|Keywords||Micronucleus test Reticulocyte Ferritin Flow cytometry X-ray Magnetic cell separation surgery Mice Human peripheral blood|
With the progress of science and technology, ray, microwave, electromagnetic radiation in human life is becoming increasingly popular, such as medical diagnosis, special contrast, medical treatment, as well as commercial irradiation. However, the progress of science and technology to the human society has brought many negative effects, one of the most important human health problems. The radiation caused a serious threat to human health, many people have noticed the damage of its genetic material of the body. The world of rays, microwaves, electromagnetic radiation are regulated, the unification of the evaluation program, which is the most common chromosomal aberrations (CA) analysis and micronucleus test (MNT). CA analysis of the advantages is that the results are accurate, reliable and able to provide information rich, but time-consuming, the need for cell culture. Traditional micronucleus test (MNT) has failed to meet the needs of actual work, still rely on manual microscopic count, too subjective; generally require counting thousands of cells, too much work and less sensitive, and can lead to some undetected or some false positive sample. The development of the automation of the micronucleus detection has become an important means for fast monitoring of population genetic damage. In recent years, with the continuous improvement and perfection of flow cytometry technology, many experts at home and abroad, scholars dedicated to the study of automated testing. 1992 Swedish scientist Grawe development of a flow cytometry mouse peripheral blood reticulocyte micronucleus automatic detection technology, 1993 my room CAO Jia development and improvement of this technology to the practical level. In 2003, my room Sun Liping establish automated detection system based on single-laser flow cytometry rat bone marrow micronucleus polychromatic erythrocytes; combined with magnetic cell separation technology, successfully used in human nasopharyngeal carcinoma (NPC) radiotherapy in patients with peripheral blood reticulocytes erythrocyte micronucleus rate of detection, preliminary attempts to estimate the cumulative radiation dose biological Meter. The present study is based on previous work of my room, mice as animal model, the establishment of a low-dose X-ray-induced mouse bone marrow erythrocyte micronucleus rate of flow cytometry automated detection method. Subsequently, using this method combined with a newly developed magnetic cell separation technology, observation of the head and face of cancer patients receiving low-dose X-ray treatment, peripheral blood the reticulocyte micronucleus rate dynamic change. The main results of the studies are as follows: 1, flow cytometry low-dose X-ray irradiation mice the bone marrow fMNPCE fMNNCE: analysis and comparison of different detection parameters finalized indication DNA FL1 (fluorescence intensity) for the longitudinal coordinates , FSC (indicate cell size) as the abscissa can well distinguish various cell populations - total nucleated cells (TNC), polychromatic erythrocytes cells (PCE) and micronucleated polychromatic erythrocytes (MNPCE) is dyed erythrocytes (NCE), and positive micronucleated polychromatic erythrocytes (MNNCE). After 20 hours, X-ray treatment, a irradiation dose 0.05Gy, 0.1Gy and 0.4Gy, mouse bone with fMNPCE were 3.01 ± 0.55 ‰, 4.58 ± ± 0.83 ‰ and 11.12 ± 1.72 ‰; for different cumulative dose of 0.1 Gy, 0.15 Gy, 0.2 Gy, 0.3Gy, 0.4Gy, 0.8Gy, 1.2Gy and 1.6Gy mouse bone marrow fMNPCE were 4.72 ± 0.95 ‰ ± 2.09 ‰, 10.26, 13.77 ± 1.66 ‰ ± 2.33 ‰, 11.77, 17.71 ± 2.19 ‰, 16 43 ± 1.33 ‰, 21.85 ± 3 25 ‰, and 22.17 ± 3.48%; The results showed a good dose - response time - effect relationship. 2, the comparison of the results of flow cytometry and artificial microscopy results: the results of the two methods for detecting showed a good dose - response time - effect relationship, but slightly higher than the artificial microscopy results by flow cytometry results, statistics The analysis shows that the results of the two methods for detecting no significant differences, and both have a good correlation (R = 0.984, P LT; 0.01). 3, magnetic cell sorting technique separation and enrichment effect: a total of 7 healthy volunteers and patients with head and facial cancer patients receiving different cumulative radiation dose peripheral blood CD71 antigen-positive reticulocytes (Trf-Ret ) enrichment and separation results show that Trf-Ret yield differences between different individuals, between 20% -90%, but this is not due to the separation method, but rather caused by individual differences, such as age, work and to accept radiation dose size. And confirmed that the method is simple, reliable, and reproducible characteristics. 4, flow cytometry low-dose X-ray irradiation descendants peripheral blood MN-Trf-Ret: FL4 (indicating RNA fluorescence) for the longitudinal coordinates, FSC (indicating cell size) as the abscissa, and the enrichment of the descendants of the outer periphery blood sample is divided into five distinct cell populations --- reticulocytes and micronucleated reticulocytes, and mature erythrocytes micronucleated mature erythrocytes and nucleated cells using flow cytometry successful detection of patients with cancer of the head and face accept low-dose X-ray irradiation of peripheral blood MN-Trf-Ret changes. The results show that the head and face of cancer patients receiving radiotherapy after fMN-Trf-Ret Quick significantly higher, has been in a high platform levels continue to increase with the cumulative dose of radiotherapy, fMN-Trf-Ret. Corresponds's cumulative radiation dose 0Gy of different 2Gy radiation., 4Gy, 6Gy 8Gy well 10Gy, fMN-Trf-Ret the average value is 2.89 ± 0.93 ‰, respectively, 16.2 ± 2.31 ‰, 23.84 ± 3.22 ‰, 32.17 ± 6.95 ‰, 30.89 ± 9.35 ‰ and 35.74 ± 9.95 ‰. 5, low-dose X-ray irradiation curve fitting equation: obtained fMN-Trf-Ret amount of cumulative radiation dose efficiency and timeliness response relationship on the basis of the best fit by SPSS13.0 curve equation Y = 2.595 8.874X-1.005X2 0.044X3 (R = 0.978, P lt; 0.05), preliminary studies suggest that, tried to use the equation to assess the exposed biological dose of radiation accidents, radiation wounded in the dose range of 0Gy-10Gy and further validation. In summary, the experiment successful use of flow cytometry in mice after total body irradiation in the low-dose X-rays, bone marrow erythrocyte micronucleus rate observed trends; and used to observe the people in the low-dose X ray irradiation peripheral network the reticulocyte micronucleus rate changes. The results show that the automated detection method of flow cytometry in the low-dose X-ray sensitive, rapid, and easy to lay the foundation for the promotion and application of automatic detection of flow cytometry micronucleus.