| Cancer is one of the serious threats to human health.According to the forecast statistics published by the Chinese Medical Journal,in 2022,the number of new cancer cases in China will be about 4,820,000,with about 3,210,000 deaths,while these two data are 237,000 and640,000 in the United States.The increase in the number of adults population and aging population are the two major factors contributing to the increase in cancer mortality.In recent years,there has been a significant increase in the incidence of prostate cancer in the male population in China,and malignancy of prostate cancer has a strong age correlation.Despite the progressively advances in cancer treatments,one of the most direct causes associated with recurrence and lethality of tumors remains the metastasis of tumor cells.Circulating Tumor Cells(CTCs)are formed when tumor cells are shed from the primary lesion and enter the peripheral bloodstream,then travel through the bloodstream to the specific target organs and grow into metastatic sites.Therefore,the detection and eliminate of CTCs during metastasis is one of the key measures used clinically to diagnose and inhibit cancer recurrence.Up to date,common in vitro flow cytometry technologies is able to detect and statistically analyze CTCs by frequently drawing blood samples,but this process has shortages such as residual error,false positives or trauma to the patients.Fluorescence in vivo flow cytometer(IVFC),as an emerging technology,allows for non-invasive,real-time,dynamic quantitative monitoring of CTCs.Based on the above analysis,this research proposes to use IVFC to quantitatively monitor the clearance kinetics of CTCs in the blood of individuals of different ages,and to assess whether aging of the body has an impact on the clearance rate of circulating tumor cells in the blood,thus providing a new technical tool for clinical investigation of the relationship between aging and tumor metastasis,and developing relevant prevention and treatment strategies.In Chapter 1,the relationship between age and cancer development and metastasis is firstly outlined;then,the common CTCs detection techniques currently used in clinical and scientific research are listed,the advantages and shortcomings of the existing detection techniques are summarized as well.This is followed by a systematic introduction to the basic principles and applications of the new CTCs detection technology IVFC,followed by a presentation of the research idea and the design of the relevant protocol.In Chapter 2,a circulating tumor cell model of prostate cancer is constructed,and the feasibility of the constructed CTCs model to be detected by IVFC is assessed using laser confocal microscopy and conventional flow cytometry.In Chapter 3,C57BL/6J mice at 3 months,6 months and 12 months of age were selected as representatives to assess the CTCs kinetics in mice at different months of age using IVFC.In Chapter 4,the propensity of CTCs to metastasize in specific tissues and target organs is analyzed using small animal live fluorescence imaging as well as histopathological sections.A comparison of the number of CTCs passed per unit time(per minute)in mice of different months of age showed that there was no significant difference in the number of CTCs in blood of 3-month-old and 6-month-old mice at the first two time points post-injection,whereas the number of CTCs in the blood of 3-month-old and 12-month-old mice as well as6-month-old and 12-month-old mice have significant difference.We then statistically compared the first time point 0-10 min post-injection with that of Day 7 and obtained the similar results.The number of individuals counted yielded significant differences p = 0.0002 and p = 0.0004.The small animal live fluorescence imaging and histopathological sectioning experiments had also confirmed that CTCs tended to select specific tissues as their target organs for metastasis.The result shows that fluorescent cells are mostly concentrated in the lungs and liver,resulting in brighter fluorescence intensity in these two organs compared to the heart,kidneys and pancreas.This result is also consistent with the finding of the pathology of the lungs and liver.In addition,immunoinflammatory reactions such as macrophage infiltration were also observed in lymph node tissue sections.In this study,the non-invasive,real-time,and dynamic detection of circulating tumor cells by fluorescence IVFC demonstrated that the rates of clearance of circulating tumor cells was slowest in the circulation of 12-month-old mice and was statistically different from that of 3and 6-month-old mice.This shows that the way in which tumor cells form metastases through bloodstream metastases at different ages correlates with the aging factors of physical body,and older the people are,the slower the blood circulation is to remove circulating tumor cells.The result of this research provides a basis for the investigation of a clinically applicable IVFC technique. |
