Identification And Establishment Of Sorting Method For Isolating CD11b~+ Myeloid Cells In Human Hepatic Metastased Colorectal Cancer

BackgroundThe colorectal cancer is one of the most common tumor types worldwide and the liver metastasis is the leading cause of death in patients.Presently,the five year survival of patients with CRLM has limited changes that the effective treatment rate remains at a low level,about10%.Tumor metastasis is a multiple and complex process,and the inhibition of the LM is an effective way to treat colorectal cancer.Mountain literatures have shown that myeloid cells are widely involved in the formation and progression of tumor metastasis,while myeloid cells play different roles in the liver metastasis of colorectal cancer.Tip-DC,the secretion of tumor necrosis factor alpha(TNF-α)and inducible nitric oxide synthase(iNOS)of dendritic cells,is a subsets of myeloid cells which surface molecule phenotype is CD11b special type.In the preliminary study,we found that in CRLM,the chemokine CCL2 binding to its receptor CCR2,recruiting a large number of CD11b~+myeloid cells in the body.And in animal experiments,selective removal of CD11b~+myeloid cells the liver metastasis was inhibited,some formed metastases were narrow or even disappear than before.Therefore,the study of myeloid cells which anticipant the metastasis of CRC will help us understand the mechanism of LM more detailed,and provide a new sight in the clinical practice.In the further study,we found that the myeloid cells exist in the stroma of tumor.We explored the extract of the myeloid cells in the tissue of surgery.This study combined mechanical digestion method and flow cytometry to select target cells,and for the sorted cells we identified and evaluated it to establish a method of obtaining the specific cells in CRLM tissue.ObjectiveTo establish a method for obtaining specific cells in solid tumor tissue by sorting of CD11b~+myeloid cells in hepatic metastases tissue of colorectal cancer.MethodsAll subjects were asked to undergo questionnaires,physical examinations,laboratory tests.The incidence in different risk factors groups were analyzed by T test.ResultsHigh purity and sufficient numbers of CD11b positive cells were isolated by sorting with flow cytometry from the single cell suspension prepared by mechanical and enzyme digestion.The purity of the cells was confirmed by statistical analysis(P<0.05).The positive rates of the cells before and after sorting were significantly different(P<0.01).The positive cells are verified by immunocytochemistry.Meanwhile,the sorted cells had complete morphology and good activity.ConclusionThe CD11b~+myeloid cells in solid tumor tissue can be isolated by sorting with flow cytometry from the machine-enzyme digestion suspension with high purity,activity and complete morphology.