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Evaluation of in vitro macrophage differentiation during space flight

ABSTRACT: We differentiated mouse bone marrow cells in the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14 days during the flight of space shuttle Space Transportation System (STS)-126. We tested the hypothesis that the receptor expression for M-CSF, c-Fms was reduced. We used flow cytometry to assess molecules on cells that were preserved during flight to define the differentiation state of the developing bone marrow macrophages; including CD11b, CD31, CD44, Ly6C, Ly6G, F4/80, Mac2, c-Fos as well as c-Fms. In addition, RNA was preserved during the flight and was used to perform a gene microarray. We found that there were significant differences in the number of macrophages that developed in space compared to controls maintained on Earth. We found that there were significant changes in the distribution of cells that expressed CD11b, CD31, F4/80, Mac2, Ly6C and c-Fos. However, there were no changes in c-Fms expression and no consistent pattern of advanced or retarded differentiation during space flight. We also found a pattern of transcript levels that would be consistent with a relatively normal differentiation outcome but increased proliferation by the bone marrow macrophages that were assayed after 14 days of space flight. There also was a surprising pattern of space flight influence on genes of the coagulation pathway. These data confirm that a space flight can have an impact on the in vitro development of macrophages from mouse bone marrow cells. MESH:Space Flight/Space Flight

ORGANISM(S): Mus musculus

PROVIDER: GSE95544 | GEO | 2017/03/15

SECONDARY ACCESSION(S): PRJNA377415

REPOSITORIES: GEO

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Project description:Tumor-associated macrophages (TAMs) are highly heterogeneous, derived from myeloid monocyte/macrophages or tissue-resident macrophages, and play vital role in tumor progression. Here we adopted a C57BL/6 mouse model imitating the late-stage colorectal liver metastasis (CRLM) by Mc38 colorectal cancer cell injection via the portal vein. We defined the critical period at 7 to 9 days post injection (dpi) for tumor neovascularization on serial sections of CRLM biopsies. The tumor neovascularization was initiated from one of the liver blood vessels via vessel cooption and extended by vascular mimicry and growth of CD34+cells. In samples with increasing sized liver metastases of CRLM, infiltrated Ly6C+ CD11b+ F4/80 monocytes steadily gained the expression of F4/80, a Kupffer cell marker, and then transformed into Ly6C CD11bint F4/80+ cells, which, the same phenotype was also adapted by Ly6C CD11b F4/80+ Kupffer cells. F4/80+ TAMs showed proximity to neovascularization and tumor vessels. Depletion of macrophages or disturbance on macrophage polarization during the crucial period of neovascularization impeded tumor growth and vascularization and resulted in greatly reduced F4/80+ TAMs, yet increased CD11b+ cells due to inhibition of TAM differentiation. In summary, we showed dynamic F4/80+ TAM differentiation within the tumor microenvironment and strengthened its role as perivascular TAMs in CRLM. A set of aliquot samples (about 0.25 cm3) from for flow cytometry preparations (the above method and Fig. 4A) were used for bulk RNA-Sequencing, among these samples, Ht1 and Ht3 were identical samples; F4/80+cells and CD11b+ cells were selected from medium-sized CRLM (Mt2) with microbeads and MS column;A total of 20 samples were sent for analysis. These collected tissues were flash-frozen in liquid nitrogen, stored at - 80°C and shipped on dry ice to the sequencing provider (Novogene, www.novogene.com), who then extracted RNA from these samples and confirmed that all sample RNA were in good quality and adequate concentrations. The resulted RNA-Sequencing data were verified as the clean reads > 95%, Q30 > 90%, and error rate < 0.05%; Square of Pearson Correlation Coefficient (R2) between the biological replicates was > 0.89; R2 of the identical samples (Ht-1 and Ht-3) was > 0.98.

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Evaluation of in vitro macrophage differentiation during space flight

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