Figure 1.
BM adipocytes support the survival and proliferation of primary AML. (A) Immunofluorescence of primary AML blasts stained for CD34+ (Alexa Fluor 568 [red]) and neutral lipids (green) using BODIPY 493/503. All images are representative of 6 AML patient samples. Original magnification ×63. Scale bar = 10 μm. (B) Freshly isolated AML, AML cultured for 1 day, and AML cultured for 2 days stained with the neutral lipid BODIPY 493/503 dye and analyzed by flow cytometry. Data are represented as mean ± standard deviation. (C) Freshly isolated nonmalignant CD34+ cells and freshly isolated AML samples stained with the neutral lipid BODIPY 493/503 dye and analyzed by flow cytometry. (D) AML patients samples in monoculture and cocultured with BM-derived adipocytes for 6 days and then stained with PI/Annexin V. The line through the data indicates the median. (E) AML blasts incubated alone or with adipocytes or BMSC for 6 days and AML blasts counted using flow cytometry and Trypan blue exclusion (n = 12). The line through the data indicates the median. (F) Nonmalignant CD34+ cells cultured alone or in coculture with BMSC or adipocytes and CD34+ cell counted using flow cytometry and Trypan blue exclusion (n = 5). The line through the data indicates the median. (G) AML blasts from 3 different patients were cultured alone or with adipocytes or BMSC for 6 days and then placed in a CFC assay for 15 days. Colonies were then counted. Data are represented as mean ± standard deviation. (H) primary AML cells (2 × 106; 4 individual patient AML) cultured on BM adipocytes or cultured alone and then 2 × 106 viable cells were injected into NSG mice. Engraftment was measured using human CD33 and human CD45. Shown in the flow figure are the characteristics of AML#12 engraftment into BM and spleen. In the dot plot, each AML engraftment into NSG mice is shown for BM and spleen, and the engraftment of the AML cultured alone is shown by a shaded circle. The line through the data indicates the median. *P < .05. Ad, adipocytes; DAPI, 4′,6-diamidino-2-phenylindole; DIC, differential interference contrast; MFI, mean fluorescent intensity.

BM adipocytes support the survival and proliferation of primary AML. (A) Immunofluorescence of primary AML blasts stained for CD34+ (Alexa Fluor 568 [red]) and neutral lipids (green) using BODIPY 493/503. All images are representative of 6 AML patient samples. Original magnification ×63. Scale bar = 10 μm. (B) Freshly isolated AML, AML cultured for 1 day, and AML cultured for 2 days stained with the neutral lipid BODIPY 493/503 dye and analyzed by flow cytometry. Data are represented as mean ± standard deviation. (C) Freshly isolated nonmalignant CD34+ cells and freshly isolated AML samples stained with the neutral lipid BODIPY 493/503 dye and analyzed by flow cytometry. (D) AML patients samples in monoculture and cocultured with BM-derived adipocytes for 6 days and then stained with PI/Annexin V. The line through the data indicates the median. (E) AML blasts incubated alone or with adipocytes or BMSC for 6 days and AML blasts counted using flow cytometry and Trypan blue exclusion (n = 12). The line through the data indicates the median. (F) Nonmalignant CD34+ cells cultured alone or in coculture with BMSC or adipocytes and CD34+ cell counted using flow cytometry and Trypan blue exclusion (n = 5). The line through the data indicates the median. (G) AML blasts from 3 different patients were cultured alone or with adipocytes or BMSC for 6 days and then placed in a CFC assay for 15 days. Colonies were then counted. Data are represented as mean ± standard deviation. (H) primary AML cells (2 × 106; 4 individual patient AML) cultured on BM adipocytes or cultured alone and then 2 × 106 viable cells were injected into NSG mice. Engraftment was measured using human CD33 and human CD45. Shown in the flow figure are the characteristics of AML#12 engraftment into BM and spleen. In the dot plot, each AML engraftment into NSG mice is shown for BM and spleen, and the engraftment of the AML cultured alone is shown by a shaded circle. The line through the data indicates the median. *P < .05. Ad, adipocytes; DAPI, 4′,6-diamidino-2-phenylindole; DIC, differential interference contrast; MFI, mean fluorescent intensity.

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