Hepatoma SK Hep-1 Cells Exhibit Characteristics of Oncogenic Mesenchymal Stem Cells with Highly Metastatic Capacity - Figure 7

<p>In vitro invasion assay A–C, the invasion of stained adipose-derived msenchymal stem cells (A), bone marrow-derived mesenchymal stem cells (B), and SK cells (C), were evaluated from transwells at 24 hours after culturing under low serum. D, based on the standard curve, the number of invading cells was quantitated. Magnifications: 100× (A–D).</p

Pathological and histological features of primary tumors and metastatic tumors.

<p>A-I, the primary tumors (human xenografts) were formed by the subcutaneous injection of SK cells, and were metastasized to different organs. H and E staining reveals that the histological and pathological features of primary subcutaneous tumor cells (A), metastatic tumor cells (arrow) in liver (B), lung (C), pancreas (D), kidney (E), spleen (F), colon (G), uterus (H), and ovary (I). The tumors consisted of spindle-like shaped cells with ovoid or elongated nuclei containing 2 or more nucleoli per nucleus (A–I). Magnifications: 200 x</p

Gene expression patterns in differentiated SK cells, MSC-ad and MSC-BM.

<p>A and B, expression levels of adipocyte genes, C/EBPα, C/EBPβ, PPAR-γ, and FABP4 (A), osteoblast genes, RBC32 and COL1 (B), and MSC genes, CD73, CD90, and CD105 (A and B) were measured by quantitative PCR in differentiated SK cells, MSC-ad and MSC-BM at day 15 after differentiation. Abbreviations: SK, SK Hep-1 cells; MSC-ad, adipocyte-derived MSC; MSC-BM, bone marrow-derived MSC; FABP4, fatty acid binding protein 4; RBC32, response gene to complement 32; COL1, collage 1A1.</p

Differentiation of three cell types into adipocytes and osteoblasts.

<p>A, SK cells, adipocyte-derived MSC (MSC-ad), and bone marrow-derived MSC (MSC-BM) were used for the differentiation. B, the three cell types were differentiated into adipocytes determined by Oil Red R staining. C, the three cell types were differentiated into osteoblasts determined by Alizarin Red S staining. D, alkaline phosphatase staining was performed after the osteoblast differentiation. Magnifications: 100X</p

Histological features of primary tumors and metastatic tumors.

<p>A, the primary tumors (human xenografts) were formed by the subcutaneous injection of SK cells, and these tumors had a sarcomatous appearance (white color) lacking angiogenesis. B-I, metastatic tumors were found in different organs, such as liver (B), lung (C), spleen, kidney and ureter (D), kidney and ureter (arrow) (E), kidney, liver, and pancreas (arrow) (F), joint at chest and front leg (G), outer peritoneum (H), under skin (arrow) (I) (non-injection site).</p

Tumourigenicity and metastasis of SK cells and its xenograft cells.

<p>The tumourigenicity was evaluated by the injection of SK cells into NOD/SCID/IL2rg mice, and serial transplantation was also performed by the injection of the tumor xenograft cells isolated from primary and metastatic tumors. The tumors were formed, and metastasis was widely found in the mice.</p><p>Tumourigenicity and metastasis of SK cells and its xenograft cells.</p

Characterization and comparison of SK cells with mesenchymal stem cells.

<p>A, flow cytometry was employed to detect classical mesenchymal stem cell (MSC) markers in three cell types, SK Hep-1 cells (SK cells), adipose-derived MSC (MSC-ad), and bone marrow-derived MSC (MSC-BM). B, expression of MSC markers, vimentin (a mesodermal origin marker), and alpha smooth muscle actin (α-SMA) was measured by quantitative PCR in three cell types. Abbreviations: HLA ABC, major histocompatibility complex (MHC) class I antigen; HLA DR, MHC class II antigen; ALP, Alkaline phosphatase.</p

Characterization of isolates from primary tumors and metastatic tumors.

<p>A–D, parental SK cells (A), re-culture of subcutaneous primary tumors (B), re-culture of metastatic tumors in liver (C) and lung (D). E, classical mesenchymal stem cell (MSC) markers were detected and measured in three isolates employing flow cytometry. F, expression of MSC markers, vimentin (a mesodermal origin marker), and alpha smooth muscle actin (α-SMA) was measured by quantitative PCR in three isolates. Abbreviations: HLA ABC, major histocompatibility complex (MHC) class I antigen; HLA DR, MHC class II antigen; ALP, Alkaline phosphatase. Magnifications: 100X</p