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The novel myxofibrosarcoma cell line MUG-Myx1 expresses a tumourigenic stem-like cell population with high aldehyde dehydrogenase 1 activity.

Lohberger B, Stuendl N, Wolf E, Liegl-Atzwanger B, Leithner A, Rinner B - BMC Cancer (2013)

Bottom Line: These neoplasms mainly affect patients in the sixth to eighth decades of life and the overall 5-year survival rate is 60-70%.In addition, MUG-Myx1 ALDH1(high) cells showed an upregulation of the ABC transporter ABCB1 and ABCG2; higher c-Myc, E-cadherin and SOX-2 expression; and a higher potential for tumourigenicity and proliferation levels.Furthermore, because of their tumourigenicity, the cell line is also suitable for in vivo experiments.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Orthopaedic Surgery, Medical University of Graz, Auenbruggerplatz 5, A-8036, Graz, Austria. birgit.lohberger@medunigraz.at.

ABSTRACT

Background: Myxofibrosarcoma comprises a spectrum of malignant neoplasms withprominent myxoid stromata, cellular pleomorphism, and distinct curvilinear vascular patterns. These neoplasms mainly affect patients in the sixth to eighth decades of life and the overall 5-year survival rate is 60-70%.

Methods: After the establishment of the novel myxofibrosarcoma cell lines MUG-Myx1, cells were characterized using short tandem repeat (STR), copy number variation (CNV), and genotype/loss-of-heterozygosity (LOH) analyses. The growth behaviour of the cells was analyzed with the xCELLigence system and an MTS assay. The tumourigenicity of MUG-Myx1 was proved in NOD/SCID mice. Additionally, a stem-like cell population with high enzymatic activity of aldehyde dehydrogenase 1 (ALDH1(high)) was isolated for the first time from myxofibrosarcoma cells using the Aldefluor® assay followed by FACS analysis.

Results: The frozen primary parental tumour tissue and the MUG-Myx1 cell line showed the same STR profile at the markers D3S1358, TH01, D21S11, D18S51, Penta E, D5S818, D13S317, D7S820, D16S539, CSF1PO, Penta D, Amelogenin, D8S1179, TPOX, and FGY. Typically, myxofibrosarcoma gain and/or amplification was mapped to 7p21.3-q31.1, q31.1-q31.33, q33-q36.2, p21.3, p21.2, p14.1-q11.23, q31.33-q33, p21.2-p14.1, q11.23-q21.3, q36.2-q36.3, which, respectively are known to harbour tumour-associated genes, including TIF, BRAF, MLL3, SMO, and MET. Typically an LOH for myxofibrosarcoma on chr5 q21 was found. In addition, MUG-Myx1 ALDH1(high) cells showed an upregulation of the ABC transporter ABCB1 and ABCG2; higher c-Myc, E-cadherin and SOX-2 expression; and a higher potential for tumourigenicity and proliferation levels.

Conclusion: The new myxofibrosarcoma cell line MUG-Myx1 was established to enrich the bank of publicly available cell lines, with respect to providing comprehensive genetic and epigenetic characterization. Furthermore, because of their tumourigenicity, the cell line is also suitable for in vivo experiments.

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Establishment and characterization of the myxofibrosarcoma cell line MUG-Myx1. (A) IHC analysis on patient tumour tissue showed poorly differentiated tumour components (left) in connection with better differentiated tumour areas with myxoid stroma and curvilinear blood vessels (right). (B) The hematoxylin and eosin (H&E) staining of the MUG-Myx1 cell line showed spindle and multinucleated tumour cells. (C) Strong expression of Vimentin of the MUG-Myx1 cell line confirmed the mesenchymal origin of the tumour cells; nuclei were stained with DAPI. (D) Dynamic proliferation curves for MUG-Myx1; 5 × 103 and 1 × 104 cells were seeded per well and measured with the xCELLigence system. (E) MTS proliferation analysis revealed a 24 h doubling time. (F) The DNA index of 1.15 indicated hyperdiploid tumour cells (left). Gating strategy of cell cycle analysis, to exclude doublets from the total population (right). (G) MUG-Myx1 Tumour formation in NOD/SCID/IL-2rγ (NSG-) mice.
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Figure 1: Establishment and characterization of the myxofibrosarcoma cell line MUG-Myx1. (A) IHC analysis on patient tumour tissue showed poorly differentiated tumour components (left) in connection with better differentiated tumour areas with myxoid stroma and curvilinear blood vessels (right). (B) The hematoxylin and eosin (H&E) staining of the MUG-Myx1 cell line showed spindle and multinucleated tumour cells. (C) Strong expression of Vimentin of the MUG-Myx1 cell line confirmed the mesenchymal origin of the tumour cells; nuclei were stained with DAPI. (D) Dynamic proliferation curves for MUG-Myx1; 5 × 103 and 1 × 104 cells were seeded per well and measured with the xCELLigence system. (E) MTS proliferation analysis revealed a 24 h doubling time. (F) The DNA index of 1.15 indicated hyperdiploid tumour cells (left). Gating strategy of cell cycle analysis, to exclude doublets from the total population (right). (G) MUG-Myx1 Tumour formation in NOD/SCID/IL-2rγ (NSG-) mice.

Mentions: Haematoxylin and eosin (HE)-stained slides of the above-mentioned patient revealed a myxofibrosarcoma G3. The tumour was composed of tumour areas showing a myxoid stroma with classic curvilinear tumour vessels, as well as areas showing a high grade tumour component. Immunohistochemical (IHC) analysis of the patient’s tumour revealed only focal SMA positivity, whereas tests for Desmin, Caldesmon, S100, CD34, EMA, and Pan-CK were negative (Figure 1A).


The novel myxofibrosarcoma cell line MUG-Myx1 expresses a tumourigenic stem-like cell population with high aldehyde dehydrogenase 1 activity.

Lohberger B, Stuendl N, Wolf E, Liegl-Atzwanger B, Leithner A, Rinner B - BMC Cancer (2013)

Establishment and characterization of the myxofibrosarcoma cell line MUG-Myx1. (A) IHC analysis on patient tumour tissue showed poorly differentiated tumour components (left) in connection with better differentiated tumour areas with myxoid stroma and curvilinear blood vessels (right). (B) The hematoxylin and eosin (H&E) staining of the MUG-Myx1 cell line showed spindle and multinucleated tumour cells. (C) Strong expression of Vimentin of the MUG-Myx1 cell line confirmed the mesenchymal origin of the tumour cells; nuclei were stained with DAPI. (D) Dynamic proliferation curves for MUG-Myx1; 5 × 103 and 1 × 104 cells were seeded per well and measured with the xCELLigence system. (E) MTS proliferation analysis revealed a 24 h doubling time. (F) The DNA index of 1.15 indicated hyperdiploid tumour cells (left). Gating strategy of cell cycle analysis, to exclude doublets from the total population (right). (G) MUG-Myx1 Tumour formation in NOD/SCID/IL-2rγ (NSG-) mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4219478&req=5

Figure 1: Establishment and characterization of the myxofibrosarcoma cell line MUG-Myx1. (A) IHC analysis on patient tumour tissue showed poorly differentiated tumour components (left) in connection with better differentiated tumour areas with myxoid stroma and curvilinear blood vessels (right). (B) The hematoxylin and eosin (H&E) staining of the MUG-Myx1 cell line showed spindle and multinucleated tumour cells. (C) Strong expression of Vimentin of the MUG-Myx1 cell line confirmed the mesenchymal origin of the tumour cells; nuclei were stained with DAPI. (D) Dynamic proliferation curves for MUG-Myx1; 5 × 103 and 1 × 104 cells were seeded per well and measured with the xCELLigence system. (E) MTS proliferation analysis revealed a 24 h doubling time. (F) The DNA index of 1.15 indicated hyperdiploid tumour cells (left). Gating strategy of cell cycle analysis, to exclude doublets from the total population (right). (G) MUG-Myx1 Tumour formation in NOD/SCID/IL-2rγ (NSG-) mice.
Mentions: Haematoxylin and eosin (HE)-stained slides of the above-mentioned patient revealed a myxofibrosarcoma G3. The tumour was composed of tumour areas showing a myxoid stroma with classic curvilinear tumour vessels, as well as areas showing a high grade tumour component. Immunohistochemical (IHC) analysis of the patient’s tumour revealed only focal SMA positivity, whereas tests for Desmin, Caldesmon, S100, CD34, EMA, and Pan-CK were negative (Figure 1A).

Bottom Line: These neoplasms mainly affect patients in the sixth to eighth decades of life and the overall 5-year survival rate is 60-70%.In addition, MUG-Myx1 ALDH1(high) cells showed an upregulation of the ABC transporter ABCB1 and ABCG2; higher c-Myc, E-cadherin and SOX-2 expression; and a higher potential for tumourigenicity and proliferation levels.Furthermore, because of their tumourigenicity, the cell line is also suitable for in vivo experiments.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Orthopaedic Surgery, Medical University of Graz, Auenbruggerplatz 5, A-8036, Graz, Austria. birgit.lohberger@medunigraz.at.

ABSTRACT

Background: Myxofibrosarcoma comprises a spectrum of malignant neoplasms withprominent myxoid stromata, cellular pleomorphism, and distinct curvilinear vascular patterns. These neoplasms mainly affect patients in the sixth to eighth decades of life and the overall 5-year survival rate is 60-70%.

Methods: After the establishment of the novel myxofibrosarcoma cell lines MUG-Myx1, cells were characterized using short tandem repeat (STR), copy number variation (CNV), and genotype/loss-of-heterozygosity (LOH) analyses. The growth behaviour of the cells was analyzed with the xCELLigence system and an MTS assay. The tumourigenicity of MUG-Myx1 was proved in NOD/SCID mice. Additionally, a stem-like cell population with high enzymatic activity of aldehyde dehydrogenase 1 (ALDH1(high)) was isolated for the first time from myxofibrosarcoma cells using the Aldefluor® assay followed by FACS analysis.

Results: The frozen primary parental tumour tissue and the MUG-Myx1 cell line showed the same STR profile at the markers D3S1358, TH01, D21S11, D18S51, Penta E, D5S818, D13S317, D7S820, D16S539, CSF1PO, Penta D, Amelogenin, D8S1179, TPOX, and FGY. Typically, myxofibrosarcoma gain and/or amplification was mapped to 7p21.3-q31.1, q31.1-q31.33, q33-q36.2, p21.3, p21.2, p14.1-q11.23, q31.33-q33, p21.2-p14.1, q11.23-q21.3, q36.2-q36.3, which, respectively are known to harbour tumour-associated genes, including TIF, BRAF, MLL3, SMO, and MET. Typically an LOH for myxofibrosarcoma on chr5 q21 was found. In addition, MUG-Myx1 ALDH1(high) cells showed an upregulation of the ABC transporter ABCB1 and ABCG2; higher c-Myc, E-cadherin and SOX-2 expression; and a higher potential for tumourigenicity and proliferation levels.

Conclusion: The new myxofibrosarcoma cell line MUG-Myx1 was established to enrich the bank of publicly available cell lines, with respect to providing comprehensive genetic and epigenetic characterization. Furthermore, because of their tumourigenicity, the cell line is also suitable for in vivo experiments.

Show MeSH
Related in: MedlinePlus