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Development and molecular characterization of polymeric micro-nanofibrous scaffold of a defined 3-D niche for in vitro chemosensitivity analysis against acute myeloid leukemia cells.

Nair MS, Mony U, Menon D, Koyakutty M, Sidharthan N, Pavithran K, Nair SV, Menon KN - Int J Nanomedicine (2015)

Bottom Line: Culturing of acute myeloid leukemia (AML) KG1a cells in FN-coated PU/PLLA 60:40 shows increased cell adhesion and cell adhesion-mediated drug resistance to the drugs cytarabine and daunorubicin without changing the original CD34(+)/CD38(-)/CD33(-) phenotype for 168 hours compared to fibronectin tissue culture plate systems.Molecularly, as seen in vivo, increased chemoresistance is associated with the upregulation of anti-apoptotic Bcl2 and the cell cycle regulatory protein p27(Kip1) leading to cell growth arrest.Abrogation of Bcl2 activity by the Bcl2-specific inhibitor ABT 737 led to cell death in the presence of both cytarabine and daunorubicin, demonstrating that the cell adhesion-mediated drug resistance induced by Bcl2 and p27(Kip1) in the scaffold was similar to that seen in vivo.

View Article: PubMed Central - PubMed

Affiliation: Amrita Centre for Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham University, Kerala, India.

ABSTRACT
Standard in vitro drug testing employs 2-D tissue culture plate systems to test anti-leukemic drugs against cell adhesion-mediated drug-resistant leukemic cells that harbor in 3-D bone marrow microenvironments. This drawback necessitates the fabrication of 3-D scaffolds that have cell adhesion-mediated drug-resistant properties similar to in vivo niches. We therefore aimed at exploiting the known property of polyurethane (PU)/poly-L-lactic acid (PLLA) in forming a micro-nanofibrous structure to fabricate unique, not presented before, as far as we are aware, 3-D micro-nanofibrous scaffold composites using a thermally induced phase separation technique. Among the different combinations of PU/PLLA composites generated, the unique PU/PLLA 60:40 composite displayed micro-nanofibrous morphology similar to decellularized bone marrow with increased protein and fibronectin adsorption. Culturing of acute myeloid leukemia (AML) KG1a cells in FN-coated PU/PLLA 60:40 shows increased cell adhesion and cell adhesion-mediated drug resistance to the drugs cytarabine and daunorubicin without changing the original CD34(+)/CD38(-)/CD33(-) phenotype for 168 hours compared to fibronectin tissue culture plate systems. Molecularly, as seen in vivo, increased chemoresistance is associated with the upregulation of anti-apoptotic Bcl2 and the cell cycle regulatory protein p27(Kip1) leading to cell growth arrest. Abrogation of Bcl2 activity by the Bcl2-specific inhibitor ABT 737 led to cell death in the presence of both cytarabine and daunorubicin, demonstrating that the cell adhesion-mediated drug resistance induced by Bcl2 and p27(Kip1) in the scaffold was similar to that seen in vivo. These results thus show the utility of a platform technology, wherein drug testing can be performed before administering to patients without the necessity for stromal cells.

No MeSH data available.


Related in: MedlinePlus

Effect of the Bcl2 inhibitor ABT 737 in inducing cell death in the presence of cytarabine (Ara C) and daunorubicin (DNR).Notes: Note the increased cell death induced by both Ara C and DNR on the polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold at 96 and 144 hours (B–D) following treatment with ABT 737 in comparison to ABT 737 alone (A).Abbreviations: FNTCPS, fibronectin-coated tissue culture plate system; TCPS, tissue culture plate system.
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f6-ijn-10-3603: Effect of the Bcl2 inhibitor ABT 737 in inducing cell death in the presence of cytarabine (Ara C) and daunorubicin (DNR).Notes: Note the increased cell death induced by both Ara C and DNR on the polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold at 96 and 144 hours (B–D) following treatment with ABT 737 in comparison to ABT 737 alone (A).Abbreviations: FNTCPS, fibronectin-coated tissue culture plate system; TCPS, tissue culture plate system.

Mentions: Since we observed the upregulation of Bcl2 and p27Kip1, we analyzed whether Bcl2 inhibition would reverse the drug resistance. Here, we used Bcl2-specific inhibitor ABT 737. As shown in Figure 6, in the presence of ABT 737 alone, the KG1a cells underwent minimal cell death and more than 90% of cells survived, even at 144 hours in culture (Figure 6A). On the contrary, the addition of Ara C and DNR singly or in combination, along with ABT 737, induced significant cell death (Figure 6B–D). Notably, by 144 hours, almost all the cells had been wiped out, in contrast to with ABT 737 alone (Figure 6B–D vs A).


Development and molecular characterization of polymeric micro-nanofibrous scaffold of a defined 3-D niche for in vitro chemosensitivity analysis against acute myeloid leukemia cells.

Nair MS, Mony U, Menon D, Koyakutty M, Sidharthan N, Pavithran K, Nair SV, Menon KN - Int J Nanomedicine (2015)

Effect of the Bcl2 inhibitor ABT 737 in inducing cell death in the presence of cytarabine (Ara C) and daunorubicin (DNR).Notes: Note the increased cell death induced by both Ara C and DNR on the polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold at 96 and 144 hours (B–D) following treatment with ABT 737 in comparison to ABT 737 alone (A).Abbreviations: FNTCPS, fibronectin-coated tissue culture plate system; TCPS, tissue culture plate system.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440427&req=5

f6-ijn-10-3603: Effect of the Bcl2 inhibitor ABT 737 in inducing cell death in the presence of cytarabine (Ara C) and daunorubicin (DNR).Notes: Note the increased cell death induced by both Ara C and DNR on the polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold at 96 and 144 hours (B–D) following treatment with ABT 737 in comparison to ABT 737 alone (A).Abbreviations: FNTCPS, fibronectin-coated tissue culture plate system; TCPS, tissue culture plate system.
Mentions: Since we observed the upregulation of Bcl2 and p27Kip1, we analyzed whether Bcl2 inhibition would reverse the drug resistance. Here, we used Bcl2-specific inhibitor ABT 737. As shown in Figure 6, in the presence of ABT 737 alone, the KG1a cells underwent minimal cell death and more than 90% of cells survived, even at 144 hours in culture (Figure 6A). On the contrary, the addition of Ara C and DNR singly or in combination, along with ABT 737, induced significant cell death (Figure 6B–D). Notably, by 144 hours, almost all the cells had been wiped out, in contrast to with ABT 737 alone (Figure 6B–D vs A).

Bottom Line: Culturing of acute myeloid leukemia (AML) KG1a cells in FN-coated PU/PLLA 60:40 shows increased cell adhesion and cell adhesion-mediated drug resistance to the drugs cytarabine and daunorubicin without changing the original CD34(+)/CD38(-)/CD33(-) phenotype for 168 hours compared to fibronectin tissue culture plate systems.Molecularly, as seen in vivo, increased chemoresistance is associated with the upregulation of anti-apoptotic Bcl2 and the cell cycle regulatory protein p27(Kip1) leading to cell growth arrest.Abrogation of Bcl2 activity by the Bcl2-specific inhibitor ABT 737 led to cell death in the presence of both cytarabine and daunorubicin, demonstrating that the cell adhesion-mediated drug resistance induced by Bcl2 and p27(Kip1) in the scaffold was similar to that seen in vivo.

View Article: PubMed Central - PubMed

Affiliation: Amrita Centre for Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham University, Kerala, India.

ABSTRACT
Standard in vitro drug testing employs 2-D tissue culture plate systems to test anti-leukemic drugs against cell adhesion-mediated drug-resistant leukemic cells that harbor in 3-D bone marrow microenvironments. This drawback necessitates the fabrication of 3-D scaffolds that have cell adhesion-mediated drug-resistant properties similar to in vivo niches. We therefore aimed at exploiting the known property of polyurethane (PU)/poly-L-lactic acid (PLLA) in forming a micro-nanofibrous structure to fabricate unique, not presented before, as far as we are aware, 3-D micro-nanofibrous scaffold composites using a thermally induced phase separation technique. Among the different combinations of PU/PLLA composites generated, the unique PU/PLLA 60:40 composite displayed micro-nanofibrous morphology similar to decellularized bone marrow with increased protein and fibronectin adsorption. Culturing of acute myeloid leukemia (AML) KG1a cells in FN-coated PU/PLLA 60:40 shows increased cell adhesion and cell adhesion-mediated drug resistance to the drugs cytarabine and daunorubicin without changing the original CD34(+)/CD38(-)/CD33(-) phenotype for 168 hours compared to fibronectin tissue culture plate systems. Molecularly, as seen in vivo, increased chemoresistance is associated with the upregulation of anti-apoptotic Bcl2 and the cell cycle regulatory protein p27(Kip1) leading to cell growth arrest. Abrogation of Bcl2 activity by the Bcl2-specific inhibitor ABT 737 led to cell death in the presence of both cytarabine and daunorubicin, demonstrating that the cell adhesion-mediated drug resistance induced by Bcl2 and p27(Kip1) in the scaffold was similar to that seen in vivo. These results thus show the utility of a platform technology, wherein drug testing can be performed before administering to patients without the necessity for stromal cells.

No MeSH data available.


Related in: MedlinePlus