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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

Comparison of sensitivity of KG1a to (A) cytarabine (Ara C), (B) daunorubicin (DNR), and (C) a combination of both on a tissue culture plate system (TCPS), fibronectin (FN)-coated tissue culture plate system (FNTCPS), and FN-coated polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold.Note: PU/PLLA 60:40 was significantly different from TCPS and FNTCPS (*P≤0.01 vs TCPS and FNTCPS) at different time points – that is, 48, 96, and 144 hours – in Ara C, DNR, and a combination of both.
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f4-ijn-10-3603: Comparison of sensitivity of KG1a to (A) cytarabine (Ara C), (B) daunorubicin (DNR), and (C) a combination of both on a tissue culture plate system (TCPS), fibronectin (FN)-coated tissue culture plate system (FNTCPS), and FN-coated polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold.Note: PU/PLLA 60:40 was significantly different from TCPS and FNTCPS (*P≤0.01 vs TCPS and FNTCPS) at different time points – that is, 48, 96, and 144 hours – in Ara C, DNR, and a combination of both.

Mentions: Following the generation of an appropriate PU/PLLA composite that supported cell proliferation and survival for a longer period without changing the cell phenotype, the stem-like KG1a cells were used for drug-testing applications. KG1a cells were treated with selected drugs or their combination, as shown in Figure 4. The percentage proliferation/viability of KG1a cells cultured on TCPSs was reduced to 81.68%±1.51%, 66.73%±2.51%, and 59.03%±5.54% following treatment with Ara C for 48, 96, and 144 hours, respectively (Figure 5A, TCPS). A similar pattern was obtained with Ara C on an FNTCPS with proliferation/viability of 83.03%±3%, 80.66%±1.64%, and 62.28%±6.39% at 48, 96, and 144 hours, respectively (Figure 5A, FNTCPS).


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)

Comparison of sensitivity of KG1a to (A) cytarabine (Ara C), (B) daunorubicin (DNR), and (C) a combination of both on a tissue culture plate system (TCPS), fibronectin (FN)-coated tissue culture plate system (FNTCPS), and FN-coated polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold.Note: PU/PLLA 60:40 was significantly different from TCPS and FNTCPS (*P≤0.01 vs TCPS and FNTCPS) at different time points – that is, 48, 96, and 144 hours – in Ara C, DNR, and a combination of both.
© Copyright Policy
Related In: Results  -  Collection

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

f4-ijn-10-3603: Comparison of sensitivity of KG1a to (A) cytarabine (Ara C), (B) daunorubicin (DNR), and (C) a combination of both on a tissue culture plate system (TCPS), fibronectin (FN)-coated tissue culture plate system (FNTCPS), and FN-coated polyurethane (PU)/poly-l-lactic acid (PLLA) 60:40 scaffold.Note: PU/PLLA 60:40 was significantly different from TCPS and FNTCPS (*P≤0.01 vs TCPS and FNTCPS) at different time points – that is, 48, 96, and 144 hours – in Ara C, DNR, and a combination of both.
Mentions: Following the generation of an appropriate PU/PLLA composite that supported cell proliferation and survival for a longer period without changing the cell phenotype, the stem-like KG1a cells were used for drug-testing applications. KG1a cells were treated with selected drugs or their combination, as shown in Figure 4. The percentage proliferation/viability of KG1a cells cultured on TCPSs was reduced to 81.68%±1.51%, 66.73%±2.51%, and 59.03%±5.54% following treatment with Ara C for 48, 96, and 144 hours, respectively (Figure 5A, TCPS). A similar pattern was obtained with Ara C on an FNTCPS with proliferation/viability of 83.03%±3%, 80.66%±1.64%, and 62.28%±6.39% at 48, 96, and 144 hours, respectively (Figure 5A, FNTCPS).

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