TY - JOUR
T1 - BAY11 enhances OCT4 synthetic mRNA expression in adult human skin cells
AU - Awe, Jason P.
AU - Crespo, Agustin Vega
AU - Li, You
AU - Kiledjian, Megerditch
AU - Byrne, James A.
N1 - Funding Information: This work is based on a research collaboration with Fibrocell Science, Inc. and the Clinical Investigations for Dermal Mesenchymally Obtained Derivatives (CIDMOD) Initiative to generate safe patient-specific stem cell-based therapeutics. This work is supported by funding from the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, The Phelps Family Foundation, and Fibrocell Science, Inc. to JAB and National Institutes of Health grant GM67005 to MK. We would like to thank Renee Reijo Pera and Anne Chang for providing the human skin cell line for this study and Patricia Phelps for her valuable editorial assistance and advice.
PY - 2013
Y1 - 2013
N2 - Introduction. The OCT4 transcription factor is involved in many cellular processes, including development, reprogramming, maintaining pluripotency and differentiation. Synthetic OCT4 mRNA was recently used (in conjunction with other reprogramming factors) to generate human induced pluripotent stem cells. Here, we discovered that BAY 11-7082 (BAY11), at least partially through an NF-κB-inhibition based mechanism, could significantly increase the expression of OCT4 following transfection of synthetic mRNA (synRNA) into adult human skin cells. Methods. We tested various chemical and molecular small molecules on their ability to suppress the innate immune response seen upon synthetic mRNA transfection. Three molecules - B18R, BX795, and BAY11 - were used in immunocytochemical and proliferation-based assays. We also utilized global transcriptional meta-analysis coupled with quantitative PCR to identify relative gene expression downstream of OCT4. Results: We found that human skin cells cultured in the presence of BAY11 resulted in reproducible increased expression of OCT4 that did not inhibit normal cell proliferation. The increased levels of OCT4 resulted in significantly increased expression of genes downstream of OCT4, including the previously identified SPP1, DUSP4 and GADD45G, suggesting the expressed OCT4 was functional. We also discovered a novel OCT4 putative downstream target gene SLC16A9 which demonstrated significantly increased expression following elevation of OCT4 levels. Conclusions: For the first time we have shown that small molecule-based stabilization of synthetic mRNA expression can be achieved with use of BAY11. This small molecule-based inhibition of innate immune responses and subsequent robust expression of transfected synthetic mRNAs may have multiple applications for future cell-based research and therapeutics.
AB - Introduction. The OCT4 transcription factor is involved in many cellular processes, including development, reprogramming, maintaining pluripotency and differentiation. Synthetic OCT4 mRNA was recently used (in conjunction with other reprogramming factors) to generate human induced pluripotent stem cells. Here, we discovered that BAY 11-7082 (BAY11), at least partially through an NF-κB-inhibition based mechanism, could significantly increase the expression of OCT4 following transfection of synthetic mRNA (synRNA) into adult human skin cells. Methods. We tested various chemical and molecular small molecules on their ability to suppress the innate immune response seen upon synthetic mRNA transfection. Three molecules - B18R, BX795, and BAY11 - were used in immunocytochemical and proliferation-based assays. We also utilized global transcriptional meta-analysis coupled with quantitative PCR to identify relative gene expression downstream of OCT4. Results: We found that human skin cells cultured in the presence of BAY11 resulted in reproducible increased expression of OCT4 that did not inhibit normal cell proliferation. The increased levels of OCT4 resulted in significantly increased expression of genes downstream of OCT4, including the previously identified SPP1, DUSP4 and GADD45G, suggesting the expressed OCT4 was functional. We also discovered a novel OCT4 putative downstream target gene SLC16A9 which demonstrated significantly increased expression following elevation of OCT4 levels. Conclusions: For the first time we have shown that small molecule-based stabilization of synthetic mRNA expression can be achieved with use of BAY11. This small molecule-based inhibition of innate immune responses and subsequent robust expression of transfected synthetic mRNAs may have multiple applications for future cell-based research and therapeutics.
UR - https://www.scopus.com/pages/publications/84876129161
UR - https://www.scopus.com/pages/publications/84876129161#tab=citedBy
U2 - 10.1186/scrt163
DO - 10.1186/scrt163
M3 - Article
C2 - 23388106
SN - 1757-6512
VL - 4
JO - Stem Cell Research and Therapy
JF - Stem Cell Research and Therapy
IS - 1
M1 - 15
ER -