MicroRNAs as Engineering Targets: Pathway Manipulation to Impact Bioprocess Phenotypes - MicroRNAs as Tools in Biopharmaceutical Production - page 73

inefficient mixing that exposes the cells to different oxygen tensions ranging from
absence of oxygen (hypoxia) to above-atmospheric concentrations (hyperoxia) (Dun-
ster et al. 1997 ). An example of a genetic engineering approach to reduce oxidative
stress is the over-expression of the antioxidant haptoglobin which increased tolerance
to oxidative stress in CHO cells (Tseng et al. 2004 ). The effects of shear stress on cell
growth, metabolite consumption, and protein production were investigated in recom-
binant CHO cell cultures producing tissue plasminogen activator (tPA) and human
growth hormone (hGH) (Keane et al. 2003 ; Senger and Karim 2003 ). In both cases,
shear stress had a negative effect on recombinant protein production. Elevated levels
of ammonia were identified as another stress condition that lowers the expression of
genes managing cell cycle and protein folding, up-regulates genes affecting energy
metabolism and induces protein degradation (Chen and Harcum 2006 ; 2007 ). Cold
stress was shown to have positive effects on recombinant protein production in CHO
cells (Fox et al. 2005 ) and the exogenous over-expression of the cold-inducible RNA-
binding protein (CIRP) increased the titer of recombinant protein (Tan et al. 2008 ).
Changes in expression profiles of several miRNAs have been observed in response
to ER and hypoxic stress conditions: (1) miR-708 was shown to be induced during
ER stress by the transcription factor CCAAT enhancer-binding homologous protein
(CHOP), and may facilitate the enhancement of ER protein-folding capacity under
the stress of accelerated protein synthesis (Behrman et al. 2011 ); (2) miR-204 sup-
ports ER and oxidative stress induction in human trabecular meshwork cells. This
miRNA inhibited two genes involved in the elimination of damaged and misfolded
proteins (SERP1/RAMP4 and M6PR) and facilitated the increase of carbonylated
proteins (Li et al. 2011 ); (3) The miR-221/222 cluster was down-regulated during
ER stress in human hepatocellular carcinoma cells. The ectopic introduction of miR-
221/222 mimics increased ER-stress induced apoptosis which was associated with
p27 Kip1 and MEK/ERK-directed cell cycle regulation (Dai et al. 2010 ); (4) miR-15a,
miR-16, and miRs-20a were down-regulated at hypoxic conditions in human car-
cinomas (Hua et al. 2006 ); (5) miR-26, miR-107, and miR-210 were up regulated
in neoplastic cells in response to low oxygen. These miRNAs are likely to decrease
the pro-apoptotic signaling in a hypoxic environment (Kulshreshtha et al. 2007 ).
miR-210 was also found to be progressively up-regulated in endothelial cells in hy-
poxic conditions and inhibited receptor tyrosine-kinase ligand Ephrin-A3 which is
critical in vascular development (Fasanaro et al. 2008 ); (6) The up-regulation of the
miR-34 family, while being part of the p53 network, can be also implicated as a
stress response to DNA damage, hyperactive cytokine signaling, and hypoxia (He
et al. 2007 ); (7) The miR 17-92 cluster was shown to target hypoxia-inducible factor
alpha (Hif-1

), a transcriptional factor known to regulate cellular response to hy-
poxia. The latter has an important role in various biological processes such as glucose
metabolism, pH regulation and angiogenesis (Taguchi et al. 2008 ); (8) miR-31 was
shown to activate Hif-1

α

via the inhibition of factor-inhibiting hypoxia -inducible
factor (HIF) (Liu et al. 2010 ).
Several miRNAs were found to be associated with oxidative stress: (1) the bi-
cistronic transcript miR-144/451 is involved in oxidative stress in erythroid cells.
miR-144 was shown to modulate the oxidative stress response in K562 and primary

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