Growth and viability of fed-batch cultures in response to AIP
As in the batch cultures, we investigated the effects of AIP on the
growth and viability profiles of CHO-aIL-1β cells in the fed-batch
cultures. AIP-treated cells were compared to the 0 µM AIP control
(Figure 4). Both the control cultures and those stimulated with 1 µM AIP
grew relatively rapidly with similar kinetics before slowing and
reaching a maximum by day 10, and then rapidly dropping off thereafter.
As in the Figure 2A batch cultures, correlated with higher additions of
AIP, an increasing lag in growth was observed (Figure 4A). After this
initial lag phase, the 2, 3 and 4 µM AIP-containing cultures grew
rapidly for an additional 6 days before reaching a maximum, with 3 and 4
µM AIP cultures exhibiting a stationary phase before declining. Compared
to the control (20.2 +/- 1.7 x 106 cells/mL),
significantly lower maximum cell densities (ANOVA, P < 0.05)
were observed at AIP concentrations of 3 µΜ (13.4 +/- 0.9 x
106 cells/mL) and 4 µΜ (10.3 +/- 0.7 x
106 cells/mL). Concomitantly, for the 3 and 4 µM AIP
cultures, the IVCD values were significantly increased (ANOVA, P
< 0.05) compared to the control, 1 µM, and 2 µM cultures,
indicating lower maximum growth but increased culture longevity.
The viability profiles for all cultures are shown in Figure 4B. Control
and 1 µM AIP-containing cultures maintained viabilities
>90%, until they decreased after day 10. Cultures
containing 2 - 4 µM AIP exhibited an initial and AIP dose-related drop
in viability by day 2. In particular, both the 3 and 4 µM AIP cultures
had viabilities of less than 50% by culture day 2. Similar to the batch
cultures, a rapid recovery in viability for all AIP cultures was
observed on day 4. The cell viability remained above 90% after day 6
until cultures reached their maximum cell concentration, before
declining again. After day 10, a rapid decline in viability was observed
for control, 1 and 2 µM cultures, while 3 and 4 µM cultures had a more
prolonged decline phase. Overall, as for the batch cultures, the initial
decline in cell viability could be due to autophagic death, and then the
autophagy induction may promote the subsequent rapid recovery and
altered growth characteristics (prolonged stationary phase), either
selecting cells or providing the conditions that enhance recombinant
protein productivity.