External modulation of HT‐1080 human fibrosarcoma cells improves urokinase production

Abstract

in Wiley InterScience (www.interscience.wiley.com).Urokinase was produced in a hollow fiber reactor using HT-1080 human fibrosarcoma cells. External modulation comprised replenishing of the medium in the extracapillary space, reducing the serum concentration in the extracapillary space from 10% to 2% and increasing flow rate of the circulating medium in the intracapillary space from 20 to 80 mL/min, each according to a specific protocol. More than sixfold increase was observed in the cumulative urokinase production for two and three medium replenishing modulations of the extracapillary space. After 15 days of continuous operation, the highest cumulative urokinase obtained was 1.63 Â 10 6 PU/mL. SDS-PAGE and zymogram study established that the urokinase obtained was in the high molecular weight range of 54 kDa. The effect of external modulation on cumulative urokinase production was visualized as trajectories with respect to the ratio of lactic acid production rate (LPR) to the glucose uptake rate (GUR). The collective external modulation data showed two separate physiological regions in the cumulative urokinase vs. LPR/GUR plane. The HT-1080 cells exhibited two distinct morphologies in these regions that may be related to acidosis and metastasis. These regions also correspond to low and high urokinase productivity.

Colorimetric Nanoparticle-Embedded Hydrogels for a Biosensing Platform

“…PNHCs without cells were estimated to have a pH of 7.6, while PNHCs with cells were assessed to have a pH of 7.2. The decrease in pH by only 0.4 is relatively reasonable because HT-1080 is well known as a malignant cancer cell; however, it depends on the oxidative phosphorylation and not glycolysis during the energy production process [ 39 , 40 ]. These results suggest the practicality of PNHC for monitoring the pH of live cells.…”

Fiber-Reinforced Silk Composite for Enhanced Urokinase Production Using High-Density Perfusion Culture and Bioactive Molecule Supplementation

“…Also, the supplementation of amino acids and growth factors during the production phase of urokinase in bioreactor culture increased the urokinase activity by 4-fold as compared to the static culture in our study. Low cell density and product degradation are the major reasons for the reduced bioreactor performance. , External modulation, comprised of formulating a suitable medium for the production phase, replenishing the formulated medium from growth phase to production phase, and using perfusion bioreactor culture for maintaining bidirectional diffusional characteristics, ameliorates the factors affecting the urokinase synthesis. Further, many previous studies have highlighted the importance of the matrix during the mammalian culture in terms of surface area, cell attachment, growth rate, sensitivity against shear stress, etc .…”

“…On the other hand, a perfusion bioreactor is more effective owing to its high surface-to-volume ratio, high cell densities, added productivity, economical operation, convenient handling, and lesser space requirement . It has been well established that a high-density perfusion culture assists maximum urokinase production. , Significantly, the presence of cells cultured on a matrix with continuous perfusion of nutrients in the bioreactor mimics the in vivo system, thus assisting the synthesized protein to display properties that closely resemble those of the native proteins . Hence, a perfusion bioreactor culture is poised to provide favorable conditions like less shear stress, substantial inflow of nutrients, and variation in culture period while enhancing yield and productivity.…”