Transgenic Expression and Functional Characterization of Human Platelet Derived Growth Factor BB (hPDGF-BB) in Tobacco (Nicotiana tabacum L.)

Abstract

Recombinant human platelet derived growth factor BB (rhPDGF-BB) is clinically approved for treating diabetic neuropathic ulcers. Plant-based expression systems offer less expensive ways of producing recombinant drugs, which do not require purification for clinical use. From this perspective, rhPDGF-BB is an ideal candidate for expression in plants as it can be applied topically. Here, we report a proof of concept study, in which rhPDGF-BB was expressed in tobacco plants, and its biological activity was tested in vitro. The mature human platelet derived growth factor BB (hPDGF-BB) gene was codon-optimized for tobacco and fused with ER targeting and retention signals, 5' and 3' UTRs of arc5-1 gene along with CaMV 35S promoter, and then, transferred by Agrobacterium-mediated transformation. Gene and protein expression of hPDGF-BB were confirmed by PCR and immunoblot studies. Bioactivity of hPDGF-BB expressed protein was determined by in vitro assays such as proliferation and migration in NIH3T3 cells. Our data reveals that total soluble proteins containing hPDGF-BB from transgenic plants showed a 4.5-fold increase in fibroblast proliferation compared to non-transgenic plants. Furthermore, plant-made rhPDGF-BB induced chemotaxis of treated cells and promoted wound healing in vitro. These results clearly demonstrate that functionally active rhPDGF-BB protein can be produced in plants and might have therapeutic benefits.

Transgenic Silkworm-Based Silk Gland Bioreactor for Large Scale Production of Bioactive Human Platelet-Derived Growth Factor (PDGF-BB) in Silk Cocoons

“…Thus, the clinical demand for PDGF-BB for a wide range of applications in biomedicine and bio-cosmetics has increased, drawing much attention to the recombinant expression of PDGF-BB. For decades, several attempts have been made to produce the recombinant PDGF-BB in the hosts of the yeasts Pichia Pink [ 5 ] and tobacco N. tabacum L. [ 6 ]. However, poor yields, complicated processing, and low bioactivity have severely limited their ability to meet the market’s demand, especially with the increasing number of applications for cell therapy and translational medicine.…”

Storage of the Recombinant Protein hPDGF-BB in the Culture of Pichia pastoris

“…The rhPDGF-BB has been approved by FDA as the treatment for diabetic foot ulcer and self-bone grafting [1] . It has been produced in a variety of heterologous systems including Escherichia coli [2][3][4] , Chinese hamster ovary cells [5] , Saccharomyces cerevisiae [6,7] , baculovirus [8] , vaccinia viruses [9] , mushroom [10] , plant [11] and Pichia pastoris [12] . Babavalian et al [12] reported a high efficiency of 30 mg/L in Pichia pink, a P. pastoris mutant cell, without optimization.…”