Abstract
Necrotizing enterocolitis (NEC) is a devastating condition of premature infants that results from the gut microbiome invading immature intestinal tissues. This results in a life-threatening disease that is frequently treated with the surgical removal of diseased and dead tissues. Epidermal growth factor (EGF), typically found in bodily fluids, such as amniotic fluid, salvia and mother’s breast milk, is an intestinotrophic growth factor and may reduce the onset of NEC in premature infants. We have produced human EGF in soybean seeds to levels biologically relevant and demonstrated its comparable activity to commercially available EGF. Transgenic soybean seeds expressing a seed-specific codon optimized gene encoding of the human EGF protein with an added ER signal tag at the N’ terminal were produced. Seven independent lines were grown to homozygous and found to accumulate a range of 6.7 +/- 3.1 to 129.0 +/- 36.7 μg EGF/g of dry soybean seed. Proteomic and immunoblot analysis indicates that the inserted EGF is the same as the human EGF protein. Phosphorylation and immunohistochemical assays on the EGF receptor in HeLa cells indicate the EGF protein produced in soybean seed is bioactive and comparable to commercially available human EGF. This work demonstrates the feasibility of using soybean seeds as a biofactory to produce therapeutic agents in a soymilk delivery platform.
Highlights
Each year in the United States, more than 530,000 babies, approximately 12% of total births, are born before 37 full weeks of gestation [1]
Recombinant human EGF (hEGF) is expressed in both cotyledonary-stage embryos regenerated in primary transformation events and subsequently in homozygous soybean seeds
The Epidermal growth factor (EGF) transgenic Line 5 produced in excess of 100 μg hEGF per gm dry seed weight, a level calculated to be much in excess of potential therapeutic requirements
Summary
Each year in the United States, more than 530,000 babies, approximately 12% of total births, are born before 37 full weeks of gestation [1]. As a growing health issue the rate of premature birth has increased by 36 percent since the early 1980s. One of the major problems associated with prematurity is the development of a condition known as neonatal necrotizing enterocolitis (NEC) [2]. This is observed clinically as the abrupt development of bloody diarrhea, abdominal swelling, and tenderness in a premature infant who is otherwise doing well [3]. Current treatment often requires surgical removal of the damaged and dead intestine, often resulting in mortality (about 40%) or, if the infant survives, to manifest significant resulting lifetime problems [3,4,5]. The direct cause of NEC is not known, the most significant contributing
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