HEPARIN-BINDING EGF–LIKE GROWTH FACTOR AS A PROSPECTIVE MEDIATOR OF TISSUE REPAIR AND REGENERATION
Journal: Biotechnologia Acta (Vol.17, No. 2)Publication Date: 2024-04-30
Authors : A.V. Didan A.A. Siromolot D.V. Kolybo;
Page : 33-35
Keywords : HB-EGF; recombinant protein; proliferation; tissue repair.;
Abstract
Aim. To obtain bioactive recombinant human HB-EGF and investigate its proliferation capacity. Methods. The pET32(a)-HB-EGF cloning plasmid was extracted from E. coli DH10B cells and transformed into E. coli BL21 (DE3) Rosetta cells. The expression of HB-EGF was induced with IPTG and the protein was purified by metal-affinity chromatography with Co²⁺-NTA. Purified HB-EGF was analyzed by SDS-PAGE. The interaction of HB-EGF with receptors was analyzed by flow cytometry using A431 cells. The effect of HB-EGF on the proliferation of 3T3 and L929 fibroblast cell lines was accessed using the MTT assay. Results. High-quality HB-EGF was obtained through an optimized protocol involving plasmid purification, E. coli transformation, and protein purification. SDS-PAGE confirmed the purity of HB-EGF. Flow cytometry validated the binding of EGFP-HB-EGF to A431 cell receptors. The MTT assay data showed that HB-EGF significantly promoted the proliferation of 3T3 cells in a dose-dependent manner, with a 67% and 112% increase in cell density in the presence of 500 ng/mL and 1000 ng/mL of HB-EGF, respectively. No significant changes in the proliferation of L929 cells were observed under the same concentrations of HB-EGF. Conclusions. We have successfully optimized a protocol to produce and purify biologically active HB-EGF in E. coli. The bioactivity of HB-EGF was validated in A431 and 3T3 cell lines, demonstrating its potential to interact with receptors and significantly increase cell proliferation. This bioactive HB-EGF can be potentially used for tissue repair and regeneration.
Other Latest Articles
- ABSORPTION AND LUMINESCENCE PROPERTIES OF ACID AND SALT FORMS OF MONONUCLEOTIDES, THEIR COMPONENTS AND COMPLEXES WITH D-MANNITOL AT ROOM TEMPERATURE
- PREVENTION OF MERCURY-INDUCED EXCITOTOXICITY IN PRESYNAPTIC BRAIN NERVE TERMINALS WITH CARBON DOTS
- CALIX[4]ARENES С-715 AND С-772 AS INSTRUMENTS OF INFLUENCE ON Са2+-TRANSPORT AND BIOENERGETICS IN MYOMETRIAL MITOCHONDRIA
- BIOCHEMICAL INDICATORS OF GASTRIC JUICE IN INCOMPETENCE OF THE PHYSIOLOGICAL GASTRIC CARDIA
- AMARANTHUS AS A SOURCE OF POLYPHENOLIC COMPOUNDS AND FLAVONOIDS FOR USE IN MEDICINE
Last modified: 2024-06-13 00:50:43