Coexpression of CD9 augments the ability of membrane-bound heparin-bindingepidermal growth factor-like growth factor (proHB-EGF) to preserve renal epithelial cell viability

Background. Transfection of renal epithelial cells (NRK 52E) with membrane- associated heparin-binding epidermal growth factor-like growth factor (proH B-EGF) increased renal epithelial cell survival by promoting cell-cell and cell-extracellular matrix interactions. ProHB-EGF has been shown to form a complex in the plasma membrane with the tetraspanin CD9, an interaction tha t significantly increases the effectiveness of proHB-EGF as a juxtacrine mi togenic agent. Methods. We examined whether the coexpression of proHB-EGF and CD9 would in crease renal epithelial cell survival. CD9 was stably transfected into NRK 52E cells, either alone (NRKCD9) Or together with proHB-EGF (NRKboth). Results. Juxtacrine mitogenic activity of NRKCD9 was no different than in c ells transfected with vector alone (NRKvector), but was increased by NRKbot h; juxtacrine mitogenic activity by NRKboth was twofold greater than when p roHB-EGF was transfected alone (NRKproHB-EGF). When grown in 10% fetal calf serum, growth rates were similar among all transfectants. However, in 1% f etal calf serum, NRKproHB-EGF grew 50% faster than NRKvector or NRKCD9, and NRKboth grew 20% to 50% faster than NRKproHB-EGF at one, two, and three da ys of culture. NRKproHB-EGF attachment to plastic substratum at one, two, a nd three hours was 250% greater than that of NRKvector, and NRKboth was 20% to 30% greater than that of NRKproHB-EGF. Coating plates with either poly 2-hydroxyethyl methacrylate or the GRGDTP peptide prevented normal cell-ext racellular matrix attachment, and NRKvector or NRKCD9 failed to attach or f orm cell-cell attachments. NRKproHB-EGF exhibited 300% and NRKboth exhibite d 600% greater cell viability under these conditions. Expression of type I and type III collagen mRNA was enhanced similarly in NRKproHB-EGF and NRKbo th, but the expression of beta(1) integrin was up-regulated only in NRKboth . Conclusions. Coexpression of proHB-EGF and CD9 may render the renal epithel ial cells more resistant to disruption of cell-cell and cell-matrix interac tions and could accelerate the re-establishment of these attachments.