The transfer of ocular cells using collagen.
The present study was designed to evaluate the use of collagen gel loaded with human retinal pigment epithelium (ARPE19) in cellular transfer and to assess its viability within the gel. Collagen solution was prepared by dissolving calfskin in hydrochloric acid to make a final concentration of 2.0 mg/ml and this was mixed with 10,000 ARPE19 cells/ml. The cell viability in gel was determined using MTT assay. van Gieson stain and proliferating cell nuclear antigen (PCNA) were used to identify the location of collagen and to localize the site of cell proliferation, respectively. The ARPE19 cells in gel appeared to be healthy with a rounded morphology. The optimal collagen concentration was 1.9 mg/ml. When this concentration was used to hold cells for over 12 days, it could be seen that the growth rate was the same between day 2 and day 8 in gel and on plastic. When the cell-loaded gels were transferred onto standard tissue culture plastics, progressive cell migrations over time resembling cell migrations in organotypic explant cultures were observed. Upon intravitreal injection of cell-containing collagen suspension into a rabbit's eye, the gel became suspended within the vitreous a few hours after injection (day 0). However, it became obvious that the gel dispersed and spread around the vitreous even after just 24 h. These cells inside the vitreous were PCNA positive, indicating that the human ARPE19 cells have the capacity to proliferate even after 11 days. The present study demonstrated the potential use of collagen gel as a tool in the transfer of cellular matrix onto other substrates. The results show that the cell seeding number must be critically balanced with the concentration of gel for it to be used as transplant material.
Yeung
Chi-Kong
CK
Department of Ophthalmology & Visual Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, China. ckyeung@ust.hk
Chiang
Sylvia Wai-Yee
SW
Chan
Kwok-Ping
KP
Lam
Dennis Shun-Chiu
DS
Pang
Chi-Pui
CP
D000818:Animals; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D002470:Cell Survival; D017690:Cell Transplantation / Q000379:methods*; D003094:Collagen / Q000737:chemistry / Q000494:pharmacology*; D003470:Culture Media; D005123:Eye / Q000166:cytology*; D006801:Humans; D006851:Hydrochloric Acid / Q000737:chemistry / Q000494:pharmacology; D007150:Immunohistochemistry; D008858:Microscopy, Phase-Contrast; D010857:Pigment Epithelium of Eye / Q000166:cytology; D018809:Proliferating Cell Nuclear Antigen / Q000096:biosynthesis; D011817:Rabbits; D013997:Time Factors; D016378:Tissue Transplantation / Q000379:methods*
D003470:Culture Media; D018809:Proliferating Cell Nuclear Antigen; D003094:Collagen; D006851:Hydrochloric Acid