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IRCT20210513051283N1 IRCT 2021-08-15 Iran University of Medical Sciences Evaluation of the expanded conjunctiva in pterygium surgery Pterygium surgery by using autologous conjunctival cells transplantation implanted on human amniotic membrane 2021-09-23 anticipated 30 Complete https://irct.ir/trial/56351 interventional Randomization: N/A, Blinding: Not blinded, Placebo: Not used, Assignment: Single, Purpose: Treatment. N/A primary pterygium-. Intervention group: We will get a sample of conjuctiva, 2x1 mm, from inferior fornix of the eye with pterygium two weeks before operation. After isolation the epithelium from tenon layer, the sample will be sent to laboratory. In laboratory this sample will be chopped and implanted over amniotic membrane (epithelial side). Then, this complex will be put in an acellular culture media, keratonocyte growth media, that consists of 10 ng/ml human epidermal growth factor (Gibco) ،5 _g/ml insulin (Sigma) ،0.5 _g/ml hydrocortisone (Sigma) 30 _g/ml bovine pituitary extract (Sigma) ،50 _g/ml Gentamicin (Gibco) ،50 ng/ml Amphotericin B (Gibco). Calcium concentration is 15/0 Millimol in culture media. Adhesion and proliferation of cells will increase by increasing the calcium concentration to 9/0 Mm. The chopped sample and 500 micro- liters culture media will have cultured in an incubator of 37 centigrade temperature, 5/5 % Co2 and 95% humidity for one to two weeks. the culture media will change every two to three days and at each change cells will evaluate by microscope. we will use of more amount of culture media by adhesion the cells to amniotic membrane. We evaluate the cells based on rate of cells growth and adhesion to amniotic membrane every day. In addition, cells growth is visible by color change of culture media. After ending the cultivation and proliferation of cells on amniotic membrane, the complex will put up in a new culture media that consists of Dulbecco’s Modified Eagle’s Medium (DMEM) and Ham’s F12 media (Gibco), supplemented with 2.5% fetal bovine serum (Hyclone, Logan, Utah), 10 ng/ml human epidermal growth factor, 5 _g/ml insulin, 0.4 _g/ml hydrocortisone, 8.4 ng/ml cholera toxin (Sigma), 50 _g/ml streptomycin (Gibco), and 50 IU/ml penicillin (Gibco) for three days.. Undecided - It is not yet known if there will be a plan to make this available Justification or reason for indecision in sharing IPD is There is no further information
public Hossein Aghaei
Rassoul Akram Hospital, Niyayesh avenue satarkhan street Tehran Iran
Tehran Iran (Islamic Republic of) 1445613131 +98 21 6650 9162 aghaei.h@iums.ac.ir Iran University of Medical Sciences scientific Hossein Aghaei
Rassoul Akram Hospital, Niyayesh avenue Sattarkhan street Tehran Iran
Tehran Iran (Islamic Republic of) 1445613131 +98 21 6650 9162 aghaei.h@iums.ac.ir Iran University of Medical Sciences Iran (Islamic Republic of) presence the primary pterygium no limit no limit Both ِDissatisfaction patients H11.0 Pterygium of eye Treatment - Surgery Intervention group: We will get a sample of conjuctiva, 2x1 mm, from inferior fornix of the eye with pterygium two weeks before operation. After isolation the epithelium from tenon layer, the sample will be sent to laboratory. In laboratory this sample will be chopped and implanted over amniotic membrane (epithelial side). Then, this complex will be put in an acellular culture media, keratonocyte growth media, that consists of 10 ng/ml human epidermal growth factor (Gibco) ،5 _g/ml insulin (Sigma) ،0.5 _g/ml hydrocortisone (Sigma) 30 _g/ml bovine pituitary extract (Sigma) ،50 _g/ml Gentamicin (Gibco) ،50 ng/ml Amphotericin B (Gibco). Calcium concentration is 15/0 Millimol in culture media. Adhesion and proliferation of cells will increase by increasing the calcium concentration to 9/0 Mm. The chopped sample and 500 micro- liters culture media will have cultured in an incubator of 37 centigrade temperature, 5/5 % Co2 and 95% humidity for one to two weeks. the culture media will change every two to three days and at each change cells will evaluate by microscope. we will use of more amount of culture media by adhesion the cells to amniotic membrane. We evaluate the cells based on rate of cells growth and adhesion to amniotic membrane every day. In addition, cells growth is visible by color change of culture media. After ending the cultivation and proliferation of cells on amniotic membrane, the complex will put up in a new culture media that consists of Dulbecco’s Modified Eagle’s Medium (DMEM) and Ham’s F12 media (Gibco), supplemented with 2.5% fetal bovine serum (Hyclone, Logan, Utah), 10 ng/ml human epidermal growth factor, 5 _g/ml insulin, 0.4 _g/ml hydrocortisone, 8.4 ng/ml cholera toxin (Sigma), 50 _g/ml streptomycin (Gibco), and 50 IU/ml penicillin (Gibco) for three days. Evaluation of re-epithelialization of cultivated tissue. Timepoint: Before conjunctival transplantation. Method of measurement: Hematoxilin-eosin staining. Evaluation of collagenesis. Timepoint: Before conjunctival transplantation. Method of measurement: Trichrome-mason staining. Evaluation of re-epithelialization and collagenesis of cultivated tissue. Timepoint: Before conjunctival transplantation. Method of measurement: Expression of integrin B 1,MUC4,CX43,Oct4,p63,CK19,CK13,CK12,CK3 by RT-PCR. Healing process of transplanted conjunctiva. Timepoint: Day 1,3,7 and months 1,3,6,12. Method of measurement: Examination and Fluorescin staining. Iran University of Medical Sciences Approved 2021-04-20 Ethics committee of Iran University of Medical Sciences Hemmat Highway near to Milad Tower Iran University of medical Sciences Tehran Tehran Iran (Islamic Republic of)