A method for constructing tissue engineered corneal limbus
A tissue engineering and limbal technology, applied in the field of bioengineering medical materials, can solve the problem of few decellularized limbal stroma, and achieve the effect of strong in vitro proliferation ability, wide source, and guaranteed stability
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Embodiment 1
[0034] Cut out the fresh porcine limbal stroma containing Descemet's membrane, including 1mm sclera, 2mm peripheral cornea, and a thickness of about 0.2mm.
[0035] 1. Preparation of carrier
[0036] The limbal tissue is protected with a protective solution throughout the process. The components of the protective solution are: PBS buffer solution with 6g / L hyaluronic acid, 7g / L chondroitin sulfate, 6g / L dextran and 2.5mg / L tobramycin , adjust the pH value to 7.2, and the osmotic pressure to 310mOsm;
[0037] Put the corneal limbus in the protective solution containing 0.2% Triton+200U / ml DNase, set the shaker speed to 100 rpm, and the temperature to 25°C, and digest for 3 hours to remove the DNA components in the cornea; the decellularized The corneal limbus was placed in the protective solution and rinsed for 4 hours to obtain the decellularized limbal carrier.
[0038] 2. Construction of tissue engineered limbus
[0039] Limbal cells or stem cells were divided into 2 × 10...
Embodiment 2
[0041] Cut out the fresh porcine limbal stroma containing Descemet's membrane, including 1mm sclera, 2mm peripheral cornea, and a thickness of about 0.2mm.
[0042] 1. Preparation of carrier
[0043] The limbal tissue was protected with protective solution throughout the whole process. The components of the protective solution were: PBS buffer solution with 8g / L hyaluronic acid, 10g / L chondroitin sulfate, 5g / L dextran and 3mg / L tobramycin, Adjust the pH value to 7.3 and the osmotic pressure to 320mOsm;
[0044] Put the limbus in a protective solution containing 0.3% sodium lauryl sulfate + 1000U / ml DNase, set the shaking table speed to 150 rpm, and the temperature to 20°C, digest for 4 hours to remove the DNA components in the cornea ; Rinse the decellularized limbus in protective solution for 3 hours to obtain the decellularized limbal carrier.
[0045] 2. Construction of tissue engineered limbus
[0046] Limbal cells or stem cells were divided into 2 × 10 4 / cm 2 The de...
Embodiment 3
[0048] Cut out the fresh porcine limbal stroma containing Descemet's membrane, including 1mm sclera, 2mm peripheral cornea, and a thickness of about 0.2mm.
[0049] 1. Preparation of carrier
[0050] The limbal tissue was protected with a protective solution throughout the process. The components of the protective solution were: 5g / L hyaluronic acid, 18g / L chondroitin sulfate, 10g / L dextran and 4mg / L tobramycin were added to the PBS buffer. Adjust the pH value to 7.4 and the osmotic pressure to 340mOsm;
[0051] Put the corneal limbus in a protective solution containing 0.5% sodium dodecylsulfonate + 500U / ml DNase, set the shaking table speed to 120 rpm, and the temperature to 30°C, digest for 2 hours to remove the DNA in the cornea Composition: the decellularized corneal limbus is placed in the protective solution and rinsed for 5 hours to obtain the decellularized limbal carrier.
[0052] 2. Construction of tissue engineered limbus
[0053] Oral mucosal epithelial cells o...
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