[0042] Example 2: Ams membrane-fibrin cement containing amniotic membranes (AM-FS: Amniotic MEMBRANE-FIBRIN Sealant), the preparation steps are as follows:
[0043] S1, the amnimeric powder prepared in Example 1 was thoroughly mixed with the catalyst solution in pig fibromy protein glue, and prepared according to mass ratio, and the amniotic powder concentration was 130 mg / ml, 65 mg / ml, 32 mg / ml, 16 mg / ml, 8 mg / ML, 4 mg / mL, 2 mg / ml, 1 mg / ml, 0.5 mg / ml, 0.25 mg / ml, 0 mg / ml of amniotic powder - catalyst mixture.
[0044] S2, sequentially take the above concentration of the amniotic powder-catalyst 50 ul, and the 24-well cell culture plate is added, and evenly placed on the bottom of the culture plate, and the equal volume main glue solution is sequentially added to the above culture holes, gently shock, mix well, formation According to the mass ratio, the amniotic powder concentration is: 65 mg / ml, 32 mg / ml, 16 mg / ml, 8 mg / ml, 4 mg / ml, 2 mg / ml, 1 mg / ml, 0.5mg / ml, 0.25 mg / ml, 0.125mg / ML, 0 mg / ml of AM-FS cement.
[0045] The cement of different concentrations of amniotic powder was selected from the 2nd generation of corneal epithelial cells (CEC: CORNEAL Epithelial Cells) to study in vitro cell experiments in vitro cell experiments, and studied the effects of CEC proliferation. Press 1 × 10 3 The CEC density of a / well was inoculated on a 24-well cell culture plate that has been paved in AM-FS cement, and 24-well cell culture plates were placed in 37 ° C, and the volume fraction was 5%. 2 Culture in the incubator for 72 h. The cells of each well were digested with 0.25% trypsin, and the cell suspension of each well was collected, 1000 r / min, centrifuged for 5 min, and the 96-well cell culture plate was added, and the cultured for 24 h was continued. 10 ul of CCK-8 solution was added to each well, and 2 h was continued to determine the absorbance (a) value at 450 nm. All 4 sub-holes per group were repeated 3 times.
[0046] According to CCK-8 results, the maximum AM-FS cement concentration of 0.25 mg / ml is the best ratio concentration, and the results are seen figure 2.
[0047] Am-fs cement concentration of amniotic powder was 0.25 mg / ml of AM-FS cement in the burn therapeutic experiment in rabbit. Building a rabbit severe eye oint base burn model, a total of 32 eyes; according to different treatments, model animals are randomly divided into 4 groups: AM-FS cement group 8 8 eyes, fresh amniotic membrane graft group 8 8 8 eyes, fibrin glue FS (FS is Fibrin Sealant abbreviation) group 8 8 eyes, 8 8 eyes in an antibiotic control group. The four sets of rabbits rows of rabbit row, compared, comparative cornea, and corneal neovascular displacement, comparative corneal. image 3 , Figure 4 Down:
[0048] At 28 days after treatment, there was no statistically significant difference between the AM-FS cement group corneal turbidity and the fresh amniotic membrane graft (P <0.05), and the two groups of corneal turbidity scores were lower than the antibiotic control group (P <0.05). The lower the corneal turbidity score, the more transparent the cornea, the AM-FS cement has the same effect of reducing the corneal turbidity after alkali burn, and the effect of restoring the corneal transparency is the same as the effect of the fresh amniotic membrane transplant. It is superior to the antibiotic control group.
[0049] After alkali burn increases with time, the AM-FS cement group, fresh amniotic membrane group, FS group, the newly ravron area is less than the antibiotic control group (P <0.05), although fresh amniotic membrane transplantation angle The new blood vessel area was less than the AM-FS cement group (P <0.05) in 7d, 21d, and 28d, but the difference was small (0.34, 0.38, 0.88), and the AM-FS cement was visible to the fresh amniotic membrane. Inhibition of the corneal neovascularization of the alkali burn, and the two methods are similar.
[0050] At the 21st, four groups of rabbits, tissue fixtures fixed eyeballs, compared the degree of corneal tissue new blood vessels and inflammatory response, Figure 5 Indicated. AM-Fs cement can alleviate corneal inflammatory response and neovascularization compared to FS group and antibiotic control group.
[0051] BRIEF DESCRIPTION OF THE DRAWINGS AM-FS gear can effectively promote the repair of rabbits with weight alkali burn. After AM-FS cement treatment, the recovery of corneal epithelial healing and transparency, inhibiting the effect of corneal neovascularization is better than antibiotic and simple fibrinogenic treatment group, which can achieve the same treatment with fresh amniotic membrane.
[0052] The wind-dried amniotic powder of the present invention is simple, stable in room temperature, easy to store, is a convenient and effective amniotic article. In this way, the amniotic powder in the amniotic membrane is small in the amniotic tissue, which can be compounded into a wound repair, a wound repair for minimally invasive intervention or special demand, can also be made with a variety of materials. Molecular polymers and synthetic polymer crosslinking are used in the field of tissue engineering, providing new ideas for the development of new amniotic membrane products.
[0053] The AM-FS cement prepared by amniotic powder is made of fibrin glue as a transfer matrix load amniotic membrane, which contains both the rich growth factor from the amniotic tissue, combining the fine biological properties of fibrin glue, and eye on the eyes The effect of forming damage is similar to the fresh amniot membrane. At the same time, it has overcome the fresh amniotic membrane needs to be stitched. The surgical operation is difficult, and it is easy to fall off, and the AM-FS cement can repeatedly drop the eye watch, form a uniform Soft gel-like substances have certain attraction and plasticity, no need to sew, easy to use, strong eye-catching, suitable for various types of eye-catching tissues and skin tissue damage.
