demineralized bone allografts
technology of manufacturing osteoplastic material
liquid preserving mediums
V.I. Saveliev, M.V. Lekishvili, А.А. Bulatov, Yu.А.Rykov, A.Yu.
RyabovDemineralized bone allografts (DBA) have received wide application in the course of treatment of patients with a various pathology of a musculoskeletal system. It is known that DBA useful prop-erties connect with the bone morphogenetic proteins, providing osteoinductive effect. Their sensi-tivity to external influence forces to observe DBA preparation and storage rules. Results of a series of experiments have shown an identical inductive bone formation at DBA storage of donor tissues within 48 hours at 5оС and 24 hours at 18±3оС. Simultaneously, storage of bone tissues at tem-perature a minus 20оС within 6 weeks does not reduce their osteoinductive and bioplastic proper-ties to a demineralization.
One of components of an optimum technique DBA manufacturing are ways of their steriliza-tion and preservation. There are the simple ways of DBA sterilization which are not demanding the difficult equipment, such, as sterilization in liquid mediums by means of chemical agents, including antibiotics. 0,25 % a solution of formalin strengthened by introduction in it of solutions of antibiotics can be that. At use of a similar admixture periods of DBA storage at 5°С this medium make 180-200 days. Formalin as the sterilizing and preserving agent has received the wide and deserved recognition among many experts. On the basis of own positive experience we do not share opinion of some authors on unfitness of formalin for DBA preparation because formalin blasts osteoinductive system and possesses carcinogenicity. Concerning cancerogenic properties it is known that any chemical substance at wrong and long application is capable to cause a cellular malignancy and development of tumors. Our technique of application of formalin for manufacturing DBA completely excludes this danger.
There is a cold-resistant liquid medium, suitable for preparation, preservation and sterilization of DBA. Into its structure enter: Citras acidic (1,0); a glucose (3,0); Furacilinum (0,001); sodium bromide (0,2); alcohol ethyl 95 ° (15-20,0); a dimethylsulfoxide (5-10,0); Amikacinum Zinci sulfas (0,1); the distilled water to 100 ml. After a preparation demineralization place in tight container with the presented liquid medium and transfer to a deep-freezer. For 10 days carry out DBA bacteriological control on sterility. Optimum periods of DBA storage in medium make 6 months at temperature a minus 10°С, and at addition in it of a dimethylsulfoxide – 12 months at temperature a minus 20°С.
We offer an original way of dehydration and sterilization implants by their chemical treatment from the frozen condition an admixture of ethanol, acetone and sulfosalicylic acids. After a demi-neralization and excision of the rests of acid bone implants drain and place in Sodium chloride normal saline solution, from 0,5 % of Dimexidum and 0,25 % glutar aldehyde. Then implants transfer to a household refrigeration cabinet with a temperature regimen 5-7°С in which them maintain within 18-24 hours. After that preparations freeze at temperature a minus 20°С and more low. In such conditions implants can be stored till the moment of their chemical treatment by the replacing admixture consisting of ethanol, acetone and sulfosalicylic acids. An admixture before use without fail cool to temperature a minus 20-30°С. The following stage – the basic. It includes processing of frozen implants the cooled replacing admixture which changes three times every 3-4 day. And, in the first change ethanol and acetone take in a proportion 2:1, in the second – 1:1, in the third 1:2. At last change in a solution add sulfosalicylic acid from calculation 0,5 g on 100 ml of a solution. Preparation process comes to an end with that implants in sterile conditions take from a replacing solution, maintain 20-30 minutes under sterile napkins and display in sterile, tight container. At this stage carries out the control on sterility. Process of evaporation of steams of acetone can be accelerated if to carry out it under vacuum. Storage of dehydrate DBA carry out in a household refrigeration cabinet (till 1 year), or at a room temperature (to 6 months).
The offered way, unlike sublimation drying, is simpler, is effective enough and popular. Its application does not break osteoinductive properties of a bone tissue as ethanol, acetone and sul-fosalicylic acid operate on it in a sparing regimen and at low temperature. As a result of bilateral diffusion there is a gradual replacement of the water transformed into ice with alcohol and acetone. At the moment of extraction implants from an admixture at a room temperature or under vacuum, these substances evaporate also tissues become dehydrate.
Some ways of DBA manufacturing surveyed in article, do not demand the difficult equipment and the big material inputs, thanks to the simplicity and general availability. Similar techniques can promote the organization in Russia to wider network of the specialized services capable in the maximum volume and in the shortest terms to provide interested medical institutions with this valuable osteoplastic material.