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Many bone diseases need bone transplantation. The best clinical effect for the patients is to obtain autologous bone but they have to suffer the pain and the risks from the bone surgery, at the same time the surgery time may be extended and the infection chance of post operation increased, the hospitalization time and the expenses as well. Besides various complications are possible after the autogenous bones are removed. Moreover, when the patients are too young or too old, the needed bone is larger or more, the autogenous bones can not meet the surgery requirements any more, the homogenous bones (or allograft bones) from human donors become the best substitutes. Therefore, many internal and overseas Tissue Banks preparing and supplying the homogenous tissue transplantation materials have been established. Those only preparing and supplying homogenous bones are called bone bank and those hospitals which prepare and supply for them are called hospital bone bank, those supply for the society are called regional bone bank.

Frequently Asked Questions

How is the clinical effect of allograft bone transplantation?

Whether can the allograft bone transplantation probably cause the immunological rejection or infection?

Whether can the allograft bone transplantation probably cause disease transmission?

Whether can the transplanted allograft bone survive?

Whether do deep frozen/freeze drying radiation allograft bones have the properties of osteoinduction and bone conduction?

Whether does the specific antigen of HLA or blood type of the donor have to be in accordance with the recipient for the allograft bone transplantation?

Which one is better, deep-frozen or freeze drying?

Question one: How is the clinical effect of allograft bone transplantation?

Autologous bone has its best clinical effect for the bone transplantation, so allograft bone should be compared with autologous bones when it`s clinical effect is measured. The survival rate of the allograft bone, the difference effect between them may decide whether allograft bone is worth using. The survive rate of allograft bone transplantation would lie on multi-factors such as the cause of bone injury, the type and the position and the size of the transplanted bone, the age of the patients, the local blood supply. Generally speaking, the survival rate of deep-freezing/freeze drying radiation allograft bone can be up to 80%-90%(Komender,1991, Malinin,1989). The younger patients are better than the older, the smaller bone transplantation better than larger ones, cancellous bones better than cortical ones, bone cavity filling better than bone bulk plugging, the transplantation without joints better than those with the joints ones, the more muscle covering better than the less. The survival rate for the smaller bone defect filling and vertebrate column fusion can both be up to 95% or higher, while the survive rate for the massive bone transplantation and knee joints injury repairing is only 70-80%(Czitrom,1992). The main reason for the failure of big section bone and bone joints transplantation is deep infection, the absorption of the transplanted bone, bone fracture, bone nonunion and bone tumor recurrence. Most complications can be improved by the conservative treatment or second surgery. To promote the allograft bone transplantation technology, our country held two bone transplantation symposium in 1995 and 2001 and many hospitals reported a good few cases with similarly good clinical effect as the report from other countries.

In animal experiments the bone transplantation conditions are easily controlled when the clinical effect of allograft bones and autologous bone defect are compared. Many similar experiments have proved that allograft bones obviously are not as good as autologous ones (Itoman,1991). However it is difficult to do so in the clinical practices, for different situation of different patients it is impossible to cut the same size autologous bone as the allograft bone from the patients as a comparison. The situation in the department of stomatology and the cases of vertebrate column fusion seems different for that the transplantation material is small and the state of illness is almost similar in different cases. Mazson(1990) once repaired the dental socket cleft with the freeze dried allograft bones and the effect was similar to the autologous bones. Allard (1987) once used the deep-freezing allograft bone, autologous bone and the mixture bone to repair the jaw bones defect and dental socket cleft, proving their effects to be no differences. For vertebrate column fusion some scholars think the autologous bones are better than allograft bones (Fernyhough,1991), while some other scholars think their effects are similar (Maschlert, 1990, Schwarzenbach, 1996). Chinese Plastic Surgery Hospital reported that the survival rate of the implantation of autogenous ilium, autogenous costal cartilage, allogenous costal cartilage grafts was respectively 66%, 78%, 90%, and are not better than allogenous ones. Donald chose the bone absorption as the index and observed that radiation allograft cartilages are better than autogenous cartilages. These clinical reports are not good enough to prove which is better between autogenous bones and allograft bones, but these reports can tell us the difference of the patients’ state and the operation conditions sometimes have more influences than the differences of the bone materials. Overall, under the conditions of being unable to obtain autogenous bones, freeze radiation allograft bone can meet the emergent need for the bone material in clinical bone transplantation.

Question Two: Whether can the allograft bone transplantation probably cause the immunological rejection or infection?

Deep-frozen (-80℃), freeze-drying and radiation can significantly reduce the immunological rejection of allograft bones (Pellet,1983). Many animal experiments have proved that the ossifying effect is obviously better than those fresh allograft bones without deep-frozen or being radiated. The patients’ clinical response to the deep-frozen radiated allograft bones transplantation is slight, comparing with the autogenous bone transplantation, slightly transitory absorption fever is observed, occasionally some abnormities such as local lymphadenectasis, white cells in the terminal blood vessels increasing, blood sedimentation accelerating occurred in few patients but these abnormal responses would disappear soon. If the draining is put in the wound these responses can eliminate in two or three days. All these prove that even the immunological rejection exists its function is also quite slight. But when a large quantity of allograft bones are used to fill those large bone cavities or the massive bones are transplanted more obvious immunological rejection with increasing oozing or bone absorption would be observed in occasional cases, which shows that the immunological rejection can be significantly reduced but can not be thoroughly eliminated through the deep-frozen and radiation. The extent of immunological rejection was relevant with the general immunity characteristics of the patients. Besides, local bad responses are relevant with local conditions, for example, the bone end connection unfit or loosening would promote local oozing and excessive absorption of transplanted bones.

The important complication of bone transplantation is local infection which almost happens in large bulk bones or massive bones transplantation. The infection rate of bone allograft is similar to autogenous bone transplantation or other bone surgeries, the bigger the surgery size or the deeper the operation position or the worse the surgery conditions, the higher the infection occurrence. Lord (1986) and Dick (1984) once reported the infection rate of the massive bone transplantation in the earlier period is respectively 11.7% and 13.3%, which is similar to that of artificial joint replacement (8-9%). Recent years with the improvement of control methods of infection the infection rate has obviously reduced (Fitzgerald, 1991). Both prosthesis and large bulk allograft bones can make the antibacterial ability in the local tissues decrease and lead to the occurrence of deep infection at terminal phase. Skin necrosis after the operation is an important inducement cause of infections. Draining after operation contributes to the prevention of the infection. To prevent the infection the bone can be dipped in the sodium water with the antibiotics before being transplanted. Once the infection occurred the antibiotics should be used. When necessary the transplanted bone should be removed and then be transplanted again, or give amputation and other treatments. Lord (1988) reported among the collected 283 bone infection cases only one cases suffered from the polluted allograft bones, the reason was that the bones which had not been secondly sterilized was used in advance before the bacterium positive report was published. For the radiation sterilized allograft bones, whether massive or small, the infection cases from the allograft bone have never been reported, which shows that the radiation sterilized allograft bones can be safely used.

Question Three: Whether can the allograft bone transplantation probably cause disease transmission?

The most concerned disease is acquired immune deficiency syndrome (AIDS). Blood transfusion and organs transplantation can transmit HIV virus of AID-A, and bone transplantation as well. The first case ever reported was a female who received the vertebrate column fusion bone transplantation and the donor was a male whose hip joint was shaped to cut off the femoral head then transplanted into the female without removing the bone marrow and with out second sterilization. 20 months after the transplantation the lymph node swelled in the recipient axillary fossa and HIV responses was diagnosed positive, next year she died. Later the male donor was checked to be HIV positive, he confessed he had been injected with the drugs and soon he died. His wife also was HIV positive (CDC, 1988). The most attracting is that the case reported by Simonds (1992). One male with HIV negative suddenly died and his four organs including heart and both kidneys and liver were donated for organs transplantation and 54 tissues were sent to 35 hospitals for tissues transplantation. The results reported that all four organs transplantation recipients were infected with HIV and died within one to two years. Those preserved tissues were checked with PCR methods again and find HIV positive. This showed that at the first check the donor was in the early phase of infection the HIV antibody reaction didn’t showed positive, in another word, “window phase” or at this period this “false negative phase” so this tragedy was caused. Among 54 transplanted tissues there were 4 fresh freezing bones, three of which were cancellous bones without cleaning bone marrow and their corresponding patients were all infected with HIV another case without being infected with HIV was transplanted the back bones which were removed the cancellous bones without bone marrow. Other 50 tissues were transplanted to 40 patients including the freeze drying bones(38),freeze drying soft tissues(4),freeze drying radiated endocranium（6）and cornea（2）,they were HIV negative. It suggests that HIV is mainly situated in bone marrow and removing the bone marrow can contribute to eliminating the transmission of HIV. To prevent the recipients and the working staff in the bone banking from being infected with HIV, the first is to check the HIV antibody reaction of donors and learn about the personal history including the residence history whether they are in high HIV prevalence regions, whether they have the history of wrenching，homosexuality，drug abuse, the purpose is to eliminate those donors having infected or probably infected with HIV. PCR technology can be used to check the HIV antigen of the donors and prevent the HIV false negative reaction, but its cost is too expensive to be used. Both pasteurization (56℃,30min) and the radiation can make the HIV inactivate.（Spire,1985).

The patients are probably transmitted with hepatitis through blood transfusion, but this kind of cases have seldom occurred in tissues transplantation. Eastlund (1991) pointed out only one case reported by Shutkin (1954) 40 years ago. One young man suffering from the bone fracture was transplanted with the bone, 10 weeks later he was diagnosed as the hepatitis with the jaundice. Through the research the donor had not the hepatitis history but he had accepted the blood transfusion 3 years ago. The cases with hepatitis transmission is scarce because the serology check (HBsAB, HCV) is feasible and effective. Obviously removing the marrow, pasteurization and radiation can contribute to prevention the hepatitis virus from transmission.

All above suggests that only if performing strict quality control procedures, removing the bone marrow and radiation sterilization, the risk of disease transmission by the allograft bones and the danger of the local infection can be controlled effectively.

Question Four: Whether can the transplanted allograft bone survive?

The reaction after transplanting the allograft bones is the same with that of transplanting homogenous bones without blood vessels. Because once removed and cut off the blood supply, except the thin layer tissues close to bone membrane homogenous bones become dead bone like the allograft bones. Soon after the dead bones are transplanted the small blood vessels may grow into the dead bone cavity, then the dead bone surface is absorbed and gradually substituted by extending new bones. Such process is called as the creep-substitution. Obviously the dead bone absorption is earlier than new bones substitution. The transplanted allograft bones are in the transition from the new and the old and from the dead and the live for a long period. The time for all dead bones to be replaced by new bones is up to the type of transplanted bones, the position and the age of the patients (Stevenson, 1991, Enneking,1991). Obviously the time for the homogenous bone to replace is shorter than that of allograft bones because the former hasn’t the immunological rejection; the time of small pieces cancellous bones is shorter than massive cortical bones because the bone trabecula of most cancellous bones can be directly covered by new bones, not necessary to experience the absorption, while for the dense cortical bones they have to be absorbed first, then they from the new apertures, and the new bone can creep on the surface of the dense cortical bones(Glimcher,1983); the creep-substitution may complete in one year for the children while for the adults the time may be over 10 years. However if the transplanted bone tissues can fulfill the needed support and connection functions, the time seems not so important.

The aim of creep-substitution is the replacement of the new bones after the absorption of the old ones, so the absorption to different extent is inevitable. But if the old new absorption can not be replaced by the newly formed bones in time, advance absorption or excessive absorption would occur, which is the osteoporosis. The bone fracture often happens in long bone about 6 to 8 months after massive bone transplantation(Glomcher,1983), which is worth of the notice. If the absorption was in advance and the bones formation is delayed the transplanted bones would disappear, which means the bone transplantation would fail, this kind of response may happen in the homogenous bone transplantation. The following are important reasons to cause excessive absorption：the transplanted bone being more massive or much more, the bone end being badly connected, local inflammation and immunological rejection. The excessive radiation for freeze drying, for instance, over 35kGy, easily cause the absorption (Dziedzio-Goclawska,1991).

Question Five: Whether do deep frozen/freeze drying radiation allograft bones have the properties of osteoinduction and bone conduction?

The bone growth after the bone transplantation can be fulfilled through the bone induction and bone conduction. The bone induction makes the connective tissues transform into the bone tissues. Bone morphogenesis proteins (BMP), tissues grow factors TGF-β are the important bone induction factors which are contained in the normal bone matrix. The important evidences of bone induction are that the demineralized bone matrix imbedded in the muscle can transform into the bone tissues. So the demineralized bone is regarded as the bone transplantation materials possessing the ability of bone induction (Urist,1975). Most bones applied in clinics are non-demineralized bone allograft bones. The ability of the bone induction of the non-demineralized allograft bone can not lie on the result of the bone morphogenesis experiment, because itself is the bone tissues. Learning about whether the deep-frozen/radiation allograft bone possesses the ability of bone induction depends on whether these preparation processes can destroy the bone induction factors.

The relevant experiments have proved that deep-frozen and freeze drying will not affect the viability of BMP. The most attracting is radiation. In the earlier years, Urist (1974) proposed that 20-35kGy radiation can destroy the BMP. But later, many scholars disagreed this idea and they thought the common sterilization dose of 25kGy was not enough to affect the viability of BMP, only when the dose up to 50kGy had the side effect of destroying BMP (Schwarz,1988,Wientroub,1988). The destruction of BMP by chemical sterilization such as ethylene oxide is more serious than that of radiation in obtaining the same sterilization effect (Munting, 1988, Thoren, 1995). Other relevant experiments have also proved that 20kGy radiation dose is not sufficient enough to affect the viability of TGF-β(Puolakkainen,1993). We can therefore deduce that the bone induction factors contained in the freezing radiated allograft bone matrix can not be reduced, only being released slowly during the bone absorption, and the rate of releasing is not faster than the demineralized bones. To increase the ability of bone induction of allograft bone the bone induction factors may be mixed into the allograft bone for clinical use. What mentioned above shows that even if the non-demineralized allograft bones are bone inductive but little. The more important bone morphogenesis mechanism than the bone induction is bone conduction (Aspenberg, 1988, Enneking, 1991). The bone conduction means the bone tissues extend and creep from the old bone tissues to the surface of the transplanted bone to realize the bone growth. So the creep-substitution belongs to the bone conduction, which is the basic form of bone growth and healing after the allograft bone transplantation. Obviously the ability of bone conduction is also up to the characteristics of the transplanted bone. The bone conduction room particularly made proved that 25kGy radiation is not enough to affect the ability of the allograft bone conduction (Thoren,1995) and neither the ability of bone induction nor the bone conduction of the allograft bone would be affected by radiation.

Question Six: Whether does the specific antigen of HLA or blood type of the donor have to be in accordance with the recipient for the allograft bone transplantation?

Same with homogenous organs transplantation, the transplanted allograft bone was expected to be compatible with the recipient without the immunological rejection. This is up to the coincidence extent of human histocompatibility antigen (HLA) in the cell surface of the recipient with the donors. Bone transplantation is different from other organs transplantation, the former being the nonliving frame transplantation not the living function transplantation. Once the immunological rejection occurred during the organs transplantation, the tissues would die and function would disappear, which pronounces the failure of the transplantation. But the bone transplantation uses the old dead bone as the frame of the creep-substitution. The slight bone absorption in bone surface caused by the immunological rejection is inevitable, What's more, this kind of absorption is often regarded to be helpful to the new bone creep-substitution (Friedlaender,1983) .Of course it could not prove that histocomatibility has no favor for the bone transplantation. A great quantity of animal experiments testified that the transplantation effect of histological compatible allograft bone obviously outweigh the incompatible ones (Bos,1983, Stevenson,1991). But like the organs transplantation the bone transplantation could not select the donors according to the HLA antigen, also it is unnecessary. Because the retrospective analysis for the histological antigen type and the clinics of the advantageous evidence for the histocompatibility has not been found(Muscolo,1987). Those cases, who are checked with the special HLA antibody after the bone transplantation, were not found the transplantation effect worse (Friedlaender,1984). These facts are possibly due to others factors which playe more important roles in affecting the prognosis of bone transplantation than the bone immunology. The bone immunogen mainly come from the bone marrow, not bone matrix, so the immunogen of allograft bone after cleaning the marrow is very low and the deep-frozen/ freezing drying and radiation can make the immunogen further decrease.

The immunological rejection may be the main reason for the failure of massive allograft bone transplantation. Therefore, the immunosuppressive agent is expected to use after the operation as in organs transplantation. The bone transplantation experiments with the rabbits and dogs showed that the immunosuppressive agent do decrease the immunological rejection (Goldberg,1984) but the animals are susceptible with some complications such as infection, leukocyte decrease even parts dying, so that at present this treatment is distant from the clinics.

The blood type needs to be considered while having blood transfusion, but the ABO blood type is not in relation to bone immunological rejection, only the Rh blood type should be given consideration. Three cases with Rh negative women, ever reported to be transplanted with Rh positive donated bones, became Rh positive and one of them was pregnant whose new born infant suffering from the haemolyticus disease (Tomford,1993). However, the bone banks do not usually check the Rh blood type because such risks can totally be avoided by cleaning the marrow and also Rh negative person are seldom among the average population (2% in Chinese). This kind of risk can only occur in procreative women. Overall, the concordance of HLA particular antigen and blood type between the receptor and the donor is not necessarily considered when the allograft bone transplantation in clinics is given.

Question Seven: Which one is better, deep-frozen or freeze drying?

Both of these allograft bones have their own advantages and disadvantages, so choices should be made according to the different needs. Animal experiment shows that freeze drying can further decrease the immunogen of allograft skin or bone on the base of the freezing (Pellet, 1983, 1984). The big advantages of freeze drying are facile for long-term preservation and transportation. The disadvantages of freeze drying are that its mechanical characteristics are not better than the deep frozen. The ability of resisting the destruction is called as the intensity; the ability of resisting the deformation is called as the rigidity (expressing the elasticity modulus). Compared with the wet bone, the strength/compression intensity extremity and elasticity modulus of the freeze drying will increase rather than decrease and the bone become too brittle to be reshaped by the saw, however it is soon recovered after rehydration (Bright, 1983). The process after the transplantation is equal to the rehydration inside the body, so it is not necessary to rehydrate beforehand. Only thing is to soak the bones with salt water before the transplantation. Yet it should be pointed out the requirement for the bone transplantation materials is mainly the ability to resist curving, twisting and shearing rather than stretching and compressing, especially for the massive bone transplantation. Compared with wet bones, such abilities would decrease to some extent after being frozen, which may be relevant with the vertical fissures in freezing bones (Pelker, 1987, Rock, 1996). All these prove that during the gravity support and massive bone transplantation, the deep frozen bones, rather than the freezing bone, should be more needed. Materials mechanics can be divided into the plastic materials and the brittle materials according to whether the extension rate of the materials given force is being above 5% or below 5%. The cortical bone, whether dry or not, belongs to brittle materials for they would be destroyed before the larger deformation when being forced; the cancellous bones which belong to the brittle material when dry, but they belong to plastic materials when wet, which proves that the wet bones and freeze drying bones after rehydration are tend to be deformed when force is given. So the drying cancellous bones, especially after the rehydration are more used for filling and fusing than for the weight-bearing. Because of this, such bones should not be excessively compressed and filled.

A great quantity of experiments show that purely freezing, even liquid nitrogen, will not affect the mechanical characteristics (Rock, 1996). Large dose radiation can decrease the bone intensity (Pelker,1987), but only when the dose is above 35kGy or 50kGy the bone intensity may have some change (Anderson,1992, Zhang,1994). Therefore, the common radiation sterilization dose (25kGy) is safe for the deep frozen bones and freeze dried bones.