Purpose of the study: Nonunion, which is a biological failure, requires revision, usually an aggressive operation. Haematopoietic bone marrow contains colony forming unit fibroblasts (CFU-F) which could favour bone healing. The purpose of this work was to determine whether a minimally invasive procedure, injection of CFU-F into the nonunion space, could favour bone healing without further procedure.

Material and methods: Our series included 43 patients: 36 male and 7 female, mean age 39.9 years. Forty-seven atrophic and aseptic nonunions of long bones were treated with percutaneous injection of concentrated autologous bone marrow: 27 tibias, 17 femurs, 3 humeri. Bone marrow was harvested from the posterior iliac crests (346 ml) then centrifuged to keep the leuko-platelet fraction (78 ml). This concentrate was injection into the nonunion space under radioscopic guidance. Efficacy was assessed on the basis of clinical criteria (complete pain-free weight-bearing, absence of contention, absence of mobility) and on radiographic criteria (healing of 3/4 corticals).

Results: Thirty nonunions healed: 19 tibias (70%, 11 femurs (65%) and 0 humerus. Mean time to healing was 5.9 months (2.4–15.6). Factors of poor prognosis were: smoking, alcohol, diabetes, corticosteroids, radiotherapy, history of sepsis (p=0.01). Early grafting increased the chances of success (p=0.04). Age, initial skin opening, type of fixation did not have a significant impact on healing. The number of CFU-F had an effect on the rate of healing.

Discussion: This technique is effective for the treatment of nonunion of the lower limb, allowing bone healing in two thirds of the cases with a minimally aggressive procedure. The method is easy to perform but requires a rigorous technique for the different phases of puncture, concentration and reinjection. Nonunions unresponsive to conventional methods, and thus corresponding to multifactorial problems, probably constitute the limitation of this method. Cell expansion or differentiation techniques could be helpful in improving the success rate but at the present time the osteogenic potential of these cells remains to be elucidated as a function of their stage of maturation.

Conclusion: Percutaneous grafts of concentrated autologous bone marrow can be a useful contribution to the therapeutic armamentarium for nonunion. Morbidity is low and the method does not compromise future options. It can be proposed as a first-intention solution for the treatment of long bone nonunion.

Purpose of the study: Monolayer cultures of chondrocytes multiply and rapidly lose their chondrocyte phenotype, limiting their potential for tissue engineering. Mesenchymatous stem cells can preserve their phenotypic characteristics after several monolayer passages, offering a promising alternative for cartilage repair. The purpose of this work was to study the influence of transforming growth factor beta-1 (TGF-beta1) and bone morphogenic protein-2 (BMP2) and/or culture supplements (hyaluronic acid) on matrix synthesis and chondrocyte differentiation of human mesenchymatous stem cells (MSC) cultured on collagen sponges.

Material and methods: MSC were isolated from bone marrow harvested during hip arthroplsty. At the third passage in monolayer culture, the MSC were reseeded on collagen sponges and cultured in vitro for 28 days under seven differ conditions: insulin transferrin selenium (ITS), foetal calf serum (FCS), ITS+TGFbeta1, ITS+ hyaluronate, ITS+TGFbeta1+hyaluronate, ITS+TGFbeta1+BMP2, ITS +TGFbeta1+BMP2+hyaluronate. The phenotypic evolution was followed using the expression of different genes of interest with PCRq (collagen2, collagen1, collagen3, collagen10, agrecanne, versicanne, COMP, Sox9). Synthesis of matrix material was assessed histologically and immunohistochemically.

Results: Used alone, hyaluronic acid did not trigger chondrocyte differentiation of MSC. For the additives FCS, ITS, or hyaluronate, the synthesis of matrix material in the sponge was weak and poor in major constituents of cartilage. Conversely, the other conditions in presence of TGFbeta1±BMP2 induced important expression of collagen2, agrecanne and COMP as well as increased matrix synthesis with a strong content in proteoglycans and collagen.

Discussion: The usefulness of MSC is growing due to their pluripotent characteristics. The conditions leading to their differentiation into the chondrocyte phenotype remains a subject of discussion. Our results show the particular importance of TGFbeta1 in the process of differentiation.

Conclusion: Chondrogenic differentiation of MSC cultured in collagen sponges as well as the synthesis of the cartilaginous matrix requires the presence of TGFbeta1 in the culture medium and to a lesser extent BMP2. These results suggest the perspective of using MSC for guided cell therapy targeting cartilage.

Purpose: Local factors such as poor vascular supply, open fracture, or infection can affect the potential for bone formation after fracture, arthrodesis or distraction. The fundamental principal for the treatment of late healing or nonunion is to supplement the local supply of the elements necessary for bone maturation. Centrifuged bone marrow is known to have a osteogenic effect in the treatment of femoral head necrosis or as a complement to conventional grafts. We examined the effect of bone marrow grafts used with conventional grafts.

Material and methods: This retrospective analysis included 14 cases where centrifuged bone marrow graft was used as complementary treatment for post-traumatic nonunion (10 cases), distraction callus (three cases) or late healing after arthrodesis (one case). Bone marrow (300 ml) was harvested from the posterior iliac crest then centrifuged to isolate the maximum number of nucleated cells and stem cells. The centrifugate (60–80 ml) was injected into the fracture site with a trocar during the same operative time. Cell concentrations (total nucleated cells, stem cells (CFU-GM), fibroblastic colonies) were noted. Patients were followed at regular visits. Bone healing was considered to be acquired when weight-bearing was possible without fixation or immobilisation.

Results: Definitive bone healing was achieved rapidly in two cases. Two patients required a conventional graft of a nonunion to achieve consolidation. For six patients, consolidation could not be achieved (three nonunions and three distraction calluses). Final outcome was good or very good in 57% of the cases. Mean delay to bone healing was 6.5 months. The infectious context had no effect on the method. The mean number of nucleated cells injected was 3.9•109 cells in successful cases and 2.8•109 cells in unsuccessful cases. These concentrations affected outcome.

Discussion: This technique for stimulating bone maturation by supplying bone generating cells is indicated for late healing or recent nonunion. It is less effective for distraction calluses or for very old nonunions. Morbidity and iatrogenic effects are minimal. A rigorous harvesting method is required since the result is highly dependent on the cell concentrations and the number of injected cells. Bone marrow injections after centrifugation should be greater than 85 ml and have a cell concentration around 45•106 cells/ml. The method is less successful for old injuries and in patients with arteritis.

Conclusion: Bone marrow grafts are indicated for the treatment of late healing or recent nonunion. Morbidity is low but a rigorous harvesting method is required. The method should be implemented shortly after the fracture without waiting for potential signs of nonunion.