TABLE 4.
Study characteristics of non‐union tissue and relevant tissue
| Author | Duration of non‐union (months) | Classification | Definition of non‐union | Isolation of tissue | Cells/material isolation |
|---|---|---|---|---|---|
| Cuthbert*, 8 | Not mentioned | Atrophic | Not mentioned | Non‐union: Fibrotic tissue lying directly between the fractured bone fragments was excised and collected; induced periosteum from centre of bone defect area; and bone marrow | Colony forming unit fibroblast (CFU‐F) assay; trilineage differentiation; histological analysis of vessel number, size and area; immunohistochemistry (CD45, SDF1, VEGF, BMP‐2); flow cytometry; qPCR; matrigel‐based angiotube formation assay |
| Wei*, 9 | Not mentioned | Atrophic | Not mentioned | Tissue samples were collected intra‐operatively from (i) non‐union tissues of atrophic bone; and (ii) healing callus around internal fixation plates in normal controls. Collected tissues were cut into “small” pieces | RNA isolation, miRNA microarray, bioinformatics of target genes, qPCR, Western blot, luciferase reporter assay |
| Burska**, 18 | Not mentioned | Not mentioned | Failure of the fracture to progress to healing radiographically with the presence of bridging callous on at least 3 cortices by a period of 9 months | Peripheral blood | ELISA |
| El‐Jawhari**, 19 | Not mentioned | Atrophic | Absence of radiological features of fracture healing (lack of callus formation in at least 3 cortices) either on plane radiographs or computed tomography scans after 9 months from fracture fixation and with ongoing pain at the NU site during ambulation | BMA; peripheral venous blood | FACS cell sorting; flow cytometry surface cytokine receptor measurement; flow cytometry—immunosuppression assay: levels of IDO, PGE2 and TGF‐β transcripts; osteogenic differentiation; RNA extraction; RT‐qPCR; proliferation (XTT colorimetric assay); ELISA |
| Ouyang**, 20 | Not mentioned | Not mentioned | Not mentioned | BMA | circRNA microarray, RNA FISH, Osteogenic differentiation assay (ALP and Alizarin red staining), cck‐8 assay, RNA pull‐down assay, double luciferase reporter assay, qPCR, RNA immunoprecipitation, Western Blot |
| McCoy**, 21 | Not mentioned | Not mentioned | Not mentioned | Peripheral blood | DNA was extracted from blood samples |
| Zhang**, 22 | Not mentioned | Not mentioned | Not mentioned | Peripheral blood | DNA was extracted from blood samples |
| Wang*, 10 | Not mentioned | Not mentioned | Not mentioned | Not applicable | Cell viability; mineralization assay; gene expression |
| Vallim*, 11 | 34 months (range 9–120 months) | Not mentioned | Lack of bone healing after 9 months of the fracture | Fibrous tissue interposed between the bone ends was excised, along with adjacent osseous fragments | Histology; population doubling; cell senescence; flow cytometry; osteogenic / adipogenic differentiation |
| Huang**, 23 | >9 months | Not mentioned | The cessation of all healing processes and failure to achieve union within 9 months without radiographic signs of progression of the fracture callus | Peripheral blood | DNA was extracted from blood samples |
| Granchi**, 24 | >3 months | Not mentioned | Not mentioned | BMA, peripheral blood | Immunoenzymatic assays |
| Takahara*, 12 | 14.8 months (range 4–26 months) | Pseudoarthrosis | (1) gross motion at the fracture site on physical examination; (2) bridging bone on 0 of 4 cortices on anteroposterior and lateral radiographs; (3) CT showing no purpose‐ ful cross‐sectional area of healing; and (4) evidence showing the existence of pseudocapsule and fluid collection between the fracture gap at the surgery | A small amount of pseudoarthrosis tissue (pseudocapsule) was obtained during the surgical treatment |
Alizarin Red S staining, ALP activity assay, and RT‐PCR after osteogenic induction. Chondrogenic differentiation capacity was assessed via Safranin O staining and RT‐PCR after chondrogenic induction. Histological analysis and cell cultures |
| Schira*, 13 | 18.3 months (range, 3–100 months) | Not mentioned | Non‐unified fractures >3 months with a resorption zone wider than 1 mm (as determined by a mandatory CT scan) with no apparent potential to heal without surgical intervention | Non‐union tissue (excluding the cortex) and cancellous bone from the ipsilateral radius has been obtained at the time of operative repair | Histology, immunohistochemistry, gene expression |
| Han*, 14 | 11 months (range, 6–30 months) | Not mentioned | Failure of the fracture to heal 6 months or more after surgery or non‐surgical treatment | Fracture and scar tissue during surgery, which was divided into bone stump tissue, marrow cavity contents, and sticking bone scars according to the sites | Histology, immunohistochemistry, gene expression |
| Wang*, 15 |
Hypertrophic non‐unions: 19.88 ± 17.88 months. Atrophic non‐unions: 14.20 ± 7.42 months |
Not mentioned | Failure of the fracture to heal 9 months or more after the injury | Intra‐operative biopsy samples | Immunohistochemistry |
| Schwabe*, 16 | Not mentioned | Not mentioned | Time span from the initial operation until the revision surgery of a least 6 months | Intra‐operative biopsy samples for the treatment of the non‐union or removal of metalwork for the control (normal healing) | Histology, immunohistochemistry, ELISA |
| Sathyendra**, 25 | Not applicable | Not applicable | Minimal callus formation 6 months after injury requiring additional surgery to achieve union | Buccal mucosal cell harvesting | SNP genotype |
| Ismail*, 17 |
37.2 ± 24.0 (range, 12–72) |
Not mentioned | Not mentioned | Intra‐operative BM from the site adjacent to the non‐union, compared to BM from iliac crest. | Not mentioned |
| Marchelli**, 26 | Atrophic non‐union: 6 to 11 months; healed: 8.5 ± 3.5 months; healing: 0.5 ± 0.5 months | Not mentioned | Not mentioned | Blood samples | ELISAs |
| Zeckey**, 27 | >9 months | Aseptic tibial and femoral shaft non‐unions | Clinically and radiologically confirmed unhealed shaft fractures >9 months following the injury and osteosynthesis treatment | Peripheral venous blood sample | DNA was extracted from blood samples |
| Dimitriou**, 28 | Required further intervention to achieve union | Atrophic | Cessation of all healing processes and failure to achieve union after the expected period of time, as seen clinically and radiologically | Peripheral venous blood sample | DNA was extracted from blood samples |
| Xiong**, 29 | Not mentioned | Not mentioned | Fracture that does not heal 6 months after injury | Normal and non‐union callous bone samples examined | Gene expression |
| Seebach**, 30 | Not mentioned | Atrophic | Not mentioned | BM cells were obtained from the iliac crest aspirate | CFU‐F; flow cytometry; osteogenic differentiation |
| Henle**, 31 | >4 months | Atrophic | No bony consolidation of the fracture in conventional X‐ray films and the patient continued to report exercise induced pain 4 months after trauma + no bone healing on CT scan | Venous blood | Immunosorbent assays |
Abbreviations: BM, bone marrow; BMA, bone marrow aspirate.
*
Non‐union tissue.
**
Relevant tissue.