Comparison of reamer irrigator aspirator (RIA) suspension versus bone marrow aspirate concentrate (BMC) for percutaneous treatment of long bone nonunions-A preclinical canine model.

OBJECTIVE: To compare the bone healing effects of percutaneously delivered bone marrow aspirate concentrate (BMC) versus reamer irrigator aspirator (RIA) suspension in a validated preclinical canine ulnar nonunion model. We hypothesized that BMC would be superior to RIA in inducing bone formation across a nonunion site after percutaneous application. The null hypothesis was that BMC and RIA would be equivalent. METHODS: A bilateral ulnar nonunion model (n= 6; 3 matched pairs) was created. Eight weeks after segmental ulnar ostectomy, RIA from the ipsilateral femur and BMC from the proximal humerus were harvested and percutaneously administered into either the left or right ulnar defect. The same volume (3 ml) of RIA suspension and BMC were applied on each side. Eight weeks after treatment, the dogs were euthanized, and the nonunions were evaluated using radiographic, biomechanical, and histologic assessments. RESULTS: All dogs survived for the intended study duration, formed radiographic nonunions 8 weeks after segmental ulnar ostectomy, and underwent the assigned percutaneous treatment. Radiographic and macroscopic assessments of bone healing at the defect sites revealed superior bridging-callous formation in BMC-treated nonunions. Histologic analyses revealed greater amount of bony bridging and callous formation in the BMC group. Biomechanical testing of the treated nonunions did not reveal any significant differences. CONCLUSION: Bone marrow aspirate concentrate (BMC) had important advantages over Reamer Irrigator Aspirator (RIA) suspension for percutaneous augmentation of bone healing in a validated preclinical canine ulnar nonunion model based on clinically relevant radiographic and histologic measures of bone formation.

Polytherapy versus monotherapy in the treatment of tibial non-unions: a retrospective study.

BACKGROUND: Treating tibial non-unions efficiently presents a challenge for orthopaedic trauma surgeons. The established gold standard involves implanting autologous bone graft with adequate fixation, but the addition of biologicals according to the so-called diamond concept has become increasingly popular in the treatment of non-unions. Previous studies have indicated that polytherapy, which involves implanting mesenchymal stem cells, bioactive factors and osteoconductive scaffolds, can improve bone healing. This study aims to evaluate the efficacy of polytherapy compared with monotherapy in treating tibial non-unions of varying severity. MATERIALS AND METHODS: Data from consecutive tibial non-unions treated between November 2014 and July 2023 were retrospectively analysed. The Non Union Scoring System (NUSS) score before non-union surgery, and the Radiographic Union Score for Tibial fractures (RUST), scored at 1, 3, 6, 9, 12 and 18 months post-surgery, were recorded. Initially, a comparison was made between the polytherapy and monotherapy groups. Subsequently, patients receiving additional surgical non-union treatment were documented, and the frequency of these treatments was tallied for a subsequent per-treatment analysis. RESULTS: A total of 34 patients were included and divided into a polytherapy group (n = 15) and a monotherapy group (n = 19). The polytherapy group demonstrated a higher NUSS score (44 (39, 52) versus 32 (29, 43), P = 0.019, z = -2.347) and a tendency towards a higher success rate (93% versus 68%, P = 0.104) compared with the monotherapy group. For the per-treatment analysis, 44 treatments were divided into the polytherapy per-treatment group (n = 20) and the monotherapy per-treatment group (n = 24). The polytherapy per-treatment group exhibited a higher NUSS score (48 (43, 60) versus 38 (30, 50), P = 0.030, z = -2.173) and a higher success rate (95% versus 58%, P = 0.006) than the monotherapy per-treatment group. Within the monotherapy per-treatment group, the NUSS score displayed excellent predictive performance (AUC = 0.9143). Setting the threshold value at 48, the sensitivity and specificity were 100.0% and 70.0%, respectively. CONCLUSIONS: Polytherapy is more effective than monotherapy for severe tibial non-unions, offering a higher success ratio. The NUSS score supports decision-making in treating tibial non-unions. LEVEL OF EVIDENCE: Level III.

Stem Cells and Their Derivatives: An Implication for the Regeneration of Nonunion Fractures.

Despite advances in biomedical research, fracture nonunion rates have remained stable throughout the years. Long-bone fractures have a high likelihood of nonunion, but the specific biological pathways involved in this severe consequence are unknown. Fractures often heal in an organized sequence, including the production of a hematoma and an early stage of inflammation, the development of a soft callus and hard callus, and eventually the stage of bone remodeling. Deficient healing can result in a persistent bone defect with instability, discomfort, and loss of function. In the treatment of nonunions, mesenchymal stem cells (MSCs) prove to be a promising and safe alternative to the standard therapeutic strategies. Moreover, novel scaffolds are being created in order to use a synergistic biomimetic technique to rapidly generate bone tissue. MSCs respond to acellular biomimetic matrices by regenerating bone. Extracellular vesicles (EVs) derived from MSCs have recently gained interest in the field of musculoskeletal regeneration. Although many of these techniques and technologies are still in the preclinical stage and have not yet been approved for use in humans, novel approaches to accelerate bone healing via MSCs and/or MSC derivatives have the potential to reduce the physical, economic, and social burdens associated with nonhealing fractures and bone defects. In this review, we focus on providing an up-to-date summary of recent scientific studies dealing with the treatment of nonunion fractures in clinical and preclinical settings employing MSC-based therapeutic techniques.

Nonunion of conservatively treated humeral shaft fractures is not associated with anatomic location and fracture pattern.

INTRODUCTION: Humeral shaft fractures make up 1-3% of all fractures and are most often treated nonoperatively; rates of union have been suggested to be greater than 85%. It has been postulated that proximal third fractures are more susceptible to nonunion development; however, current evidence is conflicting and presented in small cohorts. It is our hypothesis that anatomic site of fracture and fracture pattern are not associated with development of nonunion. MATERIALS AND METHODS: In a retrospective cohort study, 147 consecutive patients treated nonoperatively for a humeral shaft fracture were assessed for development of nonunion during their treatment course. Their charts were reviewed for demographic and radiographic parameters such as age, sex, current tobacco use, diabetic comorbidity, fracture location, fracture pattern, AO/OTA classification, and need for intervention for nonunion. RESULTS: One hundred and forty-seven patients with 147 nonoperatively treated humeral shaft fractures were eligible for this study and included: 39 distal, 65 middle, and 43 proximal third fractures. One hundred and twenty-six patients healed their fractures by a mean 16 ± 6.4 weeks. Of the 21 patients who developed a nonunion, two were of the distal third, 10 of the middle third, and nine were of the proximal third. In a binomial logistic regression analysis, there were no differences in age, sex, tobacco use, diabetic comorbidity, fracture pattern, anatomic location, and OTA fracture classification between patients in the union and nonunion cohorts. CONCLUSIONS: Fracture pattern and anatomic location of nonoperatively treated humeral shaft fractures were not related to development of fracture nonunion.

The perceptions of clinicians using low-intensity pulsed ultrasound (LIPUS) for orthopaedic pathology: A national qualitative study.

INTRODUCTION: Low-intensity pulsed ultrasound (LIPUS) is a non-invasive treatment modality for delayed union or non-union of acute fractures. We aimed to assess the current use of LIPUS at a national level in the United Kingdom, why and how clinicians use it, what treatment protocols are followed, and what the current perceptions are on this technology. METHODOLOGY: Using a detailed online survey compromised of 20 questions delivered to known LIPUS users, we were able to collect qualitative data on indication of use, type of machine used, personal views on the technology, frequency of usage, and treatment protocols. Each question was peer-reviewed to exclude bias. RESULTS: A total of 70 respondents completed the survey. LIPUS was used by most clinicians for cases of non-union (N = 55, 78.5%) and delayed union (N = 51, 72.8%). The majority of respondents personally used a LIPUS device between 1 and 5 times in 12 months (N = 38, 54.3%). Most considered LIPUS a failure after three to six months of treatment without clinical improvement (N = 39, 55.7%). A total of 32 respondents (45.7%) mentioned the need for funding approval before accessing LIPUS technology. Poor revision surgery candidates (N = 48, 68.6%) and atrophic non-union (N = 46, 65.7%) were the most frequently cited reasons for using LIPUS technology as treatment. Most participants (N = 48, 68.6%) considered LIPUS to be cost-effective. Despite most clinicians being comfortable with the use of LIPUS, some respondents did not understand the basic science underpinning the technology nor could explain the need for LIPUS to patients comfortably. CONCLUSION: LIPUS technology may have a significant role to play in the treatment of orthopaedic fracture related pathology. Regular users perceived the technology to be cost-effective and efficacious. Further research should standardize treatment protocols and aim to establish a national LIPUS registry.

Addition of shock wave therapy to nail dynamization increases the chance of long-bone non-union healing.

BACKGROUND: Long-bone non-unions after intramedullary nailing can be treated by nail dynamization or focused high-energy extracorporal shock wave therapy (fESWT). The objective of this study was to assess the effect of the combination therapy of nail dynamization and fESWT on long-bone non-unions. MATERIALS AND METHODS: 49 patients with long-bone non-unions (femur and tibia) after nailing were treated with nail dynamization (group D, n = 15), fESWT (group S, n = 17) or nail dynamization in addition to fESWT (group DS, n = 17). Patients were followed up for 6 months retrospectively. Furthermore, age, sex, Non-Union Scoring System (NUSS) score, time intervals from primary and last surgery until intervention and smoking status were analysed for their correlations to bone union. RESULTS: Union rates were 60% for group D, 64.7% for group S and 88.2% for group DS, with a significant difference between group D and DS (p = 0.024). Successful treatment was correlated with high age (OR 1.131; 95% CI 1.009-1.268; p = 0.034), female gender (OR 0.009; 95% CI 0.000-0.89; p = 0.039), low NUSS score (OR 0.839; 95% CI 0.717-0.081; p = 0.028) and negative smoking status (OR 86.018; 95% CI 3.051-2425.038; p = 0.009). CONCLUSIONS: Data from the present study indicate that the combination therapy of nail dynamization and fESWT leads to a higher union rate than dynamization or fESWT alone. LEVEL OF EVIDENCE: Level 3.

High-energy extracorporeal shockwave therapy in humeral delayed and non-unions.

INTRODUCTION: Within the last few decades, focused high-energy extracorporeal shockwave therapy (ESWT) has proven to be an effective alternative to standard of care revision surgery in delayed healing fractures or manifest non-unions in various anatomical regions. MATERIALS AND METHODS: A retrospective multi-variant analysis of an open prospective, single-armed clinical study was conducted. Patients receiving focused high-energy ESWT for a delayed healing or an apparent non-union of a humeral fracture between January 1999 and December 2015 at a single trauma center were included in the study. Bony healing was defined as cortical continuity in three of four cortices and pain-free force loading and evaluated using CT scans and clinical examination at three- and six-month follow-ups after ESWT. RESULTS: A total of 236 patients were included. N = 93 (43.8%) showed bony consolidation three months after ESWT and n = 105 (52.5%) after six months. Sub-group analysis showed significantly better healing for the proximal metaphyseal humerus (66.7% after six months, n = 42) compared to the diaphyseal region (48.1%, n = 133) and distal metaphyseal humerus (48.1%, n = 25). Regression analysis indicated significantly increased healing rates for patients of younger ages (p = 0.001) and a fracture diastasis of less than 5 mm (p = 0.002). CONCLUSION: The findings of this study indicate that ESWT can be considered as a treatment option for a well-selected patient population despite the lower healing rates compared to other anatomical regions.

Platelet-Rich Plasma, Mesenchymal Stem Cell, and Non-metallic Suture-Based Fixation Technique in a Patellar Fracture Nonunion: A Technical Note and Systematic Review.

BACKGROUD: We performed a systematic review on the management of patellar fracture nonunion and report a novel suture-based non-metallic fixation technique associated with platelet-rich plasma and mesenchymal stem cell injections in the management of this injury. METHODS: A systematic search was performed up to August 2020 in PubMed and Scopus electronic databases of scholarly articles evaluating different surgical techniques used for nonunion of patellar fractures, with no restrictions on language or year of publication. Furthermore, we describe our novel non-metallic suture fixation technique and a patient in whom this technique was applied. RESULTS: A total of 9 articles were included in the systematic review. Tension band wiring was the most commonly used procedure (62.7%). Nonoperative procedures (8.1%) resulted in nonunion in all patients. The most common complication after open reduction and internal fixation was infection (7.8%). Our patient at the latest follow-up reported full functional recovery and full extension and flexion of the affected knee with no pain and subjectively normal strength. CONCLUSIONS: The management of patella nonunions is still a challenge. The technique reported here can be used in patellar fracture nonunion, as well as in primary patellar fractures.

Minimally invasive treatment of long bone non-unions with bone marrow concentrate, demineralized bone matrix and platelet-rich fibrin in 38 patients.

To determine the efficacy of percutaneous injection of autologous bone marrow concentrated (BMC), demineralized bone matrix (DBM), and platelet rich fibrin (PRF) in the treatment of long bone non-unions. From January 2011 to January 2018 patients with non-union of the lower limbs who were on the waiting list for open grafting with established tibial or femoral non-union and minimal deformity were eligible to participate in this study. Patients were treated with a single percutaneous injection of DBM, BMC and PRF. Our study group comprised 38 patients (26 males and 12 females; mean age 39, range 18 to 65). Non-unions were located in the femur (18 cases) and in the tibia (20 cases). Clinical and imaging follow-up ranged from 4 to 60 months (mean 20 months). Bone union occurred in 30 out of 38 patients (79%) in an average of 7 months (range 3 to 12) and all healed patients had full weight bearing after 9 months on average (range 6 to 12) from injection. In 19 cases the osteosynthesis was removed 12 months on average (range 3 to 36) from surgery. One patient developed infection at the non-union site after treatment. Percutaneous injection of DBM, BMC, and PRF is an effective treatment for long-bone non-unions. This technique allows the bone to heal with a minimally invasive approach and with a hospitalization of 2 days. Key elements of bone regeneration consist of a combination of biological and biomechanical therapeutic approach.

Adolescent mid-shaft clavicular fracture displacement does not predict nonunion or inferior functional outcome at long-term follow-up.

AIMS: The aim of this study was to define the complications and long-term outcome following adolescent mid-shaft clavicular fracture. METHODS: We retrospectively reviewed a consecutive series of 677 adolescent fractures in 671 patients presenting to our region (age 13 to 17 years) over a ten-year period (2009 to 2019). Long-term patient-reported outcomes (abbreviated version of the Disabilities of the Arm, Shoulder and Hand (QuickDASH) score and EuroQol five-dimension three-level (EQ-5D-3L) quality of life score) were undertaken at a mean of 6.4 years (1.2 to 11.3) following injury in severely displaced mid-shaft fractures (Edinburgh 2B) and angulated mid-shaft fractures (Edinburgh 2A2) at a minimum of one year post-injury. The median patient age was 14.8 years (interquartile range (IQR) 14.0 to 15.7) and 89% were male (n = 594/671). RESULTS: The majority of fractures were mid-shaft (n = 606) with angulation (Edinburgh 2A2, n = 241/606, 39.8%) or displacement (Edinburgh 2B1/2, n = 263/606, 43.4%). Only 7% of the displaced mid-shaft fractures underwent acute fixation (n = 18/263). The incidence of refracture over ten years following nonoperative management of mid-shaft fractures was 3.2% (n = 19/588) and all united without surgery. Fracture type, severity of angulation, or displacement were not associated with refracture. One nonunion occurred following nonoperative management in a displaced mid-shaft fracture (0.4%, n = 1/245). Of the angulated fractures, 61 had angulation > 30°, of which 68.9% (n = 42/61) completed outcome scores with a median QuickDASH of 0.0 (IQR 0.0 to 0.6), EQ-5D-3L 1.0 (1.0 to 1.0), and 98% satisfaction with shoulder function. For the displaced fractures, 127 had displacement beyond one cortical width of bone for which completed outcome scores were provided in 72.4% (n = 92/127). Of these 15 had undergone acute fixation. Following nonoperative treatment, the median QuickDASH was 0.0 (IQR 0.0 to 2.3), EQ-5D-3L 1.0 (1.0 to 1.0), and satisfaction with shoulder function was 95%. There were no significant differences in the patients' demography or functional outcomes between operative and nonoperative treatments. CONCLUSION: Nonoperative management of adolescent mid-shaft clavicle fractures results in excellent functional outcomes at long-term follow-up. Nonunion is exceptionally rare following nonoperative management and the relative indications for surgical intervention in adults do not appear to be applicable to adolescents. Cite this article: Bone Joint J 2021;103-B(5):951-957.

Neither operative nor nonoperative approach is superior for treating displaced midshaft clavicle fractures: a partially blinded randomized controlled clinical trial.

AIMS: To compare the functionality of adults with displaced mid-shaft clavicular fractures treated either operatively or nonoperatively and to compare the relative risk of nonunion and reoperation between the two groups. METHODS: Based on specific eligibility criteria, 120 adults (median age 37.5 years (interquartile range (18 to 61)) and 84% males (n = 101)) diagnosed with an acute displaced mid-shaft fracture were recruited, and randomized to either the operative (n = 60) or nonoperative (n = 60) treatment group. This randomized controlled, partially blinded trial followed patients for 12 months following initial treatment. Functionality was assessed by the Constant score (CS) (assessor blinded to treatment) and Disability of the Arm, Shoulder and Hand (DASH) score. Clinical and radiological evaluation, and review of patient files for complications and reoperations, were added as secondary outcomes. RESULTS: At 12 months, 87.5% of patients (n = 105) were available for analysis. The two groups were well balanced based on demographic and fracture-related characteristics. At six weeks of follow-up a significant difference in DASH score (p < 0.001) was found in favour of operative treatment. The functionality at 12 months of follow-up based on CS and DASH was excellent in both groups (CS > 90 points and DASH < 10 points) with no significant difference (p = 0.277 for DASH and p = 0.184 for CS) between the two groups. The risk of symptomatic nonunion was significantly higher in the nonoperative group (p = 0.014), with a relative risk of 9.47 (95% confidence interval (CI) 1.26 to 71.53) in this group compared to the operative group. The number-needed-to-treat to avoid one symptomatic nonunion was 6.2. Initial treatment and age were factors significantly associated with nonunion in a logistic analysis. There were 26% in both groups (n = 14 in operative group and n = 15 in nonoperative group) who required secondary surgery, with most indications in the nonoperative group mandatory due to nonunion compared to most relative indications in the operative group requiring intervention due to implant irritation. CONCLUSION: Superiority was not identified with either an all-operative or all-nonoperative approach. The functionality at short term (within six weeks) seems igreater following operative treatment but was not found at one year. The risk of nonunion is significantly higher with nonoperative treatment. However, an all-operative approach to lower the nonunion risk may result in unnecessary surgery and is not recommended. Cite this article: Bone Joint J 2021;103-B(4):762-768.

Percutaneous injection of platelet-rich plasma to treat atrophic nonunion after internal fixation of ulnar fracture: a case report.

Non-union is a serious postoperative complication of fracture. Early detection and intervention can avoid revision surgery. Platelet-rich plasma releases many active tissue factors and has potential to promote fracture healing. Percutaneous injection of platelet-rich plasma at the fracture site may avoid surgical treatment when non-union occurs. We present a case of atrophic non-union of an ulna fracture treated conservatively with percutaneous injection of platelet-rich plasma.

Fracture Nonunion Treated with Low-Intensity Pulsed Ultrasound and Monitored with Ultrasonography: A Feasibility Study.

The positive effect of low-intensity pulsed ultrasound (LIPUS) on bone fracture healing has been proved. However, during the period of LIPUS therapy, it is undetermined whether LIPUS promotes the formation of heterotopic ossification (HO), which usually occurs in muscle tissues after trauma such as bone fracture and spinal cord injury. Here, we used 6-week LIPUS therapy in a 42-year-old Chinese male patient with a fracture nonunion in combination with ultrasonography for monitoring fracture healing and HO formation. After the LIPUS therapy, the mineralized bone formation in the area of defect of the distal tibia was presented in an ultrasound image, which was consistent with the outcome of plain radiography showing callus formation and the blurred fracture line in the area exposed to LIPUS. In addition, ultrasound images revealed no evidence of HO development within soft tissues during the period of LIPUS therapy. This study suggests that ultrasonography is a potential tool to guarantee the performance of LIPUS therapy with monitoring HO formation. Easy to use, the integration of the handheld ultrasound scanner and the ultrasonic therapeutic apparatus is entirely dedicated to help orthopedists make high-quality care and diagnosis.

Repurposing denosumab for recalcitrant bone healing.

Fracture healing has four phases: haematoma formation, soft callus, hard callus and remodelling. Often, non-healing fractures have an arrest of one of these phases, which need resurgery. We have repurposed denosumab for impaired fracture healing cases to avoid surgical intervention. Here, we report a series of three cases of impaired fracture healing where denosumab was given 120 mg subcutaneous dosages for 3 months to enhance healing. All the three cases have shown complete bone union at a mean follow-up of 6.7 months (5-9 months) as assessed clinically and radiologically, and have observed no adverse effect of the therapy. Denosumab given in this dose aids fracture healing by increasing callus volume, density and bridges the fracture gap in recalcitrant fracture healing cases where the callus fails to consolidate.

The Use of Platelet-Rich Plasma (PRP) for the Management of Non-union Fractures.

PURPOSE OF REVIEW: The treatment of non-union fractures represents a significant challenge for orthopaedic surgeons. In recent years, biologic agents have been investigated and utilised to support and improve bone healing. Among these agents, platelet-rich plasma (PRP) is an emerging strategy that is gaining popularity. The aim of this review is to evaluate the current literature regarding the application and clinical effectiveness of PRP injections, specifically for the treatment of non-union fractures. RECENT FINDINGS: The majority of published studies reported that PRP accelerated fracture healing; however, this evidence was predominantly level IV. The lack of randomised, clinical trials (level I-II evidence) is currently hampering the successful clinical translation of PRP as a therapy for non-union fractures. This is despite the positive reports regarding its potential to heal non-union fractures, when used in isolation or in combination with other forms of treatment. Future recommendations to facilitate clinical translation and acceptance of PRP as a therapy include the need to investigate the effects of administering higher volumes of PRP (i.e. 5-20 mL) along with the requirement for more prolonged (> 11 months) randomised clinical trials.

Comparison of bone morphogenetic protein and autologous grafting in the treatment of limb long bone nonunion: a systematic review and meta-analysis.

BACKGROUND: Bone morphogenetic proteins (BMPs) have strong bone induction properties and can promote healing of fractures and other defects. However, BMP treatment efficacy for long bone nonunion remains controversial. The aim of this meta-analysis was to synthetically evaluate the advantages and disadvantages of BMP plus bone grafting (observation group) versus autologous bone grafting (control group) for limb long bone nonunion. METHODS: PubMed, Embase, Web of Science, Cochrane Library, OVID, CNKI, Weipu Journal, Chinese Biomedical Literature, and WanFang were searched for randomized and non-randomized controlled trials published before November 2019. A meta-analysis of outcome indicators was performed using RevMan 5.3 and Stata 12.0. RESULTS: Five randomized and four non-randomized controlled trials involving 30-124 cases were included, with a total of 655 nonunion cases. There were no significant group differences in postoperative healing rate, infection, and secondary operation rates (P > 0.05), but the study group demonstrated significantly shorter mean healing time (WMD = - 1.27, 95%CI - 1.67 to - 0.88, P < 0.00001), a greater frequency of excellent/good post-treatment limb function (RR = 1.18, 95%CI 1.01-1.39, P = 0.04), and lower intraoperative blood loss (P < 0.05). Alternatively, the hospitalization cost was significantly higher in the study group (P < 0.01). CONCLUSIONS: Bone morphogenetic protein is a viable alternative to autologous bone grafting, with potential advantages of accelerated fracture healing and improved postoperative function.

Bioinformatic analysis and experimental identification of blood biomarkers for chronic nonunion.

BACKGROUND: Incomplete fracture healing may lead to chronic nonunion; thus, determining fracture healing is the primary issue in the clinical treatment. However, there are no validated early diagnostic biomarkers for assessing chronic nonunion. In this study, bioinformatics analysis combined with an experimental verification strategy was used to identify blood biomarkers for chronic nonunion. METHODS: First, differentially expressed genes in chronic nonunion were identified by microarray data analysis. Second, Dipsaci Radix (DR), a traditional Chinese medicine for fracture treatment, was used to screen the drug target genes. Third, the drug-disease network was determined, and biomarker genes were obtained. Finally, the potential blood biomarkers were verified by ELISA and qPCR methods. RESULTS: Fifty-five patients with open long bone fractures (39 healed and 16 nonunion) were enrolled in this study, and urgent surgical debridement and the severity of soft tissue injury had a significant effect on the prognosis of fracture. After the systems pharmacology analysis, six genes, including QPCT, CA1, LDHB, MMP9, UGCG, and HCAR2, were chosen for experimental validation. We found that all six genes in peripheral blood mononuclear cells (PBMCs) and serum were differentially expressed after injury, and five genes (QPCT, CA1, MMP9, UGCG, and HCAR2) were significantly lower in nonunion patients. Further, CA1, MMP9, and QPCT were markedly increased after DR treatment. CONCLUSION: CA1, MMP9, and QPCT are biomarkers of nonunion patients and DR treatment targets. However, HCAR2 and UGCG are biomarkers of nonunion patients but not DR treatment targets. Therefore, our findings may provide valuable information for nonunion diagnosis and DR treatment. TRIAL REGISTRATION: ISRCTN, ISRCTN13271153. Registered 05 April 2020-Retrospectively registered.

Early efficacy evaluation of mesenchymal stromal cells (MSC) combined to biomaterials to treat long bone non-unions.

BACKGROUND AND STUDY AIM: Advanced therapy medicinal products (ATMP) frequently lack of clinical data on efficacy to substantiate a future clinical use. This study aims to evaluate the efficacy to heal long bone delayed unions and non-unions, as secondary objective of the EudraCT 2011-005441-13 clinical trial, through clinical and radiological bone consolidation at 3, 6 and 12 months of follow-up, with subgroup analysis of affected bone, gender, tobacco use, and time since the original fracture. PATIENTS AND METHODS: Twenty-eight patients were recruited and surgically treated with autologous bone marrow derived mesenchymal stromal cells expanded under Good Manufacturing Practices, combined to bioceramics in the surgical room before implantation. Mean age was 39 ± 13 years, 57% were males, and mean Body Mass Index 27 ± 7. Thirteen (46%) were active smokers. There were 11 femoral, 4 humeral, and 13 tibial non-unions. Initial fracture occurred at a mean ± SD of 27.9 ± 31.2 months before recruitment. Efficacy results were expressed by clinical consolidation (no or mild pain if values under 30 in VAS scale), and by radiological consolidation with a REBORNE score over 11/16 points (value of or above 0.6875). Means were statistically compared and mixed models for repeated measurements estimated the mean and confidence intervals (95%) of the REBORNE Bone Healing scale. Clinical and radiological consolidation were analyzed in the subgroups with Spearman correlation tests (adjusted by Bonferroni). RESULTS: Clinical consolidation was earlier confirmed, while radiological consolidation at 3 months was 25.0% (7/28 cases), at 6 months 67.8% (19/28 cases), and at 12 months, 92.8% (26/28 cases including the drop-out extrapolation of two failures). Bone biopsies confirmed bone formation surrounding the bioceramic granules. All locations showed similar consolidation, although this was delayed in tibial non-unions. No significant gender difference was found in 12-month consolidation (95% confidence). Higher consolidation scale values were seen in non-smoking patients at 6 (p = 0.012, t-test) and 12 months (p = 0.011, t-test). Longer time elapsed after the initial fracture did not preclude the occurrence of consolidation. CONCLUSION: Bone consolidation was efficaciously obtained with the studied expanded hBM-MSCs combined to biomaterials, by clinical and radiological evaluation, and confirmed by bone biopsies, with lower consolidation scores in smokers.

Can injection of adipose stem cells to non-union zone increase bone union? Experimental rat study.

OBJECTIVES: This study aims to evaluate the effects of local adipose stem cell injection on non-union and diabetic non-union of rat femurs. MATERIALS AND METHODS: Forty-eight female Wistar albino rats (weighing mean 200 g and aged 8 weeks) were used in this study. The rats were divided into six groups. Group 1 was chosen as a reference for receptor activator of nuclear factor-kappa (κ) B (RANK), receptor activator of nuclear factor-κ B ligand (RANKL) and osteoprotegerin (OPG) genes and no femur osteotomy was performed in this group. Group 2 underwent femur osteotomy, the osteotomy was fixed with a 1.5 mm K-wire as retrograde from the knee joint, and no gap was left in the osteotomy line. In order to induce non-union, femurs underwent osteotomy fixed with K-wires in groups 3, 4, 5 and 6. In addition, the osteotomy line was measured as 1.8 mm gap with electronic calipers and the gap was fixed with U staple. Before osteotomy, streptozocin was injected intraperitoneally at a dose of 60 mg/kg in 0.1 mol/L citrate buffer solution (Ph 4.4) in groups 4 and 6, in order to induce diabetes mellitus. Left femur anteroposterior and lateral X-rays were taken 10 weeks after the operation and the union in group 2 and non-union in groups 3, 4, 5, and 6 were confirmed. To see if injection of adipose stem cells into the non-union site increases bone union, 2 mL 0.9% sodium chloride (NaCl) in groups 3 and 4 and 2×106 adipose stem cell in groups 5 and 6 were locally injected into the non-union area with fluoroscopy. Femur X-rays were taken eight weeks after the injection and all rats were sacrificed. Femurs of rats were removed for histopathological and gene expression evaluation. RESULTS: There were significant differences between the groups injected 0.9% NaCI and adipose stem cells in terms of bone healing according to radiological and histopathological evaluations (p<0.05). No statistically significant difference was observed between the groups in terms of gene expression levels. CONCLUSION: According to the results of our study, local adipose stem cell injection has positive radiological and histopathological effects in diabetic and non-diabetic femoral non-unions, independently of RANK, RANKL, or OPG gene expression pathways.