The purpose of our study was to identify possible risk factors of patients with GCT of the long bones after curettage and packing the bone cavity with bone cement or bone allografts.

We retrospectively reviewed the records of 249 patients with GCT of the limbs treated at Musculoskeletal Oncology Department of our institution between 1990 and 2013, confirmed histologically and recorded in the Bone Tumor Registry. We reviewed 219 cases located in the lower limb and 30 of the upper limb. This series includes 135 females and 114 males, with mean age 32 years (ranging 5 to 80 yrs). According to Campanacci's grading system, 190 cases were stage 2, 48 cases stage 3, and 11 cases stage 1. Treatment was curettage (intralesional surgery). Local adjuvants, such as phenol and cement, were used in 185 cases; whereas in the remaining 64 cases the residual cavity was filled with allografts or autografts only.

Oncological outcome shows 203 patients alive and continuously disease-free (CDF), 41 patients NED1 after treatment of local recurrence (LR), 2 patients NED1 after treatment of lung metastases, 2 AWD with lung metastases. One patient died of unrelated causes (DOD).

LR rate was 15.3% (38 pts). Lung metastases rate was 1.6% (4 pts). In patients treated by curettage and cement (185 cases) LR was 12% (22 pts). Conversely, in patients treated curettage and bone allografts it was higher (16/64 cases), with an incidence of 25% of cases (p=0.004). Oncological complications seemed to be related with site, more frequently occurring in the proximal femur (p=0.037). LR occurred only in stage 2 or 3 tumors without statistical significance (p>0.05). The mean interval between the first surgical treatment and LR was 22 months (range: 3–89 mos). However, in the multivariate analysis no significant statistical effect on local recurrence rate could be identified for gender, patient's age, Campanacci's grading, or cement vs allografts. The only independent risk factor related to the local recurrence was the site, with a statistical significance higher risk for patients with GCT of the proximal femur (p= 0.008).

Our observation on the correlation of tumor location and risk of local recurrence is new. Therefore, special attention must be given to GCTs in the proximal femur. In fact, primary benign bone tumors in the proximal femur are difficult to treat due to the risk of secondary osteonecrosis of the femoral head or pathologic fracture.

Numerous methods of reconstructions have been reported. Among these, total hip arthroplasty (THA) or bipolar hip arthroplasty (BHA) should be avoided when possible as more cases are observed in young patients.

Therefore, we do not suggest different approach for the proximal femur. GCT in the proximal femur is much more difficult to treat than in other sites, but if curettage is feasible, the best way is to save the joint with a higher risk of local recurrence, knowing that the sacrifice of the hip articulation in case of recurrence is always possible with THA or BHA.

Osteosarcoma (OS) is a highly malignant primary tumor frequently occurring in children and adolescents. The mainstay of therapy is neoadjuvant chemotherapy and surgical removal of the lesion yielding a 50–70% of 5-year survival rate. Unfortunately, chemotherapy is currently unable to induce complete tumor necrosis leaving residual tumor cells free to metastasize or recidivate, thus resulting in a 30% mortality. The major limitation in those patients is the development of multidrug resistance (MDR) and the low water solubility of drugs such as Paclitaxel (PTX) that is in fact not included in the majority of chemotherapy protocols for OS treatment.

We thus hypothesized to prevent the emergence of MDR and obtain significant tumor reduction, by engineering innovative nanoparticles (NPs) able to vehiculate the PTX and induce a dual synergic action: the cytostatic effect of PTX and the cytotoxicity generated by reactive oxygen species produced from light triggered photoactivation (PDT) of Chlorin e6 photosensitizer. To further improve the efficacy and reduce the side effects of NPs systemic administration, Mesenchymal Stromal Cells (MSC) are used as a “Trojan horse” to deliver the NPs directly to tumor cells, taking advantage of MSC ability to selectively recognize and efficiently engraft in OS tumor stroma.

HSA were conjugated with photosensitizer Ce6 and the functionalized protein was used to produce PTX loaded nanoparticles through desolvation technique and drug-induced protein self-assembly (PTX-Ce6@HSA NP).

Human MSC lines, isolated from the Bone marrow (BM) of different donors, were then loaded with different dosages of nanoparticles and their ability to internalize and transport the NPs, migrate and induce cytotoxic ROS upon light treatment were tested in in vitro cultures.

Preliminary results showed that MSC efficiently internalize PTX-Ce6@HSA NPs and the photosensitizer Ce6 remains active inside the cells for at least 3 days after loading.

Electron microscopy performed onto loaded MSC showed that NPs internalization take places via clathrin mediated transport, whereas HPLC analysis demonstrated a release kinetics of PTX mediated by exocytosis. Finally, PTX-Ce6@HSA NPs loaded MSC co-cultured with the OS tumor cell line SaOS-2 showed a significant tumor cell growth reduction.

So fare, advances in drug delivery have failed to produce specific tool to improve the overall survival of OS patients. However, given our preliminary in vitro data we believe that the proposed multimodal therapy will minimize the side effects of the systemic chemotherapy and enhance the efficacy through the synergic effect of PTX and PDT.

In the future, our strategy could be intended as an innovative co-adjuvant approach for OS treatment to be performed right before surgery to eliminate residual tumors cells after tumor mass removal.

Delayed bone healing and nonunion are complications of long bone fractures, with prolonged pain and disability. Regenerative therapies employing mesenchymal stromal cells (MSC) and/or bone substitutes are increasingly applied to enhance bone consolidation. The REBORNE project entailed a multi-center orthopaedic clinical trial focused on the evaluation of efficacy of expanded autologous bone marrow (BM) derived MSC combined with a CaP-biomaterial, to enhance bone healing in patients with nonunion of diaphyseal fractures. To complement the clinical and radiological examination of patients, bone turnover markers (BTM) were assayed as potential predictors of bone healing or non-union.

Peripheral blood was collected from patients at fixed time-endpoints, that is at 6,12 and 24 weeks post-surgery for implantation of expanded autologus MSC and bone-like particles. Bone-specific alkaline phosphatase (BAP), C-terminal-propeptide type I-procollagen (PICP), osteocalcin (OC), β-Cross-Laps Collagen (CTX), soluble receptor activator of NFkB (RANKL), osteoprotegerin (OPG) were measured by ELISA assays in blood samples of 22 patients at BM collection and at follow-up visits.

A significant relationship with age was found only at 6 months, with an inverse correlation for CTX, RANKL and OC, and positive for OPG. BTM levels were not related to gender. As an effect of local regenerative process, some BTM showed significant changes in comparison to the baseline value. In particular, the time course of BAP, PICP and RANKL was different in patients with a successful healing in comparison to patients with a negative outcome. The BTM profile apparently indicated a remarkable bone formation activity 12 weeks after surgery. However, the paucity of failed patients in our case series did not allow to prove statistically the role of BTM as predictors of the final outcome.

Blood markers related to bone cell function are useful to measure the efficacy of a expanded MSC-regenerative approach applied to long bone non-unions. Changes of the markers may provide a support to radiological assessment of bone healing.

Demineralized bone matrix (DBM) is a natural, collagen-based, well-established osteoinductive biomaterial. Nevertheless, there are conflicting reports on the efficacy of this product. The purpose of this study was to evaluate whether DBM collagen structure is affected by particle size and can influence DBM osteoinductivity.

Sheep cortical bone was ground and particles were divided in three fractions with different sizes, defined as large (L, 1–2 mm), medium (M, 0.5–1 mm), and small (S, < 0.5 mm). After demineralization, the three DBM samples were characterized by DTA analysis, XRD, ICP-OES, and FTIR. Data clearly showed a particle size-dependent alteration in collagen structure, with DBM-M being altered but not as much as DBM-S. The in vivo study showed that only DBM-M was able to induce new bone formation in a subcutaneous ectopic mouse model. When sheep MSC were seeded onto DBM particles before implantation, all DBM particles were able to induce new bone formation with the best incidence for DBM-M and DBM-S. Gene expression analysis performed on recovered implants supports the histological results and underlines the supportive role of MSC in DBM osteoinduction through the regulation of host cells. In conclusion, our results show a relation between DBM particle size, structural modification of the collagen and in vivo osteoinductivity. The medium particles represent a good compromise between no modification (largest particles) and excessive modification (smallest particles) of collagen structure, yielding highest osteoinduction. We believe that these results can guide researchers to use DBM particles of 0.5–1 mm size range in applications aimed at inducing new bone formation, obtaining results more comparable and reliable among different research groups. Furthermore, we suggest to carefully analyze the structure of the collagen when a collagen-based biomaterial is used alone or in association with cells to induce new bone formation.