Bone marrow and adipose tissue human mesenchymal stem cells were seeded in highly performing 3D gelatin–chitosan hybrid hydrogels of varying chitosan content in the presence of human platelet lysate and evaluated for their proliferation and osteogenic differentiation. Both bone marrow and adipose tissue human mesenchymal stem cells in gelatin–chitosan hybrid hydrogel 1 (chitosan content 8.1%) or gelatin–chitosan hybrid hydrogel 2 (chitosan 14.9%) showed high levels of viability (80%–90%), and their proliferation and osteogenic differentiation was significantly higher with human platelet lysate compared to fetal bovine serum, particularly in gelatin–chitosan hybrid hydrogel 1. Mineralization was detected early, after 21 days of culture, when human platelet lysate was used in the presence of osteogenic stimuli. Proteomic characterization of human platelet lysate highlighted 59 proteins mainly involved in functions related to cell adhesion, cellular repairing mechanisms, and regulation of cell differentiation. In conclusion, the combination of our gelatin–chitosan hybrid hydrogels with hPL represents a promising strategy for bone regenerative medicine using human mesenchymal stem cells.

We investigated the frequency of detection and the prognostic and predictive significance of circulating tumor cells (CTCs) in patients with recurrent/metastatic (R/M) head and neck carcinoma (HNC) before starting systemic therapy. Using the CellSearch technology, CTCs were assessed prospectively in peripheral blood of 53 R/M-HNC patients. We performed spiking experiments to test the diagnostic performance of the CellSearch platform in identifying squamous carcinoma cells. CTCs were identified in 14 (26%) and 22 (41%) patients at baseline and at any time point, respectively. In univariate analysis ≥2 CTCs had a poorer prognostic role than 0–1 CTC. In multivariate analysis, the presence of one CTC or more was associated with a poor prognosis both in terms of progression-free survival (PFS) [Hazard Ratio (HR): 3.068, 95% confidence interval (CI): 1.53–6.13, p 0.002] and overall survival (OS) [HR: 3.0, 95% CI: 1.48–6.0, p 0.002]. A disease control after systemic therapy was obtained in 8% of CTC-positive patients as opposed to 45% in CTC-negative ones (p 0.03). The epidermal growth factor receptor (EGFR) expression was identified in 45% of CTC-positive patients. In conclusion, CTCs are detected in one out of three patients with RM-HNC. CTC detection is a strong prognostic parameter and may be predictive of treatment efficacy. The frequency of EGFR expression in CTCs seems to be lower than that expected in the primary tumor.

Background Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is burdened by life-threatening complications, with graft-versus-host disease (GvHD) being the major cause of morbility and mortality. Recently, clinical and physiopathologic evidences showed that vascular endothelium can be a target of GvHD in the early phase and circulating endothelial cells (CECs) represent surrogate markers of endothelial damage. Methods Using the CellSearch System (Veridex LLC, Raritan, NJ), CECs were counted before (T1), after conditioning regimen (T2), at engraftment (T3), at GvHD onset (T4), and after steroid treatment (T5) in 40 patients (7 Hodgkin’s Disease, 13 Acute Myeloblastic Leukemia, 5 Acute Lymphoblastic Leukemia, 8 Multiple Myeloma, 3 Chronic Lymphocytic Leukemia, 1 Non-Hodgkin Lymphoma, 1 Chronic Myeloid Leukemia, 2 Severe Aplastic Anemia) undergoing allo-HSCT. Results The median CEC per milliliter at T1 was 20 (n=33, range 4–718), in comparison to a value of 2 (range, 1–14) in controls (P<0.001). At T3, CEC per milliliter were 47 (range, 16–148) in GvHD patients and 92 (range, 23–276) in patients without GvHD (P=0.006). This difference remained significant in multivariate analysis (odds ratio, 0.97; 95% confidence interval, 0.96–0.99; P=0.02). At GvHD onset, the relative increase of CEC counts from time of engraftment (T4 vs. T3) was 44% (range, −43% to 569%) in GvHD patients versus 0% (range, −49% to 2%) in patients without GvHD (P=0.003), being confirmed as significant in multivariate analysis (odds ratio, 1.04; 95% confidence interval, 1.0–1.08; P=0.04). Conclusion Changes in CEC count can represent a promising marker to monitor endothelial damage in patients undergoing allo-HSCT and could become a valuable tool in the diagnostic definition of GvHD.

The physio-pathologic interrelationships between endothelium and GvHD have been better elucidated and have led to definition of the entity ‘endothelial GvHD’ as an essential early phase prior to the clinical presentation of acute GvHD. Using the CellSearch system, we analyzed circulating endothelial cells (CEC) in 90 allogeneic hematopoietic stem cell transplantation (allo-HSCT) patients at the following time-points: T1 (pre-conditioning), T2 (pre-transplant), T3 (engraftment), T4 (onset of GvHD) and T5 (1 week after steroid treatment). Although CEC changes in allo-HSCT represent a dynamic phenomenon influenced by many variables (that is, conditioning, immunosuppressive treatments, engraftment syndrome and infections), we showed that CEC peaks were constantly seen at onset of acute GvHD and invariably returned to pre-transplant values after treatment response. Since we showed that CEC changes during allo-HSCT has rapid kinetics that may be easily missed if blood samples are drawn at pre-fixed time-points, we rather suggest an ‘on demand’ evaluation of CEC counts right at onset of GvHD clinical symptoms to possibly help differentiate GvHD from other non-endothelial complications. We confirm that CEC changes are a suitable biomarker to monitor endothelial damage in patients undergoing allo-transplantation and hold the potential to become a useful tool to support GvHD diagnosis (ClinicalTrials.gov NCT02064972).