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Background: Since the esophagus has no redundancy, congenital and acquired esophageal diseases often require esophageal substitution, with complicated surgery and intestinal or gastric transposition. Peri-and-post-operative complications are frequent, with major problems related to the food transit and reflux. During the last years tissue engineering products became an interesting therapeutic alternative for esophageal replacement, since they could mimic the organ structure and potentially help to restore the native functions and physiology. The use of acellular matrices pre-seeded with cells showed promising results for esophageal replacement approaches, but cell homing and adhesion to the scaffold remain an important issue and were investigated. Methods: A porcine esophageal substitute constituted of a decellularized scaffold seeded with autologous bone marrow-derived mesenchymal stromal cells (BM-MSCs) was developed. In order to improve cell seeding and distribution throughout the scaffolds, they were micro-perforated by Quantum Molecular Resonance (QMR) technology (Telea Electronic Engineering). Results: The treatment created a microporous network and cells were able to colonize both outer and inner layers of the scaffolds. Non seeded (NSS) and BM-MSCs seeded scaffolds (SS) were implanted on the thoracic esophagus of 4 and 8 pigs respectively, substituting only the muscle layer in a mucosal sparing technique. After 3 months from surgery, we observed an esophageal substenosis in 2/4 NSS pigs and in 6/8 SS pigs and a non-practicable stricture in 1/4 NSS pigs and 2/8 SS pigs. All the animals exhibited a normal weight increase, except one case in the SS group. Actin and desmin staining of the post-implant scaffolds evidenced the regeneration of a muscular layer from one anastomosis to another in the SS group but not in the NSS one. Conclusions: A muscle esophageal substitute starting from a porcine scaffold was developed and it was fully repopulated by BM-MSCs after

Chronic lymphocytic leukemia (CLL) is characterized by a low CD20 expression, in part explained by an epigenetic-driven downregulation triggered by mutations of the NOTCH1 gene. In the present study, by taking advantage of a wide and well-characterized CLL cohort (n=537), we demonstrate that CD20 expression is downregulated in SF3B1-mutated CLL in an extent similar to NOTCH1-mutated CLL. In fact, SF3B1-mutated CLL cells show common features with NOTCH1-mutated CLL cells, including a gene expression profile enriched of NOTCH1-related gene sets and elevated expression of the active intracytoplasmic NOTCH1. Activation of the NOTCH1 signaling and down-regulation of surface CD20 in SF3B1-mutated CLL cells correlate with over-expression of an alternatively spliced form of DVL2, a component of the Wnt pathway and negative regulator of the NOTCH1 pathway. These findings are confirmed by separately analyzing the CD20-dim and CD20-bright cell fractions from SF3B1-mutated cases as well as by DVL2 knock-out experiments in CLL-like cell models. Altogether, the clinical and biological features that characterize NOTCH1-mutated CLL may also be recapitulated in SF3B1-mutated CLL, contributing to explain the poor prognosis of this CLL subset and providing the rationale for expanding novel agents-based therapies to SF3B1-mutated CLL.

CD49d is a remarkable prognostic biomarker of chronic lymphocytic leukemia (CLL). The cutoff value for the extensively validated 30% of positive CLL cells is able to separate CLL patients into 2 subgroups with different prognoses, but it does not consider the pattern of CD49d expression. In the present study, we analyzed a cohort of 1630 CLL samples and identified the presence of ∼20% of CLL cases (n 5 313) characterized by a bimodal expression of CD49d, that is, concomitant presence of a CD49d1 subpopulation and a CD49d2 subpopulation. At variance with the highly stable CD49d expression observed in CLL patients with a homogeneous pattern of CD49d expression, CD49d bimodal CLL showed a higher level of variability in sequential samples, and an increase in the CD49d1 subpopulation over time after therapy. The CD49d1 subpopulation from CD49d bimodal CLL displayed higher levels of proliferation compared with the CD49d2 cells; and was more highly represented in the bone marrow compared with peripheral blood (PB), and in PB CLL subsets expressing the CXCR4dim/CD5bright phenotype, known to be enriched in proliferative cells. From a clinical standpoint, CLL patients with CD49d bimodal expression, regardless of whether the CD49d1 subpopulation exceeded the 30% cutoff or not, experienced clinical behavior similar to CD49d1 CLL, both in chemoimmunotherapy (n 5 1522) and in ibrutinib (n 5 158) settings. Altogether, these results suggest that CD49d can drive disease progression in CLL, and that the pattern of CD49d expression should also be considered to improve the prognostic impact of this biomarker in CLL.