Lieve Vanwalleghem

Human embryonic stem cells (hESC) are pluripotent stem cells. A major challenge in the field of hESC is the establishment of specific differentiation protocols that drives hESC down a particular lineage fate. So far, attempts to generate T cells from hESC in vitro were unsuccessful. In this study, we show that T cells can be generated in vitro from hESC-derived hematopoietic precursor cells present in hematopoietic zones (HZs). These zones are morphologically similar to blood islands during embryonic development, and are formed when hESC are cultured on OP9 stromal cells. Upon subsequent transfer of these HZs on OP9 cells expressing high levels of Delta-like 1 and in the presence of growth factors, cells expand and differentiate to T cells. Furthermore, we show that T cells derive exclusively from a CD34(high)CD43(low) population, further substantiating the notion that hESC-derived CD34(high)CD43(low) cells are formed in HZs and are the only population containing multipotent hematopoietic precursor cells. Differentiation to T cells sequentially passes through the physiological intermediates: CD34(+)CD7(+) T/NK committed, CD7(+)CD4(+)CD8(-) immature single positive, CD4(+)CD8(+) double positive, and finally CD3(+)CD1(-)CD27(+) mature T cell stages. TCRalphabeta(+) and TCRgammadelta(+) T cells are generated. Mature T cells are polyclonal, proliferate, and secrete cytokines in response to mitogens. This protocol for the de novo generation of T cells from hESC could be clinically and scientifically relevant.

RATIONALE: Assessment of mediastinal lymph nodes is recommended in patients with non-small cell lung cancer without distant metastases. Linear transesophageal endoscopic ultrasound with real-time guided fine-needle aspiration (EUS-FNA) is a promising, nonsurgical tool for mediastinal staging. OBJECTIVES: We conducted a randomized controlled trial comparing surgical staging with EUS-FNA. METHODS: Patients with proven or suspected non-small cell lung cancer in whom mediastinal exploration was required were randomly assigned to undergo EUS-FNA or the appropriate surgical staging procedure. When EUS-FNA did not show malignant lymph node invasion, a confirmatory surgical staging procedure was done. A negative surgical staging procedure was followed by thoracotomy with systematic lymph node sampling. The primary endpoint was the rate of surgical staging interventions. The secondary endpoints were test performance of EUS-FNA and surgical staging, morbidity, and length of hospital stay, considering surgical staging was performed as an in-patient procedure. MEASUREMENTS AND MAIN RESULTS: A total of 40 patients were randomized: 19 to EUS-FNA, and 21 to surgical mediastinal staging. Patient and tumor characteristics were well balanced between both groups. For patients allocated to EUS-FNA, surgical staging was needed in 32% (P < 0.001). The sensitivity to detect malignant lymph node invasion was 93% (95% confidence interval, 66-99%) for EUS-FNA and 73% (95% confidence interval, 39-93%) for surgical staging (P = 0.29). Complication rate was 0% for EUS-FNA and 5% for surgical staging (P = 1.0). The median hospital stay was significantly shorter for EUS-FNA than for surgical staging (0 vs. 2 nights; P < 0.001). CONCLUSIONS: EUS-FNA reduces the need for surgical staging procedures in patients with (suspected) lung cancer in whom a mediastinal exploration is needed.

1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), the active form of vitamin D, mediates antitumor effects in various cancers. The expression of key players in vitamin D signaling in thyroid tumors was investigated. Vitamin D receptor (VDR) and CYP27B1 and CYP24A1 (respectively activating and catabolizing vitamin D) expression was studied (RT-PCR, immunohistochemistry) in normal thyroid, follicular adenoma (FA), differentiated thyroid cancer (DTC) consisting of the papillary (PTC) and follicular (FTC) subtype, and anaplastic thyroid cancer (ATC). VDR, CYP27B1, and CYP24A1 expression was increased in FA and DTC compared with normal thyroid. However, in PTC with lymph node metastasis, VDR and CYP24A1 were decreased compared with non-metastasized PTC. In ATC, VDR expression was often lost, whereas CYP27B1/CYP24A1 expression was comparable to DTC. Moreover, ATC with high Ki67 expression (>30%) or distant metastases at diagnosis was characterized by more negative VDR/CYP24A1/CYP27B1 staining. In conclusion, increased expression of key players involved in local 1,25(OH)(2)D(3) signaling was demonstrated in benign and differentiated malignant thyroid tumors, but a decrease was observed for local nodal and especially distant metastasis, suggesting a local antitumor response of 1,25(OH)(2)D(3) in early cancer stages. These findings advocate further studies with 1,25(OH)(2)D(3) and analogs in persistent and recurrent iodine-refractory DTC.