The Sodium-Calcium Exchanger Controls the Membrane Potential of AFT024: A Mesenchymal Stem Cell Hematopoietic Niche Forming Line.

The aim of this study was to characterize the functional expression of sodium-calcium exchangers on AFT024 cell line, a murine model of mesenchymal stem/stromal cells (MSCs) supporting human primitive hematopoiesis. All current-clamp and voltage-clamp experiments were performed using the perforated patch whole-cell recording technique with amphotericin B. The membrane potential of -14 mV shifted to -35 mV when lowering the external sodium concentration to 0.33 mM and an increase of cytosolic calcium concentration was observed. KB-R7943, a selective blocker of cardiac sodium-calcium exchangers, also named NCX1, induced a hyperpolarization at physiological sodium concentration while it blocked the hyperpolarization observed at low sodium concentration. This demonstrates for the first time the presence of the sodium-calcium exchangers in AFT024 cells and provides initial evidence that the membrane potential of these stromal cells is maintained depolarized by this exchanger. Lowering external sodium concentration and KB-R7943 had no effect on the membrane potential of 2018 cells, a nonhematopoietic-supportive cell line. Since NCX1 is differentially expressed in AFT024 cells as compared with nonhematopoietic supportive cells with more restricted differentiation potential, this study suggests a potential role of this sodium-calcium exchanger, in the differentiation process or hematopoietic support of MSCs.

Low-Dose Pesticides Alter Primary Human Bone Marrow Mesenchymal Stem/Stromal Cells through ALDH2 Inhibition.

(1) Background: The impact of occupational exposure to high doses of pesticides on hematologic disorders is widely studied. Yet, lifelong exposure to low doses of pesticides, and more particularly their cocktail effect, although poorly known, could also participate to the development of such hematological diseases as myelodysplastic syndrome (MDS) in elderly patients. (2) Methods: In this study, a cocktail of seven pesticides frequently present in water and food (maneb, mancozeb, iprodione, imazalil, chlorpyrifos ethyl, diazinon and dimethoate), as determined by the European Food Safety Authority, were selected. Their in vitro effects at low-doses on primary BM-MSCs from healthy volunteers were examined. (3) Results: Exposure of normal BM-MSCs to pesticides for 21 days inhibited cell proliferation and promoted DNA damage and senescence. Concomitantly, these cells presented a decrease in aldehyde dehydrogenase 2 (ALDH2: mRNA, protein and enzymatic activity) and an increase in acetaldehyde levels. Pharmacological inhibition of ALDH2 with disulfiram recapitulated the alterations induced by exposure to low doses of pesticides. Moreover, BM-MSCs capacity to support primitive hematopoiesis was significantly altered. Similar biological abnormalities were found in primary BM-MSCs derived from MDS patients. (4) Conclusions: these results suggest that ALDH2 could participate in the pathophysiology of MDS in elderly people long exposed to low doses of pesticides.

Disruption of gap junctions attenuates acute myeloid leukemia chemoresistance induced by bone marrow mesenchymal stromal cells.

The bone marrow (BM) niche impacts the progression of acute myeloid leukemia (AML) by favoring the chemoresistance of AML cells. Intimate interactions between leukemic cells and BM mesenchymal stromal cells (BM-MSCs) play key roles in this process. Direct intercellular communications between hematopoietic cells and BM-MSCs involve connexins, components of gap junctions. We postulated that blocking gap junction assembly could modify cell-cell interactions in the leukemic niche and consequently the chemoresistance. The comparison of BM-MSCs from AML patients and healthy donors revealed a specific profile of connexins in BM-MSCs of the leukemic niche and the effects of carbenoxolone (CBX), a gap junction disruptor, were evaluated on AML cells. CBX presents an antileukemic effect without affecting normal BM-CD34+ progenitor cells. The proapoptotic effect of CBX on AML cells is in line with the extinction of energy metabolism. CBX acts synergistically with cytarabine (Ara-C) in vitro and in vivo. Coculture experiments of AML cells with BM-MSCs revealed that CBX neutralizes the protective effect of the niche against the Ara-C-induced apoptosis of leukemic cells. Altogether, these results suggest that CBX could be of therapeutic interest to reduce the chemoresistance favored by the leukemic niche, by targeting gap junctions, without affecting normal hematopoiesis.

Correction: Disruption of gap junctions attenuates acute myeloid leukemia chemoresistance induced by bone marrow mesenchymal stromal cells.

The original version of this Article omitted the following from the Acknowledgements: This research was also supported by grants to KZ (UL and L-CNRS). This has now been corrected in both the PDF and HTML versions of the Article.

Aerodynamical, immunological and pharmacological properties of the anticancer antibody cetuximab following nebulization.

PURPOSE: Despite an increasing interest in the use of inhalation for local delivery of molecules for respiratory diseases and systemic disorders, methods to deliver therapy through airways has received little attention for lung cancer treatment. However, inhalation of anticancer drugs is an attractive alternative route to systemic administration which results in limited concentration of the medication in the lungs, and triggers whole-body toxicity. In this study, we investigated the feasibility of nebulization for therapeutic antibodies, a new class of fully-approved anticancer drugs in oncology medicine. MATERIALS AND METHODS: Cetuximab, a chimeric IgG1 targeting the epidermal growth factor receptor (EGFR), was nebulized using three types of delivery devices: a jet nebulizer PARI LC+, a mesh nebulizer AeronebPro and an ultrasonic nebulizer SYST'AM LS290. Aerosol size distribution was measured using a cascade impactor and aerosol droplets were observed under optical microscopy. The immunological and pharmacological properties of cetuximab were evaluated following nebulization using A431 cells. RESULTS: The aerosol particle clouds generated with the three nebulizers displayed similar aerodynamical characteristics, but the IgG formed aggregates in liquid phase following nebulization with both the jet and ultrasonic devices. Flow cytometry analyses and assays of EGFR-phosphorylation and cell growth inhibitions on A431 demonstrated that both the mesh and the jet nebulizers preserved the binding affinity to EGFR and the inhibitory activities of cetuximab. CONCLUSIONS: Altogether, our results indicate that cetuximab resists the physical constraints of nebulization. Thus, airway delivery represents a promising alternative to systemic administration for local delivery of therapeutic antibodies in lung cancer treatment.