A ß1-tubulin-based megakaryocyte maturation reporter system identifies novel drugs that promote platelet production.

During maturation, megakaryocytes (MKs) express ß1-tubulin (TUBB1) and rearrange their microtubule components to enlarge, form proplatelets, and eventually release platelets. The development of a platform to identify in vitro conditions that would efficiently promote MK development could potentially enable large-scale platelet production. Here, we show that an immortalized MK cell line (imMKCL) genetically modified to express the ß1-tubulin-Venus reporter provides a practical system to efficiently monitor the in vitro production of platelet-like particles (PLPs). The Venus transgene was inserted downstream of the TUBB1 locus in imMKCLs using CRISPR/Cas9, and the expression was visualized by Venus fluorescence intensity. This imMKCL reporter line was then used for high-throughput drug screening. We identified several compounds that significantly improved the efficiency of PLP production in vitro under feeder-free conditions and showed a significant tendency to recover platelets in vivo in a mouse thrombocytopenia model induced by anti-GPIbα antibody administration. Interestingly, most of these compounds, including a WNT signaling pathway inhibitor, Wnt-C59, antagonized the aryl hydrocarbon receptor (AhR) to increase PLP production, confirming the crucial role of AhR inhibition in MK maturation. Consistently, small interfering RNA treatment against AhR increased the Venus intensity and PLP production. TCS 359, an FLT3 inhibitor, significantly increased PLP production independently of FLT3 or AhR. This study highlights the usefulness of the ß1-tubulin reporter MK line as a useful tool to study the mechanisms underlying thrombopoiesis and to identify novel inducers of ex vivo platelet production.

Anti-prediabetic effect of rose hip (Rosa canina) extract in spontaneously diabetic Torii rats.

BACKGROUND: Prediabetes, a high-risk state for developing diabetes showing impaired glucose tolerance but a normal fasting blood glucose level, has an increasing prevalence worldwide. However, no study investigating the prevention of impaired glucose tolerance at the prediabetic stage by anti-diabetic functional foods has been reported. Thus, the present study aimed to evaluate the anti-prediabetic effect of rose hip in a prediabetic rat model. RESULTS: Spontaneously diabetic Torii (SDT) rats were supplemented with hot-water extract of rose hip at a dose of 100 mg kg-1 body weight day-1 for 12 weeks. The results obtained showed that the supplementation of rose hip extract improved impaired glucose tolerance, promoted insulin secretion, preserved pancreatic beta-cell function and suppressed plasma advanced glycation end-products formation of methylglyoxal-derived hydroimidazolone (MG-H1) residue and Nϵ -carboxymethyl-lysine residues (e.g. MG-H1, control: 465.5 ± 43.8 versus rose hip: 59.1 ± 13.0 pmol mg protein-1 , P < 0.05) in SDT rats at the prediabetic stage (12-20 weeks old). CONCLUSION: The present study provides the first evidence showing that a hot-water extract of rose hip could exert an anti-prediabetic effect in a rat model. © 2017 Society of Chemical Industry.