GM-CSF and MEF-conditioned media support feeder-free reprogramming of mouse granulocytes to iPS cells

Induced pluripotent stem cells (iPSCs) are characterised by their ability to differentiate into any cell type of the body. Accordingly, iPSCs possess immense potential for disease modelling, pharmaceutical screening and autologous cell therapies. The most common source of iPSCs derivation is skin fibroblasts. However, from a clinical point of view, skin fibroblasts may not be ideal, as invasive procedures such as skin biopsies are required for their extraction. Moreover, fibroblasts are highly heterogeneous with a poorly defined developmental pathway, which makes studying reprogramming mechanistics difficult. Granulocytes, on the other hand, are easily obtainable, their developmental pathway has been extensively studied and fluorescence activated cell sorting allows for the isolation of these cells at high purity; thus iPSCs derivation from granulocytes could provide an alternative to fibroblast-derived iPSCs. Previous studies succeeded in producing iPSC colonies from mouse granulocytes but with the use of a mitotically inactivated feeder layer, restricting their use for studying reprogramming mechanistics. As granulocytes display poor survival under culture conditions, we investigated the influence of haematopoietic cytokines to stabilise this cell type in vitro and allow for reprogramming in the absence of a feeder layer. Our results show that treatment with MEF-conditioned media and/or initial exposure to GM-CSF allows for reprogramming of granulocytes under feeder-free conditions. This work can serve as a basis for future work aimed at dissecting the reprogramming mechanism as well as obtaining large numbers of iPSCs from a clinically relevant cell source.     

Changes in leptin, ghrelin, growth hormone and neuropeptide-Y after an acute model of MDMA and methamphetamine exposure in rats.

Club drug abuse is a growing problem in the United States. Beyond addiction and toxicity are endocrine effects which are not well characterized. Specifically, the changes in appetite following exposure to drugs of abuse are an interesting but poorly understood phenomenon. Serum hormones such as leptin, ghrelin, growth hormone (GH), and neuropeptide-Y (NP-Y) are known to affect appetite, but have not been studied extensively with drugs of abuse. In this work, we examine the effects of club drugs 3,4-methylenedioxymethamphetamine (MDMA) (ecstasy) and methamphetamine (METH) (doses of 5, 20 and 40 mg/kg) on serum concentrations of these hormones in adult male Sprague-Dawley rats 6, 12, 24 and 48 hours after drug administration. In a dose-dependent manner, MDMA was shown to cause transient significant decreases in serum leptin and GH followed by a base line recovery after 24 hours. Conversely, serum ghrelin increased and normalized after 24 hours. Interestingly, serum NP-Y showed a steady decrease in both treatment of MDMA and METH at different time points and dosages. In humans, abuse of these drugs reduces eating. As evident from these data, acute administration of METH and MDMA had significant effects on different serum hormone levels involved in appetite regulation. Future studies should be performed to see how chronic, low dose drug administration would affect hormone levels and try to answer questions about the physiological mechanisms involved in the anorexic paradigm observed in drug use.