Person:

Choi, Michael

MicroRNAs play a pivotal role in cellular maintenance, proliferation, and differentiation. They have also been implicated to play a key role in disease pathogenesis, and more recently, cellular reprogramming. Certain microRNA clusters can enhance or even directly induce reprogramming, while repressing key proteins involved in microRNA processing decreases reprogramming efficiency. Although microRNAs clearly play important roles in cellular reprogramming, it remains unknown whether microRNAs are absolutely necessary. We endeavored to answer this fundamental question by attempting to reprogram Dicer-null mouse embryonic fibroblasts (MEFs) that lack almost all functional microRNAs using a defined set of transcription factors. Transduction of reprogramming factors using either lentiviral or piggyBac transposon vector into two, independently derived lines of Dicer-null MEFs failed to produce cells resembling embryonic stem cells (ESCs). However, expression of human Dicer in the Dicer-null MEFs restored their reprogramming potential. Our study demonstrates for the first time that microRNAs are indispensable for dedifferentiation reprogramming.

Non-alcoholic fatty liver disease (NAFLD) encompasses a wide spectrum of disease severity, starting from pure steatosis, leading to fatty inflammation labeled as non-alcoholic steatohepatitis (NASH), and finally fibrosis leading to cirrhosis. Activated hepatic stellate cells (HSCs) are known to contribute to fibrosis, but less is known about their function during NAFLD’s early stages prior to fibrosis. We developed an ex vivo assay that cocultures primary HSCs from mouse models of liver disease with healthy hepatocytes to study their interaction. Our data indicate that chemokine Ccl5 is one of the HSC-secreted mediators in early NASH in humans and in mice fed with choline-deficient, L-amino acid defined, high fat diet. Furthermore, Ccl5 directly induces steatosis and pro-inflammatory factors in healthy hepatocytes through the receptor Ccr5. Although Ccl5 is already known to be secreted by many liver cell types including HSCs and its pro-fibrotic role well characterized, its pro-steatotic action has not been recognized until now. Similarly, the function of HSCs in fibrogenesis is widely accepted, but their pro-steatotic role has been unclear. Our result suggests that in early NASH, HSCs secrete Ccl5 which contributes to a broad array of mechanisms by which hepatic steatosis and inflammation are achieved.

Homeodomain transcription factors serve important functions in organogenesis and tissue differentiation, particularly with respect to the positional identity of individual cells. The Nkx6 subfamily controls tissue differentiation in the developing central nervous system where they function as transcriptional repressor proteins. Recent work indicates that Nkx6.3 is expressed in hindbrain V2 interneurons that co-express Nkx6.1, suggesting the possibility of functional redundancy. Here, we report that Nkx6.3 expression is specific to Chx10+ V2a interneurons but not Gata3+ V2b interneurons of the hindbrain, and that Nkx6.3 expression appears to mark cells of the prospective medullary reticular formation. Molecular analysis of Nkx6.3 null embryonic mouse hindbrain did not reveal detectable defects in progenitor markers, motor neuron or V2 interneuron sub-types. Forced expression of Nkx6.3 and Nkx6.1 promote V2 interneuron differentiation in the developing chick hindbrain. These findings indicate Nkx6.3 function is dispensable for CNS development and lead to the proposal that absence of overt defects is due to functional compensation from a related homeodomain transcription factor.