Adipose tissue is a major site of energy storage and plays a role in regulation of metabolism through release of adipokines. Here we show that mice with a fat-specific knockout of the miRNA-processing enzyme Dicer (ADicerKO), as well as humans with lipodystrophy, have major decreases in circulating exosomal miRNAs. Transplantation of white and especially brown adipose tissue (BAT) into ADicerKO mice restores circulating miRNAs associated with an improvement in glucose tolerance and a reduction of hepatic FGF21 mRNA and circulating FGF21. This gene regulation can be mimicked by administration of normal, but not AdicerKO, serum exosomes. Expression of a human-specific miRNA in BAT of one mouse in vivo can also regulate its 3’UTR-reporter in liver of another mouse through serum exosomal transfer. Thus, adipose tissue constitutes a major source of circulating exosomal miRNAs, and these miRNAs can regulate gene expression in distant tissues thereby serving as novel forms of adipokines.

Both intrinsic cell state changes and variations in the composition of stem cell populations have been implicated as contributors to aging. We used single-cell RNA-seq to dissect variability in hematopoietic stem cell (HSC) and hematopoietic progenitor cell populations from young and old mice from two strains. We found that cell cycle dominates the variability within each population and that there is a lower frequency of cells in the G1 phase among old compared with young long-term HSCs, suggesting that they traverse through G1 faster. Moreover, transcriptional changes in HSCs during aging are inversely related to those upon HSC differentiation, such that old short-term (ST) HSCs resemble young long-term (LT-HSCs), suggesting that they exist in a less differentiated state. Our results indicate both compositional changes and intrinsic, population-wide changes with age and are consistent with a model where a relationship between cell cycle progression and selfrenewal versus differentiation of HSCs is affected by aging and may contribute to the functional decline of old HSCs.

SUMMARY Remyelination is a regenerative process in the central nervous system (CNS) that produces new myelin sheaths from adult stem cells. The decline in remyelination that occurs with advancing age poses a significant barrier to therapy in the CNS, particularly for long-term demyelinating diseases such as multiple sclerosis (MS). Here we show that remyelination of experimentally-induced demyelination is enhanced in old mice exposed to a youthful systemic milieu through heterochronic parabiosis. Restored remyelination in old animals involves recruitment to the repairing lesions of blood-derived monocytes from the young parabiotic partner, and preventing this recruitment partially inhibits rejuvenation of remyelination. These data suggest that enhanced remyelinating activity requires both youthful monocytes and other factors, and that remyelination-enhancing therapies targeting endogenous cells can be effective throughout life.

The considerable potential of Cre recombinase as a tool for in vivo fate-mapping studies depends on the availability of reliable reporter mice. By targeting a tandem-dimer red fluorescent protein (tdRFP) with advanced spectral and biological properties into the ubiquitously expressed ROSA26 locus of C57BL/6-ES cells, we have generated a novel inbred Cre-reporter mouse with several unique characteristics. We directly demonstrate the usefulness of our reporter strain in inter-crosses with a "universal Cre-deleter" strain and with mice expressing Cre recombinase in a T lineage-specific manner. Cytofluorometric and histological analyses illustrate: (i) non-toxicity and extraordinary brightness of the fluorescent reporter, allowing quantitative detection and purification of labeled cells with highest accuracy, (ii) reliable Cre-mediated activation of tdRFP from an antisense orientation relative to ROSA26 transcription, effectively excluding "leaky" reporter expression, (iii) absence of gene expression variegation effects, (iv) quantitative detection of tdRFP-expressing cells even in paraformaldehyde-fixed tissue sections, and (v) full compatibility with GFP/YFP-based fluorescent markers in multicolor experiments. Taken together, the data show that our C57BL/6-inbred reporter mice are ideally suited for sophisticated lineage-tracing experiments requiring sensitive and quantitative detection/purification of live Cre-expressing cells and their progeny. IntroductionCre recombinase is a small, bacteriophage P1-derived integrase that catalyses defined DNA recombination events between specific target sites, termed loxP (locus of crossover [x] in P1) [1]. LoxP sites are composed of two 13-bp inverted repeats and an asymmetric 8-bp spacer sequence, which endows individual loxP elements with a defined orientation [2]. The result of Cremediated recombination between two loxP sites de- pends on their specific orientation relative to one another: a DNA sequence flanked by two directly repeated loxP elements is cut out as a circular molecule, an effectively irreversible reaction due to loss of the excised product. In contrast, DNA flanked by two oppositely oriented sites will be inverted, a fully reversible process. Importantly, both activity and specificity of Cre recombinase are retained in eukaryotic cells [3], permitting the use of Cre as a genetic engineering tool in any cellular context. In combination with gene targeting techniques, the Cre/loxP recombination system has revolutionized the genetic analysis of mice [4,5]. Cre-mediated removal of loxP-flanked (floxed) selection cassettes, which are invariably introduced during the process of gene targeting into the mouse genome, and which may have unpredictable effects on neighboring genes, is already a routine manipulation. Also, conditional deletion of floxed genomic DNA sequences has become a rather standard approach for assessing tissue-, cell-type-or developmental stage-specific gene functions, provided appropriate Cre-transgenic mouse strains are available. Less common ...

In chronic kidney disease, fibroblast dysfunction causes renal fibrosis and renal anemia. Renal fibrosis is mediated by the accumulation of myofibroblasts, whereas renal anemia is mediated by the reduced production of fibroblast-derived erythropoietin, a hormone that stimulates erythropoiesis. Despite their importance in chronic kidney disease, the origin and regulatory mechanism of fibroblasts remain unclear. Here, we have demonstrated that the majority of erythropoietin-producing fibroblasts in the healthy kidney originate from myelin protein zero-Cre (P0-Cre) lineage-labeled extrarenal cells, which enter the embryonic kidney at E13.5. In the diseased kidney, P0-Cre lineage-labeled fibroblasts, but not fibroblasts derived from injured tubular epithelial cells through epithelial-mesenchymal transition, transdifferentiated into myofibroblasts and predominantly contributed to fibrosis, with concomitant loss of erythropoietin production. We further demonstrated that attenuated erythropoietin production in transdifferentiated myofibroblasts was restored by the administration of neuroprotective agents, such as dexamethasone and neurotrophins. Moreover, the in vivo administration of tamoxifen, a selective estrogen receptor modulator, restored attenuated erythropoietin production as well as fibrosis in a mouse model of kidney fibrosis. These findings reveal the pathophysiological roles of P0-Cre lineage-labeled fibroblasts in the kidney and clarify the link between renal fibrosis and renal anemia.

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SummaryHematopoietic stem cells (HSCs) maintain blood homeostasis and are the functional units of bone marrow transplantation. To improve the molecular understanding of HSCs and their proximal progenitors, we performed transcriptome analysis within the context of the ImmGen Consortium data set. Gene sets that define steady-state and mobilized HSCs, as well as hematopoietic stem and progenitor cells (HSPCs), were determined. Genes involved in transcriptional regulation, including a group of putative transcriptional repressors, were identified in multipotent progenitors and HSCs. Proximal promoter analyses combined with ImmGen module analysis identified candidate regulators of HSCs. Enforced expression of one predicted regulator, Hlf, in diverse HSPC subsets led to extensive self-renewal activity ex vivo. These analyses reveal unique insights into the mechanisms that control the core properties of HSPCs.