Spatial distribution of beta-klotho mRNA in the mouse hypothalamus, hippocampal region, subiculum, and amygdala.

Beta-klotho (KLB) is a coreceptor required for endocrine fibroblast growth factor (FGF) 15/19 and FGF21 signaling in the brain. Klb is prominent within the hypothalamus, which is consistent with its metabolic functions, but diverse roles for Klb are now emerging. Central Klb expression is low but discrete and may govern FGF-targeted sites. However, given its low expression, it is unclear if Klb mRNA is more widespread. We performed in situ hybridization to label Klb mRNA to generate spatial maps capturing the distribution and levels of Klb within the mouse hypothalamus, hippocampal region, subiculum, and amygdala. Semiquantitative analysis revealed that Klb-labeled cells may express low, medium, or high levels of Klb mRNA. Hypothalamic Klb hybridization was heterogeneous and varied rostrocaudally within the same region. Most Klb-labeled cells were found in the lateral hypothalamic zone, but the periventricular hypothalamic region, including the suprachiasmatic nucleus, contained the greatest proportion of cells expressing medium or high Klb levels. We also found heterogeneous Klb hybridization in the amygdala and subiculum, where Klb was especially distinct within the central amygdalar nucleus and ventral subiculum, respectively. By contrast, Klb-labeled cells in the hippocampal region only expressed low levels of Klb and were typically found in the pyramidal layer of Ammon's horn or dentate gyrus. The Klb-labeled regions identified in this study are consistent with reported roles of Klb in metabolism, taste preference, and neuroprotection. However, additional identified sites, including within the hypothalamus and amygdala, may suggest novel roles for FGF15/19 or FGF21 signaling.

Development and characterization of a DNA aptamer for MLL-AF9 expressing acute myeloid leukemia cells using whole cell-SELEX.

Current classes of cancer therapeutics have negative side effects stemming from off-target cytotoxicity. One way to avoid this would be to use a drug delivery system decorated with targeting moieties, such as an aptamer, if a targeted aptamer is available. In this study, aptamers were selected against acute myeloid leukemia (AML) cells expressing the MLL-AF9 oncogene through systematic evolution of ligands by exponential enrichment (SELEX). Twelve rounds of SELEX, including two counter selections against fibroblast cells, were completed. Aptamer pools were sequenced, and three candidate sequences were identified. These sequences consisted of two 23-base primer regions flanking a 30-base central domain. Binding studies were performed using flow cytometry, and the lead sequence had a binding constant of 37.5 + / - 2.5 nM to AML cells, while displaying no binding to fibroblast or umbilical cord blood cells at 200 nM. A truncation study of the lead sequence was done using nine shortened sequences, and showed the 5' primer was not important for binding. The lead sequence was tested against seven AML patient cultures, and five cultures showed binding at 200 nM. In summary, a DNA aptamer specific to AML cells was developed and characterized for future drug-aptamer conjugates.

Ciliary melanin-concentrating hormone receptor 1 (MCHR1) is widely distributed in the murine CNS in a sex-independent manner.

Melanin-concentrating hormone (MCH) is a ubiquitous vertebrate neuropeptide predominantly synthesized by neurons of the diencephalon that can act through two G protein-coupled receptors, called MCHR1 and MCHR2. The expression of Mchr1 has been investigated in both rats and mice, but its synthesis remains poorly described. After identifying an antibody that detects MCHR1 with high specificity, we employed immunohistochemistry to map the distribution of MCHR1 in the CNS of rats and mice. Multiple neurochemical markers were also employed to characterize some of the neuronal populations that synthesize MCHR1. Our results show that MCHR1 is abundantly found in a subcellular structure called the primary cilium, which has been associated, among other functions, with the detection of free neurochemical messengers present in the extracellular space. Ciliary MCHR1 was found in a wide range of areas, including the olfactory bulb, cortical mantle, striatum, hippocampal formation, amygdala, midline thalamic nuclei, periventricular hypothalamic nuclei, midbrain areas, and in the spinal cord. No differences were observed between male and female mice, and interspecies differences were found in the caudate-putamen nucleus and the subgranular zone. Ciliary MCHR1 was found in close association with several neurochemical markers, including tyrosine hydroxylase, calretinin, kisspeptin, estrogen receptor, oxytocin, vasopressin, and corticotropin-releasing factor. Given the role of neuronal primary cilia in sensing free neurochemical messengers in the extracellular fluid, the widespread distribution of ciliary MCHR1, and the diverse neurochemical populations who synthesize MCHR1, our data indicate that nonsynaptic communication plays a prominent role in the normal function of the MCH system.