Endothelial cells and macrophages are known to engage in tight and specific interactions that contribute to the modulation of vascular function. Here we show that adult endothelial cells provide critical signals for the selective growth and differentiation of macrophages from several hematopoietic progenitors. The process features the formation of wellorganized colonies that exhibit progressive differentiation from the center to the periphery and toward an M2-like phenotype, characterized by enhanced expression of Tie2 and CD206/Mrc1. These colonies are long-lived depending on the contact with the endothelium; removal of the endothelial monolayer results in rapid colony dissolution. We further found that Csf1 produced by the endothelium is critical for the expansion of the macrophage colonies and that blockade of Csf1 receptor impairs colony growth. Functional analyses indicate that these macrophages are capable of accelerating angiogenesis, promoting tumor growth, and effectively engaging in tight associations with endothelial cells in vivo. These findings uncover a critical role of endothelial cells in the induction of macrophage differentiation and their ability to promote further polarization toward a proangiogenic phenotype. This work also highlights some of the molecules underlying the M2-like differentiation, a process that is relevant to the progression of both developmental and pathologic angiogenesis. (Blood. 2012;120(15):3152-3162) IntroductionThe link between the hematopoietic and the endothelial cell lineages is rooted early in development. In fact, definitive hematopoietic stem cells (HSCs) first emerge in the embryo from a specialized endothelial intermediate that holds hemogenic capacity. [1][2][3][4] Although the process of hematopoietic cells (HCs) budding from hemogenic endothelium is no longer present in the adult, the interactions between HCs and the endothelium continue to be critical for the trafficking and homing of HCs, as well as for activation and recruitment of inflammatory cells to specific tissue sites. 5 More recently, sinusoidal endothelial cells were shown to be essential for the self-renewal capacity of hematopoietic stem/ progenitor cells (HSPCs) through the production of specific angiocrine factors. 6,7 Intriguingly, bone marrow sinusoidal endothelial cells can also constitute a platform for the differentiation of HSPCs. This dual role of endothelial cells has been best exemplified by findings communicated by Kobayashi and colleagues, where the coculture of genetically modified human umbilical vein endothelial cells (HUVECs) with HSPCs supported both selfrenewal and lineage-specific differentiation of HSPCs. 8 Notably, the mechanisms by which endothelial cells mediate regeneration or differentiation of HCs depend largely on organ-specific determinants. Overall, mounting evidence supports the concept that the crosstalk between endothelial cells and HCs impacts the differentiation and stem cell properties of hematopoietic progenitors.The consequences of endothelial-hematopoietic...

Scl/Tal1 confers hemogenic competence and prevents ectopic cardiomyogenesis in embryonic endothelium by unknown mechanisms. We discovered that Scl binds to hematopoietic and cardiac enhancers that become epigenetically primed in multipotent cardiovascular mesoderm, to regulate the divergence of hematopoietic and cardiac lineages. Scl does not act as a pioneer factor but rather exploits a pre-established epigenetic landscape. As the blood lineage emerges, Scl binding and active epigenetic modifications are sustained in hematopoietic enhancers, whereas cardiac enhancers are decommissioned by removal of active epigenetic marks. Our data suggest that, rather than recruiting corepressors to enhancers, Scl prevents ectopic cardiogenesis by occupying enhancers that cardiac factors, such as Gata4 and Hand1, use for gene activation. Although hematopoietic Gata factors bind with Scl to both activated and repressed genes, they are dispensable for cardiac repression, but necessary for activating genes that enable hematopoietic stem/progenitor cell development. These results suggest that a unique subset of enhancers in lineage-specific genes that are accessible for regulators of opposing fates during the time of the fate decision provide a platform where the divergence of mutually exclusive fates is orchestrated.

The purpose of this work is to characterize microchannels created by polymeric microneedles, applied by hand, and to demonstrate enhanced delivery of topically applied formulations of lidocaine hydrochloride and methylprednisolone sodium succinate (MPSS). 3M's Microstructured Transdermal System (MTS) arrays were applied to domestic swine to demonstrate reliability of penetration, depth of penetration and durability of the structures to repeat application and high force. Tissue levels of lidocaine and MPSS following topical application with and without microneedle pretreatment were determined by HPLC-MS analysis following digestion of biopsies. Almost all microneedles penetrate the stratum corneum upon hand force application. The depth of penetration varies from <100µm to nearly 150µm depending on the application force and the firmness of the underlying tissue. The arrays show excellent durability to repeated in-vivo application, with less than 5% of the structures evidencing even minimal tip bending after 16 applications. Under extreme force against a rigid surface, the microneedles bend but do not break. A lidocaine hydrochloride formulation applied topically in-vivo showed ~340% increase in local tissue levels when the MTS arrays were used to twice pre-treat the skin prior to applying the drug. Local delivery of a topically applied formulation of MPSS was over one order of magnitude higher when the application site was twice pre-treated with the MTS array. 3M's MTS array (marketed as 3M(TM) Microchannel Skin System) provides repeatable and robust penetration of the stratum corneum and epidermis and enhances delivery of some formulations such as lidocaine hydrochloride.

DNA double strand break (DSB) repair is critical for generation of B-cell receptors, which are pre-requisite for B-cell progenitor survival. However, the transcription factors that promote DSB repair in B cells are not known. Here we show that MEF2C enhances the expression of DNA repair and recombination factors in B-cell progenitors, promoting DSB repair, V(D)J recombination and cell survival. Although Mef2c-deficient mice maintain relatively intact peripheral B-lymphoid cellularity during homeostasis, they exhibit poor B-lymphoid recovery after sub-lethal irradiation and 5-fluorouracil injection. MEF2C binds active regulatory regions with high-chromatin accessibility in DNA repair and V(D)J genes in both mouse B-cell progenitors and human B lymphoblasts. Loss of Mef2c in pre-B cells reduces chromatin accessibility in multiple regulatory regions of the MEF2C-activated genes. MEF2C therefore protects B lymphopoiesis during stress by ensuring proper expression of genes that encode DNA repair and B-cell factors.

The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), is a widespread and destructive pest of cruciferous crops. Owing to its increasing resistance to conventional pesticides, new strategies need to be developed for diamondback moth control. Here, we investigated factors that modulate juvenile hormone esterase (JHE) activity and jhe (Px004817) transcription, and determined the effects of these factors on subsequent growth and development in diamondback moth. Starvation inhibited JHE activity and jhe transcription, increased mortality, and decreased the rate of molting from the third- to the fourth-instar stages. Larvae kept at 32°C molted earlier and showed increased JHE activity and jhe transcription after 24-h treatment. Exposure to 1,325 mg/liter OTFP (3-octylthio-1,1,1-trifluoro-2-propanone) delayed molting and pupation, increased pupal weight, and decreased JHE activity and jhe transcription at both 24 and 48 h. Treatment with 500 mg/liter pyriproxyfen delayed molting, completely suppressed pupation, and significantly increased JHE activity at 48 h and jhe transcription at 24 and 48 h. A combination of OTFP (1,325 mg/liter) and pyriproxyfen (500 mg/liter) induced the highest mortality, delayed molting, completely suppressed pupation, and significantly increased JHE activity at 48 h and jhe transcription at 24 and 48 h. These effects on JHE activity and jhe transcription were similar to those in insects treated only with pyriproxyfen. The results demonstrated that JHE and jhe (Px004817) were involved in the responses of diamondback moth to external modulators and caused changes in growth and development. The combination of OTFP and pyriproxyfen increased the effectiveness of action against diamondback moth.

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Recent advances in high throughput screening technologies have accelerated the identification and characterization of potential factors involved in host-virus interactions, facilitating early detection and diagnosis of diseases, as well as providing promising drug targets. The last decade has seen a plethora of successful examples of high throughput screening approaches, especially siRNA screening. With support from protein interaction studies, mRNA expression profiling, and bioinformatics, siRNA screening has also been successfully utilized to identify host factors required for a number of viruses including HIV, West Nile virus and H1N1 virus. Such studies have raised the awareness of virologists, and have opened a new chapter of global analysis of host-pathogen interactions. However, to play a more defining role in prognostics, diagnostics and therapeutics for virus diseases, acknowledged drawbacks, including false positives and negatives, inherent in this technology, must be successfully addressed.

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