TGF-ß Signaling Plays an Essential Role in the Lineage Specification of Mesenchymal Stem/Progenitor Cells in Fetal Bone Marrow.

Mesenchymal stromal cells are key components of hematopoietic niches in the bone marrow. Here we abrogated transforming growth factor ß (TGF-ß) signaling in mesenchymal stem/progenitor cells (MSPCs) by deleting Tgfbr2 in mesenchymal cells using a doxycycline-repressible Sp7 (osterix)-Cre transgene. We show that loss of TGF-ß signaling during fetal development results in a marked expansion of CXCL12-abundant reticular (CAR) cells and adipocytes in the bone marrow, while osteoblasts are significantly reduced. These stromal alterations are associated with significant defects in hematopoiesis, including a shift from lymphopoiesis to myelopoiesis. However, hematopoietic stem cell function is preserved. Interestingly, TGF-ß signaling is dispensable for the maintenance of mesenchymal cells in the bone marrow after birth under steady-state conditions. Collectively, these data show that TGF-ß plays an essential role in the lineage specification of fetal but not definitive MSPCs and is required for the establishment of normal hematopoietic niches in fetal and perinatal bone marrow.

Bone marrow dendritic cells regulate hematopoietic stem/progenitor cell trafficking.

A resident population of dendritic cells (DCs) has been identified in murine bone marrow, but its contribution to the regulation of hematopoiesis and establishment of the stem cell niche is largely unknown. Here, we show that murine bone marrow DCs are perivascular and have a type 2 conventional DC (cDC2) immunophenotype. RNA expression analysis of sorted bone marrow DCs shows that expression of many chemokines and chemokine receptors is distinct from that observed in splenic cDC2s, suggesting that bone marrow DCs may represent a unique DC population. A similar population of DCs is present in human bone marrow. Ablation of conventional DCs (cDCs) results in hematopoietic stem/progenitor cell (HSPC) mobilization that is greater than that seen with ablation of bone marrow macrophages, and cDC ablation also synergizes with G-CSF to mobilize HSPCs. Ablation of cDCs is associated with an expansion of bone marrow endothelial cells and increased vascular permeability. CXCR2 expression in sinusoidal endothelial cells and the expression of two CXCR2 ligands, CXCL1 and CXCL2, in the bone marrow are markedly increased following cDC ablation. Treatment of endothelial cells in vitro with CXCL1 induces increased vascular permeability and HSPC transmigration. Finally, we show that HSPC mobilization after cDC ablation is attenuated in mice lacking CXCR2 expression. Collectively, these data suggest that bone marrow DCs play an important role in regulating HSPC trafficking, in part, through regulation of sinusoidal CXCR2 signaling and vascular permeability.

Linear IgA Bullous Dermatosis Preceding the Diagnosis of Primary Sclerosing Cholangitis and Ulcerative Colitis: A Case Report.

Linear IgA bullous dermatosis (LABD) is a rare autoimmune blistering disorder seen in the pediatric and adult populations that is often linked to a medication, infection, or underlying gastrointestinal, hepatobiliary, or autoimmune disease. In this study, we describe the case of a 23-year-old white man whose presentation and diagnosis of LABD ultimately led to the discovery of underlying primary sclerosing cholangitis (PSC) and ulcerative colitis (UC). His dermatitis resolved with topical steroids and dapsone, and he is undergoing systemic treatment for his UC and PSC. This exceptional case further validates the association between LABD with UC, strengthens that with PSC, and underscores the importance of alerting clinicians to consider conducting a systemic workup in addition to thorough medication history on making the diagnosis of LABD.

Osteoclasts are dispensable for hematopoietic progenitor mobilization by granulocyte colony-stimulating factor in mice.

The contribution of osteoclasts to hematopoietic stem/progenitor cell (HSPC) retention in the bone marrow is controversial. Studies of HSPC trafficking in osteoclast-deficient mice are limited by osteopetrosis. Here, we employed two non-osteopetrotic mouse models to assess the contribution of osteoclasts to basal and granulocyte colony-stimulating factor (G-CSF)-induced HSPC mobilization. We generated Rank(-/-) fetal liver chimeras using Csf3r(-/-) recipients to produce mice lacking G-CSF receptor expression in osteoclasts. Basal and G-CSF-induced HSPC mobilization was normal in these chimeras. We next acutely depleted osteoclasts in wild-type mice using the RANK ligand inhibitor osteoprotegerin. Marked suppression of osteoclasts was observed after a single injection of osteoprotegerin-Fc. Basal and G-CSF-induced HSPC mobilization in osteoprotegerin-Fc-treated mice was comparable to that in control mice. Together, these data indicate that osteoclasts are not required for the efficient retention of HSPCs in the bone marrow and are dispensable for HSPC mobilization by G-CSF.

Antitumorigenic potential of STAT3 alternative splicing modulation.

Signal transducer and activator of transcription 3 (STAT3) plays a central role in the activation of multiple oncogenic pathways. Splicing variant STAT3ß uses an alternative acceptor site within exon 23 that leads to a truncated isoform lacking the C-terminal transactivation domain. Depending on the context, STAT3ß can act as a dominant-negative regulator of transcription and promote apoptosis. We show that modified antisense oligonucleotides targeted to a splicing enhancer that regulates STAT3 exon 23 alternative splicing specifically promote a shift of expression from STAT3α to STAT3ß. Induction of endogenous STAT3ß leads to apoptosis and cell-cycle arrest in cell lines with persistent STAT3 tyrosine phosphorylation compared with total STAT3 knockdown obtained by forced splicing-dependent nonsense-mediated decay (FSD-NMD). Comparison of the molecular effects of splicing redirection to STAT3 knockdown reveals a unique STAT3ß signature, with a down-regulation of specific targets (including lens epithelium-derived growth factor, p300/CBP-associated factor, CyclinC, peroxisomal biogenesis factor 1, and STAT1ß) distinct from canonical STAT3 targets typically associated with total STAT3 knockdown. Furthermore, similar in vivo redirection of STAT3 alternative splicing leads to tumor regression in a xenograft cancer model, demonstrating how pharmacological manipulation of a single key splicing event can manifest powerful antitumorigenic properties and validating endogenous splicing reprogramming as an effective cancer therapeutic approach.

Cell cycle-regulated trafficking of human telomerase to telomeres.

Telomerase synthesizes telomeres at the ends of human chromosomes during S phase. The results presented here suggest that telomerase activity may be regulated by intranuclear trafficking of the key components of the enzyme in human cells. We examined the subcellular localization of endogenous human telomerase RNA (hTR) and telomerase reverse transcriptase (hTERT) in HeLa cervical carcinoma cells. Throughout most of the cell cycle, we found that the two essential components of telomerase accumulate at intranuclear sites separate from telomeres. However, during S phase, both hTR and hTERT are specifically recruited to subsets of telomeres. The localization of telomerase to telomeres is dynamic, peaking at mid-S phase. We also found complex associations of both hTR and hTERT with nucleoli and Cajal bodies during S phase, implicating both structures in the biogenesis and trafficking of telomerase. Our results mark the first observation of human telomerase at telomeres and provide a mechanism for the cell cycle-dependent regulation of telomere synthesis in human cells.