White Fat Progenitor Cells Reside in the Adipose Vasculature

White adipose (fat) tissues regulate metabolism, reproduction, and life span. Adipocytes form throughout life, with the most marked expansion of the lineage occurring during the postnatal period. Adipocytes develop in coordination with the vasculature, but the identity and location of white adipocyte progenitor cells in vivo are unknown. We used genetically marked mice to isolate proliferating and renewing adipogenic progenitors. We found that most adipocytes descend from a pool of these proliferating progenitors that are already committed, either prenatally or early in postnatal life. These progenitors reside in the mural cell compartment of the adipose vasculature, but not in the vasculature of other tissues. Thus, the adipose vasculature appears to function as a progenitor niche and may provide signals for adipocyte development

LRP1 Regulates Architecture of the Vascular Wall by Controlling PDGFRβ-Dependent Phosphatidylinositol 3-Kinase Activation

Low density lipoprotein receptor-related protein 1 (LRP1) protects against atherosclerosis by regulating the activation of platelet-derived growth factor receptor beta (PDGFRbeta) in vascular smooth muscle cells (SMCs). Activated PDGFRbeta undergoes tyrosine phosphorylation and subsequently interacts with various signaling molecules, including phosphatidylinositol 3-kinase (PI3K), which binds to the phosphorylated tyrosine 739/750 residues in mice, and thus regulates actin polymerization and cell movement.In this study, we found disorganized actin in the form of membrane ruffling and enhanced cell migration in LRP1-deficient (LRP1-/-) SMCs. Marfan syndrome-like phenotypes such as tortuous aortas, disrupted elastic layers and abnormally activated transforming growth factor beta (TGFbeta) signaling are present in smooth muscle-specific LRP1 knockout (smLRP1-/-) mice. To investigate the role of LRP1-regulated PI3K activation by PDGFRbeta in atherogenesis, we generated a strain of smLRP1-/- mice in which tyrosine 739/750 of the PDGFRbeta had been mutated to phenylalanines (PDGFRbeta F2/F2). Spontaneous atherosclerosis was significantly reduced in the absence of hypercholesterolemia in these mice compared to smLRP1-/- animals that express wild type PDGFR. Normal actin organization was restored and spontaneous SMC migration as well as PDGF-BB-induced chemotaxis was dramatically reduced, despite continued overactivation of TGFbeta signaling, as indicated by high levels of nuclear phospho-Smad2.Our data suggest that LRP1 regulates actin organization and cell migration by controlling PDGFRbeta-dependent activation of PI3K. TGFbeta activation alone is not sufficient for the expression of the Marfan-like vascular phenotype. Thus, regulation of PI3 Kinase by PDGFRbeta is essential for maintaining vascular integrity, and for the prevention of atherosclerosis as well as Marfan syndrome

A Study of the PDGF Signaling Pathway with PRISM

In this paper, we apply the probabilistic model checker PRISM to the analysis of a biological system -- the Platelet-Derived Growth Factor (PDGF) signaling pathway, demonstrating in detail how this pathway can be analyzed in PRISM. We show that quantitative verification can yield a better understanding of the PDGF signaling pathway.Comment: In Proceedings CompMod 2011, arXiv:1109.104

Additive Effects of PDGF Receptor β Signaling Pathways in Vascular Smooth Muscle Cell Development

The platelet-derived growth factor β receptor (PDGFRβ) is known to activate many molecules involved in signal transduction and has been a paradigm for receptor tyrosine kinase signaling for many years. We have sought to determine the role of individual signaling components downstream of this receptor in vivo by analyzing an allelic series of tyrosine–phenylalanine mutations that prevent binding of specific signal transduction components. Here we show that the incidence of vascular smooth muscle cells/pericytes (v/p), a PDGFRβ-dependent cell type, can be correlated to the amount of receptor expressed and the number of activated signal transduction pathways. A decrease in either receptor expression levels or disruption of multiple downstream signaling pathways lead to a significant reduction in v/p. Conversely, loss of RasGAP binding leads to an increase in this same cell population, implicating a potential role for this effector in attenuating the PDGFRβ signal. The combined in vivo and biochemical data suggest that the summation of pathways associated with the PDGFRβ signal transduction determines the expansion of developing v/p cells

Structural Evidence of T Cell Xeno-reactivity in the Absence of Molecular Mimicry

The T cell receptor (TCR), from a xeno-reactive murine cytotoxic T lymphocyte clone AHIII12.2, recognizes murine H-2Db complexed with peptide p1027 (FAPGVFPYM), as well as human HLA-A2.1 complexed with peptide p1049 (ALWGFFPVL). A commonly proposed model (the molecular mimicry model) used to explain TCR cross-reactivity suggests that the molecular surfaces of the recognized complexes are similar in shape, charge, or both, in spite of the primary sequence differences. To examine the mechanism of xeno-reactivity of AHIII12.2, we have determined the crystal structures of A2/p1049 and Db/p1027 to 2.5 Å and 2.8 Å resolution, respectively. The crystal structures show that the TCR footprint regions of the two class I complexes are significantly different in shape and charge. We propose that rather than simple molecular mimicry, unpredictable arrays of common and differential contacts on the two class I complexes are used for their recognition by the same TCR

Overexpression of platelet-derived growth factor receptor α in breast cancer is associated with tumour progression

INTRODUCTION: Receptor tyrosine kinases have been extensively studied owing to their frequently abnormal activation in the development and progression of human cancers. Platelet-derived growth factor receptors (PDGFRs) are receptors with intrinsic tyrosine kinase activity that regulate several functions in normal cells and are widely expressed in a variety of malignancies. After the demonstration that gastrointestinal stromal tumours without c-Kit mutations harbour PDGFR-α-activating mutations and that PDGFR-α is also a therapeutic target for imatinib mesylate, the interest for this receptor has increased considerably. Because breast cancer is one of the most frequent neoplasias in women worldwide, and only one study has reported PDGFR-α expression in breast carcinomas, the aim of this work was to investigate the potential significance of PDGFR-α expression in invasive mammary carcinomas. METHODS: We used immunohistochemistry to detect PDGFR-α overexpression on a series of 181 formalin-fixed paraffin-embedded invasive ductal breast carcinomas and in two breast cancer cell lines: MCF-7 and HS578T. We associated its expression with known prognostic factors and we also performed polymerase chain reaction–single-stranded conformational polymorphism and direct sequencing to screen for PDGFR-α mutations. RESULTS: PDGFR-α expression was observed in 39.2% of the breast carcinomas and showed an association with lymph node metastasis (P = 0.0079), HER-2 expression (P = 0.0265) and Bcl2 expression (P = 0.0121). A correlation was also found with the expression of platelet-derived growth factor A (PDGF-A; P = 0.0194). The two cell lines tested did not express PDGFR-α. Screening for mutations revealed alterations in the PDGFR-α gene at the following locations: 2500A→G, 2529T→A and 2472C→T in exon 18 and 1701G→A in exon 12. We also found an intronic insertion IVS17-50insA at exon 18 in all sequenced cases. None of these genetic alterations was correlated with PDGFR-α expression. The cell lines did not reveal any alterations in the PDGFR-α gene sequence. CONCLUSION: PDGFR-α is expressed in invasive breast carcinomas and is associated with biological aggressiveness. The genetic alterations described were not correlated with protein expression, but other mechanisms such as gene amplification or constitutive activation of a signalling pathway inducing this receptor could still sustain PDGFR-α as a potential therapeutic target

Epiblastic Cited2 deficiency results in cardiac phenotypic heterogeneity and provides a mechanism for haploinsufficiency

AIMS: Deletion of the transcription factor Cited2 causes penetrant and phenotypically heterogenous cardiovascular and laterality defects and adrenal agenesis. Heterozygous human CITED2 mutation is associated with congenital heart disease, suggesting haploinsufficiency. Cited2 functions partly via a Nodal-->Pitx2c pathway controlling left-right patterning. In this present study we investigated the primary site of Cited2 function and mechanisms of haploinsufficiency. METHODS AND RESULTS: A Cited2 conditional allele enabled its deletion in particular cell lineages in mouse development. A lacZ reporter cassette allowed indication of deletion. Congenic Cited2 heterozygous mice were used to investigate haploinsufficiency. Embryos were examined by magnetic resonance imaging, by sectioning and by quantitative real-time polymerase chain reaction (qRT-PCR). Epiblast-specific deletion of Cited2 using Sox2Cre recapitulated penetrant and phenotypically heterogenous cardiovascular and laterality defects. Neural crest-specific deletion using Wnt1Cre affected cranial ganglia but not cardiac development. Mesodermal deletion with Mesp1Cre resulted in low penetrance of septal defect. Mesodermal deletion with T-Cre resulted in adrenal agenesis, but infrequent cardiac septal and laterality defects. beta-Galatactosidase staining and qRT-PCR demonstrated the efficiency and location of Cited2 deletion. Murine Cited2 heterozygosity is itself associated with cardiac malformation, with three of 45 embryos showing ventricular septal defect. Cited2 gene expression in E13.5 hearts was reduced 2.13-fold in Cited2(+/-) compared with wild-type (P = 2.62 x 10(-6)). The Cited2 target gene Pitx2c was reduced 1.5-fold in Cited2(+/-) (P = 0.038) hearts compared with wild-type, and reduced 4.9-fold in Cited2(-/-) hearts (P = 0.00031). Pitx2c levels were reduced two-fold (P = 0.009) in Cited2(+/-) embryos, in comparison with wild-type. Cited2 and Pitx2c expression were strongly correlated in wild-type and Cited2(+/-) hearts (Pearson rank correlation = 0.68, P = 0.0009). Cited2 expression was reduced 7474-fold in Sox2Cre deleted hearts compared with controls (P = 0.00017) and Pitx2c was reduced 3.1-fold (P = 0.013). Deletion of Cited2 with Mesp1Cre resulted in a 130-fold reduction in cardiac Cited2 expression compared with control (P = 0.0002), but Pitx2c expression was not affected. CONCLUSION: These results indicate that phenotypically heterogenous and penetrant cardiac malformations in Cited2 deficiency arise from a primary requirement in epiblast derivatives for left-right patterning, with a secondary cell-autonomous role in the mesoderm. Cardiac malformation associated with Cited2 haploinsufficiency may occur by reducing expression of key Cited2 targets such as Pitx2c