An X-linked Myh11-CreERT2 mouse line resulting from Y to X chromosome-translocation of the Cre allele.

The Myh11-CreERT2 mouse line (Cre+ ) has gained increasing application because of its high lineage specificity relative to other Cre drivers targeting smooth muscle cells (SMCs). This Cre allele, however, was initially inserted into the Y chromosome (X/YCre+ ), which excluded its application in female mice. Our group established a Cre+ colony from male ancestors. Surprisingly, genotype screening identified female carriers that stably transmitted the Cre allele to the following generations. Crossbreeding experiments revealed a pattern of X-linked inheritance for the transgene (k > 1000), indicating that these female carries acquired the Cre allele through a mechanism of Y to X chromosome translocation. Further characterization demonstrated that in hemizygous X/XCre+ mice Cre activity was restricted to a subset arterial SMCs, with Cre expression in arteries decreased by 50% compared to X/YCre+ mice. This mosaicism, however, diminished in homozygous XCre+ /XCre+ mice. In a model of aortic aneurysm induced by a SMC-specific Tgfbr1 deletion, the homozygous XCre+ /XCre+ Cre driver unmasked the aortic phenotype that is otherwise subclinical when driven by the hemizygous X/XCre+ Cre line. In conclusion, the Cre allele carried by this female mouse line is located on the X chromosome and subjected to X-inactivation. The homozygous XCre+ /XCre+ mice produce uniform Cre activity in arterial SMCs.

Trends and Predictors of Imaging Utilization by Modality within an Academic Health System's Active Patient Population.

RATIONALE AND OBJECTIVES: Evaluate trends and demographic predictors of imaging utilization at a university-affiliated health system. MATERIALS AND METHODS: In this single-institution retrospective study, per capita estimates of imaging utilization among patients active in the health system were computed by cross-referencing all clinical encounters (2004-2016) for 1,628,980 unique patients with a listing of 6,157,303 diagnostic radiology encounters. Time trends in imaging utilization and effects of gender, race/ethnicity, and age were assessed, with subgroup analyses performed by imaging modality. Utilization was analyzed as both a continuous and binary outcome variable. RESULTS: Over 13 years, total diagnostic exams rose 6.8% a year (285,947-622,196 exams per annum), while the active population size grew 7.0% a year (244,238-543,290 active patients per annum). Per capita utilization peaked in 2007 at 1.33 studies/patient/year before dropping to 1.06 from 2011 to 2015. Latest per capita utilization was 0.22 for computed tomography, 0.10 for MR, 0.20 for US, 0.03 for NM, 0.51 for radiography, and 0.07 for mammography. Over the study period, ultrasound utilization doubled, whereas NM and radiography utilization decreased. computed tomography, MR, and mammography showed no significant net change. Univariate analysis of utilization as a continuous variable showed statistically significant effects of gender, race/ethnicity, and age (P < 0.0001), with utilization higher in males and Blacks and lower in Asian/Pacific Islanders and Hispanics. Utilization increased with age, except for a decline after age 75. Many of the effects of age, gender, and race/ethnicity were also found when analyzing the binarized utilization variable. CONCLUSIONS: Although absolute counts of imaging studies more than doubled, the net change in per capita utilization over the study period was minimal. Variations in utilization across age, gender, and race/ethnicity may reflect differential health needs and/or access disparities, warranting future studies.

Smooth muscle cell-specific Tgfbr1 deficiency attenuates neointimal hyperplasia but promotes an undesired vascular phenotype for injured arteries.

Neointimal hyperplasia (NIH) and inward wall remodeling cause arterial restenosis and failure of bypass vein grafts. Previous studies from our group suggest that transforming growth factor (TGF) ß promotes these pathologies via regulating cell kinetics at the early stage and matrix metabolism at the late stage. Although these temporal TGFß effects may result from its signaling in different cell groups, the responsible cell type has not been identified. In the current study, we evaluated the effect of smooth muscle cell (SMC)-specific TGFß signaling through its type I receptor TGFBR1 on NIH and wall remodeling of the injured femoral arteries (FAs). An inducible Cre/loxP system was employed to delete SMC Tgfbr1 (Tgfbr1iko). Mice not carrying the Cre allele (Tgfbr1f/f) served as controls. The injured FAs were evaluated on d3, d7, and d28 postoperatively. Tgfbr1iko attenuated NIH by 92%, but had insignificant influence on arterial caliber when compared with Tgfbr1f/f controls on d28. This attenuation correlated with greater cellularity and reduced collagen content. Compared with Tgfbr1f/f FAs, however, Tgfbr1iko FAs exhibited persistent neointimal cell proliferation and cell apoptosis, with both events at a greater rate on d28. Tgfbr1iko FAs additionally contained fewer SMCs and more inflammatory infiltrates in the neointima and displayed a thicker adventitia than did Tgfbr1f/f FAs. More MMP9 proteins were detected in the adventitia of Tgfbr1iko FAs than in that of Tgfbr1f/f controls. Our results suggest that disruption of SMC Tgfbr1 inhibits arterial NIH in the short term, but the overall vascular phenotype may not favor long-term performance of the injured arteries.

Crosslinking of collagen scaffolds promotes blood and lymphatic vascular stability.

The low stiffness of reconstituted collagen hydrogels has limited their use as scaffolds for engineering implantable tissues. Although chemical crosslinking has been used to stiffen collagen and protect it against enzymatic degradation in vivo, it remains unclear how crosslinking alters the vascularization of collagen hydrogels. In this study, we examine how the crosslinking agents genipin and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide alter vascular stability and function in microfluidic type I collagen gels in vitro. Under moderate perfusion (∼10 dyn/cm(2) shear stress), tubes of blood endothelial cells (ECs) exhibited indistinguishable stability and barrier function in untreated and crosslinked scaffolds. Surprisingly, under low perfusion (∼5 dyn/cm(2) shear stress) or nearly zero transmural pressure, microvessels in crosslinked scaffolds remained stable, while those in untreated gels rapidly delaminated and became poorly perfused. Similarly, tubes of lymphatic ECs under intermittent flow were more stable in crosslinked gels than in untreated ones. These effects correlated well with the degree of mechanical stiffening, as predicted by analysis of fracture energies at the cell-scaffold interface. This work demonstrates that crosslinking of collagen scaffolds does not hinder normal EC physiology; instead, crosslinked scaffolds promote vascular stability. Thus, routine crosslinking of scaffolds may assist in vascularization of engineered tissues.

Artificial lymphatic drainage systems for vascularized microfluidic scaffolds.

The formation of a stably perfused microvasculature continues to be a major challenge in tissue engineering. Previous work has suggested the importance of a sufficiently large transmural pressure in maintaining vascular stability and perfusion. Here we show that a system of empty channels that provides a drainage function analogous to that of lymphatic microvasculature in vivo can stabilize vascular adhesion and maintain perfusion rate in dense, hydraulically resistive fibrin scaffolds in vitro. In the absence of drainage, endothelial delamination increased as scaffold density increased from 6 to 30 mg/mL and scaffold hydraulic conductivity decreased by a factor of 20. Single drainage channels exerted only localized vascular stabilization, the extent of which depended on the distance between vessel and drainage as well as scaffold density. Computational modeling of these experiments yielded an estimate of 0.40-1.36 cm H2O for the minimum transmural pressure required for vascular stability. We further designed and constructed fibrin patches (0.8 × 0.9 cm(2)) that were perfused by a parallel array of vessels and drained by an orthogonal array of drainage channels; only with the drainage did the vessels display long-term stability and perfusion. This work underscores the importance of drainage in vascularization, especially when a dense, hydraulically resistive scaffold is used.