Recovery of altered diabetic myofibroblast heterogeneity and gene expression associated with CD301B+ macrophages

Diabetic wound healing is associated with impaired function and reduced numbers of myofibroblasts, a heterogeneous cell population with varying capacities to promote repair. To determine how diabetes alters myofibroblast composition, we performed flow cytometry and spatial tissue analysis of myofibroblast subsets throughout the healing process in diabetic (db/db) and control (db/+) mouse skin. We observed reduced numbers of profibrotic SCA1+; CD34+; CD26+ myofibroblasts in diabetic wounds five days after injury, with decreased expression of fibrosis-associated genes compared to myofibroblasts from db/+ mouse wounds. While the abundance of myofibroblasts remained reduced in db/db mouse wounds compared to controls, the altered myofibroblast heterogeneity and gene expression in diabetic mice was improved seven days after injury. The natural correction of myofibroblast composition and gene expression in db/db wound beds temporally corresponds with a macrophage phenotypic switch. Correlation analysis from individual wound beds revealed that wound healing in control mice is associated with CD206+ macrophages, while the rescued myofibroblast phenotypes in diabetic wounds are correlated with increased CD301b+ macrophage numbers. These data demonstrate how diabetes impacts specific subsets of myofibroblasts and indicate that signaling capable of rescuing impaired diabetic wound healing could be different from signals that regulate wound healing under nonpathological conditions

Characterization of Cre recombinase models for the study of adipose tissue

The study of adipose tissue in vivo has been significantly advanced through the use of genetic mouse models. While the aP2-CreBI and aP2-CreSalk lines have been widely used to target adipose tissue, the specificity of these lines for adipocytes has recently been questioned. Here we characterize Cre recombinase activity in multiple cell populations of the major adipose tissue depots of these and other Cre lines using the membrane-Tomato/membrane-GFP (mT/mG) dual fluorescent reporter. We find that the aP2-CreBI and aP2-CreSalk lines lack specificity for adipocytes within adipose tissue, and that the aP2-CreBI line does not efficiently target adipocytes in white adipose depots. Alternatively, the Adiponectin-CreERT line shows high efficiency and specificity for adipocytes, while the PdgfRα-CreERUCL and PdgfRα-CreERJHU lines do not efficiently target adipocyte precursor cells in the major adipose depots. Instead, we show that the PdgfRα-Cre line is preferable for studies targeting adipocyte precursor cells in vivo

DYX1C1 is required for axonemal dynein assembly and ciliary motility

DYX1C1 has been associated with dyslexia and neuronal migration in the developing neocortex. Unexpectedly, we found that deleting exons 2–4 of Dyx1c1 in mice caused a phenotype resembling primary ciliary dyskinesia (PCD), a disorder characterized by chronic airway disease, laterality defects and male infertility. This phenotype was confirmed independently in mice with a Dyx1c1 c.T2A start-codon mutation recovered from an N-ethyl-N-nitrosourea (ENU) mutagenesis screen. Morpholinos targeting dyx1c1 in zebrafish also caused laterality and ciliary motility defects. In humans, we identified recessive loss-of-function DYX1C1 mutations in 12 individuals with PCD. Ultrastructural and immunofluorescence analyses of DYX1C1-mutant motile cilia in mice and humans showed disruptions of outer and inner dynein arms (ODAs and IDAs, respectively). DYX1C1 localizes to the cytoplasm of respiratory epithelial cells, its interactome is enriched for molecular chaperones, and it interacts with the cytoplasmic ODA and IDA assembly factor DNAAF2 (KTU). Thus, we propose that DYX1C1 is a newly identified dynein axonemal assembly factor (DNAAF4)

The Design and Implementation of a Multi-Chip Power Module Layout Synthesis Tool

This thesis presents the design and implementation of a software tool, referred to as PowerSynth, which is used to synthesize multi-chip power module (MCPM) layouts and also aids in the overall design process. MCPMs allow power electronics systems to achieve higher efficiencies and reduced size, weight, and cost through the compact integration of devices into a single package. MCPM design is highly multidisciplinary which involves electrical, thermal, and mechanical performance considerations. Conventionally, the design process may require several iterations and multiple software modeling tools to create high performance MCPMs. The layout of an MCPM consists of a pattern of metal traces and the placement of components on these traces. The design of an MCPM layout and the selection of materials, dimensions, and components used in an MCPM affect its performance. PowerSynth incorporates thermal and electrical models into a single tool and provides a unified system for designing MCPMs and synthesizing layouts for them. A user is guided through the design process with graphical user interfaces and is able to select from a set of solutions which trade-off thermal and electrical performance. Final MCPM designs can be exported to two different commercial modeling tools for further performance verification or for manufacturing artwork generation

The Design and Implementation of a Multi-Chip Power Module Layout Synthesis Tool

This thesis presents the design and implementation of a software tool, referred to as PowerSynth, which is used to synthesize multi-chip power module (MCPM) layouts and also aids in the overall design process. MCPMs allow power electronics systems to achieve higher efficiencies and reduced size, weight, and cost through the compact integration of devices into a single package. MCPM design is highly multidisciplinary which involves electrical, thermal, and mechanical performance considerations. Conventionally, the design process may require several iterations and multiple software modeling tools to create high performance MCPMs. The layout of an MCPM consists of a pattern of metal traces and the placement of components on these traces. The design of an MCPM layout and the selection of materials, dimensions, and components used in an MCPM affect its performance. PowerSynth incorporates thermal and electrical models into a single tool and provides a unified system for designing MCPMs and synthesizing layouts for them. A user is guided through the design process with graphical user interfaces and is able to select from a set of solutions which trade-off thermal and electrical performance. Final MCPM designs can be exported to two different commercial modeling tools for further performance verification or for manufacturing artwork generation

Age-Related Changes to the Subventricular Zone Stem Cell Niche

Through adulthood the rodent subventricular zone (SVZ) stem cell niche generates new olfactory bulb interneurons. The SVZ stem cell niche is regulated by many components. In particular the ependymal monolayer that lines the ventricle surface is critical to maintain niche organization and function. The studies presented here examined changes to the ependymal monolayer and ventricle-contacting neural stem cells (NSCs) through aging. ^ While neurogenesis declines with age, specific changes to the NSC population have not been determined. Here, I conducted a spatio-temporal evaluation of adult SVZ NSCs, describing a decline in NSC numbers with age. Regional analysis along the lateral ventricle surface revealed that NSC loss is spatially uniform. Surprisingly, I found no significant change in the number of actively proliferating NSCs. Instead, my data reveal that although the total NSC number declines with age, the percentage of actively, mitotic NSCs increases, indicating that age-related declines in SVZ-mediated olfactory bulb neurogenesis occur downstream of NSC proliferation. ^ Following mild disruption of an intact ependyma in young adult mice, I have detected that proliferative cells from the SVZ generate new ependymal cells in an effort to maintain the integrity of the barrier. Additionally, extensive damage to the ependymal layer caused glial scarring at the ventricle surface and a subsequent deterioration of the underlying SVZ. ^ Comparative analysis of lateral ventricle volume and surface composition was performed using mouse and human tissue. I observed an intact, contiguous ependymal monolayer in mice throughout aging and mouse lateral ventricle size was maintained through aging. However, an age-related increase in human ventricle volume was observed. In addition, glial scarring at the ventricle surface was associated with the enlarged ventricles, indicating that humans typically do not maintain constant ventricle size or a contiguous ependymal barrier with age. Together this work contributes to our knowledge of age-related changes that occur at the ventricle surface in mice and humans and how these changes impact the underlying SVZ.

Dermal Drivers of Injury-Induced Inflammation: Contribution of Adipocytes and Fibroblasts

Irregular inflammatory responses are a major contributor to tissue dysfunction and inefficient repair. Skin has proven to be a powerful model to study mechanisms that regulate inflammation. In particular, skin wound healing is dependent on a rapid, robust immune response and subsequent dampening of inflammatory signaling. While injury-induced inflammation has historically been attributed to keratinocytes and immune cells, a vast body of evidence supports the ability of non-immune cells to coordinate inflammation in numerous tissues and diseases. In this review, we concentrate on the active participation of tissue-resident adipocytes and fibroblasts in pro-inflammatory signaling after injury, and how altered cellular communication from these cells can contribute to irregular inflammation associated with aberrant wound healing. Furthering our understanding of how tissue-resident mesenchymal cells contribute to inflammation will likely reveal new targets that can be manipulated to regulate inflammation and repair