Following fibroblast lineages in dermal development and scars

Fibroblast heterogeneity studies have shown that it vastly influences the outcome of wound repair (Griffin et al., 2020; Rinkevich et al., 2015). We aimed to characterize the origins of these heterogeneous populations in the publication I. Here we have shown that during dermal development there is an inherent inversion of fibroblast populations from abundant regenerative fibroblasts (ENFs) at early fetal stages of development, to abundant scar-producing fibroblasts (EPFs) during perinatal and adult life. Using novel imaging and analysis approaches, we have charted the dermal maturation dynamics of EPFs during the transition from scar-less (E12) to scarring stages (E16.5) of development. We then followed up on the role of scar-forming fibroblasts in postnatal and adult stages. In publication III, we identified the subcutaneous fascia as the main anatomical contributor of scars upon deep skin injury. Next, we followed the role of fascial scar producing cells, EPFs, and its contribution to scar formation in publication II. Here, we developed a relevant ex-vivo model called “scar-like tissue in a dish”- termed SCAD. We show that scars on SCADs emulate the bona fide in-vivo scar phenotype. Using this model, we visualize and chart live migration dynamics of EPFs at all stages of scar development. Further, using antibody screening and CRISPR-Cas9 based genetic approaches, we identified that N-Cadherin is the adhesion molecular that orchestrates EPF and fascial response to scarring. Finally, to check the clinical relevance, we validated our N-cadherin mechanistic findings in human skin biopsies from various anatomical locations. These findings provide a range of therapeutic avenues in modulating subcutaneous fascial response and prevention of pathological scars

AAV-mediated gene delivery in the cutaneous wound healing

Cutaneous wound healing is a complex physiological process. Abnormal skin damage repair such as chronic wounds caused by diabetes, or excessive scar formation places a huge burden on the patient and society. AAV is widely used in biological research and is an ideal tool for human gene therapy. But AAV is rarely used in the research of cutaneous wound repair due to several technical issues. Here, we identified that sodium butyrate, a histone deacetylase, can greatly enhance the viral transduction of AAV6 serotypes in the wound healing model – SCAD (scar in a dish). We also optimized the transgenic plasmid using 2A self-cleavage peptide instead of the flexible linker, achieving robust co-expression of GFP and our target gene. Using the optimized system, we investigated the function of Akr1c18 in cutaneous wound healing. Akr1c18 is a steroid dehydrogenase and is mainly involved in the progesterone signal pathway. We found that Akr1c18 overexpression could slightly affect the fibroblast migration in the SCAD model and Akr1c18 overexpressed fibroblast have longer cell shape. However, the overall effect of Akr1c18 overexpression in wound healing is limited. Single-cell RNA analysis showed some elevated levels of progesterone response genes in mature wound fibroblast having reduced Akr1c18 expression. Based on the function of Akr1c18 as a progesterone inactivator, reduced expression of Akr1c18 in mature wound fibroblasts may lead to elevated progesterone levels in the skin, leading to increased progesterone responsive gene expression in wound fibroblasts

Aeronautical Engineering. A continuing bibliography with indexes, supplement 156

This bibliography lists 288 reports, articles and other documents introduced into the NASA scientific and technical information system in December 1982

Connexin43 gap junction drives fascia matrix mobilization and repair of deep skin wounds

Tiefe und voluminöse Hautwunden werden repariert, indem Fibroblasten und extrazelluläre Matrix aus der Faszie tief unter der Haut austreten und die verletzten Oberflächen mit Narben verschließen. Der molekulare Auslöser dieses neuartigen Reparaturmechanismus ist nur unvollständig verstanden. Hier zeigen wir, dass Cx43 der Schlüssel zur Patch-Reparatur von tiefen Wunden ist. Durch die Kombination von Full-Thickness-Wundmodellen mit Fibroblastenlinien-spezifischen transgenen Linien zeigen wir, dass Cx43 ausschließlich in tiefen Wunden hochreguliert wird, und zwar in spezialisierten Fibroblasten der Faszie tief unter der Haut, die für die Narbenbildung verantwortlich sind. Mit Hilfe von Live-Imaging von Faszienfibroblasten und Fate Tracing der extrazellulären Matrix der Faszien zeigen wir, dass die Hemmung von Cx43 die Oszillationen des Kalziumspiegels in tiefen Faszienfibroblasten unterbricht und dass dies ihre kollektive Wanderung hemmt, die notwendig sind, um die Faszienmatrix in offene Wunden auszuschütten. Cx43 und die Zell-Zell-Kommunikation sind somit Vehikel der Matrixausschüttung und Narbenbildung und notwendig für die Patch-Reparatur von voluminösen Wunden. Diese Erkenntnisse haben weitreichende klinische Implikationen für die Behandlung von Fibrose, überschießender Narbenbildung und gestörter Wundheilung.Deep and voluminous skin wounds are repaired, by outpouring of fibroblasts and extracellular matrix from fascia, deep below the skin, plugging breached surfaces with scars. The molecular trigger of this novel repair mechanism is incompletely understood. Here we reveal that Cx43 is the key to patch repair of deep wounds. By combining full-thickness wound models with fibroblast lineage specific transgenic lines, we show Cx43 is upregulated exclusively in deep wounds, in specialised fibroblasts of the fascia deep beneath the skin that are responsible for scar formation. Using live imaging of fascia fibroblasts and fate tracing of the fascia extracellular matrix, we show that Cx43 inhibition disrupts calcium level oscillations in deep fascia fibroblasts and that this inhibits their collective migrations necessary to outpour fascia matrix into open wounds. Cx43 and cell-cell communication are thus vehicles of matrix outpouring and scar formation, and necessary for patch repair of voluminous wounds. These findings have broad clinical implications for treating fibrosis, aggravated scarring and impaired wound healing

Developing a New Mechanical Model for Swarm Development in Myxococcus xanthus & Establishing a Genotype-to-Phenotype Correlation Between Swarm Pattern Formation and Gene Homology

The primary goal of systems biologists is to understand the mechanics underlying complex, collective, self-organizing behaviors displayed by all living systems, from biofilm formation and wound healing to embryogenesis. Myxococcus xanthus is a soil bacterium used as a model organism to study biofilm self-organization. It has a relatively large genome and a complex life cycle that involves two distinct phases. M. xanthus cells can move on agar, and a few million cells will organize to form a predatory biofilm or “swarm” that grows and expands if placed on a nutrient-rich agar surface. If placed on a nutrient-poor agar surface, the same swarm will turn inward and contract, aggregating to form spore-filled multicellular fruiting bodies designed to survive periods of starvation. Extensive progress has been made in identifying genes and genetic pathways that regulate fruiting body formation. However, an accurate description of the dynamics that underlie the process of aggregation is still lacking, and there is still debate and disagreement on the subject. This dissertation provides some explanation regarding individual M. xanthus cell behavior during fruiting body formation, as well as the behavior of aggregates. In the first part of this work, we show that the transition from individual cells to the formation of multicellular aggregates can be controlled through relatively small changes in M. xanthus cell behavior; complicated cell-to-cell signaling, stigmergy (where a trace formed by a cell on an agar surface influences the movement of nearby cells that contact the trace), and cell differentiation are not required for aggregation. We propose that M. xanthus aggregation matches a physical phenomenon that has been characterized in non-living systems, called motility induced phase separation (MIPS). By studying non-reversing mutant cells and manipulating their velocities, we show that cell movement can be made to fall within the boundary of the phase region so that cells succeed in forming aggregates. Alternately, cell movement can be made to fall outside the boundary of the phase region so that cells fail to form aggregates. After the initial stages of aggregation, an M. xanthus swarm actively rearranges the number and relative positions of aggregates by causing some of them to move, merge, or disappear. In the second part of this work, we demonstrate that equations describing Ostwald ripening within a thin liquid film were able to predict aggregate behavior with high accuracy. Consistent with this theory, both relative aggregate size and the distance between aggregates influence the likelihood that a given aggregate will disappear. In general, in neighboring aggregates, the ones that are small and in close proximity will tend to shrink and disappear, while the larger, more isolated aggregates will likely persist and become permanent. By tracking individual cells around aggregates, we show that more cells are leaving shrinking, disappearing aggregates than entering them, while more cells are entering growing, persistent aggregates than leaving them. All of these data are in good agreement with the Ostwald ripening equations. In the last part of this work, we show that aggregation can break down in only a certain number of ways by analyzing single gene disruption of four paralogous gene families representing almost 400 genes. We found that gene families correlate with phenotype, suggesting possible redundancy. In conclusion, my dissertation data provides possible answers to some of the persistent questions regarding M. xanthus developmental dynamics

Animal people

Animal People is an ethnography of contemporary animal-centric subcultures including furry fans, zoophiles, exotic animal keepers, trophy hunters, taxidermists, horse show competitors, and pit bull advocates, inviting the reader into their hidden worlds, their desires, their culture, activities, and gatherings. These essays examine the impacts of these subcultures on individual people and animals as well as the human and animal worlds, exploring how animals function as vehicles for human desires and why humans are drawn to extremes in our relationships with them. Why do some people want to kill a lion and others want to be a lion? Why do some people want a pet lion and others want to have sex with a lion? What makes people respond to the same animal in such vastly different ways? Sociologists have largely overlooked animal-related subcultures, but there’s a lot to be to learned from knowing them. The nine subcultures described in these chapters represent the enormous variety, breadth, and depth of human relationships to animals. They have taken their relationships with animals to the edge

To use or not to use? influences of list presentation format and working memory capacity on older adults' semantic clustering

The goal of the present study was to examine effects of list presentation format (study list presented as a whole vs. words presented briefly individually) on younger and older adults' semantic clustering of study words. Spontaneous clustering use did not differ between format conditions in either age group. Older adults spontaneously clustered to a similar extent as younger adults, evidencing no production deficiency. When clustering use was instructed, the whole-list format conditions clustered more successfully, resulting in greater recall than in the individual-words conditions, even under dual-task demands. Older adults clustered less successfully than younger adults, evidencing a utilization deficiency, with no overall recall improvements after clustering instructions in the individual-words format. Clustering interfered with performance on a simple tone-discrimination task, indicating its general cognitive resource demands; absolute interference was greater for older adults. Working memory capacity (WMC) predicted clustering success and mediated age-related reductions therein when clustering use was instructed but not for spontaneous use. WMC-clustering correlations were similar across presentation formats but adjusted means differed such that individuals at the same level of WMC clustered more successfully in the whole-list format. Beliefs about clustering difficulty correlated with its spontaneous use but did not evidence metacognitive awareness of presentation-format effects in either age group. These results suggest that a simple change in presentation format can facilitate encoding strategy use, particularly for older adults, but these benefits do not necessarily translate into spontaneous use differences. Thereby, presentation format alone cannot explain mixed findings regarding age-related differences in spontaneous clustering

The coastal environmental profile of Brunei Darussalam: resource assessment and management issues

Coastal zone management, Resource development, Coastal zone, Environmental profile, Brunei Darussalam, Resource /Energy Economics and Policy,

The coastal environmental profile of Brunei Darussalam: resource assessment and management issues

Coastal zone management, Resource development, Coastal zone, Environmental profile, Brunei Darussalam,

Acoustic Standing Wave Manipulation of Particles and Cells in Microfluidic Chips

The rise of MEMS and µTAS techniques has created a whole new family of microfluidic devices for a wide range of chemical and biomedical analyses to be performed on small Lab-on-a-chip platforms. The operations often include small samples of particle or cell suspensions on which separation, mixing, trapping or sorting is performed. External fields and forces are used for these operations, and this thesis is specifically focused the development of ultrasonic standing wave technology and the use of acoustic force fields to perform bioanalytical unit operations. The combination of acoustic standing waves and the laminar flow in microfluidics has proven to be well suited for performing particle and cell separation. The fundamental acoustic separator used in this thesis consists of a microfluidic flow channel with a three way flow splitter (trifurcation) in the end of the channel. An acoustic standing wave field is applied to the main flow channel by attaching the transducer underneath the chip. The acoustic standing wave is however obtained perpendicular to the axial propagation of the wave field and the direction of the flow. The half wavelength resonance affects rigid particles or cells driving them into the acoustic pressure node while liquid spheres having other density and compressibility properties may move to the pressure antinode. This enables acoustic separation of different particle types. Blood has proven to be very suitable for acoustic cell manipulation. An application where lipid particles can be removed acoustically from shed blood from open heart surgery is demonstrated. An application for acoustic plasmapheresis is also shown where high quality blood plasma is generated. Different separator designs, device material, and the influence of the separation channel cross-section design are also investigated