Perfusion Changes by Hyperspectral Imaging in a Burn Model

BACKGROUND: Early excision and skin grafting of full-thickness and deep-dermal burns is therapeutically advantageous. However, while full-thickness burns are clinically evident, differentiating between superficial versus deep partial-thickness burns presents a diagnostic challenge, with only 50-75% accuracy. Superficial-dermal burns heal, while deep-dermal burns often require excision and skin grafting. Decision of surgical treatment is often delayed until burn depth is definitively identified. This study’s aim is to establish a thermal burn model in mice in order to assess the ability of Hyperspectral Imaging (HSI) in differentiating burn depth. METHODS: Burns of graded severity were generated on the dorsum of seventy-six hairless mice with a brass rod heated to 50, 60, 70, 80, or 90°C. Perfusion and oxygenation parameters of the injured skin were measured with HSI, a non-invasive method of wide-field, diffuse reflectance spectroscopy at 2 minutes, 1 hour, 24 hours, 48 hours, and 72 hours after wounding. Burn depth was measured histologically (n=44) at 72 hours post injury using Masson’s trichrome staining. RESULTS: Three discrete levels of burn depth were verified histologically, as follows in order of increasing depth: intermediate-dermal, deep-dermal, and full-thickness injury. At 24 hours post injury, total hemoglobin increased by 67% and 18% in intermediate and deep dermal burns, respectively. In contrast, total hemoglobin decreased by 64% in full-thickness burns. Differences in deoxygenated hemoglobin, total hemoglobin, and oxygen saturation for all group comparisons were statistically significant (p CONCLUSION: HSI was able to differentiate among three discrete levels of burn injury. This is likely due to its correlation with skin perfusion: superficial burn injury causes an inflammatory response and increased perfusion to the burn site, while deeper burns destroy the dermal microvasculature and a decrease in perfusion follows. This study supports further investigation in the use of HSI in early burn depth assessment

Translational Model for External Volume Expansion in Irradiated Skin

Introduction: External Volume Expansion (EVE) treatment has gained popularity in breast reconstruction, enriching recipient sites for fat grafting. For patients receiving radiotherapy (XRT), results of EVE use vary, partly because the effects of EVE on irradiated tissue are not well understood. Based on our previous work with EVE and XRT, we developed a new translational model to investigate the effects of EVE in the setting of chronic radiation skin injury. Methods: Twenty-Eight SKH1-E mice received 50Gy of beta-radiation to each flank. Animals were monitored until chronic radiation fibrosis developed (8 weeks). EVE was then applied to one side for 6hrs on 5 consecutive days. The opposite side served as control. Hyperspectral Imaging (HSI) was used to assess perfusion changes before and after EVE. Mice were sacrificed at 5 days (n=14) and 15 days (n=14) after last application for histological analysis. Tissue samples were stained for vascularity (CD31) and collagen composition (Picro-Sirius red). Results: All animals developed skin fibrosis 8 weeks post-radiation, and changes in perfusion verified skin damage. EVE application induced edema on treated sides. Five days post-application, both sides were hypo-perfused as seen by HSI; with the EVE side 13% more ischemic than the untreated side (p\u3c0.001). Perfusion returned to control side levels by day 15. Blood vessels increased 20% by day 5 in EVE versus control. Collagen composition showed no difference in scar index analysis. Conclusion: EVE temporarily augments radiation-induced hypo-perfusion, likely due to transient edema. Fibrosis remained unchanged after EVE, possibly accounting for the limited expansion seen in patients. It appears that EVE induces angiogenic effect but does not affect dermal collagen composition. Future efforts should focus on reducing fibrosis post radiation to allow EVE to achieve its full potential, to benefit irradiated patients

Exploring hypotheses of the actions of TGF-beta 1 in epidermal wound healing using a 3D computational multiscale model of the human epidermis

In vivo and in vitro studies give a paradoxical picture of the actions of the key regulatory factor TGF-beta 1 in epidermal wound healing with it stimulating migration of keratinocytes but also inhibiting their proliferation. To try to reconcile these into an easily visualized 3D model of wound healing amenable for experimentation by cell biologists, a multiscale model of the formation of a 3D skin epithelium was established with TGF-beta 1 literature-derived rule sets and equations embedded within it. At the cellular level, an agent-based bottom-up model that focuses on individual interacting units ( keratinocytes) was used. This was based on literature-derived rules governing keratinocyte behavior and keratinocyte/ECM interactions. The selection of these rule sets is described in detail in this paper. The agent-based model was then linked with a subcellular model of TGF-beta 1 production and its action on keratinocytes simulated with a complex pathway simulator. This multiscale model can be run at a cellular level only or at a combined cellular/subcellular level. It was then initially challenged ( by wounding) to investigate the behavior of keratinocytes in wound healing at the cellular level. To investigate the possible actions of TGF-beta 1, several hypotheses were then explored by deliberately manipulating some of these rule sets at subcellular levels. This exercise readily eliminated some hypotheses and identified a sequence of spatial-temporal actions of TGF-beta 1 for normal successful wound healing in an easy-to-follow 3D model. We suggest this multiscale model offers a valuable, easy-to-visualize aid to our understanding of the actions of this key regulator in wound healing, and provides a model that can now be used to explore pathologies of wound healing

The effect of the swietenia mahagoni seed extracts on the production of collagen in human fibroblast cell (HSF1184)

Abstract:We have used the in vitro sircol collagen test to investigate the effects of Swietenia mahagoni seed extract on collagen production in human fibroblast cells (HSF1184). The results showed that Swietenia mahagoni seed extract with concentrations of 0.001, 0.01 and 0.1 mg / mL caused stimulation in the synthesis of type I collagen in HSF1184 cells. Although, three concentration of SC-CO2 and soxhlet that showed no statistically significant of collagen on the HSF1184 when compared with the negative control, while LA (fibroblast cells with pure Linoleic acid as supplement in culture medium) gave significant effect on the collagen production

Contractility Dominates Adhesive Ligand Density in Regulating Cellular De-adhesion and Retraction Kinetics

Cells that are enzymatically detached from a solid substrate rapidly round up as the tensile prestress in the cytoskeleton is suddenly unopposed by cell–ECM adhesions. We recently showed that this retraction follows sigmoidal kinetics with time constants that correlate closely with cortical stiffness values. This raises the promising prospect that these de-adhesion measurements may be used for high-throughput screening of cell mechanical properties; however, an important limitation to doing so is the possibility that the retraction kinetics may also be influenced and potentially rate-limited by the time needed to sever matrix adhesions. In this study, we address this open question by separating contributions of contractility and adhesion to cellular de-adhesion and retraction kinetics. We first develop serum-free conditions under which U373 MG glioma cells can be cultured on substrates of fixed fibronectin density without direct matrix contributions from the medium. We show that while spreading area increases with ECM protein density, cortical stiffness and the time constants of retraction do not. Conversely, addition of lysophosphatidic acid (LPA) to stimulate cell contractility strongly speeds retraction, independent of the initial matrix protein density and LPA’s contributions to spreading area. All of these trends hold in serum-rich medium commonly used in tissue culture, with the time constants of retraction much more closely tracking cortical stiffness than adhesive ligand density or cell spreading. These results support the use of cellular de-adhesion measurements to track cellular mechanical properties

Development of a Three Dimensional Multiscale Computational Model of the Human Epidermis

Transforming Growth Factor (TGF-β1) is a member of the TGF-beta superfamily ligand-receptor network. and plays a crucial role in tissue regeneration. The extensive in vitro and in vivo experimental literature describing its actions nevertheless describe an apparent paradox in that during re-epithelialisation it acts as proliferation inhibitor for keratinocytes. The majority of biological models focus on certain aspects of TGF-β1 behaviour and no one model provides a comprehensive story of this regulatory factor's action. Accordingly our aim was to develop a computational model to act as a complementary approach to improve our understanding of TGF-β1. In our previous study, an agent-based model of keratinocyte colony formation in 2D culture was developed. In this study this model was extensively developed into a three dimensional multiscale model of the human epidermis which is comprised of three interacting and integrated layers: (1) an agent-based model which captures the biological rules governing the cells in the human epidermis at the cellular level and includes the rules for injury induced emergent behaviours, (2) a COmplex PAthway SImulator (COPASI) model which simulates the expression and signalling of TGF-β1 at the sub-cellular level and (3) a mechanical layer embodied by a numerical physical solver responsible for resolving the forces exerted between cells at the multi-cellular level. The integrated model was initially validated by using it to grow a piece of virtual epidermis in 3D and comparing the in virtuo simulations of keratinocyte behaviour and of TGF-β1 signalling with the extensive research literature describing this key regulatory protein. This research reinforces the idea that computational modelling can be an effective additional tool to aid our understanding of complex systems. In the accompanying paper the model is used to explore hypotheses of the functions of TGF-β1 at the cellular and subcellular level on different keratinocyte populations during epidermal wound healing

A review of the current treatment methods for posthaemorrhagic hydrocephalus of infants

Posthaemorrhagic hydrocephalus (PHH) is a major problem for premature infants, generally requiring lifelong care. It results from small blood clots inducing scarring within CSF channels impeding CSF circulation. Transforming growth factor – beta is released into CSF and cytokines stimulate deposition of extracellular matrix proteins which potentially obstruct CSF pathways. Prolonged raised pressures and free radical damage incur poor neurodevelopmental outcomes. The most common treatment involves permanent ventricular shunting with all its risks and consequences

Identification of ejaculated proteins in the house mouse (Mus domesticus) via isotopic labeling

<p>Abstract</p> <p>Background</p> <p>Seminal fluid plays an important role in successful fertilization, but knowledge of the full suite of proteins transferred from males to females during copulation is incomplete. The list of ejaculated proteins remains particularly scant in one of the best-studied mammalian systems, the house mouse (<it>Mus domesticus</it>), where artificial ejaculation techniques have proven inadequate. Here we investigate an alternative method for identifying ejaculated proteins, by isotopically labeling females with ¹⁵N and then mating them to unlabeled, vasectomized males. Proteins were then isolated from mated females and identified using mass spectrometry. In addition to gaining insights into possible functions and fates of ejaculated proteins, our study serves as proof of concept that isotopic labeling is a powerful means to study reproductive proteins.</p> <p>Results</p> <p>We identified 69 male-derived proteins from the female reproductive tract following copulation. More than a third of all spectra detected mapped to just seven genes known to be structurally important in the formation of the copulatory plug, a hard coagulum that forms shortly after mating. Seminal fluid is significantly enriched for proteins that function in protection from oxidative stress and endopeptidase inhibition. Females, on the other hand, produce endopeptidases in response to mating. The 69 ejaculated proteins evolve significantly more rapidly than other proteins that we previously identified directly from dissection of the male reproductive tract.</p> <p>Conclusion</p> <p>Our study attempts to comprehensively identify the proteins transferred from males to females during mating, expanding the application of isotopic labeling to mammalian reproductive genomics. This technique opens the way to the targeted monitoring of the fate of ejaculated proteins as they incubate in the female reproductive tract.</p

Peritoneal changes due to laparoscopic surgery

Item does not contain fulltextBACKGROUND: Laparoscopic surgery has been incorporated into common surgical practice. The peritoneum is an organ with various biologic functions that may be affected in different ways by laparoscopic and open techniques. Clinically, these alterations may be important in issues such as peritoneal metastasis and adhesion formation. METHODS: A literature search using the Pubmed and Cochrane databases identified articles focusing on the key issues of laparoscopy, peritoneum, inflammation, morphology, immunology, and fibrinolysis. Results : Laparoscopic surgery induces alterations in the peritoneal integrity and causes local acidosis, probably due to peritoneal hypoxia. The local immune system and inflammation are modulated by a pneumoperitoneum. Additionally, the peritoneal plasmin system is inhibited, leading to peritoneal hypofibrinolysis. CONCLUSION: Similar to open surgery, laparoscopic surgery affects both the integrity and biology of the peritoneum. These observations may have implications for various clinical conditions.1 januari 201