Growth factor delivery may be useful to accelerate the rate of tendon healing. We studied Platelet Concentrate, which in effect can be regarded as a cocktail of growth factors relevant for tendon healing. In a rat Achilles tendon transection model, one postoperative injection of Platelet Concentrate resulted in increased strength even 3 weeks later. Mechanical stimulation improves the repair of ruptured tendons. We studied the effects of platelets upon Achilles tendon regenerates in rats 3, 5 and 14 days after transection, either unloaded or mechanically stimulated. At 14 days, physical activity and platelets increased repair independently. Unloading decreased the mechanical properties of the repair tissue to less than half of normal. Moreover, the platelets had no effect without loading. Thrombin, which we used for platelet activation, improved healing of the rat Achilles tendon by itself. Conversely, continuous inhibition of thrombin by low molecular weight heparin (LMWH) inhibited tendon repair. However, intermittent inhibition, similar to clinical thromboprophylaxis, had no effect on tendon healing. Cartilage Derived Morphogenetic Protein-2 (CDMP-2) can improve tendon healing in loaded defect models. We now studied unloaded repair in a rabbit patellar tendon model. Two hours postoperative, the rabbits received CDMP-2 injected into the haematoma. The healing tendon became 65 % stronger than controls. We then studied Achilles tendon healing with CDMP-2 injections in sheep, to get a bigger animal model. There was an unexpectedly high variation of repair in these animals, and the study turned out to be underpowered. Spontaneous ruptures in humans have a more variable geometry than in our sheep model, so humans can also be expected to vary a lot in mechanical characteristics of Achilles tendon repair. This accentuates the importance of individualized rehabilitation programs. In conclusion, both platelet concentrate and CDMP-2 injections might be of interest for clinical use as a complement to surgical or conservative treatment of tendon ruptures. Platelet treatment for tendon ruptures should probably be combined with early physiotherapy.

Cutaneous reparative processes, including wound healing, are highly developed procedures in which a chain of actions occurs to reconstitute the function of the wounded tissue. To prevent a delayed or excessive reparative process it is important to understand how this procedure develops and is maintained. One of the major extracellular matrix components of the skin is the glycosaminoglycan hyaluronan (HA). HA contributes to an extracellular environment, which is permissive for cell motility and proliferation, features that may account for HA’s unique properties observed in scarless foetal wound healing. The molecule is found at high concentration whenever proliferation, regeneration and repair of tissue occur. The aims of the present studies were to analyse the distribution of HA and to investigate its possible role in various cutaneous conditions associated with an impaired reparative process like in scar tissue formation in healing wounds, changed skin characteristics in diabetes mellitus and proliferating activity in basal cell carcinomas. Tissue biopsies were obtained from healthy human skin, type-I diabetic skin and various scar tissues. The samples were analysed in the light microscope with a hyaluronan-binding-probe, antibodies for collagen I, III, PCNA and Ki-67. Ultrastructural analyses were performed on the same tissue samples. In normal skin HA was present mainly in the papillary dermis. In epidermis HA was located in between the keratinocytes in the spinous layer. In the different scar tissues the localization of HA varied, with an HA distribution in mature scar type resembling that in normal skin. In keloids the papillary dermis lacked HA, but the thickened epidermis contained more HA than the other scar types. Ultrastructural studies of keloids revealed an altered collagen structure in the dermal layers, with an abundance of thin collagen fibers in the reticular dermis and thicker collagen fibers in the papillary dermis. Furthermore, the keloids displayed epidermal changes, which involved the basement membrane (BM), exhibiting fewer hemidesmosomes, and an altered shape of desmosomes in the entire enlarged spinous layer. These alterations in epidermis are suggested to influence the hydrodynamic and cell regulatory properties of the wounded skin. In diabetic patients, a reduced HA staining in the basement membrane zone was seen. The staining intensity of HA correlated to the physical properties of the skin reflected by their grades of limited joint mobility (LJM). Furthermore, the HA staining correlated with serum concentration of the HbA1c. In basal cell carcinomas (BCC), HA occurred predominantly in the tumour stroma. The distribution was most intense in the highly developed superficial BCC type, and resembled that of the papillary dermis of normal skin. In contrast, in the infiltrative BCC type, the tumour stroma stained weakly in the infiltrative part of the tumour. Moreover, the surrounding dermal layer was deranged and devoid of HA. The findings suggest that the tumour stroma in superficial BCC causes a slow, well-regulated cell growth in which the tumour cells do not substantially disturb the normal skin function. In the infiltrative BCC type, the tumour cells cause a disintegration of the tumour stroma as well as the normal surrounding dermis, which permits further spreading of the tumour. In fact, the behaviour of the infiltrative BCC tumour, growing beyond its boundaries, resembles that of the keloid. The mapping of the distribution of HA could be a useful tool for prognostic information, for evaluating the degree of progress and for deciding the choice of treatment in various diseases of the skin. In skin malignancies such as BCC it can be used to determine the radicality at the surgical excision of the tumour. Keywords: Hyaluronan, scar tissue, diabetes mellitus, basal cell carcinoma, skin, wound healing

Angiogenesis, the formation of new blood vessels from preexisting ones through creation of new vessel branch points by sprouting or vessel splitting, is an important part of tissue growth in both physiological processes like wound healing and pathological conditions such as cancer. Growth factors like VEGF-A, FGF-2 and PDGF-BB are involved in both types of angiogenesis. Screening for genes regulated by VEGF-A stimulation in endothelial cells revealed up regulation of the endothelial cell specific glycoprotein endocan. Endocan itself did not stimulate angiogenesis. VEGF was a specific inducer since FGF-2, PDGF-BB, HGF and EGF did not alter expression. The signaling molecule PI3K was a negative regulator of endocan expression. Endocan was expressed in tumor cells and vessels, suggesting that although endocan did not directly regulate angiogenesis it can serve as a marker for angiogenic tumors. In two models of wound healing angiogenesis, the chick extra-embryonal CAM assay and the mouse cornea assay, we observed that blood vessels grew into avascular areas as functional mural cell covered loops by elongation of preexisting vessels. Loop formation was simultaneous with contraction of the avascular matrix mediated by proto/myofibroblasts. Reducing the contractibility of the stroma reduced vessel ingrowth, showing that contraction was necessary for mediating and directing growth of the vascular loops. These findings suggest a model for biomechanical regulation of vascularization that is complementary to sprouting angiogenesis which is guided by gradients of growth factors. In defining the role of growth factors, in the CAM assay, we found that FGF-2 and PDGF-BB induced vessel ingrowth, while VEGF-A, EGF and HGF did not. TGF-beta reduced the effect of FGF-2. By use of specific receptor kinase inhibitors we found an absolute requirement VEGF- and PDGF-receptor activity for vascularization while FGF- and TGF-beta-receptor function was dispensable. This suggests that functional VEGF- and PDGF-receptors are needed for vessel elongation.

Background: Vacuum assisted closure, VAC, is a method which can benefit wound closure. There are not many reliable studies about the effects of VAC in different types of wounds. Previous studies have shown that patients have experienced pain during dressing changes. Aim: The aim of the study was to survey in what kind of wounds the vascular surgery ward have been using VAC, to study the treatment process of VAC and how it has been documented and also to investigate if patients with VAC experienced pain during dressing changes. Method: The patients were chosen from a ledger, which contained 77 patients that had been treated with VAC on the vascular surgery ward. Data were collected with a journal survey protocol. Results: The result is based on 67 patients with a total of 79 wounds. VAC was shown to have a good effect on wound healing and infected surgical wounds were the most commonly wound category undergoing treatment with VAC. There was a lack of documentation concerning VAC and the number of wounds, which were not being measured or photographed, before, during and after VAC was 41, 44 and 56. The complications that occurred was bleeding, the suction cup came off or that the patient experienced great pain. The number of days the wound was treated with VAC differed from one to 70 days. The average of numbers of days treated with VAC was 18,5 days. The dressings were changed most commonly two to three times a week. In 18 wounds (23 %), the patients reported pain during dressing changes. Due to lack of documentation, it was impossible, in 45 cases (57 %) to see if the patient had experienced pain during dressing change. Conclusion: Despite the lack in the documentation about the VAC treatment the study shows good results in the majority of the wounds treated with VAC. Some patients experienced pain during dressing changes and therefore it is important to recognize, treat, evaluate and document this. / Bakgrund: Vacuumassisterad sårbehandling, VAC, är en metod vilken kan underlätta sårläkning. Det har utförts alldeles för få studier med hög kvalitet som har studerat effekterna av behandlingen på olika typer av sår. Tidigare studier har visat att patienter upplever smärta vid omläggning av VAC-svamp. Syfte: Studiens syften var att kartlägga vilka typer av sår den kärlkirurgiska avdelningen behandlar med VAC, studera behandlingsprocessen vid VAC och hur den dokumenteras samt undersöka om patienter upplever smärta i samband med byte av VAC-svamp. Metod: Patienterna valdes ut från en liggare på den kärlkirurgiska avdelningen vilken innehöll 77 patienter. Data samlades in med hjälp av ett journalgranskningsprotokoll. Resultat: Resultatet baseras på 67 patienter vilket innefattar 79 sår. VAC som behandlingsmetod visade sig ha bra effekt på sårläkning av de flesta sår och infekterade operationssår var den sårkategori som genomgick flest behandlingar med VAC. Dokumentationen om VAC var sparsam och de antal sår som varken var mätta eller fotograferade innan, under och efter behandling med VAC upp gick till 41 respektive 44 och 56 sår. De komplikationer som uppstod var blödning, att sugproppen lossnade eller att patienten upplevde stor smärta. Omläggning skedde vanligen två till tre gånger per vecka. Medelvärdet på antalet dagar såret behandlades med VAC uppgick till 18,5 dagar. Antalet dagar såret behandlades med VAC varierade från en till 70 dagar. I 18 fall (23 %) har patienten rapporterat smärta vid omläggning av VAC. I 45 fall (57 %) gick det inte att se om patienten hade känt någon smärta vid omläggning. Slutsats: Trots att dokumentationen om behandlingen var bristfällig på flera plan visar studien ett gott resultat av VAC-behandlingen i majoriteten av de sår som behandlades. Vissa patienter upplevde smärta vid omläggning och därför är det viktigt att uppmärksamma, behandla, utvärdera och dokumentera denna.

Bakgrund: Trycksår är vanligt förekommande inom vården. En koppling har gjorts mellan försämrad nutritionsstatus och ökad risk för trycksår. Ett sätt att påverka näringsstatusen kan vara att ge näringstillskott och genom detta indirekt påverka läkningen av trycksår, detta omvårdnadsansvar ligger hos sjuksköterskan. Ett problem kan dock vara att kunskapen hos sjuksköterskor gällande näringstillskott kan variera. Syfte: Syftet var att beskriva näringstillskotts effekt på läkningen av trycksår. Metod: Studien gjordes som en litteraturöversikt (n=10) baserade på kvantitativa artiklar. Resultat: Studiens resultat visade att de flesta näringstillskott hade positiv effekt på läkningen av trycksår. Tillskott i form av arginin, vitamin C och zink i kombination och hydrolyserat protein gav en förbättrad sårläkning. Tillskott i form av fiskolja visade ingen signifikant förbättring. Slutsats: Näringstillskott innehållande arginin, vitamin C och zink och i kombination har visat sig kunna påverka läkningsprocessen vid trycksår positivt. Studier visar dock att kunskapen hos sjuksköterskor gällande näringens roll i vården är bristande. Skulle detta bero på att informationen är svårförståelig, hoppas vi att den föreliggande studien kan bidra till ökat intresse och kunskap kring näringstillskotts effekt på läkningen av trycksår. / Background: Pressure ulcers are common in health care. A connection has been made between poor nutritional status and pressure ulcers. One way of affecting the nutritional status could be by giving nutritional supplements and by this affect the healing of pressure ulcers positively. The registered nurse is responsible of providing this care. An issue could be the lack of knowledge regarding nutritional supplements. Aim: The purpose of this study was to describe the nutritional supplements effect on pressure ulcer healing. Method: The study was executed as a literature review (n=10) based on quantitative studies. Result: The result of the study showed that most of the nutritional supplements have had a positive effect. Supplements such as arginine, vitamin C and zinc in combination and hydrolysated protein gave an improved wound healing. The supplement containing fish oil showed no significant improvements. Conclusion: Nutritional supplements containing arginine, vitamin C and zinc showed a positive effect on the healing of pressure ulcers. Studies show that nurses’ knowledge regarding nutrition in ulcer care is incomplete. If the reason behind this is that the information is difficult to understand our hope is that this study will make it easier to learn more about nutritional supplements, what supplements that should be used and its role in pressure ulcer care.

Angiogenesis is the formation of new blood vessels from pre-existing vasculature and is important during development as well as wound healing and tissue remodeling. Angiogenesis also occurs during pathological conditions such as diabetic retinopathy and cancer. This thesis is centered on the biology of endothelial cells, lining the blood vessels, and myofibroblasts, important for wound healing. We investigated an endothelial cell specific gene, ExoC3l2, and its role in VEGFR2 signaling and migration. EXOC3L2 co-localize with members of the exocyst complex, involved in vesicular transport, as well as VEGFR2. Reducing the level of EXOC3L2 in microvascular endothelial cells results in reduced VEGFR2 signaling and subsequently reduced chemotactic response to VEGF-A. MicroRNA (miRNA) have been shown to be regulators of gene transcription and cell type specific miRNAs have been identified. We investigated two miRNAs, miR-145 and miR-24. miR-145 is expressed in pericytes and fibroblasts but was shown to regulate fli1, an endothelial transcription factor. miR-145 overexpression reduced chemotaxis in both fibroblasts and endothelial cells, as did suppression of the endogenous miR-145 level in fibroblasts. miR-24 in contrast is expressed by endothelial cells and are able to target Ndst1, important for heparan sulfate (HS) sulfation. Sulfation of HS is important for many processes, amongst them growth factor signaling. Overexpression of miR-24 resulted in lower sulfation of HS chains, decreasing the ability of HS to interact with VEGF-A. Overexpressing miR-24 resulted in disturbed chemotaxis, similar to suppressing Ndst1 using siRNA. Myofibroblast recruitment is an important step in wound healing. The myofibroblasts contract the wound, synthesize new extracellular matrix and contribute to revascularization by looping angiogenesis. Maturation from resting fibroblast to myofibroblast is dependent on TGF-β. We found that fibrin fragment E (FnE), a degradation product of fibrin, potentiated the response of fibroblasts to TGF-β thus enhancing TGF-β-induced myofibroblast differentiation. FnE was also found to influence the migration of fibroblasts. These responses are dependent on integrins and toll-like receptors. These findings may serve to further increase the understanding of angiogenesis and wound healing to develop new therapies against pathological conditions.

<p>The ability of cells to migrate is of fundamental importance to a diverse array of biological processes, both physiological and pathological, such as development, the immune response and cancer cell metastasis, to name a few. The process of cell movement is a complicated cycle of coordinated steps involving dynamic and precise rearrangement of the actin-myosin cytoskeleton. As a critical component of the migration machinery, the molecular motor protein nonmuscle myosin II (myosin II) has long been a subject of scientific inquiry. It is now generally accepted that the contractile forces generated by myosin II contribute directly or indirectly to every step in migration. Interestingly, three isoforms of myosin II (myosin IIA, IIB and IIC) have been identified, and although each isoform performs the same basic molecular functions, recent findings suggest that the different myosin II isoforms make unique contributions to the motile process. In this dissertation work I used RNA interference technology to specifically deplete cells of myosin IIA and IIB in order to characterize the distinct migration phenotypes associated with loss-of-function of each individual isoform. Surprisingly, I found that the two myosin II isoforms perform not only distinct but opposing functions in cell migration, with myosin IIA and IIB normally inhibiting and facilitating proper cell movement, respectively. Furthermore, using pharmacological and microscopy techniques, I investigated the cellular mechanisms allowing for isoform-specific function. My results provide evidence for at least two isoform-specific regulatory mechanisms, namely selectivity in signaling pathways and subcellular distribution. A particularly significant finding is the identification of the different assembly properties of myosin IIA and IIB as the key element responsible for directing isoform-distinct distribution. Together the data presented herein represent a considerable advance in our understanding of the distinct functions and regulation of myosin II in cell motility.</p> / Dissertation

Epithelial to mesenchymal transition (EMT), the transdifferentation of an epithelial cell into a mesenchymal fibroblast, is a cellular process necessary for embryonic development and wound healing. However, uncontrolled EMT can result in accumulation of myofibroblasts and excessive deposition of ECM, contributing to the pathological progression of fibrotic diseases such as pulmonary fibrosis. The ability to control EMT is important for development of novel therapeutics for fibrotic pathologies and for designing novel biomaterials for tissue engineering applications seeking to promote EMT for development of complex tissues. EMT is a highly orchestrated process involving the integration of biochemical signals from specific integrin-mediated interactions with extracellular matrix (ECM) proteins and soluble growth factors such as TGFβ. TGFβ, a potent inducer of EMT, is activated via cell contraction-mediated mechanical release of the growth factor from a macromolecular latency complex. Thus TGFβ activity and subsequent EMT may be influenced by the biochemical and biophysical state of the surrounding ECM. Based on these knowns, it was hypothesized that both changes in integrin engagement and increases in substrate rigidity would modulate EMT due to changes in epithelial cell contraction and TGFβ activation. Here we show that integrin-specific interactions with fibronectin (Fn) fragments displaying both the RGD and PHSRN binding sites facilitate cell binding through α5β1 and α3β1 integrins, and lead to maintenance of epithelial phenotype, while Fn fragments displaying only the RGD site facilitate cell binding through αv integrins and lead to EMT. An in depth investigation into α3β1 binding to Fn fragments indicates that binding is dependent on both the presence and orientation of the PHSRN site. Studies investigating the contribution of ECM stiffening on EMT responses show that increasingly rigid Fn substrates are sufficient to induce spontaneous EMT. Analysis of TGFβ-responsive genes implicate TGFβ-expression, activation or signaling as a mechanism for the observed EMT responses. Together these results suggest that the ECM micromechanical environment is a significant contributor to the onset of EMT responses and provide insights into the design of biomaterial-based microenvironments for the control of epithelial cell phenotype.

"December 2009" The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from PDF of title page (University of Missouri--Columbia, viewed on January 5, 2010). Thesis advisor: David A. Wilson. Includes bibliographical references.

Dissertation (Ph.D.)--University of South Florida, 2009. / Title from PDF of title page. Document formatted into pages; contains 91 pages. Includes vita. Includes bibliographical references.