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1 oktober 2018, Contains fulltext : 196295.pdf (publisher's version ) (Closed access), Changing wound dressings inflicts pain and may disrupt wound repair. Novel synthetic thermosensitive hydrogels based on polyisocyanopeptide (PIC) offer a solution. These gels are liquid below 16 degrees C and form gels beyond room temperature. The architecture and mechanical properties of PIC gels closely resemble collagen and fibrin, and include the characteristic stiffening response at high strains. Considering the reversible thermo-responsive behavior, we postulate that PIC gels are easy to apply and remove, and facilitate healing without eliciting foreign body responses or excessive inflammation. Biocompatibility may be higher in RGD-peptide-functionalized PIC gels due to enhanced cell binding capabilities. Full-thickness dorsal skin wounds in mice were compared to wounds treated with PIC gel and PIC-RGD gel for 3 and 7 days. No foreign body reactions and similar wound closure rates were found in all groups. The level of macrophages, myofibroblasts, epithelial migration, collagen expression, and blood vessels did not significantly differ from controls. Surprisingly, granulocyte populations in the wound decreased significantly in the PIC gel-treated groups, likely because foreign bacteria could not penetrate the gel. RGD-peptides did not further improve any effect observed for PIC. The absence of adverse effects, ease of application, and the possibilities for bio-functionalization make the biomimetic PIC hydrogels suitable for development into wound dressings.

Contains fulltext : 169649.pdf (publisher's version ) (Open Access), Skin wounds may lead to scar formation and impaired functionality. Remote ischemic preconditioning (RIPC) can induce the anti-inflammatory enzyme heme oxygenase-1 (HO-1) and protect against tissue injury. We aim to improve cutaneous wound repair by RIPC treatment via induction of HO-1. RIPC was applied to HO-1-luc transgenic mice and HO-1 promoter activity and mRNA expression in skin and several other organs were determined in real-time. In parallel, RIPC was applied directly or 24h prior to excisional wounding in mice to investigate the early and late protective effects of RIPC on cutaneous wound repair, respectively. HO-1 promoter activity was significantly induced on the dorsal side and locally in the kidneys following RIPC treatment. Next, we investigated the origin of this RIPC-induced HO-1 promoter activity and demonstrated increased mRNA in the ligated muscle, heart and kidneys, but not in the skin. RIPC did not change HO-1 mRNA and protein levels in the wound 7 days after cutaneous injury. Both early and late RIPC did not accelerate wound closure nor affect collagen deposition. RIPC induces HO-1 expression in several organs, but not the skin, and did not improve excisional wound repair, suggesting that the skin is insensitive to RIPC-mediated protection.

Contains fulltext : 171101.pdf (publisher's version ) (Open Access), Orthodontic forces disturb the microenvironment of the periodontal ligament (PDL), and induce craniofacial bone remodeling which is necessary for tooth movement. Unfortunately, orthodontic tooth movement is often hampered by ischemic injury and cell death within the PDL (hyalinization) and root resorption. Large inter-individual differences in hyalinization and root resorption have been observed, and may be explained by differential protection against hyalinization. Heme oxygenase-1 (HO-1) forms an important protective mechanism by breaking down heme into the strong anti-oxidants biliverdin/bilirubin and the signaling molecule carbon monoxide. These versatile HO-1 products protect against ischemic and inflammatory injury. We postulate that orthodontic forces induce HO-1 expression in the PDL during experimental tooth movement. Twenty-five 6-week-old male Wistar rats were used in this study. The upper three molars at one side were moved mesially using a Nickel-Titanium coil spring, providing a continuous orthodontic force of 10 cN. The contralateral side served as control. After 6, 12, 72, 96, and 120 h groups of rats were killed. On parasagittal sections immunohistochemical staining was performed for analysis of HO-1 expression and quantification of osteoclasts. Orthodontic force induced a significant time-dependent HO-1 expression in mononuclear cells within the PDL at both the apposition- and resorption side. Shortly after placement of the orthodontic appliance HO-1 expression was highly induced in PDL cells but dropped to control levels within 72 h. Some osteoclasts were also HO-1 positive but this induction was shown to be independent of time- and mechanical stress. It is tempting to speculate that differential induction of tissue protecting- and osteoclast activating genes in the PDL determine the level of bone resorption and hyalinization and, subsequently, "fast" and "slow" tooth movers during orthodontic treatment.

Item does not contain fulltext, Excessive extracellular matrix (ECM) deposition and tissue contraction after injury can lead to esthetic and functional problems. Fibroblasts and myofibroblasts activated by transforming growth factor (TGF)-beta1 play a key role in these processes. The persistence of (myo)fibroblasts and their excessive ECM production and continuous wound contraction have been linked to pathological scarring. The identification of compounds reducing myofibroblast survival and function may thus offer promising therapeutic strategies to optimize impaired wound healing. The plant-derived polyphenol curcumin has shown promising results as a wound healing therapeutic in vivo; however, the exact mechanism is still unclear. In vitro, curcumin induces apoptosis in various cell types via a reactive oxygen species (ROS)-dependent mechanism. Here we treated human dermal fibroblasts with TGF-beta1 to induce myofibroblast differentiation, and compared the responses of fibroblasts and myofibroblasts to 25 microM curcumin. Curcumin induced caspase-independent apoptosis in both fibroblasts and myofibroblasts in a ROS-dependent manner. Oxidative stress leads to the induction of several antioxidant systems to regain cellular homeostasis. We detected stress-induced induction of heme oxygenase (HO)-1 in fibroblasts but not in myofibroblasts following curcumin exposure. Instead, myofibroblasts expressed higher levels of heat shock protein (HSP)72 compared to fibroblasts in response to curcumin, suggesting that TGF-beta1 treatment alters the stress-responses of the cells. However, we did not detect any differences in curcumin toxicity between the two populations. The differential stress responses in fibroblasts and myofibroblasts may open new therapeutic approaches to reduce myofibroblasts and scarring.

Item does not contain fulltext, Wound healing is a complex process that involves the well-coordinated interactions of different cell types. Topical application of high doses of curcumin, a plant-derived polyphenol, enhances both normal and diabetic cutaneous wound healing in rodents. For optimal tissue repair interactions between epidermal keratinocytes and dermal fibroblasts are essential. We previously demonstrated that curcumin increased reactive oxygen species (ROS) formation and apoptosis in dermal fibroblasts, which could be prevented by pre-induction of the cytoprotective enzyme heme oxygenase (HO)-1. To better understand the effects of curcumin on wound repair, we now assessed the effects of high doses of curcumin on the survival of HaCaT keratinocytes and the role of the HO system. We exposed HaCaT keratinocytes to curcumin in the presence or absence of the HO-1 inducers heme (FePP) and cobalt protoporphyrin (CoPP). We then assessed cell survival, ROS formation, and caspase activation. Curcumin induced caspase-dependent apoptosis in HaCaT keratinocytes via a ROS-dependent mechanism. Both FePP and CoPP induced HO-1 expression, but only FePP protected against curcumin-induced ROS formation and caspase-mediated apoptosis. In the presence of curcumin, FePP but not CoPP induced the expression of the iron scavenger ferritin. Together, our data show that the induction of ferritin, but not HO, protects HaCaT keratinocytes against cytotoxic doses of curcumin. The differential response of fibroblasts and keratinocytes to high curcumin doses may provide the basis for improving curcumin-based wound healing therapies.

Item does not contain fulltext, Mutations in the CRB1 gene lead to retinal dystrophies ranging from Leber congenital amaurosis (LCA) to early-onset retinitis pigmentosa (RP), due to developmental defects or loss of adhesion between photoreceptors and Muller glia cells, respectively. Whereas over 150 mutations have been found, no clear genotype-phenotype correlation has been established. Mouse Crb1 knockout retinas show a mild phenotype limited to the inferior quadrant, whereas Crb2 knockout retinas display a severe degeneration throughout the retina mimicking the phenotype observed in RP patients associated with CRB1 mutations. Crb1Crb2 double mutant retinas have severe developmental defects similar to the phenotype observed in LCA patients associated with CRB1 mutations. Therefore, CRB2 is a candidate modifying gene of human CRB1-related retinal dystrophy. In this study, we studied the cellular localization of CRB1 and CRB2 in human retina and tested the influence of the Crb2 gene allele on Crb1-retinal dystrophies in mice. We found that in contrast to mice, in the human retina CRB1 protein was expressed at the subapical region in photoreceptors and Muller glia cells, and CRB2 only in Muller glia cells. Genetic ablation of one allele of Crb2 in heterozygote Crb1(+/-) retinas induced a mild retinal phenotype, but in homozygote Crb1 knockout mice lead to an early and severe phenotype limited to the entire inferior retina. Our data provide mechanistic insight for CRB1-related LCA and RP.

Contains fulltext : 137751.pdf (publisher's version ) (Open Access), Impaired wound healing can lead to scarring, and aesthetical and functional problems. The cytoprotective haem oxygenase (HO) enzymes degrade haem into iron, biliverdin and carbon monoxide. HO-1 deficient mice suffer from chronic inflammatory stress and delayed cutaneous wound healing, while corneal wound healing in HO-2 deficient mice is impaired with exorbitant inflammation and absence of HO-1 expression. This study addresses the role of HO-2 in cutaneous excisional wound healing using HO-2 knockout (KO) mice. Here, we show that HO-2 deficiency also delays cutaneous wound closure compared to WT controls. In addition, we detected reduced collagen deposition and vessel density in the wounds of HO-2 KO mice compared to WT controls. Surprisingly, wound closure in HO-2 KO mice was accompanied by an inflammatory response comparable to WT mice. HO-1 induction in HO-2 deficient skin was also similar to WT controls and may explain this protection against exaggerated cutaneous inflammation but not the delayed wound closure. Proliferation and myofibroblast differentiation were similar in both two genotypes. Next, we screened for candidate genes to explain the observed delayed wound closure, and detected delayed gene and protein expression profiles of the chemokine (C-X-C) ligand-11 (CXCL-11) in wounds of HO-2 KO mice. Abnormal regulation of CXCL-11 has been linked to delayed wound healing and disturbed angiogenesis. However, whether aberrant CXCL-11 expression in HO-2 KO mice is caused by or is causing delayed wound healing needs to be further investigated.

Contains fulltext : 137723.pdf (publisher's version ) (Open Access)

Contains fulltext : 125632.pdf (publisher's version ) (Open Access), During mammalian cortical development, division of progenitor cells occurs at the apical ventricular zone. Apical complex proteins and adherens junctions regulate the different modes of division. Here, we have identified the membrane-associated guanylate kinase protein membrane palmitoylated protein 3 (MPP3) as an essential protein for the maintenance of these complexes. MPP3 localizes at the apical membrane in which it shows partial colocalization with adherens junction proteins and apical proteins. We generated Mpp3 conditional knock-out mice and specifically ablated Mpp3 expression in cortical progenitor cells. Conditional deletion of Mpp3 during cortical development resulted in a gradual loss of the apical complex proteins and disrupted adherens junctions. Although there is cellular disorganization in the ventricular zone, gross morphology of the cortex was unaffected during loss of MPP3. However, in the ventricular zone, removal of MPP3 resulted in randomization of spindle orientation and ectopically localized mitotic cells. Loss of MPP3 in the developing cortex resulted in delayed migration of progenitor cells, whereas the rate of cell division and exit from the cell cycle was not affected. This resulted in defects in cortical stratification and ectopically localized layer II-IV pyramidal neurons and interneurons. These data show that MPP3 is required for maintenance of the apical protein complex and adherens junctions and for stratification and proper migration of neurons during the development of the cortex.

Contains fulltext : 117735.pdf (publisher's version ) (Open Access), Reactive oxygen species (ROS) can be both beneficial and deleterious. Under normal physiological conditions, ROS production is tightly regulated, and ROS participate in both pathogen defense and cellular signaling. However, insufficient ROS detoxification or ROS overproduction generates oxidative stress, resulting in cellular damage. Oxidative stress has been linked to various inflammatory diseases. Inflammation is an essential response in the protection against injurious insults and thus important at the onset of wound healing. However, hampered resolution of inflammation can result in a chronic, exaggerated response with additional tissue damage. In the pathogenesis of several inflammatory skin conditions, e.g., sunburn and psoriasis, inflammatory-mediated tissue damage is central. The prolonged release of excess ROS in the skin can aggravate inflammatory injury and promote chronic inflammation. The cellular redox balance is therefore tightly regulated by several (enzymatic) antioxidants and pro-oxidants; however, in case of chronic inflammation, the antioxidant system may be depleted, and prolonged oxidative stress occurs. Due to the central role of ROS in inflammatory pathologies, restoring the redox balance forms an innovative therapeutic target in the development of new strategies for treating inflammatory skin conditions. Nevertheless, the clinical use of antioxidant-related therapies is still in its infancy.

1 december 2013, Contains fulltext : 126303.pdf (publisher's version ) (Open Access), Development in the central nervous system is highly dependent on the regulation of the switch from progenitor cell proliferation to differentiation, but the molecular and cellular events controlling this process remain poorly understood. Here, we report that ablation of Crb1 and Crb2 genes results in severe impairment of retinal function, abnormal lamination and thickening of the retina mimicking human Leber congenital amaurosis due to loss of CRB1 function. We show that the levels of CRB1 and CRB2 proteins are crucial for mouse retinal development, as they restrain the proliferation of retinal progenitor cells. The lack of these apical proteins results in altered cell cycle progression and increased number of mitotic cells leading to an increased number of late-born cell types such as rod photoreceptors, bipolar and Muller glia cells in postmitotic retinas. Loss of CRB1 and CRB2 in the retina results in dysregulation of target genes for the Notch1 and YAP/Hippo signaling pathways and increased levels of P120-catenin. Loss of CRB1 and CRB2 result in altered progenitor cell cycle distribution with a decrease in number of late progenitors in G1 and an increase in S and G2/M phase. These findings suggest that CRB1 and CRB2 suppress late progenitor pool expansion by regulating multiple proliferative signaling pathways.

Contains fulltext : 98182.pdf (publisher's version ) (Open Access)

Udgivelsesdato: 2006/10