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<p>Epidermolytic hyperkeratosis is a rare inherited disease of the skin caused by a dominant-negative mutation in keratin 1 (K1) or 10 (K10). Keratins are the major structural protein in epidermis and mutations causes instability of intermediate filament and keratinocyte fragility. No curative treatment is available, but some patients benefit from retinoid therapy. More knowledge is needed about the genotype/phenotype correlation in epidermolytic hyperkeratosis and the mechanism of action of retinoids including regulation of keratin expression. </p><p>Fifteen patients were identified in Scandinavia, 13 with a generalised disease and 2 with localised lesions. Different types of mutation were identified such as point, splice site, deletion, and deletion-insertion mutations. An association was found between mutation in K1 and the appearance of palmoplantar keratoderma. Only the patients with K10 mutation benefited from the treatment, although no differences in the mRNA levels for K1 and K10 were detected. However, retinoids caused a pronounce down-regulation of K2e in upper epidermis and upregulation of K4 not normally present in the skin. This was further investigated in normal healthy skin and in keratinocytes grown in a reconstructed skin model. By adding retinoids with different affinity for the nuclear receptors RAR and RXR to the culture, the most potent retinoids were found to be RARα agonist, the effect of which could be inhibited by addition of a pan RAR antagonist. </p><p>In conclusion, several novel keratin mutations have been shown to cause epidermolytic hyperkeratosis, and genotype/phenotype correlations have been found. Treatment with retinoids is only useful for patients with a K10 mutation, possibly because they are less vulnerable too the pronounce down-regulation of K2e also seen in normal skin. This effect and the upregulation of K4 seem to be mediated through RARα , expressed in the keratinocytes.</p>

Epidermolytic hyperkeratosis is a rare inherited disease of the skin caused by a dominant-negative mutation in keratin 1 (K1) or 10 (K10). Keratins are the major structural protein in epidermis and mutations causes instability of intermediate filament and keratinocyte fragility. No curative treatment is available, but some patients benefit from retinoid therapy. More knowledge is needed about the genotype/phenotype correlation in epidermolytic hyperkeratosis and the mechanism of action of retinoids including regulation of keratin expression. Fifteen patients were identified in Scandinavia, 13 with a generalised disease and 2 with localised lesions. Different types of mutation were identified such as point, splice site, deletion, and deletion-insertion mutations. An association was found between mutation in K1 and the appearance of palmoplantar keratoderma. Only the patients with K10 mutation benefited from the treatment, although no differences in the mRNA levels for K1 and K10 were detected. However, retinoids caused a pronounce down-regulation of K2e in upper epidermis and upregulation of K4 not normally present in the skin. This was further investigated in normal healthy skin and in keratinocytes grown in a reconstructed skin model. By adding retinoids with different affinity for the nuclear receptors RAR and RXR to the culture, the most potent retinoids were found to be RARα agonist, the effect of which could be inhibited by addition of a pan RAR antagonist. In conclusion, several novel keratin mutations have been shown to cause epidermolytic hyperkeratosis, and genotype/phenotype correlations have been found. Treatment with retinoids is only useful for patients with a K10 mutation, possibly because they are less vulnerable too the pronounce down-regulation of K2e also seen in normal skin. This effect and the upregulation of K4 seem to be mediated through RARα , expressed in the keratinocytes.

Studies were conducted using cultured immortalized wild type (non-CF) and mutant (CF) DeltaF508 cystic fibrosis transmembrane conductance regulator (CFTR) tracheal epithelial cells on the anti-inflammatory impact of agents that may alter ceramide and glutathione (GSH) metabolism. The CF cells demonstrated abnormally high levels of GSH and glutathione disulfide (GSSG), which could diminish intracellular production of ceramide, a key modulator of inflammation and apoptosis. Hence, additional cell culture studies were carried out with a known inducer of in situ ceramide synthesis, N-4(4-hydroxyphenyl) retinamide (fenretinide) on interleukin (IL)-8 release, intracellular ceramide content, and cellular proliferation in both the basal state and following the inflammatory stimuli of tumor necrosis factor (TNF) -alpha. Fenretinide treatment was associated with a dose-dependent increase in the cellular content of ceramide in both CF and non CF cells. Also, an inhibition of IL-8 release in the inflammatory condition of TNF-alpha treatment was observed following fenretinide treatment in the CF cells. As hyperbaric treatment of whey proteins was previously associated with improved survivability and higher GSH content in a Pseudomonas aeruginosa murine model of cystic fibrosis (CF), the anti-inflammatory role of low molecular weight peptides (&lt; 1kDa) generated from enzymatic hydrolysis of native and pressurized whey protein isolates (WPI) was examined. Pressure treatment of WPI was associated with an enhanced protein digestibility and an altered peptide profile following in vitro digestion. The whey peptides were tested CF and non-CF lung epithelial cells to identify for their effects on GSH metabolism. The impact of the combined treatment of fenretinide and WPH was also tested in terms of apoptosis and cytokine release in cell culture. As opposed to non-CF cells, CF cells showed a strong downtrend in release of IL-8 following the combined fenretinide and whey peptide treatment. In addition, whey peptides protected wild type epithelial cells from the apoptotic effect of fenretinide. Our results suggest the usefulness of these agents as a pharmacological treatment in CF.

While carotenoids are essential for visual function, their potential role in diabetic retinopathy is not known. By providing carrot powder, this study examined carotenoid metabolism and visual function in Type 1 diabetes. Wistar rats (n=30) were assigned to diet either with or without carrot enrichment (15%, w/w) for 12 weeks. Type 1 diabetes was induced with streptozotocin at 3 weeks. Retinal function and anatomical integrity were assessed along with retinoid and carotenoid levels in the serum, liver, and retina. Loss of ERG oscillatory potentials, with normal histology indicated early stage retinopathy. Healthy animals fed carrot diet showed highest b-wave amplitudes; reflecting higher phototransduction. Diabetic animals fed carrot diet had the lowest b-wave amplitudes, reduced retinoids liver reserves, and highest α- and β-carotene, suggesting disturbance of conversion during diabetes. Consequently carrot powder at concentrations used by this study cannot be recommended for diabetic retinopathy.

While carotenoids are essential for visual function, their potential role in diabetic retinopathy is not known. By providing carrot powder, this study examined carotenoid metabolism and visual function in Type 1 diabetes. Wistar rats (n=30) were assigned to diet either with or without carrot enrichment (15%, w/w) for 12 weeks. Type 1 diabetes was induced with streptozotocin at 3 weeks. Retinal function and anatomical integrity were assessed along with retinoid and carotenoid levels in the serum, liver, and retina. Loss of ERG oscillatory potentials, with normal histology indicated early stage retinopathy. Healthy animals fed carrot diet showed highest b-wave amplitudes; reflecting higher phototransduction. Diabetic animals fed carrot diet had the lowest b-wave amplitudes, reduced retinoids liver reserves, and highest α- and β-carotene, suggesting disturbance of conversion during diabetes. Consequently carrot powder at concentrations used by this study cannot be recommended for diabetic retinopathy.

Bovine fibroblasts and epithelial cells were isolated from surgically biopsied mammary tissue. Characterization of population doubling time, cytoskeletal intermediate filaments, cryopreservation survival, and viability were performed on all fibroblast and epithelial cells. Several clonal fibroblast cell lines were cotransfected with a plasmid bearing the SV-40 Large-T-antigen, and the pSV-2 neo plasmid. Transfected cells were subsequently selected with G418 sulfate and cloned. / MAC-T cells and non-clonal primary bovine mammary epithelial cells proliferated in response to IGF-I, insulin, serum and serum albumin. MAC-T cells did not proliferate when cultured in EGF, estrogen, progesterone, estrogen+progesterone, growth hormone, prolactin, and only modest proliferation was obtained after TGF-$ alpha$ treatment. Subsequent experiments used serum, insulin or IGF-I (and its analogues) to stimulate cellular proliferation. Serum albumin was not added to serum-free media preparations since it stimulated cellular proliferation. / TGF-$ beta$ receptors were characterized in MAC-T cells and normal fibroblasts. Affinity labelling studies revealed that MAC-T and MF-2 cells contained type I, II, and III autoregulatable receptors. Fibroblast proliferation, was inhibited 50% by TGF-$ beta$. TGF-$ beta$ inhibited MAC-T cellular proliferation at concentrations among the lowest ever reported, ED$ sb{ rm 50}$ = 4 pm. TGF-$ beta$ was not cytotoxic at concentrations 1000-fold higher. / Retinoic acid (RA) also inhibited proliferation of MAC-T cells. Inhibition of proliferation did not occur when cells were growth stimulated by IGF-I analogues that do not bind IGFBPs. Unlike TGF-$ beta$, RA treatment increased IGFBP-2 and decreased IGFBP-3 protein expression by cells into media and on the cell's membrane. RA was cytotoxic at concentrations 10-fold higher than ED$ sb{100}$. / Fibroblasts and epithelial cells expressed the gap junction (GJ) protein, connexin43, with transformed fibroblasts expressing significantly less connexin43. Perinuclear and cell surface connexin43 was immunodetected in epithelial and fibroblasts cells. TGF-$ beta$, RA or cAMP, increased connexin43 protein expression, especially phosphorylated species. Only cAMP noticeably altered immunolocalization patterns of connexin43, causing a shift from perinuclear pools to the cell surface. None of the growth inhibitors affected GJ communication as measured by dye transfer. Therefore, mammary epithelial cells are growth inhibited by TGF-$ beta$ and RA by distinct mechanisms and both growth inhibitors significantly enhance the gap junction protein, connexin43, without increasing GJ communication.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2002. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (M. Phil.)--University of Hong Kong, 2009. / Includes bibliographical references (leaves 100-109) Also available in print.