siRNA Silencing of Proteasome Maturation Protein (POMP) Activates the Unfolded Protein Response and Constitutes a Model for KLICK Genodermatosis

Keratosis linearis with ichthyosis congenita and keratoderma (KLICK) is an autosomal recessive skin disorder associated with a single-nucleotide deletion in the 5′untranslated region of the proteasome maturation protein (POMP) gene. The deletion causes a relative switch in transcription start sites for POMP, predicted to decrease levels of POMP protein in terminally differentiated keratinocytes. To investigate the pathophysiology behind KLICK we created an in vitro model of the disease using siRNA silencing of POMP in epidermal air-liquid cultures. Immunohistochemical analysis of the tissue constructs revealed aberrant staining of POMP, proteasome subunits and the skin differentiation marker filaggrin when compared to control tissue constructs. The staining patterns of POMP siRNA tissue constructs showed strong resemblance to those observed in skin biopsies from KLICK patients. Western blot analysis of lysates from the organotypic tissue constructs revealed an aberrant processing of profilaggrin to filaggrin in samples transfected with siRNA against POMP. Knock-down of POMP expression in regular cell cultures resulted in decreased amounts of proteasome subunits. Prolonged silencing of POMP in cultured cells induced C/EBP homologous protein (CHOP) expression consistent with an activation of the unfolded protein response and increased endoplasmic reticulum (ER) stress. The combined results indicate that KLICK is caused by reduced levels of POMP, leading to proteasome insufficiency in differentiating keratinocytes. Proteasome insufficiency disturbs terminal epidermal differentiation, presumably by increased ER stress, and leads to perturbed processing of profilaggrin. Our findings underline a critical role for the proteasome in human epidermal differentiation

“Magnetic Force Microscopy and Energy Loss Imaging of Superparamagnetic Iron Oxide Nanoparticles”

We present quantitative, high spatially resolved magnetic force microscopy imaging of samples based on 11 nm diameter superparamagnetic iron oxide nanoparticles in air at room temperature. By a proper combination of the cantilever resonance frequency shift, oscillation amplitude and phase lag we obtain the tip-sample interaction maps in terms of force gradient and energy dissipation. These physical quantities are evaluated in the frame of a tip-particle magnetic interaction model also including the tip oscillation amplitude. Magnetic nanoparticles are characterized both in bare form, after deposition on a flat substrate, and as magnetically assembled fillers in a polymer matrix, in the form of nanowires. The latter approach makes it possible to reveal the magnetic texture in a composite sample independently of the surface topography

Novel inhibitors of the calcineurin/NFATc hub - alternatives to CsA and FK506?

The drugs cyclosporine A (CsA) and tacrolimus (FK506) revolutionized organ transplantation. Both compounds are still widely used in the clinic as well as for basic research, even though they have dramatic side effects and modulate other pathways than calcineurin-NFATc, too. To answer the major open question - whether the adverse side effects are secondary to the actions of the drugs on the calcineurin-NFATc pathway - alternative inhibitors were developed. Ideal inhibitors should discriminate between the inhibition of (i) calcineurin and peptidyl-prolyl cis-trans isomerases (PPIases; the matchmaker proteins of CsA and FK506), (ii) calcineurin and the other Ser/Thr protein phosphatases, and (iii) NFATc and other transcription factors. In this review we summarize the current knowledge about novel inhibitors, synthesized or identified in the last decades, and focus on their mode of action, specificity, and biological effects

Best practices, new perspectives and the perfect emollient: optimizing the management of contact dermatitis

Contact dermatitis (CD) is caused by environmental agents, irritants, and allergens that penetrate the epidermis and lead to inflammation. An intact skin barrier prevents penetration and is important in maintaining healthy skin. Classical diagnosis of CD is made using the patch test, and traditional treatment strategies for CD promote skin barrier integrity and resolve the inflammatory component of the condition. This can be achieved by using emollient-based therapy, which is most important for skin barrier repair, and in addition to topical glucocorticosteroids, which are used in severe cases of CD and are most effective in reducing inflammation. Preventative measures, such as irritant and allergen avoidance in the workplace, also play a pivotal role in effective CD management. Moreover, CD management necessitates a holistic approach that incorporates prevention, barrier repair, and inflammatory resolution to ensure optimized efficacy. It is also important to consider potential barriers to optimal management when evaluating individuals with CD, such as limited patient education or poor access to care. Finally, key literature and our own clinical practice experience have highlighted the value of patient preference, as well as safety, efficacy and simplicity, in building the perfect emollient

Magnetosome Formation in Prokaryotes

Magnetotactic bacteria were discovered almost 30 years ago, and for many years and many different reasons, the number of researchers working in this field was few and progress was slow. Recently, however, thanks to the isolation of new strains and the development of new techniques for manipulating these strains, researchers from several laboratories have made significant progress in elucidating the molecular, biochemical, chemical and genetic bases of magnetosome formation and understanding how these unique intracellular organelles function. We focus here on this progress

Optical magnetic imaging of living cells.

Magnetic imaging is a powerful tool for probing biological and physical systems. However, existing techniques either have poor spatial resolution compared to optical microscopy and are hence not generally applicable to imaging of sub-cellular structure (for example, magnetic resonance imaging), or entail operating conditions that preclude application to living biological samples while providing submicrometre resolution (for example, scanning superconducting quantum interference device microscopy, electron holography and magnetic resonance force microscopy). Here we demonstrate magnetic imaging of living cells (magnetotactic bacteria) under ambient laboratory conditions and with sub-cellular spatial resolution (400 nanometres), using an optically detected magnetic field imaging array consisting of a nanometre-scale layer of nitrogen-vacancy colour centres implanted at the surface of a diamond chip. With the bacteria placed on the diamond surface, we optically probe the nitrogen-vacancy quantum spin states and rapidly reconstruct images of the vector components of the magnetic field created by chains of magnetic nanoparticles (magnetosomes) produced in the bacteria. We also spatially correlate these magnetic field maps with optical images acquired in the same apparatus. Wide-field microscopy allows parallel optical and magnetic imaging of multiple cells in a population with submicrometre resolution and a field of view in excess of 100 micrometres. Scanning electron microscope images of the bacteria confirm that the correlated optical and magnetic images can be used to locate and characterize the magnetosomes in each bacterium. Our results provide a new capability for imaging bio-magnetic structures in living cells under ambient conditions with high spatial resolution, and will enable the mapping of a wide range of magnetic signals within cells and cellular networks