Identification of biomarkers involved in differential profiling of hypertrophic and keloid scars versus normal skin.

Among raised dermal scar types, keloid (KS) and hypertrophic scars (HS) are considered to present clinical similarities, but there are no known specific biomarkers that allow both scar types to be easily distinguished. Development and progression of raised dermal scars comprises the activation of several molecular pathways and cell defence mechanisms leading to elevated extracellular matrix component synthesis, delayed apoptosis, altered migration and differentiation. Therefore, the aim here was to identify biomarkers that may differentiate between KS and HS compared to normal skin (NS). To achieve this aim, NS (n = 14), KS (n = 14) and HS (n = 14) biopsies were evaluated using histology by H&E staining. Tissue biopsies and primary fibroblasts (passages 0-4) were employed to assess the gene expression levels of 21 biomarkers selected from our previous microarray studies using qRT-PCR. Finally, protein expression was evaluated using In-Cell Western Blotting in primary fibroblasts (p 0-4). Our results demonstrated that out of the 21 biomarkers screened at mRNA and protein levels, α2ß1-integrin, Hsp27, PAI-2, MMP-19 and CGRP showed significantly higher expression (p < 0.05) in KS compared to NS and HS. Additionally, these five key biomarkers were found to be significantly higher (p < 0.05) at mRNA level in KS taken from the sternum, a region known to be subjected to high mechanical forces in the body during the performance of daily movements. In conclusion, our findings offer potential molecular targets in raised dermal scars differentiation. Future targeted research may allow provision of diagnostic and prognostic markers in keloid versus hypertrophic scars.

Use of dental impression materials in the analysis of tool marks to identify causal elements / Uso de materiales de impresión dental para el análisis de marcas de corte para identificación de elemento causal

When faced with an element which is the suspected cause of a cut or lesion, dental casting can be made to compare characteristics observed on the surface of the tool mark with those found on the bones. There is a large variety of materials available on national and international markets to record and analyze marks or signs that an element leaves on any given surface (bite marks, tool marks on bones, finger prints, etc.), all of which are useful in forensic investigation (1). However, the unconventional application of odontological techniques and the use of materials such as alginate and silicone have been found useful recovering tool marks on bones, which allows the forensic scientist to clearly establish when an element was used to cause cuts and injuries in an individual. This technique has been applied in five cases of possible homicide with the use of a cutting element and, as a result, has generated highly accurate casts. It also shows that both materials are appropriate for this purpose, and although silicone offers greater detail in the impression, either material can be used depending on the commercial availability.

En caso de contar con el elemento debitado (con el que se sospecha se causó la lesión), se pueden generar impresiones con materiales de uso odontológico para comparar las características observadas en la superficie de corte de la herramienta con las encontradas en la estructura ósea. En el mercado nacional e internacional existe gran variedad de materiales para el registro y análisis de las marcas o señales que deja un elemento sobre una superficie dada (impresiones de estructuras bucales, marcas de herramientas en huesos, huellas dactilares, etc.), y son útiles en estudios forenses (1). Con la aplicación de técnicas odontológicas y el empleo de materiales tales como el alginato y la silicona, se observó que en esta aplicación no convencional es útil para tomar impresiones de marcas de corte sobre estructuras óseas, lo que a su vez permite establecer con claridad cuando un elemento dubitado ha sido utilizado para causar cortes y lesiones en un individuo. Esta técnica se aplicó en 5 casos de posible homicidio con empleo de elemento cortante y se encontró no solo la generación de improntas de alta fidelidad sino también que ambos materiales son apropiados para ello, y aunque la silicona ofreció mayor detalle en la impresión, cualquiera de los dos puede ser utilizado dependiendo de su disponibilidad comercial.

Skin equivalent tensional force alters keloid fibroblast behavior and phenotype.

Skin tension may influence keloid scar behavior, development, and spreading, e.g., butterfly-shaped keloid disease in the sternum. Here, we developed a three-dimensional (3D) in vitro model to mimic in vivo tension and evaluate keloid fibroblast (KF) behavior and extracellular matrix synthesis under tension. In vivo skin tension measured in volunteers (n = 4) using 3D image photogrammetry enabled prediction of actual force (35 mN). A novel cell force monitor applied tension in a fibroblast-populated 3D collagen lattice replicating the in vivo force. The effect of tension on keloid (n = 10) fibroblast (KF) and normal skin (n = 10) fibroblasts (NF) at set time points (6, 12, and 24 hours) was measured in Hsp27, PAI-2, and α2ß1 integrin, tension-related genes demonstrating significant (p < 0.05) time-dependent regulation of these genes in NF vs. KF with and without tension. KF showed higher (p < 0.05) proliferation post-tension. Knockdown of all three genes in 24 and 48 hours with and without tension showed significant down-regulation in NF vs. KF. Additionally, we show significant (p < 0.05) modification of the expression of extracellular matrix-related genes post-tension following down-regulation of Hsp27, PAI-2, or α2ß1 integrin. Finally, we demonstrate significant alteration in NF compared with KF morphology following knockdown. In conclusion, this study shows induction of tension-related genes expression following mechano-regulation in KFs, with potential relevance to its development and therapy.

Up-regulation of tension-related proteins in keloids: knockdown of Hsp27, α2ß1-integrin, and PAI-2 shows convincing reduction of extracellular matrix production.

BACKGROUND: Keloid disease is a fibroproliferative disorder, with an ill-defined treatment that is characterized by excessive extracellular matrix deposition. Mechanical tension promotes deposition of extracellular matrix and overexpression of tension-related proteins, which is associated with keloid disease. The aim of this study was to investigate the effect of tension-related proteins on extracellular matrix steady-state synthesis in primary keloid fibroblasts. METHODS: Keloid fibroblasts (n = 10) and normal skin (n = 4) fibroblast cultures were established from passages 0 to 3. A panel of 21 tension-related genes from microarray data were assessed at mRNA (quantitative reverse-transcriptase polymerase chain reaction) and protein (in-cell Western blotting) levels. Three genes were significantly altered in keloid tissue and fibroblasts, and their functional role was assessed using siRNA knockdown. RESULTS: Hsp27, α2ß1-integrin, and PAI-2 were significantly up-regulated (p < 0.05)in keloid tissue and fibroblasts compared with normal skin. Hsp27, α2ß1-integrin, and PAI-2 expression was inhibited by RNA interference. Both the mRNA and protein levels of Hsp27, α2ß1-integrin, and PAI-2 significantly decreased (p < 0.05) in keloid fibroblasts at 48 hours after transfection. After down-regulation of Hsp27, α2ß1-integrin, and PAI-2, the expression of intracellular extracellular matrix was significantly reduced (p < 0.05). Water-soluble tetrazolium salt-1 assay showed that transfection of Hsp27, α2ß1-integrin, and PAI-2 did not influence the viability/metabolic activity of keloid fibroblasts. CONCLUSIONS: This study demonstrates overexpression of key tension-related proteins in keloid tissue and keloid fibroblasts. Knockdown of Hsp27, PAI-2, and α2ß1-integrin by RNA interference attenuates the expression of mRNA and protein levels and certain other extracellular matrix molecules.

Models and diagrams as thinking tools: the case of satellite-DNA.

The aim of the paper is to present some ideas on the transition between the material representations of phenomena in experimental systems, commonly referred as material traces or inscriptions (for instance, in an electrophoretic gel or a paper chromatography), and the more "public" and "general" representations of phenomena (including tables, diagrams, graphs and other visual and textual representations). To do so, the story of the early representations of satellite-DNA, a fraction of about 400 base-pair-long repeated sequences in eukaryotic DNA stabilized in the mid 1960s, will be presented. The author argues that the practices associated to different moments of research (such as the stabilization of phenomena, the reconstruction of a process, the modeling of mechanisms or the communication of results, among others) require the construction and the translocation of meanings between different ways of representing. Special attention is given to a kind of scientific diagram used in experimental tradition to understand processes and mechanisms; following Dewey's conception of tools these diagrams are called "thinking tools".

Identification of genes in cassava that are differentially expressed during infection with Xanthomonas axonopodis pv. manihotis.

SUMMARY The cDNA-amplified fragment length polymorphism approach was used to identify differentially expressed transcripts from cassava infected by Xanthomonas axonopodis pv. manihotis (Xam). Approximately 3600 transcript-derived fragments (TDFs) were screened of which 340 were isolated. The nucleotide sequences of 250 TDFs were analysed and assembled into contigs and singletons. The amino acid sequences of their predicted products were compared with entries in databases and 63 of these clones showed homology to known plant genes. Of these, 32 showed similarity to plant defence proteins. Fifty-one TDFs corresponded to proteins of unknown function and 106 did not match any sequence in the public databases. Quantitative reverse transcription PCR was carried out with a selected set of gene transcripts that demonstrated an increase of expression during the infection. These results point out candidate genes that are associated with cassava resistance to Xam and reinforce the idea of a complex process occurring during this plant-pathogen interaction.