Reducing the effects of intracellular accumulation of thermolabile collagen II mutants by increasing their thermostability in cell culture conditions.

Mutations in collagen II are associated with spondyloepiphyseal dysplasia, a group of heritable diseases whose common features include aberrations of skeletal growth. The mechanisms through which mutations in collagen II affect the cartilaginous tissues are complex and include both intracellular and extracellular processes. One of those mechanisms involves cellular stress caused by excessive accumulation of misfolded collagen II mutants. We investigated whether stabilizing the structure of thermolabile R789C and R992C collagen II mutants would improve their secretion from cells, thereby reducing cellular stress and apoptosis. Employing glycerol and trimethylamine N-oxide (TMAO), chemicals that increase the thermostability of collagen triple helices, we demonstrated that those compounds function as chaperones and stabilize the R789C and R992C mutants, accelerate their secretion, and improve cell survival. Our study provides a scientific basis for considering misfolded triple helices of collagen mutants a target for reducing the deleterious effects caused by their excessive intracellular accumulation

In vitro testing the potential of a novel chimeric IgG variant for inhibiting collagen fibrils formation in recurrent hereditary gingival fibromatosis: chimeric antibody in a gingival model

Gingival fibromatosis is a progressive enlargement of the gingiva. It may hinder oral cavity hygiene and result in underlying bone loss. The long-term benefits of surgery cannot be predicted. On the other hand, alternative, efficient and non-invasive methods are not available at present. The aim of this study was to test the inhibitory effects of a chimeric IgG variant on collagen fibril formation in the cell culture of gingival fibroblasts taken from a patient with hereditary gingival fibromatosis with a high propensity for recurrence. Gingival biopsies were collected from the mandibular gingiva and used for histological evaluation as well as to establish a fibroblast culture. A histological evaluation was made in haematoxylin-eosin and Heidenhain's trichrome stained tissue sections. The inhibitory effect of a chimeric antibody on collagen fibril formation was determined in fibroblast cultures by using a collagen-specific Western blot and immunofluorescent staining. A histological evaluation revealed epithelial acanthosis with singular elongated rete pegs extending into the underlying connective tissue stroma that consisted of locally abundant, irregular collagen bundles. Based on observations with an in vitro model we conclude that a chimeric anti-collagen antibody efficiently inhibits collagen fibril accumulation in cell culture derived from diffuse, hereditary gingival fibromatosis that is characterized by a high propensity for recurrence (high proliferation index). Employing cell cultures from standardized group of patients with recurrent hereditary gingival fibromatosis as well as standarizing relevant 3D (tissue-like) models will be crucial for further tests of the antibody

Niejęzykowe deficyty poznawcze u dzieci z dysleksją rozwojową i jej ryzykiem – przegląd badań

Learning to read depends on the development of a number of cognitive functions. Along with poor phonology skills, many dyslexic children show difficulties in several specific nonverbal abilities. Their identification during preschool or early primary school should enhance efficient prediction of dyslexia. The article presents research evidence on the deficits of dyslexic children in coherent motion detection, automatisation, attention shifting, visual attention span, and anchoring. Exemplar tasks for their assessment are described

R992C (p.R1192C) Substitution in collagen II alters the structure of mutant molecules and induces the unfolded protein response.

We investigated the molecular bases of spondyloepiphyseal dysplasia (SED) associated with the R992C (p.R1192C) substitution in collagen II. At the protein level, we analyzed the structure and integrity of mutant molecules, and at the cellular level, we specifically studied the effects of the presence of the R992C collagen II on the biological processes taking place in host cells. Our studies demonstrated that mutant collagen II molecules were characterized by altered electrophoretic mobility, relatively low thermostability, the presence of atypical disulfide bonds, and slow rates of secretion into the extracellular space. Analyses of cellular responses to the presence of the mutant molecules showed that excessive accumulation of thermolabile collagen II was associated with the activation of an unfolded protein response and an increase in apoptosis of host cells. Collectively, these data suggest that molecular mechanisms of SED may be driven not only by structural changes in the architecture of extracellular collagenous matrices, but also by intracellular processes activated by the presence of mutant collagen II molecules

The use of morphometric and fractal parameters to assess the effects of 5-fluorouracil, interferon and dexamethasone treatment on colonic anastomosis healing: an experimental study in rats

Adjuvant chemotherapy and steroid therapy have been demonstrated to interfere with the wound healing process. The aim of this study was to evaluate the effects of 5-fluorouracil, interferon, and dexamethasone, on the healing of colon anastomosis by assessing morphometric and fractal parameters of the colonic wall. An experimental anastomosis of the ascending colon was performed in 60 male Wistar rats, which were then randomly assigned to four groups. On the second to sixth post-operative days, the rats were administered 5-fluorouracil, interferon-α, dexamethasone, or 0.9% NaCl solution as a control. Macroscopic, histomorphometric and microbiological evaluation was performed in order to assess healing of the anastomosis. In three animals from the dexamethasone group, there was leakage of anastomosis; adhesion formation was highest in the interferon group, and significantly higher than in the control and 5-fluorouracil groups. Histomorphometric parameter alterations were most pronounced on the seventh and fourteenth post-operative days in all treatment groups, with submucosal thickness the most affected parameter. Connective tissue fractal dimension was significantly decreased in those animals treated with interferon and dexamethasone. All three pharmaceutical agents impaired healing of anastomosis, and promoted infection in the anastomosis and skin wound sites. As dexamethasone induced both morphometric and macroscopic alterations, it was considered the most detrimental in this study. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 1, pp. 80–89

BET bromodomain inhibitors suppress inflammatory activation of gingival fibroblasts and epithelial cells from periodontitis patients

BET bromodomain proteins are important epigenetic regulators of gene expression that bind acetylated histone tails and regulate the formation of acetylation-dependent chromatin complexes. BET inhibitors suppress inflammatory responses in multiple cell types and animal models, and protect against bone loss in experimental periodontitis in mice. Here, we analyzed the role of BET proteins in inflammatory activation of gingival fibroblasts (GFs) and gingival epithelial cells (GECs). We show that the BET inhibitors I-BET151 and JQ1 significantly reduced expression and/or production of distinct, but overlapping, profiles of cytokine-inducible mediators of inflammation and bone resorption in GFs from healthy donors (IL6, IL8, IL1B, CCL2, CCL5, COX2, and MMP3) and the GEC line TIGK (IL6, IL8, IL1B, CXCL10, MMP9) without affecting cell viability. Activation of mitogen-activated protein kinase and nuclear factor-κB pathways was unaffected by I-BET151, as was the histone acetylation status, and new protein synthesis was not required for the anti-inflammatory effects of BET inhibition. I-BET151 and JQ1 also suppressed expression of inflammatory cytokines, chemokines, and osteoclastogenic mediators in GFs and TIGKs infected with the key periodontal pathogen Porphyromonas gingivalis. Notably, P. gingivalis internalization and intracellular survival in GFs and TIGKs remained unaffected by BET inhibitors. Finally, inhibition of BET proteins significantly reduced P. gingivalis-induced inflammatory mediator expression in GECs and GFs from patients with periodontitis. Our results demonstrate that BET inhibitors may block the excessive inflammatory mediator production by resident cells of the gingival tissue and identify the BET family of epigenetic reader proteins as a potential therapeutic target in the treatment of periodontal disease