Interactive effects of surface topography and pulsatile electrical field stimulation on orientation and elongation of fibroblasts and cardiomyocytes

In contractile tissues such as myocardium, functional properties are directly related to the cellular orientation and elongation. Thus, tissue engineering of functional cardiac patches critically depends on our understanding of the interaction between multiple guidance cues such as topographical, adhesive or electrical. The main objective of this study was to determine the interactive effects of contact guidance and electrical field stimulation on elongation and orientation of fibroblasts and cardiomyocytes, major cell populations of the myocardium. Polyvinyl surfaces were abraded using lapping paper with grain size 1-80 microm, resulting in V-shaped abrasions with the average abrasion peak-to-peak width in the range from 3 to 13 microm, and the average depth in the range from 140 to 700 nm (AFM). The surfaces with abrasions 13 microm wide and 700 nm deep, exhibited the strongest effect on neonatal rat cardiomyocyte elongation and orientation as well as statistically significant effect on orientation of fibroblasts, thus they were utilized for electrical field stimulation. Electrical field stimulation was performed using a regime of relevance for heart tissue in vivo as well as for cardiac tissue engineering. Stimulation (square pulses, 1 ms duration, 1 Hz, 2.3 or 4.6 V/cm) was initiated 24 h after cell seeding and maintained for additional 72 h. The cover slips were positioned between the carbon rod electrodes such that the abrasions were either parallel or perpendicular to the field lines. Non-abraded surfaces were utilized as controls. Field stimulation did not affect cell viability. The presence of a well-developed contractile apparatus in neonatal rat cardiomyocytes (staining for cardiac Troponin I and actin filaments) was identified in the groups cultivated on abraded surfaces in the presence of field stimulation. Overall we observed that (i) fibroblast and cardiomyocyte elongation on non-abraded surfaces was significantly enhanced by electrical field stimulation, (ii) electrical field stimulation promoted orientation of fibroblasts in the direction perpendicular to the field lines when the abrasions were also placed perpendicular to the field lines and (iii) topographical cues were a significantly stronger determinant of cardiomyocyte orientation than the electrical field stimulation. The orientation and elongation response of cardiomyocytes was completely abolished by inhibition of actin polymerization (Cytochalasin D) and only partially by inhibition of phosphatidyl-inositol 3 kinase (PI3K) pathway (LY294002).     

Obtaining patient-specific point model of the human ilium bone in the case of incomplete volumetric data using the method of parametric regions

In this paper, we present the methodology for determining the point model of the ilium bone in cases when volumetric data of the whole bone are not available. An extreme traumatic bone damage, osteoporosis, destruction of bone tissue by malignant bone tumors or the existence of only 2D medical image (X-ray) can be the reason for the lack of complete volumetric data. Points on the bone surface were defined at the curves that run through 26 previously defined parameters, at the edges of anteroposterior (A–P) and lateral projections and at the parts of the surface between some parameters. Those parts of the surface, enclosed by parameters, represent ten parametric regions. The values of coordinates, which represent the input data in the statistical program, were measured in a uniquely defined coordinate system. After establishing the correlations between the values of coordinates, 8869 different linear and nonlinear regression models were obtained. The prediction values for point coordinates were calculated and exported to a CAD program. Results obtained were tested on a randomly chosen male right ilium bone, applying the methodology for creating the prediction model using the method of parametric regions, which allows creating a complete polygonal model, for each region separately or just for some parts of the region. Results obtained in the form of regression equations for the right ilium bone can be applied to the left ilium bone. The results of the research were verified using a comparative deviation and distance analysis between the initial and obtained polygonal models.     

Immunoglobulins from sera of antiphospholipid syndrome patients are internalized in the HTR-8/SVneo cell line and cytotrophoblast in culture

Women with antiphospholipid syndrome (APS) experience pregnancy complications mostly due to impaired trophoblast cell functions. Antiphospholipid antibodies (aPL) affect extravillous trophoblast in vivo and in culture, but the mechanisms are still poorly understood. Previously, syncytiotrophoblast was shown to bind and internalize aPL, which was not replicated for extravillous cytotrophoblast in short term culture. Here, aPL binding and time dependent internalization was demonstrated with exposure to aPL in the extravillous cell line HTR-8/SVneo and isolated first trimester of pregnancy cytotrophoblast (CT) using immunocytochemistry and flow cytometry. Intracellular aPL were detectable from 2?h of culture, reaching 30.7?±?3.1% (p?<?0.001) positive cells in CT and 24.8?±?7% (p?<?0.01) in HTR-8/SVneo cells at 24?h and 33?±?4.2% (p?<?0.01) at 48?h. The data presented show that extravillous trophoblast cells internalize aPL in a time-dependent manner significantly more than control immunoglobulins after 24?h of exposure.     

Generation of a poor prognostic chronic lymphocytic leukemia-like disease model: PKCα subversion induces up-regulation of PKCβII expression in B lymphocytes

Overwhelming evidence identifies the microenvironment as a critical factor in the development and progression of chronic lymphocytic leukemia, underlining the importance of developing suitable translational models to study the pathogenesis of the disease. We previously established that stable expression of kinase dead protein kinase C alpha in hematopoietic progenitor cells resulted in the development of a chronic lymphocytic leukemia-like disease in mice. Here we demonstrate that this chronic lymphocytic leukemia model resembles the more aggressive subset of chronic lymphocytic leukemia, expressing predominantly unmutated immunoglobulin heavy chain genes, with upregulated tyrosine kinase ZAP-70 expression and elevated ERK-MAPK-mTor signaling, resulting in enhanced proliferation and increased tumor load in lymphoid organs. Reduced function of PKCα leads to an up-regulation of PKCβII expression, which is also associated with a poor prognostic subset of human chronic lymphocytic leukemia samples. Treatment of chronic lymphocytic leukemia-like cells with the selective PKCβ inhibitor enzastaurin caused cell cycle arrest and apoptosis both in vitro and in vivo, and a reduction in the leukemic burden in vivo. These results demonstrate the importance of PKCβII in chronic lymphocytic leukemia-like disease progression and suggest a role for PKCα subversion in creating permissive conditions for leukemogenesis.     

New protic ionic liquids for fungi and bacteria removal from paper heritage artefacts

In this work, new protic ionic liquids (PILs) with 1-ammonium-2-propanol cation and nine different anions: formate (For), acetate (Ac), lactate (Lac), trifluoroacetate (TFA), chloroacetate (ClA), trichloroacetate (TClA), 3-chloropropionate (3-ClP), 4-chlorobutyrate (4-ClB) and mandelate (Man) were prepared in order to study their antimicrobial activity and possible application for fungi and bacteria removal from deteriorated paper heritage. Ten filamentous fungal strains isolated from specific pigmented area of the damaged books: Trichoderma sp., Cladosporium sp., Penicillium sp.⁽¹⁻³⁾, Penicillium citrinum, Aspergillus sp.^(1,2), Aspergillus flavus, Fusarium graminearum, eight Gram positive and Gram negative ATCC bacterial strains: B. subtilis⁽⁶⁶³³⁾, S. aureus⁽⁶⁵³⁸⁾, E. faecalis^(19433), K. rhizophila⁽⁹³⁴¹⁾, E. coli^(11229), S. enteritidis^(13076), P. mirabilis^(12453), P. aeruginosa^(15692) and two yeast Candida strains: Candida albicans^{(ATCC 10231)} and Candida albicans^(L) were used in this study. The results indicated that antimicrobial activity of selected ionic liquids is significantly affected by the size and specific functional groups in the anion structure. These facts opened the possibility for molecular design of new ionic liquids with strong inhibition properties against the specific bacterial, mould and yeast strains. The significant antimicrobial properties observed in this research suggest that studied PILs may have potential applications in the paper art and artefact cleaning and conservation replacing thus, conventional solvents and organic substances that are toxic for humans and environment.

New protic ionic liquids with 1-ammonium-2-propanol cation and nine different anions were prepared in order to study their antimicrobial activity and possible application for fungi and bacteria removal from deteriorated paper heritage.