Abstract: Mechanical index (MI) is used for quantifying acoustic cavitation and the relationship between acoustic pressure and the frequency. In this study, modeling of the MI was applied to provide treatment protocol and to understand the effective physical processes on reproducibility of stem cells. The acoustic pressure and MI equations are modeled and solved to estimate optimal MI for 28, 40, 150 kHz and 1 MHz frequencies. Radial and axial acoustic pressure distribution was extracted. To validate the results of the modeling, the acoustic pressure in the water and near field depth was measured by a piston hydrophone. Results of modeling and experiments show that the model is consistent well to experimental results with 0.91 and 0.90 correlation of coefficient (p
Abstract: Tissue engineering includes combination of materials and techniques used for the improvement, repair or replacement of the tissue. Scaffolds, permanent or temporally material, are used as support for the creation of the "new cell structures". For this important component (scaffold), a variety of materials can be used. The advantage of some polymeric materials is their cytocompatibility and possibility of biodegradation. Poly(L-lactic acid) (PLLA) is a biodegradable, semi-crystalline thermoplastic polymer. PLLA can be fully degraded into H2O and CO2. In this experiment, the effect of the surface modification of biodegradable polymer (performed by plasma treatment) on the various cell types was studied. The surface parameters and changes of the physicochemical properties of modified PLLA substrates were studied by different methods. Surface wettability was determined by goniometry, surface morphology and roughness study were performed with atomic force microscopy and chemical composition was determined using photoelectron spectroscopy. The physicochemical properties were studied in relation to cytocompatibility of human osteoblast (MG 63 cells), rat vascular smooth muscle cells (VSMC), and human stem cells (ASC) of the adipose tissue in vitro. A fluorescence microscopy was chosen to study and compare cell-material interaction. Important parameters of the cytocompatibility like adhesion, proliferation, viability, shape, spreading of the cells were evaluated. It was found that the modification leads to the change of the surface wettability depending on the time of modification. Short time of exposition (10-120 s) can reduce the wettability of the aged samples, exposition longer than 150 s causes to increase of contact angle of the aged PLLA. The surface morphology is significantly influenced by duration of modification, too. The plasma treatment involves the formation of the crystallites, whose number increases with increasing time of modification. On the basis of physicochemical properties evaluation, the cells were cultivated on the selected samples. Cell-material interactions are strongly affected by material chemical structure and surface morphology. It was proved that the plasma treatment of PLLA has a positive effect on the adhesion, spreading, homogeneity of distribution and viability of all cultivated cells. This effect was even more apparent for the VSMCs and ASCs which homogeneously covered almost the whole surface of the substrate after 7 days of cultivation. The viability of these cells was high (more than 98% for VSMCs, 89-96% for ASCs). This experiment is one part of the basic research, which aims to easily create scaffolds for tissue engineering with subsequent use of stem cells and their subsequent "reorientation" towards the bone cells or smooth muscle cells.
Abstract: This paper investigates a possible practical solution for reasonable oxygen supply during the pluripotent stem cells expansion processes, where the stem cells propagate as aggregates in stirred-suspension bioreactors. Low glucose and low oxygen concentrations are preferred for efficient proliferation of pluripotent stem cells. However, strong oxygen limitation, especially inside of cell aggregates, can lead to cell starvation and death. In this research, the oxygen concentration profile inside of stem cell aggregates in a stem cell expansion process was predicted using a modified oxygen diffusion model. This profile can be realized during the stem cells cultivation process by manipulating the oxygen concentration in inlet gas or inlet gas flow. The proposed approach is relatively simple and may be attractive for installation in a real pluripotent stem cell expansion processes.
Abstract: In this manuscript, we produced neat electrospun poly(o-methoxyaniline) (POMA) fibers and utilized it for applying the growth of neural stem cells. The transparency and morphology of as-prepared POMA fibers was characterized by UV-visible spectroscopy and scanning electron microscopy, respectively. It was found to have no adverse effects on the long-term proliferation of the neural stem cells (NSCs), retained the ability to self-renew, and exhibit multipotentiality. Results of immunofluorescence staining studies confirmed that POMA electrospun fibers could provide a great environment for NSCs and enhance its differentiation.
Abstract: Image mosaicing techniques are usually employed to offer researchers a wider field of view of microscopic image of biological samples. a mosaic is commonly achieved using automated microscopes and often with one “color" channel, whether it refers to natural or fluorescent analysis. In this work we present a method to achieve three subsequent mosaics of the same part of a stem cell culture analyzed in phase contrast and in fluorescence, with a common non-automated inverted microscope. The mosaics obtained are then merged together to mark, in the original contrast phase images, nuclei and cytoplasm of the cells referring to a mosaic of the culture, rather than to single images. The experiments carried out prove the effectiveness of our approach with cultures of cells stained with calcein (green/cytoplasm and nuclei) and hoechst (blue/nuclei) probes.
Abstract: Image mosaicing is a technique that permits to enlarge the field of view of a camera. For instance, it is employed to achieve panoramas with common cameras or even in scientific applications, to achieve the image of a whole culture in microscopical imaging. Usually, a mosaic of cell cultures is achieved through using automated microscopes. However, this is often performed in batch, through CPU intensive minimization algorithms. In addition, live stem cells are studied in phase contrast, showing a low contrast that cannot be improved further. We present a method to study the flat field from live stem cells images even in case of 100% confluence, this permitting to build accurate mosaics on-line using high performance algorithms.
Abstract: Bone marrow-derived stem cells have been widely
studied as an alternative source of stem cells. Mesenchymal stem
cells (MSCs) were mostly investigated and studies showed MSCs can
promote neurogenesis. Little is known about the non-mesenchymal
mononuclear cell fraction, which contains both hematopoietic and
nonhematopoietic cells, including monocytes and endothelial
progenitor cells. This study focused on unfractionated bone marrow
mononuclear cells (BMMCs), which remained 72 h after MSCs were
adhered to the culture plates. We showed that BMMC-conditioned
medium promoted morphological changes of human SH-SY5Y
neuroblastoma cells from an epithelial-like phenotype towards a
neuron-like phenotype as indicated by an increase in neurite
outgrowth, like those observed in retinoic acid (RA)-treated cells.
The result could be explained by the effects of trophic factors
released from BMMCs, as shown in the RT-PCR results that
BMMCs expressed nerve growth factor (NGF), brain-derived
neurotrophic factor (BDNF), and ciliary neurotrophic factor (CNTF).
Similar results on the cell proliferation rate were also observed
between RA-treated cells and cells cultured in BMMC-conditioned
medium, suggesting that cells creased proliferating and differentiated
into a neuronal phenotype. Using real-time RT-PCR, a significantly
increased expression of tyrosine hydroxylase (TH) mRNA in SHSY5Y
cells indicated that BMMC-conditioned medium induced
catecholaminergic identities in differentiated SH-SY5Y cells.
Abstract: Understanding the consumption and production of
various metabolites of fibroblast conditioned media is needed for its
proper and optimized use in expansion of pluripotent stem cells. For
this purpose, we have used the HPLC method to analyse the
consumption of glucose and the production of lactate over time by
mouse embryonic fibroblasts. The experimental data have also been
compared with mathematical model fits. 0.025 moles of lactate was
produced after 72 hrs while the glucose concentration decreased from
0.017 moles to 0.011 moles. The mathematical model was able to
predict the trends of glucose consumption and lactate production.
Abstract: The mosaicing technique has been employed in more and more application fields, from entertainment to scientific ones. In the latter case, often the final evaluation is still left to human beings, that assess visually the quality of the mosaic. Many times, a lack of objective measurements in microscopic mosaicing may prevent the mosaic from being used as a starting image for further analysis. In this work we analyze three different metrics and indexes, in the domain of signal analysis, image analysis and visual quality, to measure the quality of different aspects of the mosaicing procedure, such as registration errors and visual quality. As the case study we consider the mosaicing algorithm we developed. The experiments have been carried out by considering mosaics with very different features: histological samples, that are made of detailed and contrasted images, and live stem cells, that show a very low contrast and low detail levels.