Differential binding cell-SELEX method to identify cell-specific aptamers using high-throughput sequencing

AbstractAptamers have in recent years emerged as a viable alternative to antibodies. High-throughput sequencing (HTS) has revolutionized aptamer research by increasing the number of reads from a few (using Sanger sequencing) to millions (using an HTS approach). Despite the availability and advantages of HTS compared to Sanger sequencing, there are only 50 aptamer HTS sequencing samples available on public databases. HTS data in aptamer research are primarily used to compare sequence enrichment between subsequent selection cycles. This approach does not take full advantage of HTS because the enrichment of sequences during selection can be due to inefficient negative selection when using live…

Nano-engineered skin mesenchymal stem cells: potential vehicles for tumour-targeted quantum-dot delivery

Nanotechnology-based drug design offers new possibilities for the use of nanoparticles in imaging and targeted therapy of tumours. Due to their tumour-homing ability, nano-engineered mesenchymal stem cells (MSCs) could be utilized as vectors to deliver diagnostic and therapeutic nanoparticles into a tumour. In the present study, uptake and functional effects of carboxyl-coated quantum dots QD655 were studied in human skin MSCs. The effect of QD on MSCs was examined using a cell viability assay, Ki67 expression analysis, and tri-lineage differentiation assay. The optimal conditions for QD uptake in MSCs were determined using flow cytometry. The QD uptake route in MSCs was examined via fluore…

Effects of malvidin, cyanidin and delphinidin on human adipose mesenchymal stem cell differentiation into adipocytes, chondrocytes and osteocytes.

Abstract Background Anthocyanidins are plant phytochemicals found at high concentrations in berries, vegetables and flowers. Anthocyanidins have been extensively investigated due to their antioxidative, antidiabetic and anti-inflammatory effects. Few studies show that anthocyanidins decrease obesity and improve bone density. However, the effects of anthocyanidins on tissue regeneration have not been sufficiently clarified. Human mesenchymal stem cells (MSCs) are multipotent adult stem cells responsible for the regeneration of fat, bone and cartilage. Although MSCs are often used for screening of biologically active compounds, so far, the effect of anthocyanidins on MSC differentiation has n…

Injury-activated glial cells promote wound healing of the adult skin in mice

Cutaneous wound healing is a complex process that aims to re-establish the original structure of the skin and its functions. Among other disorders, peripheral neuropathies are known to severely impair wound healing capabilities of the skin, revealing the importance of skin innervation for proper repair. Here, we report that peripheral glia are crucially involved in this process. Using a mouse model of wound healing, combined with in vivo fate mapping, we show that injury activates peripheral glia by promoting de-differentiation, cell-cycle re-entry and dissemination of the cells into the wound bed. Moreover, injury-activated glia upregulate the expression of many secreted factors previously…

Photoluminescence ZnO nanorod biosensors for medical and food safety applications

Photoluminescence (PL) ZnO nanorods (NR) are prospect materials for biosensor applications. They can be deposited on solid state substrates and/or dispersed in liquid media. ZnO NRs demonstrate strong UV emission (3.28–3.3 eV), which depends on surface charge, formed by adsorbed molecules. In the present work we report on ZnO NRs, deposited on glass slides, used as biosensor templates for detection food pathogens. Change of ZnO PL after interaction with biomolecules was recorded. Dependence of PL intensity vs concentration of biomolecules was obtained. In the second part, ZnO nanorods were used as PL biomarkers. A suspension of ZnO NRs, conjugated with SSEA-4 antibodies (aSSEA-4) in PBS buf…

Betulin binds to melanocortin receptors and antagonizes alpha-melanocyte stimulating hormone induced cAMP generation in mouse melanoma cells.

Betulin is a principal component of birch bark and is known to possess a broad range of biological activities, including antiinflammatory, antiviral and anticancer actions. The present study was carried out in vitro to clarify the influence of betulin on melanocortin (MC) receptor-ergic signalling by using COS-7 cells transfected with corresponding human MC receptor DNA. The results showed that betulin binds to the human melanocortin MC1, three to five receptors with selectivity to the MC1 subtype (K(i) value 1.022 +/- 0.115 microM). Betulin binds to the MC receptors with the following potency order-MC > MC3 > MC5 > MC4. Betulin itself does not stimulate cAMP generation, however, it slightl…

Antioxidative, hypoglycaemic and hepatoprotective properties of five Vaccinium spp. berry pomace extracts

Wild

Graphical abstract

Flow cytometric analysis of glyoxalase-1 expression in human leukocytes.

Altered glyoxalase-1 (GLO-1) activity and expression is associated with the development of late diabetic complications, malignancy and oxidative stress- and aging-related diseases. In the present study, we developed a flow cytometry method for GLO-1 detection in human leukocytes isolated from peripheral blood samples to investigate GLO-1 expression in leukocyte subsets from type 1 and 2 diabetes mellitus patients (n = 11) and healthy subjects (n = 8). The flow cytometry analysis of GLO-1 in leukocytes showed that expression index of GLO-1-positive cells was slightly increased in mononuclear leukocytes from diabetic patients. This result correlated with the increase in GLO-1 activity in the …

Adult Stem Cell-Derived Extracellular Vesicles in Cancer Treatment: Opportunities and Challenges

Adult stem cells (SCs) participate in tissue repair and homeostasis regulation. The relative ease of SC handling and their therapeutic effect has made of these cell popular candidates for cellular therapy. However, several problems interfere with their clinical application in cancer treatment, like safety issues, unpredictable pro-tumour effects, and tissue entrapment. Therefore cell-free therapies that exhibit SC properties are being investigated. It is now well known that adult SCs exhibit their therapeutic effect via paracrine mechanisms. In addition to secretory proteins, SCs also release extracellular vesicles (EV) that deliver their contents to the target cells. Cancer treatment is on…

In vivophage display: identification of organ-specific peptides using deep sequencing and differential profiling across tissues

ABSTRACTIn vivophage display is widely used for identification of organ- or disease-specific homing peptides. However, the currentin vivophage biopanning approaches fail to assess biodistribution of specific peptide phages across tissues during the screen, thus necessitating laborious and time-consuming post-screening validation studies on individual peptide phages. Here, we adopted bioinformatics tools used for RNA sequencing for analysis of high throughput sequencing (HTS) data to estimate the representation of individual peptides during biopanningin vivo. The data fromin vivophage screen were analyzed using differential binding – relative representation of each peptide in the target orga…

Nanoparticle delivery to metastatic breast cancer cells by nanoengineered mesenchymal stem cells

We created a 3D cell co-culture model by combining nanoengineered mesenchymal stem cells (MSCs) with the metastatic breast cancer cell line MDA-MD-231 and primary breast cancer cell line MCF7 to explore the transfer of quantum dots (QDs) to cancer cells. First, the optimal conditions for high-content QD loading in MSCs were established. Then, QD uptake in breast cancer cells was assessed after 24 h in a 3D co-culture with nanoengineered MSCs. We found that incubation of MSCs with QDs in a serum-free medium provided the best accumulation results. It was found that 24 h post-labelling QDs were eliminated from MSCs. Our results demonstrate that breast cancer cells efficiently uptake QDs that a…

In vivo phage display: identification of organ-specific peptides using deep sequencing and differential profiling across tissues.

Abstract In vivo phage display is widely used for identification of organ- or disease-specific homing peptides. However, the current in vivo phage biopanning approaches fail to assess biodistribution of specific peptide phages across tissues during the screen, thus necessitating laborious and time-consuming post-screening validation studies on individual peptide phages. Here, we adopted bioinformatics tools used for RNA sequencing for analysis of high-throughput sequencing (HTS) data to estimate the representation of individual peptides during biopanning in vivo. The data from in vivo phage screen were analyzed using differential binding—relative representation of each peptide in the target…

Crystallized nano-sized alpha-tricalcium phosphate from amorphous calcium phosphate: microstructure, cementation and cell response

New insight on the conversion of amorphous calcium phosphate (ACP) to nano-sized alpha tricalcium phosphate (α-TCP) provides a faster pathway to calcium phosphate bone cements. In this work, synthesized ACP powders were treated with either water or ethanol, dried, crystallized between 700 and 800 °C, and then cooled at different cooling rates. Particle size was measured in a scanning electron microscope, but crystallite size calculated by Rietveld analysis. Phase composition and bonding in the crystallized powder was assessed by x-ray diffraction and Fourier-transform infrared spectroscopy. Results showed that 50 nm sized α-TCP formed after crystallization of lyophilized powders. Water trea…

Primary culture of avian embryonic heart forming region cells to study the regulation of vertebrate early heart morphogenesis by vitamin A

Background: Important knowledge about the role of vitamin A in vertebrate heart development has been obtained using the vitamin A-deficient avian in ovo model which enables the in vivo examination of very early stages of vertebrate heart morphogenesis. These studies have revealed the critical role of the vitamin A-active form, retinoic acid (RA) in the regulation of several developmental genes, including the important growth regulatory factor, transforming growth factor-beta2 (TGFβ2), involved in early events of heart morphogenesis. However, this in ovo model is not readily available for elucidating details of molecular mechanisms determining RA activity, thus limiting further examination o…

Formation of Calcium-Deficient Hydroxyapatite via Hydrolysis of Nano-Sized Pure Α-Tricalcium Phosphate

Nano-sized pure α-tricalcium phosphate (α-TCP) fabricated by a novel synthesis approach shows great potential for a faster transformation into calcium-deficient hydroxyapatite (CDHA) than conventionally prepared α-TCP. In this work, amorphous tricalcium phosphate precursors were precipitated and treated with a solvent (water or ethanol), and dried (freeze-dried and oven-dried) before heating at 775 °C. Nanosized α-TCP powders were investigated for their phase composition and crystallinity, particle shape and size, reactivity and cellular biocompatibility. Reaction with water showed faster CDHA formation for freeze-dried powder, at 6 hours, compared to ethanol treated powders, whereas a high…

Pharmacological research on natural substances in Latvia: Focus on lunasin, betulin, polyprenol and phlorizin

In this concise review the current research in plant bioactive compound studies in Latvia is described. The paper summarizes recent studies on substances from edible plants (e.g., cereals and apples) or their synthetic analogues, such as peptide lunasin, as well as substances isolated from inedible plants (e.g., birch and conifer), such as pentacyclic triterpenes (e.g., betulin, betulinic acid, and lupeol) and polyprenols. Latvian researchers have been first to demonstrate the presence of lunasin in triticale and oats. Additionally, the impact of genotype on the levels of lunasin in cereals was shown. Pharmacological studies have revealed effects of lunasin and synthetic triterpenes on the …

Bioanalytical system for detection of cancer cells with photoluminescent ZnO nanorods

Using photoluminescent ZnO nanorods and carbohydrate marker SSEA-4, a novel cancer cell recognition system was developed. Immobilization of SSEA-4 antibodies (αSSEA-4) on ZnO nanorods was performed in buffer solution (pH = 7.1) over 2 h. The cancer cell line probes were fixed on the glass slide. One hundred microliters of ZnO-αSSEA-4 conjugates were deposited on the cell probe and exposed for 30 min. After washing photoluminescence spectra were recorded. Based on the developed methodology, ZnO-αSSEA-4 probes were tested on patient-derived breast and colorectal carcinoma cells. Our data clearly show that the carbohydrate SSEA-4 molecule is expressed on cancer cell lines and patient-derived c…

Isolation, cultivation and characterization of human somatic stem cells from adult skin, adipose tissue and bone marrow

Isolation, cultivation and characterization of human somatic stem cells from adult skin, adipose tissue and bone marrow

Pharmacy Practice and Education in Latvia

Pharmacies in Bulgaria have a monopoly on the dispensing of medicinal products that are authorized in the Republic of Bulgaria, as well as medical devices, food additives, cosmetics, and sanitary/hygienic articles. Aptekari (pharmacists) act as responsible pharmacists, pharmacy owners, and managers. They follow a five year Masters of Science in Pharmacy (M.Sc. Pharm.) degree course with a six month traineeship. Pomoshnik-farmacevti (assistant pharmacists) follow a three year degree with a six month traineeship. They can prepare medicines and dispense OTC medicines under the supervision of a pharmacist. The first and second year of the M.Sc. Pharm. degree are devoted to chemical sciences, ma…

DNA damage causes TP53-dependent coupling of self-renewal and senescence pathways in embryonal carcinoma cells.

Recent studies have highlighted an apparently paradoxical link between self-renewal and senescence triggered by DNA damage in certain cell types. In addition, the finding that TP53 can suppress senescence has caused a re-evaluation of its functional role in regulating these outcomes. To investigate these phenomena and their relationship to pluripotency and senescence, we examined the response of the TP53-competent embryonal carcinoma (EC) cell line PA-1 to etoposide-induced DNA damage. Nuclear POU5F1/OCT4A and P21CIP1 were upregulated in the same cells following etoposide-induced G 2M arrest. However, while accumulating in the karyosol, the amount of OCT4A was reduced in the chromatin fract…

Dispensing of the psychotropic drugs via community pharmacies during 2005–2006 years

Skin-derived mesenchymal stem cells as quantum dot vehicles to tumors

Dominyka Dapkute,1,2 Simona Steponkiene,1 Danute Bulotiene,1 Liga Saulite,3 Una Riekstina,3 Ricardas Rotomskis1,4 1Biomedical Physics Laboratory, National Cancer Institute, Vilnius, Lithuania; 2Institute of Biosciences, Vilnius University, Vilnius, Lithuania; 3Faculty of Medicine, University of Latvia, Riga, Latvia; 4Biophotonics Group of Laser Research Center, Faculty of Physics, Vilnius University, Vilnius, Lithuania Purpose: Cell-mediated delivery of nanoparticles is emerging as a new method of cancer diagnostics and treatment. Due to their inherent regenerative properties, adult mesenchymal stem cells (MSCs) are naturally attracted to wounds and sites of inflammation, as well as tumors.…