Citotoxicidad in vitro del polipéptido antimicrobiano nisina sobre células tumorales sanguíneas / In vitro cytotoxicity of the antimicrobial polypeptide nisin on blood tumor cells

RESUMEN: Las bacteriocinas son péptidos antimicrobianos de síntesis ribosomal secretadas por bacterias. Dentro de estas destaca nisina que posee potenciales usos en terapias antibióticas, como biopreservante de alimentos y probióticos. También se ha descrito que nisina posee citotoxicidad sobre líneas celulares neoplásicas, pero existe poca información de su efecto sobre células tumorales sanguíneas. Debido al potencial uso que presenta nisina, es relevante determinar la toxicidad que presenta sobre líneas celulares tumorales del tipo sanguíneo. Para esto, se realizaron ensayos de actividad hemolítica sobre eritrocitos humanos y de toxicidad sobre células mononucleares de sangre periférica humanas, determinándose que nisina no posee efecto citotóxico sobre este tipo de células normales humanas sanguíneas. Se realizaron también, ensayos de citotoxicidad con líneas celulares tumorales (K562 y U937), con el fin de determinar dosis, tiempo de exposición y selectividad en el efecto tóxico de nisina sobre las células tumorales humanas. Estos ensayos muestran que nisina presenta actividad citotóxica sobre líneas celulares K562 y U937 a las 72 h de exposición, a una concentración de 40 µg/mL, que corresponde a 100 veces la concentración mínima inhibitoria (MIC) usada para su acción sobre bacterias. Al comparar el efecto de nisina sobre células mononucleares de sangre periférica humanas con las líneas tumorales linfoides y mieloides (K562 y U937 respectivamente), se observa un efecto selectivo de nisina sobre las células tumorales sanguíneas.

SUMMARY: Bacteriocins are antimicrobial peptides of ribosomal synthesis secreted by bacteria. Among these, nisin stands out, which has potential uses in antibiotic therapies, as a food bio preservative and probiotics. Nisin has also been reported to have cytotoxicity on neoplastic cell lines, but there is little information on its effect on blood tumor cells. Due to the potential use that nisin presents, it is relevant to determine the toxicity it presents on tumor cell lines of the blood type. For this, hemolytic activity tests were carried out on human erythrocytes and toxicity on human peripheral blood mononuclear cells, determining that nisin does not have a toxic effect on this type of normal human blood cells. Cytotoxicity tests were also carried out with tumor cell lines (K562 and U937), to determine dose, exposure time and selectivity in the toxic effect of nisin on human tumor cells. These tests show that nisin shows cytotoxic activity on K562 and U937 cell lines at 72 h of exposure, at a concentration of 40 µg / mL, which corresponds to 100 times the minimum inhibitory concentration (MIC) used for its action on bacteria. When comparing the effect of nisin on human peripheral blood mononuclear cells with lymphoid and myeloid tumor lines (K562 and U937 respectively), a selective effect of nisin on blood tumor cells is observed.

[Study of the photocatalytic effects of nitrogen-doped titanium dioxide nanoparticles on growth inhibition and apoptosis induction in K562 cell line]

Titanium dioxide [TiO[2]] nanoparticles have shown anti-tumor activity in several cancer cell lines. TiO[2] is an effective photocatalyst that needs ultraviolet [UV] light for activation. Many efforts are in progress to improve the TiO[2] photocatalytic effects by carrying out superficial alterations in its electronic structure. For this purpose, nitrogen [N] element seems to be a proper dopant to improve photocatalytic activity of this nanoparticle. In the present study, the growth inhibitory and apoptotic effects of N-doped TiO[2] nanoparticles on human K562 cells, as an experimental model of chronic myeloid leukemia [CML], were investigated and compared with those of undoped TiO[2] in both light and dark conditions. Nanocrystalline powders of N-doped TiO[2] [80 nm] were synthesized by mechanical mixing of urea with TiO[2] powders under special experimental conditions. The structure and properties of N-doped TiO[2] nanoparticles were confirmed by transmission electron microscopy [TEM], X-ray diffraction [XRD] and Brunauer-Emmett-Teller [BET] analysis. Then we investigated the growth inhibitory and apoptotic effects of these nanoparticles on leukemia K562 cells. The results showed that visible light-irradiated N-doped TiO[2] induced profound growth inhibition and apoptosis in K562 cells in a time- and dose- dependent manner. For example, N-doped TiO[2] at concentrations of 0.01 micro g/ml, 0.1microg/ml, 1 microg/ml, 5 microg/ml and 10microg/ml inhibited growth of K562 cells by 25%, 34%, 39%, 44% and 47% and viability by 11%, 22%, 30%, 35% and 38%, respectively after 48h. Furthermore, the results of DNA fragmentation assay confirmed apoptosis in treated-K562 cells after 48h. While un-doped TiO[2] did not show any inhibitory effects on the growth and viability of K562 cells. Considering the growth inhibitory and apoptotic effects of N-doped TiO[2] in human K562 cells, this nanoparticle can be a potential candidate for photocatalytic therapy in CML in the future studies.

Phorbol myristate acetate [PMA] induced megakaryocytic differentiation of K562 cells from human chronic myelogenous leukaemia patient

To observe the K562 cell line derived from a patient of chronic myelogenous leukemia differentiated into megakaryocytes by growing in the presence of phorbol myristate acetate [PMA]. The differentiation process of K562 cells was monitored by the expression of a platelet cell marker, CD61 through immunocytochemistry using mouse alkaline phosphatase antialkaline phosphatase [APAAP] complex employing fast red TR as substrate, crystal violet and MTT assay used for cell growth analysis. The crystal in the presence of PMA, cells obtained were of large size and less in number as compared to cells incubated without PMA where they were of smaller size and more in number and immunochemical reaction used to detect the presence of CD61, a platelet cell marker that is expressed during differentiation of K562 cells to megakaryocytes. The results showed that the addition of PMA to the growing culture of K562 cell lines induced differentiation, observed through CD61 expression and increase in cell size and cessation of proliferation

Multidrug resistance in tumour cells: characterization of the multidrug resistant cell line K562-Lucena 1

Multidrug resistance to chemotherapy is a major obstacle in the treatment of cancer patients. The best characterised mechanism responsible for multidrug resistance involves the expression of the MDR-1 gene product, P-glycoprotein. However, the resistance process is multifactorial. Studies of multidrug resistance mechanisms have relied on the analysis of cancer cell lines that have been selected and present cross-reactivity to a broad range of anticancer agents. This work characterises a multidrug resistant cell line, originally selected for resistance to the Vinca alkaloid vincristine and derived from the human erythroleukaemia cell K562. This cell line, named Lucena 1, overexpresses P-glycoprotein and have its resistance reversed by the chemosensitisers verapamil, trifluoperazine and cyclosporins A, D and G. Furthermore, we demonstrated that methylene blue was capable of partially reversing the resistance in this cell line. On the contrary, the use of 5-fluorouracil increased the resistance of Lucena 1. In addition to chemotherapics, Lucena 1 cells were resistant to ultraviolet A radiation and hydrogen peroxide and failed to mobilise intracellular calcium when thapsigargin was used. Changes in the cytoskeleton of this cell line were also observed