Recombinant human lactoferrin induces apoptosis, disruption of F-actin structure and cell cycle arrest with selective cytotoxicity on human triple negative breast cancer cells.

Breast cancer is the most frequently diagnosed cancer among women worldwide. Here, recombinant human lactoferrin (rhLf) expressed in Pichia pastoris was tested for its potential cytotoxic activity on a panel of six human breast cancer cell lines. The rhLf cytotoxic effect was determined via a live-cell HTS imaging assay. Also, confocal microscopy and flow cytometry protocols were employed to investigate the rhLf mode of action. The rhLf revealed an effective CC50 of 91.4 and 109.46 µg/ml on non-metastatic and metastatic MDA-MB-231 cells, with favorable selective cytotoxicity index values, 11.68 and 13.99, respectively. Moreover, rhLf displayed satisfactory SCI values on four additional cell lines, MDA-MB-468, HCC70, MCF-7 and T-47D (1.55-3.34). Also, rhLf provoked plasma membrane blebbing, chromatin condensation and cell shrinkage in MDA-MB-231 cells, being all three apoptosis-related morphological changes. Also, rhLf was able to shrink the microfilaments, forming a punctuated cytoplasmic pattern in both the MDA-MB-231 and Hs-27 cells, as visualized in confocal photomicrographs. Moreover, performing flow cytometric analysis, rhLf provoked significant phosphatidylserine externalization, cell cycle arrest in the S phase and apoptosis-induced DNA fragmentation in MDA-MB-231 cells. Hence, rhLf possesses selective cytotoxicity on breast cancer cells. Also, rhLf caused apoptosis-associated morphologic changes, disruption of F-actin cytoskeleton organization, phosphatidylserine externalization, DNA fragmentation, and arrest of the cell cycle progression on triple-negative breast cancer MDA-MB-231 cells. Overall results suggest that rhLf is using the apoptosis pathway as its mechanism to inflict cell death. Findings warranty further evaluation of rhLf as a potential anti-breast cancer drug option.

Lactoferrin, chitosan and Melaleuca alternifolia-natural products that show promise in candidiasis treatment

Abstract The evolution of microorganisms resistant to many medicines has become a major challenge for the scientific community around the world. Motivated by the gravity of such a situation, the World Health Organization released a report in 2014 with the aim of providing updated information on this critical scenario. Among the most worrying microorganisms, species from the genus Candida have exhibited a high rate of resistance to antifungal drugs. Therefore, the objective of this review is to show that the use of natural products (extracts or isolated biomolecules), along with conventional antifungal therapy, can be a very promising strategy to overcome microbial multiresistance. Some promising alternatives are essential oils of Melaleuca alternifolia (mainly composed of terpinen-4-ol, a type of monoterpene), lactoferrin (a peptide isolated from milk) and chitosan (a copolymer from chitin). Such products have great potential to increase antifungal therapy efficacy, mitigate side effects and provide a wide range of action in antifungal therapy.

Lactoferrin, chitosan and Melaleuca alternifolia-natural products that show promise in candidiasis treatment.

The evolution of microorganisms resistant to many medicines has become a major challenge for the scientific community around the world. Motivated by the gravity of such a situation, the World Health Organization released a report in 2014 with the aim of providing updated information on this critical scenario. Among the most worrying microorganisms, species from the genus Candida have exhibited a high rate of resistance to antifungal drugs. Therefore, the objective of this review is to show that the use of natural products (extracts or isolated biomolecules), along with conventional antifungal therapy, can be a very promising strategy to overcome microbial multiresistance. Some promising alternatives are essential oils of Melaleuca alternifolia (mainly composed of terpinen-4-ol, a type of monoterpene), lactoferrin (a peptide isolated from milk) and chitosan (a copolymer from chitin). Such products have great potential to increase antifungal therapy efficacy, mitigate side effects and provide a wide range of action in antifungal therapy.

Bovine lactoferrin purification from whey using Yellow HE-4R as the chromatographic affinity ligand.

The worldwide production of whey increases by around 186 million tons each year and it is generally considered as a waste, even when several whey proteins have important economic relevance. For its valorization, inexpensive ligands and integrated chromatography methods need to be developed for specific and low-cost protein purification. Here, we describe a novel affinity process with the dye Yellow HE-4R immobilized on Sepharose for bovine lactoferrin purification. This approach based on a low-cost ligand showed an efficient performance for the recovery and purification of bovine lactoferrin directly from whey, with a yield of 71% and a purification factor of 61.

Free secretory component and lactoferrin of human milk inhibit the adhesion of enterotoxigenic Escherichia coli.

The non-immunoglobulin component of human milk responsible for the inhibition of Escherichia coli cell adhesion (haemagglutination) mediated by colonisation factor antigen 1 (CFA1) was determined by chromatographic fractionation of human whey proteins with Sephadex G-200, DEAE cellulose and heparin-sepharose. Pure free secretory component (fSC) and pure lactoferrin (Lf) were isolated and both compounds inhibited the haemagglutination induced by E. coli CFA1+. The lowest concentrations of purified fSC and Lf able to inhibit the haemagglutination induced by E. coli strain TR50/3 CFA1+ were 0.06 mg/ml and 0.1 mg/ml respectively. Commercially available lactoferrin from human milk and transferrin from human serum, which has a great structural analogy to lactoferrin, also inhibited the haemagglutination. The lowest concentrations of the commercial lactoferrin and transferrin able to inhibit the haemagglutination induced by E. coli TR50/3 CFA1+ were 0.03 mg/ml and 0.4 mg/ml, respectively. These results indicate that fSC and Lf may be important non-specific defence factors against enterotoxigenic E. coli infections.

Purificación de lactoferrina humana

La lactoferrina, una glicoproteína capaz de unir 2 átomos de hierro, fuepurificada de la leche materna. El procedimiento descrito recientemente incluye el empleo de una cromatografía de Sepharosa-heparina, la cual separó la lactoferrina del resto de las proteínas del suero con un gradiente lineal de cloruro de sodio. La presencia de un contaminante hizo necesaria la realización de una cromatografía en Sephadex G75, recomendada en otros procedimientos. Una sola banda en la electroforesis en gel de poliacrilamida con SDS, así como una banda única de precipitación en la inmunoelectroforesis, demostraron la pureza de esta proteína. Esta técnica permite obtener la lactoferrina pura, lo que facilitará la obtención de un anticuerpo para el desarrollo de procedimientos para su cuantificación, así como profundizar el estudio de su estructura y función y su uso con fines terapeúticos.