Bromelain-loaded nanocomposites decrease inflammatory and cytotoxicity effects of gliadin on Caco-2 cells and peripheral blood mononuclear cells of celiac patients.

Enzyme therapy can be an appropriate treatment option for celiac disease (CeD). Here, we developed Bromelain-Loaded Nanocomposites (BLNCs) to improve the stability and retention of bromelain enzyme activity. After the characterization of BLNCs, the cytotoxicity of BLNCs was determined on the Caco-2 cell line. The effect of BLNCs on gliadin degradation and the production of pro-inflammatory cytokines and anti-inflammatory molecules in peripheral blood mononuclear cells (PBMCs) obtained from celiac patients were assessed. Furthermore, the expression of CXCR3 and CCR5 genes was measured in CaCo-2 cells treated with gliadin, gliadin-digested with BLNCs, and bromelain. Our study demonstrated that the Bromelain entrapment efficiency in these nanoparticles was acceptable, and BLNCs have no toxic effect on cells. SDS-PAGE confirmed the digestion effect of bromelain released from nanocomposites. When Caco-2 cells were treated with gliadin digested by free bromelain and BLNCs, the expression of CXCR3 and CCR5 genes was significantly decreased. PBMCs of celiac patients treated with Bromelain and BLNCs decreased inflammatory cytokines (IL-1ß, IL-6, TNF-α, and IFN-γ) production compared to untreated PBMCs. This treatment also increased IL-10 and CTLA-4 in PBMCs of CeD patients. According to the promising results of this study, we can hope for the therapeutic potential of BLNCs for CeD.

Bromelain and ficin proteolytic effects on gliadin cytotoxicity and expression of genes involved in cell-tight junctions in Caco-2 cells.

Enzyme therapy for celiac disease (CeD), which digests gliadin into non-immunogenic and non-toxic peptides, can be an appropriate treatment option for CeD. Here, we have investigated the effectiveness of bromelain and ficin on gliadin digestion using in vitro, such as SDS-PAGE, HPLC, and circular dichroism (CD). Furthermore, the cytotoxicity of gliadin and 19-mer peptide before and after digestion with these enzymes was evaluated using the MTT assay in the Caco-2 cell line. Finally, we examined the effect of these treatments along with Larazotide Acetate on the expression of genes involved in cell-tight junctions, such as Occludin, Claudin 3, tight junction protein-1, and Zonulin in the Caco-2 cell line. Our study demonstrated bromelain and ficin digestion effects on the commercial and wheat-extracted gliadin by SDS-PAGE, HPLC, and CD. Also, the cytotoxicity results on Caco-2 showed that toxicity of the gliadin and synthetic 19-mer peptide was decreased by adding bromelain and ficin. Furthermore, the proteolytic effects of bromelain and ficin on gliadin indicated the expression of genes involved in cell-tight junctions was improved. This study confirms that bromelain and ficin mixture could be effective in improving the symptoms of CeD.

Pectin-cellulose hydrogel, silk fibroin and magnesium hydroxide nanoparticles hybrid nanocomposites for biomedical applications.

Natural polymers are at the center of materials development for biomedical and biotechnological applications based on their biocompatibility, low-toxicity and biodegradability. In this study, a novel nanobiocomposite based on cross-linked pectin-cellulose hydrogel, silk fibroin, and Mg(OH)2 nanoparticles was designed and synthesized. After extensive physical-chemical characterization, the biological response of pectin-cellulose/silk fibroin/Mg(OH)2 nanobiocomposite scaffolds was evaluated by cell viability, red blood cells hemolytic and anti-biofilm assays. After 3 days and 7 days, the cell viability of this nanobiocomposite scaffold was 65.5% and 60.5% respectively. The hemolytic effect was below 20%. Furthermore, the presence of silk fibroin and Mg(OH)2 nanoparticles allowed to enhance the anti-biofilm activity, inhibiting the P. aeruginosa biofilm formation.

Alignment-based design and synthesis of new antimicrobial Aurein-derived peptides with improved activity against Gram-negative bacteria and evaluation of their toxicity on human cells.

Considering the worldwide increasing prevalence of resistance to traditional antibiotics, it is necessary to find new antibiotics to deal with this issue. Recently, antimicrobial peptides (AMPs) have been proposed as new antimicrobial agents. Aureins are a family of AMPs that are isolated from Green and Golden Bell Frogs. These peptides have a favorable antibacterial activity against Gram-positive bacteria. We designed two peptides derived from natural Aurein enjoying alignment-based design method. After synthesis of the peptides, their secondary structure was checked by circular dichroism. Consequently, the antibacterial effects of these peptides were investigated by determining the minimum inhibitory concentration (MIC) and bactericidal concentration. Eventually, the toxicity of these peptides was determined by MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) assay on normal human skin cells (Hu02 cell line). Natural Aurein1.2 was used as a natural control to compare the properties in all stages. The results indicated that these new peptides had medium-upward antimicrobial activity against Escherichia coli, Pseudomonas aeruginosa, and Bacillus subtilis (MIC of 8-64 µg/mL) and weak bactericidal activity against Staphylococcus aureus (MIC of 128-256 µg/mL). Also, MTT assays results showed that AureinN2 is less toxic than AureinN1 and Aurein1.2. Toxicity of AureinN2 for Hu02 cell lines was between 20 and 40% at the concentration of 8-500 µg/mL. In this study, we were able to improve antimicrobial activity of two synthetic derivatives of the Aurein family against Gram-negative bacteria by using machine-learning algorithm and other in silico methods.