Marzena Jaworska-Kik

Phytic Acid Modulates In Vitro IL-8 and IL-6 Release from Colonic Epithelial Cells Stimulated with LPS and IL-1β

Phytic acid (PA), a major fiber-associated component of wheat bran and legumes, is physiologically present in the human large gut. The aim of this study was to examine the role of PA in immunologic function of intestinal epithelial cells by analyzing its effect on interleukin (IL)-8 and IL-6 secretion by colonocytes and its role in the response of these cells to bacterial lipopolysaccharides (LPS) and IL-1β. The human colon cell line Caco-2 was exposed to LPS isolated from two strains of Desulfovibrio desulfuricans, wild intestinal and type soil strains, as well as to LPS from E. coli. Cells were also treated with IL-1β and with a combination of LPS and IL-1β. PA had a suppressive effect on IL-8 basal release and it dose dependently reduced IL-8 secretion by colonocytes stimulated with LPS and IL-1β. On the contrary, PA increased constitutive IL-6 secretion and exhibited differentiated effects on LPS responsiveness of cells depending on its concentration and LPS origin. PA was also an efficient down-regulator of IL-6 secretion stimulated by binary actions of LPS and IL-1β. The ability of PA to modulate IL-8 and IL-6 release suggests that PA present in the intestinal milieu may exert immunoregulatory effects on colonic epithelium under physiological conditions or during microbe-induced infection/inflammation in order to maintain the colonic mucosa in a noninflammatory state or to counteract infection.

Biological Activity of Desulfovibrio desulfuricans Lipopolysaccharides Evaluated via Interleukin-8 Secretion by Caco-2 Cells

BACKGROUND Although Desulfovibrio desulfuricans species, besides existing in the natural environment, is also found in the human digestive tract, no information is currently available on its role in the intestinal ecosystem and its activity in regard to the intestinal mucosa. Bacterial products (lipopolysaccharides, LPSs) are generally known for their ability to trigger inflammatory response by stimulating cytokine expression, such as interleukin-8 (IL-8). METHODS Colonic Caco-2 cells were exposed to LPSs isolated from the soil type and intestinal wild strains of D. desulfuricans bacteria. The amount of IL-8 secreted was measured by ELISA. The effects of sodium butyrate and cell preincubation with sodium butyrate on the IL-8 secretion in response to LPSs were also analysed. RESULTS LPSs from D. desulfuricans down-regulated IL-8 secretion by the cells. Incubation of these cells with butyrate alone resulted in a dose-dependent stimulation of IL-8 release. Butyrate also modulated IL-8 secretion by cells stimulated with LPSs. CONCLUSIONS Our findings suggest the lack of inflammatory response of intestinal mucosa in the presence of LPSs of D. desulfuricans. This response can be conditioned by the natural bacterial product, butyrate, which exerts a stimulatory effect on the IL-8 secretion and modulates its release in response to LPSs.Background: Although Desulfovibrio desulfuricans species, besides existing in the natural environment, is also found in the human digestive tract, no information is currently available on its role in the intestinal ecosystem and its activity in regard to the intestinal mucosa. Bacterial products (lipopolysaccharides, LPSs) are generally known for their ability to trigger inflammatory response by stimulating cytokine expression, such as interleukin-8 (IL-8). Methods: Colonic Caco-2 cells were exposed to LPSs isolated from the soil type and intestinal wild strains of D. desulfuricans bacteria. The amount of IL-8 secreted was measured by ELISA. The effects of sodium butyrate and cell preincubation with sodium butyrate on the IL-8 secretion in response to LPSs were also analysed. Results: LPSs from D. desulfuricans down-regulated IL-8 secretion by the cells. Incubation of these cells with butyrate alone resulted in a dose-dependent stimulation of IL-8 release. Butyrate also modulated IL-8 secretion by cells stimulated with LPSs. Conclusions: Our findings suggest the lack of inflammatory response of intestinal mucosa in the presence of LPSs of D. desulfuricans. This response can be conditioned by the natural bacterial product, butyrate, which exerts a stimulatory effect on the IL-8 secretion and modulates its release in response to LPSs.

Chemical composition of Desulfovibrio desulfuricans lipid A

Lipopolysaccharides also called endotoxins are an integral component of the outer membrane of Gram-negative bacteria. When released from the bacterial surface, they interact with a host immune system, triggering excessive inflammatory response. Lipid A is the biologically most active part of endotoxin, and its activity is modulated by the quantity, quality and arrangement of its fatty acids. Desulfovibrio desulfuricans is sulfate-reducing, Gram-negative bacterium that is supposed to be opportunistic pathogens of humans and animals. In the present study, chemical composition of lipid A from various strains of D. desulfuricans was analyzed by gas chromatography/mass spectrometry. It was found that the fatty acid component of the lipid A contains dodecanoic, tetradecanoic, 3-hydroxytetradecanoic and hexadecanoic acids, and its carbohydrate core is composed of glucosamine. The analysis of 3-acyloxyacyl residue of the lipid A revealed the presence of amide-bound 3-(dodecanoyloxy)tetradecanoic and 3-(hexadecanoyloxy)tetradecanoic acids and ester-bound 3-(tetradecanoyloxy)tetradecanoic acid. It was concluded that both fatty acid and 3-acyloxyacyl residue profiles of the lipid A from the studied bacteria were similar to those of E. coli and S.enterica.

The Chemical Composition of Endotoxin Isolated from Intestinal Strain of Desulfovibrio desulfuricans

Desulfovibrio desulfuricans anaerobes are constituents of human alimentary tract microflora. There are suggestions that they take part in the pathogenesis of periodontitis and some gastrointestinal inflammatory disorders, such as ulcerative colitis or Crohns disease. Endotoxin is one of Gram-negative bacteria cellular components that influence these microorganisms pathogenicity. Endotoxin is a lipid-polisaccharide heteropolymer consisting of three elements: lipid A, core oligosaccharide, and O-specific polysaccharide, also called antigen-O. The biological activity of lipopolysaccharide (LPS) is determined by its structure. In this study, we show that rhamnose, fucose, mannose, glucose, galactose, heptose, and 2-keto-3-deoxyoctulosonic acid (Kdo) are constituents of D. desulfuricans endotoxin oligosaccharide core and O-antigen. Lipid A of these bacteria LPS is composed of glucosamine disaccharide substituted by 3-acyloxyacyl residues: ester-bound 3-(dodecanoyloxy)tetradecanoic, 3-(hexadecanoyloxy)tetradecanoic acid, and amide-bound 3-(tetradecanoyloxy)tetradecanoic acid.

Quantitative evaluation of transcriptional activation of NF-κB p65 and p50 subunits and IκBα encoding genes in colon cancer cells by Desulfovibrio desulfuricans endotoxin

Quantification of p65, p50 and IκBα mRNAs was performed by real time QRT-PCR in Caco-2 cells treated with 10, 50, and 100 μg/mL of Desulfovibrio desulfuricans LPS for 1, 6, 12, and 24 h. A strong increase in expression of p65 and IλBα genes was induced by 10 and 100 μg/mL of LPS at 1 h; after 6 h higher transcript amounts of both genes were observed at 100 μg/mL LPS. The p65 expression level was significantly increased by 50 and 100 μg/mL at 12 h and lowered by all LPS doses at 24 h. No significant differences between IκBα mRNA quantity in cells exposed to LPS at 12 and 24 h were observed. No changes in expression of p50 mRNA were induced by LPS. The expression of p65 gene positively correlated with IκBα gene expression. D. desulfuricans LPS is capable of modulating transcriptional activity of p65 and IκBα genes in intestinal epithelial cells.

Biofilm Formation on NiTi Surface by Different Strains of Sulphate Reducing Bacteria (Desulfovibrio desulfuricans)

Bacteria of Desulfovibrio genus belong to group of widespread sulphate-reducing bacteria (SRB). D. desulfuricans is considered one among many bacterial species involved in microbiologically influenced corrosion (MIC) of metals, mainly of stainless steels and other alloys. SRB can produce gaseous hydrogen sulphide. This gas is released into the environment leading to formation of metal sulphides that significantly influence electrochemical processes and ultimately enhance the corrosion of materials. Biofilms formed by these bacteria are especially harmful for highly alloyed steels and many alloys. The aim of this work was to compare the character of growth and biofilm formation by three strains of D. desulfuricans (standard soil strain DSM and two wild intestinal strains: DV/A and DV/B) on the surface of NiTi alloy.

Effect of Artificial and Inflammatory Saliva on Desulfovibrio desulfuricans Growth and Biofilm Formation on NiTi Alloy

NiTi alloys are used for both medical and veterinary purposes, and also for production of surgical instruments. Sulphate-reducing bacteria (SRB) colonize various anaerobic environments, including human oral cavity. Desulfovibrio desulfuricans is the SRB species responsible for corrosion of many metals including highly alloyed steels as well as titanium and its alloys. The aim of this work was to compare growth of D. desulfuricans biofilms on NiTi alloy submerged in artificial saliva or in inflammatory saliva. The results of investigations showed differences between D. desulfuricans biofilms formed on NiTi alloy in the presence of artificial saliva and inflammatory saliva. The growth medium influenced biofilm structure; inflammatory saliva promotes its formation. The biofilms grown on samples immersed in inflammatory saliva were much thicker as compared with samples emerged in artificial saliva. After 28 days of incubation in inflammatory saliva, plentiful mature biofilm was present on alloy surface.

Initial stage of the biofilm formation on the NiTi and Ti6Al4V surface by the sulphur-oxidizing bacteria and sulphate-reducing bacteria

The susceptibility to the fouling of the NiTi and Ti6Al4V alloys due to the adhesion of microorganisms and the biofilm formation is very significant, especially in the context of an inflammatory state induced by implants contaminated by bacteria, and the implants corrosion stimulated by bacteria. The aim of this work was to examine the differences between the sulphur-oxidizing bacteria (SOB) and sulphate-reducing bacteria (SRB) strains in their affinity for NiTi and Ti6Al4V alloys. The biofilms formed on alloy surfaces by the cells of five bacterial strains (aerobic SOB Acidithiobacillus thiooxidans and Acidithiobacillus ferrooxidans, and anaerobic SRB Desulfovibrio desulfuricans—3 strains) were studied using scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). The protein concentrations in liquid media have also been analyzed. The results indicate that both alloys tested may be colonized by SOB and SRB strains. In the initial stage of the biofilm formation, the higher affinity of SRB to both the alloys has been documented. However, the SOB strains have indicated the higher (although differentiated) adaptability to changing environment as compared with SRB. Stimulation of the SRB growth on the alloys surface was observed during incubation in the liquid culture media supplemented with artificial saliva, especially of lower pH (imitated conditions under the inflammatory state, for example in the periodontitis course). The results point to the possible threat to the human health resulting from the contamination of the titanium implant alloys surface by the SOB (A. thiooxidans and A. ferrooxidans) and SRB (D. desulfuricans).Graphical abstract