Biomedical subjects

Bogdan J Nowicki

Publications and source records attributed to Bogdan J Nowicki.

6 recordsLinked to original sources

PubMed2006

The role of decay-accelerating factor as a receptor for Helicobacter pylori and a mediator of gastric inflammation.

Persistent gastritis induced by Helicobacter pylori is the strongest known risk factor for peptic ulcer disease and distal gastric adenocarcinoma, a process for which adherence of H. pylori to gastric epithelial cells is critical. Decay-accelerating factor (DAF), a protein that protects epithelial cells from complement-mediated lysis, also functions as a receptor for several microbial pathogens. In this study, we investigated whether H. pylori utilizes DAF as a receptor and the role of DAF within H. pylori-infected gastric mucosa. In vitro studies showed that H. pylori adhered avidly to Chinese hamster ovary cells expressing human DAF but not to vector controls. In H. pylori, disruption of the virulence factors vacA, cagA, and cagE did not alter adherence, but deletion of DAF complement control protein (CCP) domains 1-4 or the heavily O-glycosylated serine-threonine-rich COOH-terminal domain reduced binding. In cultured gastric epithelial cells, H. pylori induced transcriptional up-regulation of DAF, and genetic deficiency of DAF attenuated the development of inflammation among H. pylori-infected mice. These results indicate that DAF may regulate H. pylori-epithelial cell interactions relevant to pathogenesis.

Animals↗

PubMed2005

Pyelonephritic Escherichia coli expressing P fimbriae decrease immune response of the mouse kidney.

P fimbriae are proteinaceous appendages on the surface of Escherichia coli bacteria that mediate adherence to uroepithelial cells. E. coli that express P fimbriae account for the majority of ascending urinary tract infections in women with normal urinary tracts. The hypothesis that P fimbriae on uropathic E. coli attach to renal epithelia and may regulate the immune response to establish infection was investigated. The polymeric Ig receptor (pIgR), produced by renal epithelia, transports IgA into the urinary space. Kidney pIgR and urine IgA levels were analyzed in a mouse model of ascending pyelonephritis, using E. coli with (P+) and without (P-) P fimbriae, to determine whether P(+) E. coli regulate epithelial pIgR expression and IgA transport into the urine. (P+) E. coli establish infection and persist to a greater amount than P(-) E. coli. P(+)-infected mice downregulate pIgR mRNA and protein levels compared with P(-)-infected or PBS controls at > or =48 h. The decrease in pIgR was associated with decreased urinary IgA levels in the P(+)-infected group at 48 h. pIgR mRNA and protein also decline in P(+) E. coli-infected LPS-hyporesponsive mice. These studies identify a novel virulence mechanism of E. coli that express P fimbriae. It is proposed that P fimbriae decrease pIgR expression in the kidney and consequently decrease IgA transport into the urinary space. This may explain, in part, how E. coli that bear P fimbriae exploit the immune system of human hosts to establish ascending pyelonephritis.

Animals↗

PubMed2005

Hydrophilic domain II of Escherichia coli Dr fimbriae facilitates cell invasion.

Uropathogenic and diarrheal Escherichia coli strains expressing adhesins of the Dr family bind to decay-accelerating factor, invade epithelial cells, preferentially infect children and pregnant women, and may be associated with chronic or recurrent infections. Thus far, no fimbrial domain(s) that facilitates cell invasion has been identified. We used alanine scanning mutagenesis to replace selected amino acids in hydrophilic domain II of the structural fimbrial subunit DraE and evaluated recombinant mutant DraE for attachment, invasion, and intracellular compartmentalization. The mutation of amino acids V28, T31, G33, Q34, T36, and P40 of DraE reduced or abolished HeLa cell invasion but did not affect attachment. Electron micrographs showed a stepwise entry and fusion of vacuoles containing Escherichia coli mutants T36A and Q34A or corresponding beads with lysosomes, whereas vacuoles with wild-type Dr adhesin showed no fusion. Mutants T31A and Q34A, which were deficient in invasion, appeared to display a reduced capacity for clustering decay-accelerating factor. Our findings suggest that hydrophilic domain II may be involved in cell entry. These data are consistent with the interpretation that in HeLa cells the binding and invasion phenotypes of Dr fimbriae may be separated.

Adhesins, Bacterial↗

PubMed2004

Epithelial invasion by Escherichia coli bearing Dr fimbriae is controlled by nitric oxide-regulated expression of CD55.

We previously reported that inhibition of nitric oxide (NO) increases the rate of bacteremia and maternal mortality in pregnant rats with uterine infection by Escherichia coli expressing the Dr fimbria (Dr(+)). Epithelial binding and invasion by Dr(+) E. coli has also been shown to be dependent upon the expression level of the cellular receptor decay-accelerating factor (DAF; CD55). Here, we hypothesize that NO-related severity of infection could be mediated by changes in DAF expression and in the rate of epithelial invasion. The cellular basis of NO effects on epithelial invasion with Dr(+) E. coli was studied using Ishikawa endometrial carcinoma cells as an in vitro model of the human endometrial epithelium. Initially, we show that Ishikawa cells produce NO and express both NO synthase enzymes, NOS II and NOS III, and DAF protein. We next tested the abilities of both Dr(+) E. coli and a Dr(-) E. coli mutant to invade Ishikawa cells, and invasion was seen only with Dr(+) E. coli. Invasion by Dr(+) E. coli was decreased by elevated NO production and increased by NO inhibition. Elevated NO production significantly decreased DAF protein and mRNA expression in Ishikawa cells in a time- and dose-dependent manner. Here, we propose that in vitro invasion of an epithelial cell line is directly related to NO-regulated expression of DAF. The significance of NO-regulated receptor-ligand invasion is that it may represent a novel unrecognized phenomenon of epithelial defense against infection.

Base Sequence↗

PubMed2002

Structure-function analysis of decay-accelerating factor: identification of residues important for binding of the Escherichia coli Dr adhesin and complement regulation.

Decay-accelerating factor (DAF), a complement regulatory protein, also serves as a receptor for Dr adhesin-bearing Escherichia coli. The repeat three of DAF was shown to be important in Dr adhesin binding and complement regulation. However, Dr adhesins do not bind to red blood cells with the rare polymorphism of DAF, designated Dr(a(-)); these cells contain a point mutation (Ser165-Leu) in DAF repeat three. In addition, monoclonal antibody IH4 specific against repeat three was shown to block both Dr adhesin binding and complement regulatory functions of DAF. Therefore, to identify residues important in binding of Dr adhesin and IH4 and in regulating complement, we mutated 11 amino acids-predominantly those in close proximity to Ser165 to alanine-and expressed these mutations in Chinese hamster ovary cells. To map the mutations, we built a homology model of repeat three based on the poxvirus complement inhibitory protein, using the EXDIS, DIAMOD, and FANTOM programs. We show that perhaps Ser155, and not Ser165, is the key amino acid that interacts with the Dr adhesin and amino acids Gly159, Tyr160, and Leu162 and also aids in binding Dr adhesin. The IH4 binding epitope contains residues Phe148, Ser155, and L171. Residues Phe123 and Phe148 at the interface of repeat 2-3, and also Phe154 in the repeat three cavity, were important for complement regulation. Our results show that residues affecting the tested functions are located on the same loop (148 to 171), at the same surface of repeat three, and that the Dr adhesin-binding and complement regulatory epitopes of DAF appear to be distinct and are approximately 20 A apart.

Adhesins, Escherichia coli↗

PubMed2002

In vivo and in vitro evidence suggest that HB-EGF regulates endometrial expression of human decay-accelerating factor.

Human endometrium expresses the critical complement component C3 in a cyclic fashion, with the highest expression in the secretory phase. As activated complement can kill cells, self or foreign, the secretory endometrial epithelium protects itself by concomitant expression of complement-protective proteins. The objectives of our present study were to describe the spatial and temporal regulation of the complement-protective protein decay-accelerating factor (DAF) in human endometrium and to identify local regulators of its expression. To describe the cyclic regulation of DAF, immunohistochemistry was performed using the IH4 monoclonal antibody on secretory phase endometrial biopsies taken from normal fertile volunteers in LH-timed cycles (n = 114). DAF expression in human endometrium was predominantly localized to the apical membrane of glandular and luminal epithelium. DAF expression, as assessed by histological scoring analysis, was minimal in the proliferative and early secretory phases and increased markedly on approximately day LH +7 (lumen) and LH +8 (glands). Maximal expression was seen in both glands and lumen by LH +8, and this persisted into menses. Using the RL95-2 endometrial epithelial cancer cell line as a model system, we next examined the cellular regulation of DAF. Treatment with E2 and progesterone, alone or in combination, had little effect on DAF expression. Heparin-binding epidermal growth factor-like growth factor (HB-EGF) treatment increased cell surface and total DAF protein, increasing the signal by 260% on flow cytometry and by 200% on Western blot analysis. Stimulation of DAF protein expression was dose dependent, with maximal expression seen at 1 ng/ml. The stimulatory effects of HB-EGF were also observed at the mRNA level. EGF had effects similar to those of HB-EGF on DAF mRNA and protein expression, suggesting that the HB-EGF effect was mediated at least in part by the Her1 EGF receptor subunit. These studies suggest that DAF expression in the midsecretory phase is stimulated by HB-EGF or other members of the EGF family and may function to protect the epithelial integrity of human endometrium in the face of increased complement expression.

CD55 Antigens↗