No abstract

Binding of cells of Staphylococcus aureus to fibronectin, which may represent a mechanism of host tissue adherence, involves a fibronectin-receptor protein present on the bacterial surface. Cloning of a gene coding for a staphylococcal fibronectin-binding protein and construction of a fusion protein with fibronectin-binding properties was previously reported from our laboratory. We have now sequenced the gene and deduced a primary sequence of the fibronectin-binding protein. The protein resembles other cell-wall-associated proteins on Gram-positive bacteria in that it (i) appears to be anchored in the cell membrane via its C-terminal end, (ii) contains a proline-rich repeating unit outside the membrane anchor, and (iii) contains a long (36-amino acid) signal sequence at the N terminus. The fibronectin-binding activity has been localized to a domain composed of a 38-amino acid unit repeated completely three times and partially a fourth time; the identity between the three 38-amino acid sequences varies from 42 to 87%. Three synthetic peptides mimicking the structure of each 38-amino acid unit were constructed. All three peptides interacted with fibronectin, as indicated by their ability to inhibit binding of fibronectin to staphylococcal cells, whereas an unrelated 37-amino acid peptide showed no inhibitory activity.

A gene encoding a fibronectin binding protein (FnBP) has recently been isolated and sequenced from Staphylococcus uureus strain 8325-4. In the same bacterial strain, 682 bp downstream to the stop codon of this gene VizhA), a second gene termed,fnhB has now been discovered, encoding another FnBP (FnBPB). The two genes show in large parts striking sequence homologies. The complete amino acid sequence encoded by fnbB has been deduced and compared to that deduced fromfizhA. In FnBPB a stretch of 66 amino acids downstream to the signal peptide has 75% identity with the corresponding region in FnBPA. At the C-terminal site another 394 amino acid stretch is almost identical in both gene products. This stretch contains the 38 amino acid long D repeats, the wall spanning Wr repeats and the hydrophobic membrane spanning domain. In FnBPA each of the three D repeats has been identified as a fibronectin binding structure. These structures are highly conserved in

The present investigation reports an animal experiment the objective of which was to study the vascular topography of the periodontium and the peri-implant soft and hard tissues in the beagle dog. 2 beagle dogs were used. The right mandibular premolars were extracted and healing allowed for 3 months. 2 titanium fixtures a.m. Brånemark were installed in the right premolar region. Abutment connection was performed 3 months later. The contra-lateral mandibular sites were used as tooth harboring control units. A 4-month period of meticulous plaque control was initiated. A clinical and radiographical examination was performed towards the end of this period and revealed that the gingiva and peri-implant mucosa were clinically healthy and that the bone tissue at teeth and implants had a normal height. The carotid arteries were perfused with a mixture of carbon and calf serum. 2 bucco-lingual sections, about 100-150 microns thick, and one mesio-distal section comprising the junctional epithelium and underlying connective tissue were sampled. The sections were treated, cleared and examined in a microscope. The results of the present investigation demonstrated that the vasculature of the gingiva and the supracrestal connective tissue at teeth is derived from two sources, namely the supraperiosteal vessels lateral of the alveolar process and the vessels of the periodontal ligament. The blood vessels of the peri-implant mucosa were found to be terminal branches of larger vessels originating from the periosteum of the bone of the implant site.(ABSTRACT TRUNCATED AT 250 WORDS)

Six months of prophylactic oral anticoagulation after a first episode of venous thromboembolism led to a lower recurrence rate than did treatment lasting for six weeks. The difference between the two groups occurred between 6 weeks and 6 months after the start of treatment, and the rates of recurrence remained nearly parallel for 1 1/2 years thereafter.

The gene encoding the fibronectin‐binding protein (FNBP) from Staphylococcus aureus strain 8325‐4 was isolated from a gene bank in pBR322. The original clone, containing a 6.5‐kb insert, gave a functional product present in the periplasm of Escherichia coli. Analysis of polypeptides isolated after affinity chromatography on fibronectin‐Sepharose followed by ion‐exchange chromatography revealed two gene products, 87 and 165 kd in mol. wt. The amino acid compositions of these two polypeptides and a native FNBP from S. aureus strain Newman were very similar. Antibodies raised against the native FNBP from strain Newman precipitated the 125I‐labelled 165‐kd polypeptide, and unlabeled 165‐ and 87‐kd polypeptides as well as native FNBP inhibited the immunoprecipitation reactions. The region of the fnbp‐gene encoding the fibronectin‐binding activity has been identified and subcloned in an expression vector based on the staphylococcal protein A gene. The resulting product in E. coli is an extracellular fusion protein consisting of two IgG‐binding domains of protein A followed by a fibronectin‐binding region. The fusion protein binds to fibronectin and completely inhibits the binding of fibronectin to intact cells of S. aureus.

The mucosal immune system identifies and fights invading pathogens, while allowing non-pathogenic organisms to persist. Mechanisms of pathogen/non-pathogen discrimination are poorly understood, as is the contribution of human genetic variation in disease susceptibility. We describe here a new, IRF3-dependent signaling pathway that is critical for distinguishing pathogens from normal flora at the mucosal barrier. Following uropathogenic E. coli infection, Irf3−/− mice showed a pathogen-specific increase in acute mortality, bacterial burden, abscess formation and renal damage compared to wild type mice. TLR4 signaling was initiated after ceramide release from glycosphingolipid receptors, through TRAM, CREB, Fos and Jun phosphorylation and p38 MAPK-dependent mechanisms, resulting in nuclear translocation of IRF3 and activation of IRF3/IFNβ-dependent antibacterial effector mechanisms. This TLR4/IRF3 pathway of pathogen discrimination was activated by ceramide and by P-fimbriated E. coli, which use ceramide-anchored glycosphingolipid receptors. Relevance of this pathway for human disease was supported by polymorphic IRF3 promoter sequences, differing between children with severe, symptomatic kidney infection and children who were asymptomatic bacterial carriers. IRF3 promoter activity was reduced by the disease-associated genotype, consistent with the pathology in Irf3−/− mice. Host susceptibility to common infections like UTI may thus be strongly influenced by single gene modifications affecting the innate immune response.

BackgroundPolymorphisms affecting Toll-like receptor (TLR) structure appear to be rare, as would be expected due to their essential coordinator role in innate immunity. Here, we assess variation in TLR4 expression, rather than structure, as a mechanism to diversify innate immune responses.Methodology/Principal FindingsWe sequenced the TLR4 promoter (4,3 kb) in Swedish blood donors. Since TLR4 plays a vital role in susceptibility to urinary tract infection (UTI), promoter sequences were obtained from children with mild or severe disease. We performed a case-control study of pediatric patients with asymptomatic bacteriuria (ABU) or those prone to recurrent acute pyelonephritis (APN). Promoter activity of the single SNPs or multiple allelic changes corresponding to the genotype patterns (GPs) was tested. We then conducted a replication study in an independent cohort of adult patients with a history of childhood APN. Last, in vivo effects of the different GPs were examined after therapeutic intravesical inoculation of 19 patients with Escherichia coli 83972. We identified in total eight TLR4 promoter sequence variants in the Swedish control population, forming 19 haplotypes and 29 genotype patterns, some with effects on promoter activity. Compared to symptomatic patients and healthy controls, ABU patients had fewer genotype patterns, and their promoter sequence variants reduced TLR4 expression in response to infection. The ABU associated GPs also reduced innate immune responses in patients who were subjected to therapeutic urinary E. coli tract inoculation.ConclusionsThe results suggest that genetic variation in the TLR4 promoter may be an essential, largely overlooked mechanism to influence TLR4 expression and UTI susceptibility.