Validation of an algorithm for semiautomated surveillance to detect deep surgical site infections after primary total hip or knee arthroplasty-A multicenter study.

OBJECTIVE: Surveillance of healthcare-associated infections is often performed by manual chart review. Semiautomated surveillance may substantially reduce workload and subjective data interpretation. We assessed the validity of a previously published algorithm for semiautomated surveillance of deep surgical site infections (SSIs) after total hip arthroplasty (THA) or total knee arthroplasty (TKA) in Dutch hospitals. In addition, we explored the ability of a hospital to automatically select the patients under surveillance. DESIGN: Multicenter retrospective cohort study. METHODS: Hospitals identified patients who underwent THA or TKA either by procedure codes or by conventional surveillance. For these patients, routine care data regarding microbiology results, antibiotics, (re)admissions, and surgeries within 120 days following THA or TKA were extracted from electronic health records. Patient selection was compared with conventional surveillance and patients were retrospectively classified as low or high probability of having developed deep SSI by the algorithm. Sensitivity, positive predictive value (PPV), and workload reduction were calculated and compared to conventional surveillance. RESULTS: Of 9,554 extracted THA and TKA surgeries, 1,175 (12.3%) were revisions, and 8,378 primary surgeries remained for algorithm validation (95 deep SSIs, 1.1%). Sensitivity ranged from 93.6% to 100% and PPV ranged from 55.8% to 72.2%. Workload was reduced by ≥98%. Also, 2 SSIs (2.1%) missed by the algorithm were explained by flaws in data selection. CONCLUSIONS: This algorithm reliably detects patients with a high probability of having developed deep SSI after THA or TKA in Dutch hospitals. Our results provide essential information for successful implementation of semiautomated surveillance for deep SSIs after THA or TKA.

Tolerance to factor VIII in a transgenic mouse expressing human factor VIII cDNA carrying an Arg(593) to Cys substitution.

Inhibitory antibodies develop in approximately 25% of patients with severe hemophilia. A following treatment with factorVIII. In E-16KO or E-17KO mice, in which the factor VIII gene has been inactivated by insertion of a neo cassette, inhibitors develop following administration of factor VIII. Here, we describe the generation of transgenic mice expressing human factor VIII-R593C (huFVIII-R593C). Human factor VIII-R593C cDNA under control of a mouse albumin enhancer/promoter was injected into fertilized oocytes. Analysis of transgenic mice revealed that human factor VIII-R593C was expressed in the liver. Transgenic mice were crossed with factor VIII-deficient mice (E-16KO mice). In plasma of E-16KO mice antibodies were detected after five serial intravenous injections of factor VIII, while plasma of huFVIII-R593C/E-16KO mice did not contain detectable levels of antibodies. No antibody secreting cells were observed in either spleen or bone marrow of huFVIII-R593C/E-16KO mice. Also, factor VIII-specific memory B cells were not observed in the spleen of huFVIII-R593C/E-16KO mice. Analysis of T cell responses revealed that splenocytes derived of E-16KO mice secreted IL-10 and IFN-gamma following restimulation with factor VIII in vitro. In contrast, no factor VIII-specific T cell responses were observed in huFVIII-R593C/E-16KO mice. These results indicate that huFVIII-R593C/E-16KO mice are tolerant to intravenously administered factor VIII. It is anticipated that this model may prove useful for studying immune responses in the context of factor VIII gene therapy.

The mouse homologue of the leukocyte-associated Ig-like receptor-1 is an inhibitory receptor that recruits Src homology region 2-containing protein tyrosine phosphatase (SHP)-2, but not SHP-1.

We report the molecular cloning and characterization of the first leukocyte-associated Ig-like receptor 1 (LAIR-1) homologue in mice that we have named mouse LAIR-1 (mLAIR-1). The mLAIR-1 gene maps to the proximal end of mouse chromosome 7 in a region syntenic with human chromosome 19q13.4 where the leukocyte receptor cluster is located. The protein shares 40% sequence identity with human LAIR-1, has a single Ig-like domain, and contains two immunoreceptor tyrosine-based inhibitory motif-like structures in its cytoplasmic tail. Mouse LAIR-1 is broadly expressed on various immune cells, and cross-linking of the molecule on stably transfected RBL-2H3 and YT.2C2 cells results in strong inhibition of their degranulation and cytotoxic activities, respectively. Upon pervanadate stimulation, the mLAIR-1 cytoplasmic tail becomes phosphorylated, thereby recruiting Src homology region 2-containing tyrosine phosphatase-2. Interestingly, unlike human LAIR-1, Src homology region 2-containing tyrosine phosphatase-1 is not recruited to the mLAIR-1 cytoplasmic tail. Screening human and mouse cell lines for mLAIR-1 and human LAIR-1 binding partners identified several lines expressing putative ligand(s) for both receptors.

Immunobiology of inhibitor development in hemophilia A.

After treatment with factor (F) VIII concentrate a significant number of patients with hemophilia A develop inhibitory antibodies that neutralize FVIII. Epitope mapping revealed that antibodies bind to selected regions within the A2, A3, and C2 domains of FVIII. Anti-A2 and anti-A3 antibodies interfere with assembly of FVIIIa with FIXa, whereas anti-C2 antibodies impede the interaction of FVIII with phospholipids. The immunologic mechanisms underlying inhibitor development in hemophilia A have not been fully elucidated. FVIII is recognized by the immune system as a foreign antigenic substance that evokes the T cell-dependent formation of high-affinity antibodies. Clonal analysis of B cell responses in hemophilia A patients has given further insight into the epitope specificity and molecular characteristics of FVIII inhibitors. Costimulatory blockade of FVIII-reactive T cells in a mouse model for hemophilia A has suggested new approaches for treatment of inhibitor patients. In this article, recent studies on the immunobiology of FVIII inhibitors are summarized and discussed with reference to their potential impact on treatment and prevention of immune responses in patients with hemophilia.

Analysis of factor VIII inhibitors in a haemophilia A patient with an Arg593-->Cys mutation using phage display.

We characterized anti-factor VIII antibodies in a mild haemophilia A patient with an Arg593-->Cys mutation in the A2 domain, using V gene phage-display technology. All isolated single-chain variable-domain antibody fragments were directed against residues Arg484-Ile508, a binding site for factor VIII inhibitors in the A2 domain. After a further period of replacement therapy, a transient rise in inhibitor titre was observed. These antibodies were directed against the A2 domain. Activation of a pre-existing pool of B cells, which express antibodies against residues Arg484-Ile508, could explain the rapid anamnestic response.

Two classes of germline genes both derived from the V(H)1 family direct the formation of human antibodies that recognize distinct antigenic sites in the C2 domain of factor VIII.

Most plasmas from patients with inhibitors contain antibodies that are reactive with the C2 domain of factor VIII. Previously, we have shown that the variable heavy chain (V(H)) regions of antibodies to the C2 domain are encoded by the closely related germline gene segments DP-10, DP-14, and DP-88, which all belong to the V(H)1 gene family. Here, we report on the isolation and characterization of additional anti-C2 antibodies that are derived from V(H) gene segments DP-88 and DP-5. Competition experiments using murine monoclonal antibodies CLB-CAg 117 and ESH4 demonstrated that antibodies derived from DP-5 and DP-88 bound to different sites within the C2 domain. Epitope mapping studies using a series of factor VIII/factor V hybrids revealed that residues 2223 to 2332 of factor VIII are required for binding of the DP-10-, DP-14-, and DP-88-encoded antibodies. In contrast, binding of the DP-5-encoded antibodies required residues in both the amino- and carboxy-terminus of the C2 domain. Inspection of the reactivity of the antibodies with a series of human/porcine hybrids yielded similar data. Binding of antibodies derived from germline gene segments DP-10, DP-14, and DP-88 was unaffected by replacement of residues 2181 to 2243 of human factor VIII for the porcine sequence, whereas binding of the DP-5-encoded antibodies was abrogated by this replacement. Together these data indicate that antibodies assembled from V(H) gene segments DP-5 and the closely related germline gene segments DP-10, DP-14, and DP-88 target 2 distinct antigenic sites in the C2 domain of factor VIII.