TOLERANCE AND IMMUNITY TO MATERNALLY DERIVED INCOMPATIBLE IgG2a-GLOBULIN IN MICE

Progeny mice were confronted with maternal γ-globulin of a different allotype by either back-cross mating, intercross mating, or by foster nursing. In all cases, many mice subsequently produced alloantibodies directed against the incompatible maternal type of IgG2a-globulin. In one series of experiments, immunologic tolerance to the maternally derived γ-globulin was demonstrated to exist in the period before formation of spontaneous antibody. The state of tolerance was then lost, unless maintenance injections of foreign γ-globulin were given. These studies demonstrate in a natural situation that maternally derived foreign proteins can first induce a state of immunological tolerance which is followed, after disappearance of the antigen, by a state of immunity. As such, this parallels the experimental induction of tolerance to foreign proteins by neonatal injections

DEFECTS IN HEMATOPOIETIC DIFFERENTIATION IN NZB AND NZC MICE

Hematopoietic stem cell activity in inbred NZB and NZC mice has been determined by transplantation and endogenous spleen colony assays. Whereas NZB mice show normal colony-forming unit (CFU) activity in the transplantation assay, they show markedly elevated endogenous CFU. NZC mice also show this markedly elevated endogenous CFU activity, but in the transplantation assay show only about 5–10% of normal CFU counts. When NZC stem cells are tested for CFU activity in irradiated recipients of the H-2d type, almost normal colony numbers occur. NZB stem cells however also cannot form colonies in NZC mice. These results suggest that NZC mice have a defect in the micro-environment of the spleen which renders them incapable of allowing transplanted CFU to form colonies. Genetic analysis of both the NZC defect as a CFU recipient, and the elevated endogenous count in NZB and NZC, shows that both are controlled by single recessive genes which are not linked to either coat color, agouti, H-2 or Ig loci. Of even more relevance is the finding that these hematopoietic abnormalities are not linked to the genes involved in controlling autoantibody formation to red cells in the NZB mice. These mice therefore appear to show two distinct hematopoietic abnormalities, the analysis of which may be of considerable value in understanding the detailed events of hematopoietic stem cell differentiation

SURFACE IMMUNOGLOBULINS ON THYMUS AND THYMUS-DERIVED LYMPHOID CELLS

Lymphoid cells from thymus, thoracic duct lymph (TDL), and thoracic duct lymph in irradiated animals reconstituted with allogeneic thymus cells (TTDL) were labeled with radioiodinated anti-immunoglobulins using radioautographic techniques. Thymus and TTDL were labeled (14.4 and 37.0%, respectively) with anti-light chain protein after prolonged exposures (30–60 days). No labeling was observed on thymus and TTDL with anti-polyheavy chain globulin. In contrast 18.5–19.0% of TDL labeled on short exposure (6 days) with anti-polyheavy chain and anti-light chain materials. It is proposed that the difference between the labeling observed on short exposures versus long exposures can be related to the difference in surface density of immunoglobulins between nonthymus-derived (B) and thymus-derived (T) cells. The distribution of labeled cells in the thymus was preferentially among the larger cells (greater than 10 µ diameter). The TTDL population was mostly composed of a larger, blast-like population and the distribution of label was independent of size. As the thymus and TTDL preparations contain almost exclusively T cells, this represents a direct demonstration of surface immunoglobulin light chains on T lymphoid cells

Quantitation of cell surface antigen density by flow cytometry

Correlations between two cellular parameters measured by flow cytometry can be visualized rapidly in contour or isometric data displays. In certain cases, the ratio of two measured parameters has biological meaning and yields the distribution of a new cellular property that cannot be directly measured. In this study, the parameters measured were cell volume and the fluorescence of FITC-labeled antibody to cell surface antigen sites. Cell volume raised to the two-thirds power yields a measure of surface area. On a cell-by-cell basis, the density of a surface antigen is determined by computing the ratio of fluorescence to surface area. Statistical comparisons between calculated density distributions can be made on an absolute as well as a relative basis

Ratio of neutrophilic CD64 and monocytic HLA-DR: a novel parameter in diagnosis and prognostication of neonatal sepsis

Objective: Approaches to monitoring of sepsis have traditionally relied upon the pro-inflammatory component of the sepsis response. This study evaluated the diagnostic and prognostic potential of the ratio of neutrophilic CD64 (nCD64) and monocytic HLA-DR (mHLA-DR) median fluorescence index in monitoring of neonatal sepsis. Methods: Blood from 100 neonates suspected of sepsis and 29 healthy controls was collected on clinical suspicion of sepsis, and the expression of nCD64, mHLA-DR was evaluated by Flow Cytometry; thereby, a derived parameter “Sepsis index,” SI = nCD64/mHLA-DR × 100 was estimated. Results: At day 1, sensitivity and specificity to detect sepsis using nCD64 was 73.01% and 89.18%, respectively, while for SI it was 73.01% and 72.22%, respectively. On Kaplan-Meier analysis, neonates with SI > cut-off showed a higher 30 day-mortality than those with low SI (P = 0.096). On multivariate analysis, the factor associated with mortality in our cohort was Apgar score ≤3, while SI showed a trend toward significance. Conclusions: At day1, nCD64 is useful for the diagnosis of neonatal sepsis whereas mHLA-DR is beneficial for monitoring patients at a later time point. The SI is a marker of moderate diagnostic sensitivity and supplements the current arsenal of laboratory investigations to detect neonatal sepsis. As a marker of prognosis, a high SI shows a trend towards greater mortality

Predictive value of cell-surface markers in infections in critically ill patients: protocol for an observational study (ImmuNe FailurE in Critical Therapy (INFECT) Study).

INTRODUCTION: Critically ill patients are at high risk of nosocomial infections, with between 20% and 40% of patients admitted to the intensive care unit (ICU) acquiring infections. These infections result in increased antibiotic use, and are associated with morbidity and mortality. Although critical illness is classically associated with hyperinflammation, the high rates of nosocomial infection argue for an importance of effect of impaired immunity. Our group recently demonstrated that a combination of 3 measures of immune cell function (namely neutrophil CD88, monocyte HLA-DR and % regulatory T cells) identified a patient population with a 2.4-5-fold greater risk for susceptibility to nosocomial infections. METHODS AND ANALYSIS: This is a prospective, observational study to determine whether previously identified markers of susceptibility to nosocomial infection can be validated in a multicentre population, as well as testing several novel markers which may improve the risk of nosocomial infection prediction. Blood samples from critically ill patients (those admitted to the ICU for at least 48 hours and requiring mechanical ventilation alone or support of 2 or more organ systems) are taken and undergo whole blood staining for a range of immune cell surface markers. These samples undergo analysis on a standardised flow cytometry platform. Patients are followed up to determine whether they develop nosocomial infection. Infections need to meet strict prespecified criteria based on international guidelines; where these criteria are not met, an adjudication panel of experienced intensivists is asked to rule on the presence of infection. Secondary outcomes will be death from severe infection (sepsis) and change in organ failure. ETHICS AND DISSEMINATION: Ethical approval including the involvement of adults lacking capacity has been obtained from respective English and Scottish Ethics Committees. Results will be disseminated through presentations at scientific meetings and publications in peer-reviewed journals. TRIAL REGISTRATION NUMBER: NCT02186522; Pre-results.Innovate UK (formerly Technology Strategy Board) (Grant ID: 15457-108136), Becton Dickinson bioscience, NHS Lothian via the Edinburgh Health Services Research Unit, National Institute of Academic AnaesthesiaThis is the final version of the article. It first appeared from BMJ Publishing Group via http://dx.doi.org/10.1136/bmjopen-2016-01132