Annette J. Schlueter

Absence of cross-reactivity between murine Ly-6C and Ly-6G

The Ly‐6 family has many members, including Ly‐6C and Ly‐6G. A previous study suggested that the anti‐Ly‐6G antibody, RB6‐8C5, may react with Ly‐6Chi murine bone marrow (BM) cells. This finding has been interpreted as cross‐reactivity of RB6‐8C5 with the Ly‐6C antigen, and has been generalized to many hematopoietic cell types, using the terminology Ly‐6G/C. The present study was undertaken to determine whether anti‐Ly‐6G antibodies truly cross‐react with the Ly‐6C antigen on multiple hematopoietic cell types.

Primed innate immunity leads to autoinflammatory disease in PSTPIP2-deficient cmo mice

The mouse Lupo (I282N) mutation in proline-serine-threonine phosphatase-interacting protein 2 (PSTPIP2) leads to reduced expression of PSTPIP2 that is associated with a macrophage-mediated autoinflammatory disease. Another mutation in PSTPIP2, L98P, termed chronic multifocal osteomyelits (cmo), leads to a disease in mice that resembles chronic recurrent multifocal osteomyelits in humans. The cellular basis of cmo disease was investigated. cmo disease develops independently of lymphocytes and is cured by bone marrow transplantation. Macrophages, mast cells, and osteoclasts from cmo mice fail to express detectable PSTPIP2 protein. Asymptomatic Pstpip2(cmo/cmo) mice have increased circulating levels of macrophage inflammatory protein 1-alpha and interleukin-6, and their macrophages exhibit increased production of these inflammatory mediators, which is normalized by retroviral expression of wild-type PSTPIP2. Spleens of asymptomatic cmo mice contain increased numbers of macrophage precursors, and cmo mice mobilize more macrophage precursors in response to a sterile inflammatory stimulus. Signal transducer and activator of transcription 1 is elevated in cmo splenic macrophages, which also exhibit increased colony-stimulating factor-1-stimulated proliferation and increased extracellular signal-regulated kinase 1/2 phosphorylation. PSTPIP2 overexpression in macrophages leads to the opposite phenotype. Thus, PSTPIP2 deficiency causes both an expansion of macrophage progenitors and increased responsiveness of mature macrophages to activating stimuli, which together prime the organism for exaggerated and sustained responses leading to autoinflammatory disease.

Case Report: Eculizumab Rescue of Severe Accelerated Antibody-Mediated Rejection After ABO-Incompatible Kidney Transplant

ABO-incompatible (ABOI) living donor kidney transplantation has become a well-accepted practice with standard protocols using perioperative antibody-depleting therapies to lower blood group titers to an acceptable threshold for transplantation. However, a subset of patients will experience accelerated antibody-mediated rejection (AMR) after ABOI kidney transplantation and require aggressive intervention to prevent allograft loss. Here in we report the successful use of terminal complement inhibition with eculizumab to rescue an ABOI kidney allograft with accelerated AMR refractory to salvage splenectomy and daily plasmapheresis. This case emphasizes the fact that, despite close postoperative surveillance and aggressive intervention, graft loss from accelerated AMR after ABOI kidney transplantation remains a very real risk. Eculizumab may offer a graft-saving therapeutic option for isolated cases of severe AMR after ABOI kidney transplantation refractory to standard treatment.

Fetal exposure to ethanol has long-term effects on the severity of influenza virus infections.

Alcohol use by pregnant women is a significant public health issue despite well-described risks to the fetus including physical and intellectual growth retardation and malformations. Although clinical studies are limited, they suggest that in utero alcohol exposure also results in significant immune deficiencies in naive neonates. However, little is known about fetal alcohol exposure (FAE) effects on adult infections. Therefore, to determine the long-term effects of FAE on disease susceptibility and the adult immune system, we infected FAE adult mice with influenza virus. In this study, we demonstrate that mice exposed to ethanol during gestation and nursing exhibit enhanced disease severity as well as increased and sustained pulmonary viral titers following influenza virus infection. Secondary exposure to alcohol as an adult further exacerbates these effects. Moreover, we demonstrate that FAE mice have impaired adaptive immune responses, including decreased numbers of virus-specific pulmonary CD8 T cells, a decreased size and frequency of pulmonary B cell foci, and reduced production of influenza-specific Ab following influenza infection. Together, our results suggest that FAE induces significant and long-term defects in immunity and susceptibility to influenza virus infection and that FAE individuals could be at increased risk for severe and fatal respiratory infections.

Characterization of primitive hematopoietic cells from patients with dyskeratosis congenita

Dyskeratosis congenita (DC) is an inherited bone marrow (BM) failure syndrome associated with mutations in telomerase genes and the acquisition of shortened telomeres in blood cells. To investigate the basis of the compromised hematopoiesis seen in DC, we analyzed cells from granulocyte colony-stimulating factor mobilized peripheral blood (mPB) collections from 5 members of a family with autosomal dominant DC with a hTERC mutation. Premobilization BM samples were hypocellular, and percentages of CD34(+) cells in marrow and mPB collections were significantly below values for age-matched controls in 4 DC subjects. Directly clonogenic cells, although present at normal frequencies within the CD34(+) subset, were therefore absolutely decreased. In contrast, even the frequency of long-term culture-initiating cells within the CD34(+) DC mPB cells was decreased, and the telomere lengths of these cells were also markedly reduced. Nevertheless, the different lineages of mature cells were produced in normal numbers in vitro. These results suggest that marrow failure in DC is caused by a reduction in the ability of hematopoietic stem cells to sustain their numbers due to telomere impairment rather than a qualitative defect in their commitment to specific lineages or in the ability of their lineage-restricted progeny to execute normal differentiation programs.

Chronic Ethanol Consumption Decreases Murine Langerhans Cell Numbers and Delays Migration of Langerhans Cells as Well as Dermal Dendritic Cells

BACKGROUND Chronic alcoholics experience increased incidence and severity of infections, the mechanism of which is incompletely understood. Dendritic cells (DC) migrate from peripheral locations to lymph nodes (LN) to initiate adaptive immunity against infection. Little is known about how chronic alcohol exposure affects skin DC numbers or migration. METHODS Mice received 20% EtOH in the drinking water for up to 35 weeks. Baseline Langerhans cell (LC) and dermal DC (dDC) numbers were enumerated by immunofluorescence (IF). LC repopulation after inflammation was determined following congenic bone marrow (BM) transplant and ultraviolet (UV) irradiation. Net LC loss from epidermis was determined by IF following TNF-alpha or CpG stimulation. LC and dDC migration into LN was assessed by flow cytometry following epicutaneous FITC administration. RESULTS Chronic EtOH consumption caused a baseline reduction in LC but not dDC numbers. The deficit was not corrected following transplantation with non-EtOH-exposed BM and UV irradiation, supporting the hypothesis that the defect is intrinsic to the skin environment rather than LC precursors. Net loss of LC from epidermis following inflammation was greatly reduced in EtOH-fed mice versus controls. Ethanol consumption for at least 4 weeks led to delayed LC migration into LN, and consumption for at least 8 weeks led to delayed dDC migration into LN following epicutaneous FITC application. CONCLUSIONS Chronic EtOH consumption causes decreased density of epidermal LC, which likely results in decreased epidermal immunosurveillance. It also results in altered migratory responsiveness and delayed LC and dDC migration into LN, which likely delays activation of adaptive immunity. Decreased LC density at baseline appears to be the result of an alteration in the skin environment rather than an intrinsic LC defect. These findings provide novel mechanisms to at least partially explain why chronic alcoholics are more susceptible to infections, especially those following skin penetration.

Clinical significance of positive cranial bone flap cultures and associated risk of surgical site infection after craniotomies or craniectomies.

OBJECT The risk of surgical site infection (SSI) after craniotomies or craniectomies in patients in whom contaminated bone flaps have been reimplanted has not been determined. The objectives of this study were to identify the prevalence of bone flaps with positive cultures--especially those contaminated with Propionibacterium acnes--to assess the risk of SSI after reimplanting (either during the initial operation or subsequently) bone flaps with positive cultures, and to identify risk factors for SSI following the initial craniotomies or craniectomies. METHODS The authors conducted a retrospective review of cases in which patients underwent craniotomy/craniectomy procedures between January and October 2007 in the neurosurgery department at the University of Iowa Hospitals and Clinics. They also reviewed processes and procedures and did pulsed field gel electrophoresis of P. acnes isolates to look for a common source of contamination. They then conducted a prospective cohort study that included all patients who underwent craniotomy/craniectomy procedures between November 2007 and November 2008 and met the study criteria. For the cohort study, the authors obtained cultures from each patients bone flap during the craniotomy/craniectomy procedures. Data about potential risk factors were collected by circulating nurses during the procedures or by a research assistant who reviewed medical records after the procedures. An infection preventionist independently identified SSIs through routine surveillance using the Centers for Disease Control and Preventions definitions. Univariate and bivariate analyses were performed to determine the association between SSI and potential risk factors. RESULTS The retrospective review did not identify specific breaks in aseptic technique or a common source of P. acnes. Three hundred seventy-three patients underwent 393 craniotomy/craniectomy procedures during the cohort study period, of which 377 procedures met the study criteria. Fifty percent of the bone flaps were contaminated by microorganisms, primarily skin flora such as P. acnes, coagulase-negative staphylococci, and Staphylococcus aureus. Reimplanting bone flaps that had positive culture results did not increase the risk of infection after the initial craniotomy/craniectomy procedures and the subsequent cranioplasty procedures (p = 0.80). Allowing the skin antiseptic to dry before the procedures (p = 0.04, OR 0.26) was associated with lower risk of SSIs. Female sex (p = 0.02, OR = 3.49) was associated with an increased risk of SSIs; Gliadel wafer implants (p = 0.001, OR = 8.38) were associated with an increased risk of SSIs after procedures to treat tumors. CONCLUSIONS Operative factors such as the way the skin is prepared before the incision rather than the skin flora contaminants on the bone flaps may play an important role in the pathogenesis of SSIs after craniotomy/craniectomy. Gliadel wafers significantly increased the risk of SSI after procedures to treat tumors.

Transfusions via hand‐held syringes and small‐gauge needles as risk factors for hyperkalemia

BACKGROUND: Pediatric emergency RBC transfusions are often infused rapidly through 22‐gauge (ga) or smaller needles or catheters using hand‐held syringes. Data relating needle size, unit age, and infusion rate are needed to assess the risk of hemolysis and hyperkalemia in this setting.

Effects of Chronic Ethanol Feeding on Murine Dendritic Cell Numbers, Turnover Rate, and Dendropoiesis

BACKGROUND Chronic alcoholics have increased susceptibility to and severity of infection, which are likely to be a result of impaired immune defense mechanisms. The contribution of dendritic cells (DC) to these immune defense changes is not well understood. Alterations in DC numbers, dendropoiesis, and lifespan have not been specifically studied in vivo in chronic ethanol (EtOH) exposure models. As DC play an essential role in initiating immune responses, alterations in these DC characteristics would help explain changes observed in adaptive immune responses. METHODS Mice received 20% EtOH (w/v) in the drinking water for up to 28 weeks, with mouse chow ad libitum. In EtOH-fed and water control mice, DC were enumerated by flow cytometry. The effect of EtOH on DC precursor numbers was determined by differentiation in vitro in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4, and the effect of an EtOH environment on untreated DC differentiation was measured following bone marrow transfer to irradiated hosts. DC turnover rate was also examined by bromodeoxyuridine incorporation and loss. RESULTS The percentage and absolute numbers of DC were decreased in spleen and increased in thymus beginning as early as 4 weeks of EtOH feeding. In addition, the overall cellularity of spleen and thymus were altered by this regimen. However, chronic EtOH consumption did not adversely affect DC precursor numbers, differentiation abilities, or turnover rates. CONCLUSIONS Decreased splenic DC numbers observed following chronic murine EtOH consumption are not because of altered DC precursor numbers or differentiation, nor increased DC turnover rate. Similarly, increased thymic DC numbers are not the result of alterations in DC precursor differentiation or turnover rate. Compartment size plays a role in determining splenic and thymic DC numbers following chronic EtOH feeding. EtOH-induced alterations in total DC numbers provide several mechanisms to partially explain why chronic alcoholics have increased susceptibility to infections.

Mechanisms by which chronic ethanol feeding limits the ability of dendritic cells to stimulate T cell proliferation

BACKGROUND As initiators of immune responses, dendritic cells (DCs) are required for antigen (Ag)-specific activation of naïve T cells in the defense against infectious agents. The increased susceptibility to and severity of infection seen in chronic alcoholics could be because of impaired DCs initiation of naïve T-cell responses. Specifically, these DCs may not provide adequate Signals 1 (Ag presentation), 2 (costimulation), or 3 (cytokine production) to these T cells. METHODS Using the Meadows-Cook murine model of chronic alcohol abuse, the ability of ethanol (EtOH)-exposed DCs to stimulate T-cell proliferation, acquire and process Ag, express costimulatory molecules, and produce inflammatory cytokines was assessed. RESULTS Normal naïve T cells primed by EtOH-exposed DCs showed decreased proliferation in vitro and in vivo, compared to water-fed control mice. These EtOH-exposed DCs, after activation by CpG or tumor necrosis factor alpha (TNFα), were less able to upregulate costimulatory molecules CD40, CD80, or CD86, and produced less IL-12 p40, TNFα, and IFNα than DCs from water-fed mice. TLR9 and TNF receptor expression were also reduced in/on EtOH-exposed DCs. No evidence of defective Ag acquisition or processing as a result of EtOH feeding was identified. CONCLUSIONS Inadequate proliferation of normal T cells following stimulation by EtOH-exposed DCs is likely a result of diminished Signal 2 and Signal 3. Lack of adequate inflammatory stimulation of EtOH-exposed DCs because of diminished receptors for inflammatory mediators appears to be at least partially responsible for their dysfunction. These findings provide a mechanism to explain increased morbidity and mortality from infectious diseases in alcoholics and suggest targets for therapeutic intervention.