Toll-like receptor activation as a biomarker in traumatically injured patients.

Background: Surgical insult and trauma have been shown to cause dysregulation of the immune and inflammatory responses. Interaction of damage-associated molecular patterns (DAMPs) with toll-like receptors (TLRs) initiates innate immune response and systemic inflammatory responses. Given that surgical patients produce high levels of circulating damage-associated molecular patterns, we hypothesized that plasma-activated TLR activity would be correlated to injury status and could be used to predict pathological conditions involving tissue injury.

Expression of Mitochondrial-Encoded Genes in Blood Differentiate Acute Renal Allograft Rejection.

Despite potent immunosuppression, clinical and biopsy confirmed acute renal allograft rejection (AR) still occurs in 10-15% of recipients, ~30% of patients demonstrate subclinical rejection on biopsy, and ~50% of them can show molecular inflammation, all which increase the risk of chronic dysfunction and worsened allograft outcomes. Mitochondria represent intracellular endogenous triggers of inflammation, which can regulate immune cell differentiation, and expansion and cause antigen-independent graft injury, potentially enhancing the development of acute rejection. In the present study, we investigated the role of mitochondrial DNA encoded gene expression in biopsy matched peripheral blood (PB) samples from kidney transplant recipients.

Premature T Cell Senescence in Pediatric CKD.

An individual's immune function, susceptibility to infection, and response to immunosuppressive therapy are influenced in part by his/her T cell maturation state. Although childhood is the most dynamic period of immune maturation, scant information regarding the variability of T cell maturation in children with renal disease is available. In this study, we compared the T cell phenotype in children with renal failure (n=80) with that in healthy children (n=20) using multiparameter flow cytometry to detect markers of T cell maturation, exhaustion, and senescence known to influence immune function.

CMV reactivation drives posttransplant T-cell reconstitution and results in defects in the underlying TCRβ repertoire.

Although cytomegalovirus (CMV) reactivation has long been implicated in posttransplant immune dysfunction, the molecular mechanisms that drive this phenomenon remain undetermined. To address this, we combined multiparameter flow cytometric analysis and T-cell subpopulation sorting with high-throughput sequencing of the T-cell repertoire, to produce a thorough evaluation of the impact of CMV reactivation on T-cell reconstitution after unrelated-donor hematopoietic stem cell transplant. We observed that CMV reactivation drove a >50-fold specific expansion of Granzyme B(high)/CD28(low)/CD57(high)/CD8(+) effector memory T cells (Tem) and resulted in a linked contraction of all naive T cells, including CD31(+)/CD4(+) putative thymic emigrants.

The kSORT assay to detect renal transplant patients at high risk for acute rejection: results of the multicenter AART study.

Background: Development of noninvasive molecular assays to improve disease diagnosis and patient monitoring is a critical need. In renal transplantation, acute rejection (AR) increases the risk for chronic graft injury and failure. Noninvasive diagnostic assays to improve current late and nonspecific diagnosis of rejection are needed.

Multicenter comparison of laboratory performance in cytomegalovirus and Epstein-Barr virus viral load testing using international standards.

Background: Infections with cytomegalovirus (CMV) and Epstein-Barr virus (EBV) remain important in solid organ transplantation. Quantitative viral nucleic acid testing is a major advance to patient management. These assays are limited by a lack of standardization, resulting in viral load measurements that differ among clinical laboratories.

In vivo T cell costimulation blockade with abatacept for acute graft-versus-host disease prevention: a first-in-disease trial.

We performed a first-in-disease trial of in vivo CD28:CD80/86 costimulation blockade with abatacept for acute graft-versus-host disease (aGVHD) prevention during unrelated-donor hematopoietic cell transplantation (HCT). All patients received cyclosporine/methotrexate plus 4 doses of abatacept (10 mg/kg/dose) on days -1, +5, +14, +28 post-HCT. The feasibility of adding abatacept, its pharmacokinetics, pharmacodynamics, and its impact on aGVHD, infection, relapse, and transplantation-related mortality (TRM) were assessed.

Post-transplant lymphoproliferative disorder associated with immunosuppressive therapy for renal transplantation in rhesus macaques (Macaca mulatta).

Human post-transplant lymphoproliferative disorder (PTLD) is an abnormal lymphoid proliferation that arises in 1-12% of transplant recipients as a consequence of prolonged immunosuppression and Epstein-Barr viral infection (EBV). Nonhuman primates, primarily rhesus macaques (Macaca mulatta), have been used extensively in research models of solid organ transplantation, as the nonhuman primate immune system closely resembles that of the human. Lymphocryptovirus of rhesus monkeys has been characterized and shown to be very similar to EBV in humans in regards to its cellular tropism, host immune response, and ability to stimulate B lymphocyte proliferation and lymphomagenesis.

Limiting the amount and duration of antigen exposure during priming increases memory T cell requirement for costimulation during recall.

Donor-reactive memory T cells (Tmem) can play an important role in mediating graft rejection after transplantation. Transplant recipients acquire donor-reactive Tmem not only through prior sensitization with alloantigens but also through previous exposure to environmental pathogens that are cross-reactive with allogeneic peptide-MHC complexes. Current dogma suggests that most, if not all, Tmem responses are independent of the requirement for CD28 and/or CD154/CD40-mediated costimulation to mount a recall response.

Migration and differentiation of canine bone marrow stromal cells transplanted into the developing mouse brain.

To evaluate whether canine bone marrow stromal cells (BMSCs) can migrate and adopt neural phenotypes in the developing mouse brain we transplanted fluorescently labeled BMSCs into the lateral ventricle of immunocompromised neonatal mice. Most fibroblasts, used as a control, and BMSCs isolated from adult dogs remained around the injection site and exhibited a spindle-shaped appearance. A small number of BMSCs from young dogs were found in the subventricular zone, rostral migratory stream, and olfactory bulbs, and retained expression of neuron marker.

The effects of canine bone marrow stromal cells on neuritogenesis from dorsal root ganglion neurons in vitro.

The present in vitro study was designed to evaluate whether canine bone marrow stromal cells (BMSCs) promote neurite outgrowth from dorsal root ganglion (DRG) neurons. Bone marrow aspirates were collected from iliac crests of three young adult dogs. DRG neurons were cultured on BMSCs, fibroblasts, or laminin substrates.

The frequency, growth kinetics, and osteogenic/adipogenic differentiation properties of canine bone marrow stromal cells.

Bone marrow stromal cells (BMSCs) have gained considerable attention as a potential source for cell transplantation therapies for a variety of diseases due to their accessibility, proliferative capacity, and multilineage differentiation properties. Canine BMSCs have been shown to contribute to regeneration of osseous tissues, but knowledge about their biology is currently limited. In the present study, we investigated the frequency of adult canine BMSCs in bone marrow, morphological features, growth kinetics, and osteogenic as well as adipogenic differentiation properties in vitro.

Salmonella Enteritidis-induced alteration of inflammatory CXCL chemokine messenger-RNA expression and histologic changes in the ceca of infected chicks.

To understand better the events in early avian host immune responses to Salmonella Enteritidis (SE), we examined messenger-RNA (mRNA) expression for eight genes: CXCLi1[K60], CXCLi2 [IL-8/CAF], interferon (IFN)-gamma, interleukin (IL)-1beta, IL-6, IL-12alpha, IL-12beta, and gallinacin (Gal)-2 in the ceca of young chicks 1 wk postinoculation with SE. Cecum tissue sections were stained and evaluated for the presence of macrophages, lymphocytes, heterophils, and apoptotic cells following SE infection. With the use of quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), SE infection was associated with a significant (P < 0.

Reduced nitric oxide production and iNOS mRNA expression in IFN-gamma-stimulated chicken macrophages transfected with iNOS siRNAs.

Utilizing RNA interference technology with siRNA in the HD11 macrophage cell line, we determined how the inhibition or knock-down of the iNOS (inducible nitric oxide synthase) gene affected IFN-gamma-induced macrophage production of nitric oxide (NO) and mRNA expression of genes involved in this biological pathway in the chicken. Chicken macrophages produce NO when stimulated with recombinant chicken IFN-gamma, however, when transfected with iNOS siRNAs, the production of NO is significantly decreased. We observed a 14-28% reduction in NO production by IFN-gamma-stimulated HD11 cells at 48h after initial siRNA transfection compared to non-transfected IFN-gamma-stimulated macrophages.

Detection of oligoclonal bands in cerebrospinal fluid from German Shepherd dogs with degenerative myelopathy by isoelectric focusing and immunofixation.

Background: Detection of intrathecal IgG synthesis is important in evaluating inflammatory diseases in the central nervous system. Isoelectric focusing (IEF) is currently the most sensitive method to demonstrate intrathecal IgG synthesis and may have diagnostic value for German Shepherd degenerative myelopathy (GSDM).

Objective: The objective of this study was to adapt a modified IEF and immunofixation method for the detection of oligoclonal bands in cerebrospinal fluid (CSF) from dogs with GSDM.

Nestin-positive spheres derived from canine bone marrow stromal cells generate cells with early neuronal and glial phenotypic characteristics.

Bone marrow stromal cells (BMSCs) isolated from humans and rodents have been shown to generate neural cells under specific culture conditions and after transplantation in the central nervous system. The apparent plasticity of BMSCs has therefore been a target of intensive research in attempt to develop a novel therapy for neurological diseases. Canines sustain neurological disorders (e.

Measurement of myelin basic protein in the cerebrospinal fluid of dogs with degenerative myelopathy.

Background: Analysis of cerebrospinal fluid (CSF) is part of a routine clinical workup in veterinary patients when neurologic disease is suspected. However, knowledge of particular protein markers of disease in CSF is limited. The concentration of myelin basic protein (MBP) in CSF is used as a biochemical marker in humans to evaluate demyelinating lesions in the central nervous system (CNS).

Expression of neural markers on bone marrow-derived canine mesenchymal stem cells.

Objective: To evaluate cell surface markers of bone marrow-derived canine mesenchymal stem cells (MSCs) by use of flow cytometric analysis and determine whether canine MSCs express proteins specific to neuronal and glial cells.

Sample Population: Bone marrow aspirates collected from iliac crests of 5 cadavers of young adult dogs.

Procedures: Flow cytometric analysis was performed to evaluate cell surface markers and homogeneity of third-passage MSCs.

Breed effect on early cytokine mRNA expression in spleen and cecum of chickens with and without Salmonella enteritidis infection.

We examined mRNA expression of 11 genes: BAK, Bcl-x, Interferon [IFN]-gamma, Interleukin [IL]-1beta, IL-6, IL-10, IL-12alpha, IL-12beta, IL-18, CXCLi2 [IL-8/CAF], and a MIP family chemokine, CCLi2, in the spleen and cecum of day-old chicks after oral inoculation with Salmonella enteritidis (SE) or medium. Three distinct chicken breeds (broiler, Fayoumi, and Leghorn) were evaluated for mRNA expression levels at 2 and 18h post-inoculation using quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). SE exposure significantly increased splenic IL-18 and IFN-gamma expression.