Pathogenicity of Nc-Bahia and Nc-1 strains of Neospora caninum in experimentally infected cows and buffaloes in early pregnancy

Neospora caninum is a protozoan parasite known as an important cause of bovine abortion worldwide. Little is currently known about how different strains of N. caninum vary in their pathogenicity. In this study, we compared a Brazilian strain, Nc-Bahia, with the first isolate of this coccidian, Nc-1. Eight cows and seven buffaloes were submitted to fixed-time artificial insemination protocols for a better control of pregnancy. Group 1 was inoculated with Nc-Bahia (n?=?8; five cows and three buffaloes), and Group 2 was inoculated with Nc-1 (n?=?5; two cows and three buffaloes). One nonpregnant female of each species was left uninfected as sentinel controls for potential environmental infection. All inoculated animals received 5?×?10⁸ tachyzoites of N. caninum, by intravenous route, on the 70th day of gestation. Uninfected animals remained seronegative throughout the experiment, indicating no exogenous infection, whereas all inoculated animals became seropositive to N. caninum. In Group 1, abortion was found in only one cow on 42 days postinfection (dpi; frequency of abortion?=?12.5 %), whilst all animals from Group 2 aborted on 35 dpi (frequency of abortion?=?100 %). Parasite DNA was detected by seminested PCR in maternal, foetal and placental tissues, confirming vertical transmission in Groups 1 and 2, although histological lesions had different frequencies and degrees of severity between the groups. There was evidence of lower pathogenicity of Nc-Bahia compared to Nc-1 when used in experimental infection, as it caused fewer abortions, as well as less frequent and milder histological lesions. This was the first time Nc-Bahia has been used for experimental infection.     

Conditional ablation of NKp46+ cells using a novel Ncr1greenCre mouse strain: NK cells are essential for protection against pulmonary B16 metastases

To study gene functions specifically in NKp46⁺ cells we developed novel Cre mice allowing for conditional gene targeting in cells expressing Ncr1 (encoding NKp46). We generated transgenic Ncr1^greenCre mice carrying an EGFPcre fusion under the control of a proximal Ncr1 promoter that faithfully directed EGFPcre expression to NKp46⁺ cells from lymphoid and nonlymphoid tissues. This approach allowed for direct detection of Cre‐expressing NKp46⁺ cells via their GFP signature by flow cytometry and histology. Cre was functional as evidenced by the NKp46⁺ cell‐specific expression of RFP in Ncr1^greenCreRosa‐dtRFP reporter mice. We generated Ncr1^greenCreIl2rg^fl/fl mice that lack NKp46⁺ cells in an otherwise intact hematopoietic environment. Il2rg encodes the common gamma chain (γ_c), which is an essential receptor subunit for cytokines (IL‐2, ‐4, ‐7, ‐9, ‐15, and ‐21) that stimulate lymphocyte development and function. In Ncr1^greenCreIl2rg^fl/fl mice, NK cells are severely reduced and the few remaining NKp46⁺ cells escaping γc deletion failed to express GFP. Using this new NK‐cell‐deficient model, we demonstrate that the homeostasis of NKp46⁺ cells from all tissues (including the recently described intraepithelial ILC1 subset) requires Il2rg. Finally, Ncr1^greenCreIl2rg^fl/fl mice are unable to reject B16 lung metastases demonstrating the essential role of NKp46⁺ cells in antimelanoma immune responses.     

European consensus proposal for immunoglobulin therapies

The use of immunoglobulin (Ig) preparations (intravenous, IVIg, subcutaneous, SCIg) for replacement and immunomodulation therapy worldwide has tripled in the past 20 years and represents an ever‐increasing cost factor for healthcare organizations. The limited access to the starting material of this essential medicinal product is currently the driving force for human plasma collection. Increasing awareness and improved diagnosis of human primary immunodeficiencies and a broadening of immunomodulatory indications are responsible for this development, and on a longer run might lead to plasma supply shortages. Consensus recommendations for the optimal use of Ig in clinical practice, including priority rankings for the most urgent indications, are therefore urgently needed. During a recent meeting in Kreuth, Germany, expert nominees from 36 Council of Europe states, together with colleagues from observer countries and regulatory agencies came up with this consensus statement.

     

The burden and consequences of inherited blood disorders among young children in western Kenya

Although inherited blood disorders are common among children in many parts of Africa, limited data are available about their prevalence or contribution to childhood anaemia. We conducted a cross‐sectional survey of 858 children aged 6–35 months who were randomly selected from 60 villages in western Kenya. Haemoglobin (Hb), ferritin, malaria, C‐reactive protein (CRP) and retinol binding protein (RBP) were measured from capillary blood. Using polymerase chain reaction (PCR), Hb type, ?3.7 kb alpha‐globin chain deletion, glucose‐6‐phosphate dehydrogenase (G6PD) genotype and haptoglobin (Hp) genotype were determined. More than 2 out of 3 children had at least one measured blood disorder. Sickle cell trait (HbAS) and disease (HbSS) were found in 17.1% and 1.6% of children, respectively; 38.5% were heterozygotes and 9.6% were homozygotes for α⁺‐thalassaemia. The Hp 2‐2 genotype was found in 20.4% of children, whereas 8.2% of males and 6.8% of children overall had G6PD deficiency. There were no significant differences in the distribution of malaria by the measured blood disorders, except among males with G6PD deficiency who had a lower prevalence of clinical malaria than males of normal G6PD genotype (P = 0.005). After excluding children with malaria parasitaemia, inflammation (CRP > 5 mg L^?1), iron deficiency (ferritin < 12 μg L^?1) or vitamin A deficiency (RBP < 0.7 μg L^?1), the prevalence of anaemia among those without α⁺‐thalassaemia (43.0%) remained significantly lower than that among children who were either heterozygotes (53.5%) or homozygotes (67.7%, P = 0.03). Inherited blood disorders are common among pre‐school children in western Kenya and are important contributors to anaemia.     

Thrombospondin-1 Deficiency Causes a Shift from Fibroproliferative to Inflammatory Kidney Disease and Delays Onset of Renal Failure

Thrombospondin-1 (TSP1) is a multifunctional matricellular protein known to promote progression of chronic kidney disease. To gain insight into the underlying mechanisms through which TSP1 accelerates chronic kidney disease, we compared disease progression in Col4a3 knockout (KO) mice, which develop spontaneous kidney failure, with that of Col4a3;Tsp1 double-knockout (DKO) mice. Decline of excretory renal function was significantly delayed in the absence of TSP1. Although Col4a3;Tsp1 DKO mice did progress toward end-stage renal failure, their kidneys exhibited distinct histopathological lesions, compared with creatinine level–matched Col4a3 KO mice. Although kidneys of both Col4a3 KO and Col4a3;Tsp1 DKO mice exhibited a widened tubulointerstitium, predominant lesions in Col4a3 KO kidneys were collagen deposition and fibroblast accumulation, whereas in Col4a3;Tsp1 DKO kidney inflammation was predominant, with less collagen deposition. Altered disease progression correlated with impaired activation of transforming growth factor-β1 (TGF-β1) in vivo and in vitro in the absence of TSP1. In summary, our findings suggest that TSP1 contributes to progression of chronic kidney disease by catalyzing activation of latent TGF-β1, resulting in promotion of a fibroproliferative response over an inflammatory response. Furthermore, the findings suggest that fibroproliferative and inflammatory lesions are independent entities, both of which contribute to decline of renal function.Progression of chronic kidney disease (CKD) toward end-stage renal failure (ESRF) is a prominent problem in clinical nephrology.¹ The incidence of CKD is rising, but effective therapies to halt progression of disease remain elusive.² Progression of CKD results from a complex interplay of pathologies that involve all constituents of the kidney, which makes it difficult to single out targets for effective therapeutic strategies.³The extent of so-called tubulointerstitial fibrosis is often considered to be the rate-limiting step in progression of CKD.¹ This idea is founded on histopathological analysis of large cohorts of kidney biopsies, which demonstrated that only tubulointerstitial fibrosis (which at the time was determined as the relative volume of the interstitium within a kidney biopsy section) correlates with and also predicts progression of CKD toward ESRF, irrespective of the underlying primary disease.^{4, 5, 6, 7} Widening of the tubulointerstitium, which is referred to as tubulointerstitial fibrosis, is caused by a composite of extracellular matrix (ECM) accumulation, sterile inflammation, accumulation of activated fibroblasts, and rarefaction of microvessels.¹ Although the relevance of each of these events to progression of fibrosis and CKD is hotly debated, this knowledge led to the concept that tubulointerstitial fibrosis is a common pathway of all chronic progressive kidney diseases and that effective antifibrotic therapies could potentially halt progression of CKD irrespective of the underlying disease. However, such therapies are not yet available.¹Our aim was to gain insight into mechanisms that underlie the contribution of thrombospondin-1 (TSP1) to progression of CKD. TSP1 is the most-studied member of the thrombospondin family of matricellular proteins.⁸ Previous studies have demonstrated that pharmacological suppression or genetic depletion of TSP1 attenuates disease progression in animal models of CKD.^{9, 10, 11, 12, 13} TSP1 is a 450-kDa trimeric ECM protein, which does not fulfill primarily structural roles in the matrix, but instead functions as an extracellular modulator of cell function.^{8, 14} Most prominently, TSP1 is known to inhibit angiogenesis, inhibit inflammation, activate MMP-dependent ECM turnover, and facilitate fibroblast migration and activation, all of which are considered important contributors to progression of CKD.^{8, 10} To delineate through which of its known biological activities TSP1 impacts progression of CKD, we compared progression of kidney disease of Col4a3 knockout (KO) mice (deficient in type IV collagen α3 chain) with that of Col4a3;Tsp1 double-knockout (DKO) mutant mice.¹⁵Here, we demonstrate that decrease of excretory renal function is delayed if TSP1 is absent. Furthermore, tissue analysis of plasma creatinine level–matched kidneys of Col4a3 KO and of Col4a3;Tsp1 DKO revealed that in Col4a3 KO mice disease progression is predominantly associated with fibrosis, whereas inflammation is the predominant interstitial pathology in Col4a3;Tsp1 DKO mice. We provide evidence that this altered disease progression is due to impaired activation of latent transforming growth factor-β1 (TGF-β1) in the absence of TSP1. Our findings provide evidence that both fibroproliferative injury and inflammation can independently cause expansion of the interstitium, leading to decline of excretory renal function.     

Interleukin-3-Deficient Mice Have Increased Resistance to Blood-Stage Malaria

The contribution of interleukin-3 (IL-3), a hematopoietic growth factor and immunoregulatory cytokine, to resistance to blood-stage malaria was investigated by infecting IL-3-deficient (knockout [KO]) mice with Plasmodium berghei NK65. Male IL-3 KO mice, but not female mice, were more resistant to infection than wild-type (WT) mice, as evidenced by lower peak parasitemia and prolonged survival. Both male and female IL-3 KO mice had increased splenomegaly and were more anemic than corresponding WT mice. Anemia was compensated for by an increase in bone marrow and splenic erythropoiesis in IL-3 KO mice, as evidenced by higher levels of erythroid progenitors. Plasma levels of gamma interferon (IFN-γ) and CXCL9 (monokine induced by IFN-γ [MIG]) were found to be significantly reduced in IL-3 KO mice during early stages of infection. In contrast, granulocyte colony-stimulating factor (G-CSF) levels were significantly higher, and the percentage of peripheral blood neutrophils lower, in infected IL-3 KO mice than in WT counterparts. Overall, our results indicate that IL-3 plays a critical role in suppressing protective immunity to P. berghei NK65 infection and that it is involved in inhibiting the development of splenomegaly, anemia, and erythropoiesis. IL-3 also influences IFN-γ, CXCL9, and G-CSF production in response to infection. The abnormal responses seen in infected IL-3 KO mice may be due to the lack of IL-3 during development, to the lack of IL-3 in the infected mature mice, or to both.     

Direct acute tubular damage contributes to Shigatoxin‐mediated kidney failure

The pathogenesis and therapy of Shigatoxin 2 (Stx2)‐mediated kidney failure remain controversial. Our aim was to test whether, during an infection with Stx2‐producing E. coli (STEC), Stx2 exerts direct effects on renal tubular epithelium and thereby possibly contributes to acute renal failure. Mice represent a suitable model because they, like humans, express the Stx2‐receptor Gb3 in the tubular epithelium but, in contrast to humans, not in glomerular endothelia, and are thus free of glomerular thrombotic microangiopathy (TMA). In wild‐type mice, Stx2 caused acute tubular dysfunction with consequent electrolyte disturbance, which was most likely the cause of death. Tubule‐specific depletion of Gb3 protected the mice from acute renal failure. In vitro, Stx2 induced secretion of proinflammatory cytokines and apoptosis in human tubular epithelial cells, thus implicating a direct effect of Stx2 on the tubular epithelium. To correlate these results to human disease, kidney biopsies and outcome were analysed in patients with Stx2‐associated kidney failure (n = 11, aged 22–44 years). The majority of kidney biopsies showed different stages of an ongoing TMA; however, no glomerular complement activation could be demonstrated. All biopsies, including those without TMA, showed severe acute tubular damage. Due to these findings, patients were treated with supportive therapy without complement‐inhibiting antibodies (eculizumab) or immunoadsorption. Despite the severity of the initial disease [creatinine 6.34 (1.31–17.60) mg/dl, lactate dehydrogenase 1944 (753–2792) U/l, platelets 33 (19–124)/nl and haemoglobin 6.2 (5.2–7.8) g/dl; median (range)], all patients were discharged after 33 (range 19–43) days with no neurological symptoms and no dialysis requirement [creatinine 1.39 (range 0.84–2.86) mg/dl]. The creatinine decreased further to 0.90 (range 0.66–1.27) mg/dl after 24 months. Based on these data, one may surmise that acute tubular damage represents a separate pathophysiological mechanism, importantly contributing to Stx2‐mediated acute kidney failure. Specifically in young adults, an excellent outcome can be achieved by supportive therapy only. © 2014 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Laughter as a social rejection cue: Gelotophobia and transient cardiac responses to other persons' laughter and insult

Other persons' laughter, normally perceived as a signal that persons are friendly and inviting others to approach, can also be perceived as a cue of social rejection. In this study, prerecorded laughter was placed in a realistic and personally relevant context, and participants' responses were related to gelotophobia, a trait predisposing to perceiving laughter as a cue of social rejection. Individuals with gelotophobia showed marked heart rate deceleration in response to the laughter stimulus, possibly indicating a “freezing‐like” response. Moreover, cardiac responses to anger provocation by overtly insulting statements indicated heightened aggressive anger in response to cumulated social threat. The study adds to recent research showing specific cardiac responses to social rejection and to the literature on social rejection sensitivity by demonstrating the value of using well interpretable physiological measures in this research context.