Humanized mice in cutaneous leishmaniasis-Suitability analysis of human PBMC transfer into immunodeficient mice.

Humanized mice represent a suitable preclinical test system for example therapeutic interventions in various disease settings, including infections. Here, we intended to establish such system for cutaneous leishmaniasis by infecting T, B and NK cell-deficient mice adoptively transferred with human peripheral blood mononuclear cells (PBMC). L major infection led to the establishment of parasite lesions harbouring viable parasites and human T cells, but parasite elimination was not seen due to a species-specific activity of T cell-derived human IFNγ. In addition, up to 50% of infected mice succumbed to severe graft-versus-host disease. In summary, even though long-term disease outcome assessments are impossible, this model of humanized mice can be used for studying lesion development and generation of oligoclonal anti-parasite human T cell responses in vivo.

Vitamin D deficiency and degree of coronary artery luminal stenosis in women undergoing coronary angiography: a prospective observational study.

BACKGROUND AND AIMS: Low serum 25-OH D levels are associated with increased cardiovascular morbidity and mortality. Recent studies have linked 25-OH D deficiency with the presence of CAD. Women, especially post-menopausal, tend to suffer from accelerated atherosclerosis, along with vitamin D deficiency. In the present study we sought to investigate whether there is a direct association of coronary artery luminal stenosis with 25-OH D deficiency in women. PATIENTS AND METHODS: We enrolled women aged >40 who were scheduled to undergo elective coronary angiography between 3/2011 and 10/2016 in a prospective observational study. RESULTS: We included a total of 105 women. Patients had hypertension (73%), hyperlipidemia (54%), diabetes (29%), smoking (31%), family history of CAD (62%), and known CAD (21%). Median 25-OH D levels were 15.8 ng/mL (range, 3.9-79). Patients had left-anterior descending (31%), left circumflex (22%), and right coronary artery disease (26%); 27% had 2-vessel and 11% had 3-vessel disease. There was a significant inverse correlation between 25-OH D levels and the degree of maximum luminal stenosis. The burden of CAD increased across categories of worsening 25-OH D deficiency. CONCLUSIONS: Vitamin D deficiency is associated with the degree of luminal stenosis and burden of CAD in women undergoing coronary angiography. Future studies should investigate if the repletion of 25-OH D impacts the progression of CAD and cardiovascular mortality.

Myeloid cells do not contribute to gender-dependent differences in disease outcome in murine cutaneous leishmaniasis.

Gender-associated differences in the outcome of infections are well known. Apart from behavior-released differences in their incidence, immunological factors also contribute to disease outcome. The underlying mechanisms are often unknown. Here, we show that in murine experimental leishmaniasis, female mice develop larger skin lesions that harbor significantly more parasites, exhibit increased parasite dissemination to visceral organs associated with a shift towards T helper (Th) 2 immunity with increased levels of IL-4. Antigen presenting cells (APC) responsible for T cell priming, such as macrophages or dendritic cells, were not involved in the process. Additionally, in adoptive transfer experiments, we show that differences in the lymphoid lineage are also not critical for mediating these gender-dependent effects. In summary, neither myeloid nor lymphoid cells contribute to disease outcome against this important human pathogen, but stromal cells influenced by e.g. hormonal effects in addition to other parts of the immune system might play a role.

Immune modulating effects of NKT cells in a physiologically low dose Leishmania major infection model after αGalCer analog PBS57 stimulation.

Leishmaniasis is a parasitic infection affecting ∼12 million people worldwide, mostly in developing countries. Treatment options are limited and no effective vaccines exist to date. Natural Killer T (NKT) cells are a conserved innate-like lymphocyte population with immunomodulating effects in various settings. A number of reports state a role of NKT cells in different models of Leishmania infection. Here, we investigated the effect of NKT cells in a physiologically relevant, intradermal low dose infection model. After inoculation of 103 infectious-stage L. major, comparable numbers of skin-immigrating NKT cells in both susceptible BALB/c mice and resistant C57BL/6 mice were noted. Compared to their wild type counterparts, NKT cell-deficient mice on a C57BL/6 background were better able to contain infection with L. major and showed decreased IL-4 production in cytokine analysis performed 5 and 8 weeks after infection. Low doses of the NKT cell stimulating αGalCer analog PBS57 applied at the time of infection led to disease exacerbation in C57BL/6 wild-type, but not NKT-deficient mice. The effect was dependent both on the timing and amount of PBS57 administered. The effect of NKT cell stimulation by PBS57 proved to be IL-4 dependent, as it was neutralized in IL-4-deficient C57BL/6 or anti-IL-4 antibody-treated wild-type mice. In contrast to C57BL/6 mice, administration of PBS57 in susceptible BALB/c mice resulted in an improved course of disease. Our results reveal a strain- and cytokine-dependent regulatory role of NKT cells in the development of immunity to low dose L. major infections. These effects, probably masked in previous studies using higher parasite inocula, should be considered in future therapy and immunization approaches.

Blue light irradiation suppresses dendritic cells activation in vitro.

Blue light is a UV-free irradiation suitable for treating chronic skin inflammation, for example, atopic dermatitis, psoriasis, and hand- and foot eczema. However, a better understanding of the mode of action is still missing. For this reason, we investigated whether dendritic cells (DC) are directly affected by blue light irradiation in vitro. Here, we report that irradiation neither induced apoptosis nor maturation of monocyte-derived and myeloid DC. However, subsequent DC maturation upon LPS/IFNγ stimulation was impaired in a dose-dependent manner as assessed by maturation markers and cytokine release. Moreover, the potential of this DC to induce cytokine secretion from allogeneic CD4 T cells was reduced. In conclusion, unlike UV irradiation, blue light irradiation at high and low doses only resulted in impaired DC maturation upon activation and a reduced subsequent stimulatory capacity in allogeneic MLRs with strongest effects at higher doses.

Animal model for cutaneous leishmaniasis.

Using cutaneous leishmaniasis of mice, the existence of so-called T helper (Th) cells type 1 and type 2 had been identified more than 20 years ago. Nowadays, it is well accepted that additional T cell populations as well as B cell-mediated immunity is required for immunity against Leishmania major. Finally, using inbred mouse strains, the relevance of genetical factors that influence anti-pathogen immunity as well as elements of the skin-immune system have been identified. This protocol describes a model for murine experimental leishmaniasis that tries to mimic natural parasite transmission by several means: (1) utilization of only infectious-stage parasites that are found in sand fly saliva, (2) intradermal inoculation, and (3) infection with only 1,000 parasites similar to the numbers inoculated by an infected sand fly.

Model for generation of large numbers of primary, inflammatory skin-derived neutrophils, and macrophages.

Tissue macrophages and inflammatory neutrophils represent important cells of the innate immune system responsible for various important tasks, i.e., elimination of pathogens and/or granuloma formation. Isolation of large numbers of primary phagocytes is vital for research with these cells. Within this protocol, we present a strategy for isolation of large numbers of inflammatory neutrophils and macrophages from murine skin that allows for follow-up in vivo or in vitro studies.

Dendritic cells in Leishmania major infections: mechanisms of parasite uptake, cell activation and evidence for physiological relevance.

Leishmaniasis is one of the most important infectious diseases worldwide; a vaccine is still not available. Infected dendritic cells (DC) are critical for the initiation of protective Th1 immunity against Leishmania major. Phagocytosis of L. major by DC leads to cell activation, IL-12 release and (cross-) presentation of Leishmania antigens by DC. Here, we review the role of Fcγ receptor- and B cell-mediated processes for parasite internalization by DC. In addition, the early events after parasite inoculation that consist of mast cell activation, parasite uptake by skin-resident macrophages (MΦ), followed by neutrophil and monocyte immigration and DC activation are described. All these events contribute significantly to antigen processing in infected DC and influence resulting T cell priming in vivo. A detailed understanding of the role of DC for the development of efficient anti-Leishmania immunity will aid the development of potent anti-parasite drugs and/or vaccines.

Induced normothermia attenuates intracranial hypertension and reduces fever burden after severe traumatic brain injury.

INTRODUCTION: Hyperthermia following a severe traumatic brain injury (TBI) is common, potentiates secondary injury, and worsens neurological outcome. Conventional fever treatment is often ineffective. An induced normothermia protocol, utilizing intravascular cooling, was used to assess the impact on fever incidence and intracranial pressure (ICP) in patients with severe TBI. METHODS: A comparative cohort study of 21 adult patients with severe TBI (GCS 25 mmHg was calculated for the initial 72-h monitoring period. Non-parametric rank tests were performed. RESULTS: Mean (+/-SD) or median [range] demographics did not differ between groups [total N = 42 (6 female, 36 male, age 36.4 +/- 14.8 years and initial GCS 7 [3-8], median and range]. Fever burden in the first 3 days (time >38 degrees C) in the induced normothermia versus control group was significantly less at 1.6% versus 10.6%, respectively (P = 0.03). Mean ICP for patients with induced normothermia versus control was 12.74 +/- 4.0 and 16.37 +/- 6.9 mmHg, respectively. Furthermore, percentage of time with ICP > 25 mmHg was significantly less in the induced normothermia group (P = 0.03). CONCLUSION: Induced normothermia (fever prophylaxis via intravascular cooling catheter) is effective in reducing fever burden and may offer a means to attenuate secondary injury, as evidenced by a reduction in the intracranial hypertension burden.

Serum and cerebrospinal fluid magnesium in severe traumatic brain injury outcome.

Serum magnesium concentration has a neuroprotective effect in experimental models of traumatic brain injury (TBI). This study was designed to assess the relationship between initial serum magnesium, cerebrospinal fluid (CSF) magnesium, neurological outcome and the efficacy of magnesium replacement therapy (MgSO4). A retrospective analysis was performed on a prospectively collected dataset from 216 patients admitted during 1996-2006 to the University of Pittsburgh Medical Center with severe TBI. Admission serum and CSF magnesium were dichotomized into low and normal magnesium concentration groups for serum and normal and high concentration groups for CSF. A logistic-regression analysis was performed with 6-month Glasgow Outcome Scale (GOS) scores as outcome variable. The outcome of a subset of 31 patients who presented with low serum magnesium and who were rapidly corrected within 24 h of admission was also analyzed. Low initial serum magnesium was measured in 56.67% of all patients. Patients with an initial serum magnesium of <1.3 mEq/L were 2.37 times more likely to have a poor outcome (CI: 1.18-4.78, p = 0.016). The prognostic significance of depressed serum magnesium remained, even in patients whose serum magnesium levels were corrected within 24 h (OR = 11.03, CI: 1.87-68.14, p = 0.008). Patients with an initial high CSF magnesium were 7.63 more likely to have a poor outcome (p = 0.05). Elevated CSF magnesium correlated with depressed serum magnesium only in patients with poor outcome (p = 0.013). Patients with low serum magnesium and high CSF magnesium are most likely to have poor outcome after severe TBI. Rapid correction of serum magnesium levels does not reverse the prognostic value of these markers.