bluuebunny/crossref_metadata_2025 huggingface dataset:
Absence of Auto‐Antibodies against Cardiac Troponin I Predicts Improvement of Left Ventricular Function after Acute Myocardial Infarction
Authors: Kaya Ziya , Li Jin , Leuschner Florian , Göser Stefan , Öttl Renate , Reinhardt Lars , Remppis Andrew , Giannitsis Evangelos , Katus Hugo A.
Published: June 2021

Background: Application of antibodies (Ab) against cardiac troponin I (cTnI) can induce dilation and dysfunction of the heart in mice. But little is known about the clinical relevance of detected cTnI‐Ab.

Methods: First, anti‐cTnI and anti‐cTnT Ab titers have been measured in sera from 272 patients with dilated‐ (DCM) and 185 with ischemic‐ (ICM) cardiomyopathy. Secondly, 108 patients with acute myocardial infarction (AMI) were included for a follow‐up study. All 108 patients were treated according to current guidelines. Heart characteristics were determined by magnetic resonance imaging 3–4 days and 6–9 months after AMI.

Results: Altogether, in 14,3% of patients with DCM and in 17,3% with ICM an anti‐cTnI IgG‐Ab titer ≥1:80 was measured. In contrast, only in 1,8% of patients with DCM and in 0,5% with ICM an anti‐cTnT IgG‐Ab titer ≥1:80 was detected. Ten out of 108 patients included in the follow‐up study were tested positive for cTnI‐Ab with IgG Ab titers ≥1:160. TnI‐Ab negative patients showed a significant increase in LVF and stroke volume 6–9 months after AMI. In contrast, there was no significant increase in LVF and stroke volume in TnI‐Ab positive patients.

Conclusion: We demonstrate for the first time that the prevalence of cTnI‐Abs in patients with AMI has an impact on the improvement of the LVF.
Autoantibodies against Cardiac Troponin I in Patients with Dilated Cardio‐myopathy Predict Improvement of Cardiac Function by Immunoadsorption
Authors: Kaya Ziya , Leuschner Florian , Trimpert Christiane , Göser Stefan , Öttl Renate , Li Jin , Felix Stephan , Dengler Thomas J. , Hugo Katus A. , Staudt Alexander
Published: June 2021

Aims: Cardiac troponins in blood are the most preferred markers of myocardial damage. It has been shown that the application of antibodies against cardiac troponin I (cTnI) can induce dilation and dysfunction of the heart in mice. In the present study we investigated the presence of cTnI‐antibody in patients with dilated cardiomyopathy (DCM) and the effect of immunoadsorption (IA) on left ventricular function (LVF).

Methods: 51 patients with DCM were included. Every patient underwent heart catheterisation including myocardial biopsy and received IA. LVF was determined by echocardiography before and 6 months after IA. Serum samples from each patient (before, 5 days and 6 month after IA) were screened for the presence of cTnI‐Ab. 22 of the 51 patients had positive cTnI‐Ab‐titers. Whereas LVF of patients with positive cTnI‐Ab‐titers improved after IA by +8.1% during follow up of 6 month (p<0.001 vs. baseline), patients with negative cTnI‐Ab‐titers showed an improvement of +4,1% only (p<0.04 vs. baseline). Furthermore, Patients with positive cTnI‐antibody‐titers showed a significantly better clinical improvement after IA (+1 NYHA‐class) than patients without cTnI‐antibodies (+0.5 NYHA‐class; p<0.05).

Conclusion: Immunoadsortion therapy is associated with a pronounced improvement of LVF and NYHA‐Class in patients with detectable antibodies against cardiac troponin I.
Lymphotoxin beta receptor is required for the migration and selection of autoreactive T cells in thymic medulla
Authors: Zhu Mingzhao , Chin Robert K , Tumanov Alexei V , Liu Xiaojuan , Fu Yang‐Xin
Published: June 2021

How organ‐specific central tolerance is established and regulated has been an intriguing question. Lymphotoxin beta receptor (LTβR) deficiency is associated with autoimmune phenotypes characterized by humoral and cellular auto‐reactivity to peripheral organs. We sought to determine the role of LTβR in central tolerance of T cells directed at tissue‐restricted antigens. By tracing the development of OT‐I thymocytes in RIP‐mOVA transgenic mice on either Ltbr+/+ or Ltbr−/− background, we demonstrate that LTβR is necessary for thymic negative selection. LTβR deficiency resulted in a dramatic escape of “neo‐self” specific OT‐I cells that persist in circulation and lead to development of peri‐insulitis. When the underlying mechanism was further explored, we found interestingly that LTβR deficiency did not result in reduced thymic expression of mOVA. Instead, LTβR was revealed to control the expression of thymic medullary chemokines (SLC and ELC) which are required for thymocytes migration and negative selection in medulla as was confirmed by using SLC/ELC deficient mice. On the other hand, by tracing the development of OT‐II cells, we did not find any influence of LTβR deficiency on natural Treg positive selection, another arm of central tolerance. Thus, LTβR has been revealed to play an important role in thymic negative selection of organ‐specific thymocytes through thymic medullary chemokines regulation.

This research was supported by grants from National Institutes of Health (NIH; AI062026, DK58897 to YXF)
LAG‐3 (Lymphocyte Activation Gene‐3) Negatively Regulates Environmentally‐Induced Autoimmune Disease
Authors: Jha Vibha , Workman Creg J. , Vignali Dario , Monestier Marc
Published: June 2021

After exposure to subtoxic levels of mercury (Hg), genetically susceptible mouse strains develop an autoimmune disease characterized by the production of highly specific IgG anti‐nucleolar autoantibodies, hyperglobulinemia and nephritis. However, mice can be tolerized to the disease by a single low dose administration of Hg. Previous studies from our lab demonstrated that CD4+ CD25+ Foxp3+ regulatory T cells (Tregs) control the induction and maintenance of tolerance to Hg. Lymphocyte Activation Gene‐3 (LAG‐3) is a CD4‐related, MHC‐class II binding molecule expressed on activated T cells and NK cells. LAG‐3 is an inhibitory molecule that maintains lymphocyte homeostatic balance by controlling effector T cell expansion and contributing to the suppressive functions of Tregs. In our model, administration of anti‐LAG‐3 monoclonal antibody broke tolerance to Hg resulting in autoantibody production and an increase in the levels of serum IgE. In addition, LAG‐3‐deficient B6.H2s mice had increased susceptibility to Hg‐induced autoimmunity, as illustrated by an increase in autoantibody levels and polyclonal activation when compared to the wild‐type controls. Therefore, LAG‐3 exerts an important regulatory effect on autoimmunity elicited by a common environmental pollutant.
B cells promote Th2 cytokine production and pathophysiology during chronic allergic lung disease
Authors: Lindell Dennis Michael , Lukacs Nicholas W
Published: June 2021

The role of B cells in chronic allergic lung disease, or asthma, remains controversial. Our goal in these studies was to characterize the role of B cells in a clinically relevant murine model of chronic allergic lung disease, with a focus on B cell antigen presentation.

To this end, mice were chronically challenged (eight times total) with cockroach allergen extract, and the lung B cell response was determined. Chronic allergen challenge resulted in the accumulation of large numbers of B cells to the lungs, which expressed MHC II and the costimulatory molecules CD40, CD80 and CD86. In cocultures with CD4+ T cells, purified lung B cells promoted T cell proliferation and antigen‐specific Th2 cytokine production (IL‐4, IL‐5, and IL‐13, but not IFNg). B cell −/− mice (muMT) had attenuated airway hyperresponsiveness, and decreased mucus hypersecretion. The levels of Th2 cytokines in the lungs of B cell −/− mice were decreased relative to B cell sufficient controls, demonstrating that B cells play a role at promoting pathophysiology and Th2 cytokine responses in vivo. Repopulation of B cells into muMT mice significantly increased CD4 T cell responses in the lungs and increased mucus.

Collectively, these results suggest that B cell antigen presentation plays an important role in promoting Th2 cytokine and pathophysiology in allergic lung disease.

(NIH‐HL031963, HL059178, and T32HL07749)
Role of TSLP in allergen sensitization and allergy elicitation
Authors: Zhou Baohua , Ziegler Steven F
Published: June 2021

Thymic stromal lymphopoietin (TSLP) plays an important role in the pathogenesis of asthma so that TSLP receptor knockout (TSLPR−/−) mice are protected from ovalbumin (OVA)‐induced asthma model. However, questions regarding which cells it acts on and when it is needed for the allergic pathogenesis remain to be answered. We demonstrated that a TSLP neutralizing antibody was able to significantly reduce OVA‐induced airway inflammation only when given at both sensitization and challenge phase. Furthermore, TSLPR−/− mice with adoptively transferred wild type dendritic cells (DCs) at either OVA sensitization or challenge showed intermediate levels of eosinophilic airway inflammation and goblet cell metaplasia with no significant airway hyperresponsiveness (AHR). In contrast, the TSLPR−/− mice receiving DCs at both sensitization and challenge showed fully developed eosinophilic airway inflammation, goblet cell metaplasia and AHR similar to wild type animals undergo OVA sensitization and challenge. Taken together, these data suggest that the major target downstream of TSLP is dendritic cells and TSLP acting through DCs is needed for both allergen sensitization and allergy elicitation.
Cockroach Allergen Induced Asthma in Outbred Mice
Authors: Vaickus Louis Joseph , Natarajan Sudha , Kim Jiyoun , Remick Daniel G
Published: June 2021

Our lab has previously demonstrated the ability of German cockroach allergens (CRA) to induce allergic‐type airways inflammation in BALB/c mice. This model resembles human allergic asthma in its histology, symptomatology and airways hyperreactivity. We reproduced this model in outbred (ICR) mice to extend our observations and establish a more applicable model to the extremely outbred human population. Mice were given 4.75ug, 2.4ug and 2.4ug of CRA intratracheally on days 0, 14, and 21. Airways hyperresponsiveness (AHR) and cell content of bronchoalveolar lavage fluid was measured 24 hours post final challenge and the results compared to BALB/c mice.

Because of the encouraging data this model has produced, and the greater applicability of its genetically diverse background, we believe outbred animal experiments are highly appropriate in simulating human disease.
Novel allergen‐specific therapy for allergy through immunomodulation by CD40‐silenced dendritic cells
Authors: Suzuki Motohiko , Zheng Xiufen , Zhang Xusheng , Li Mu , Vladau Costin , Garcia Bertha , Min Rui , Kubo Norihiko , Min Wei‐Ping
Published: June 2021

Small interfering RNA (siRNA) is a potent, selective, and easily‐inducible method for blocking the expression of specifically targeted genes. siRNA is reported to be more efficient than traditional therapy such as antisense oligonucleotides. However, the use of siRNA in allergen‐specific therapy for allergy has not yet been reported. Therefore, we attempted to develop a new therapy for allergy using allergen‐pulsed dendritic cells (DCs) in which CD40 is silenced by siRNA. Bone marrow‐derived DCs were silenced with CD40 siRNA and pulsed with ovalbumin (OVA). Following administration of CD40‐silenced and OVA‐pulsed DCs (CD40‐silenced DCs), mice were sensitized with OVA intraperitoneally, and then intranasally challenged with OVA. CD40‐silenced DCs remarkably reduced allergic symptoms, and decreased eosinophilia in the nasal tissue. CD40‐silenced DCs inhibited OVA‐specific T cell responses. Additionally, anti‐OVA IgE and IgG1 in sera were significantly decreased in the mice that were treated with CD40‐silenced DCs. CD40‐silenced DCs also suppressed IL‐4 and IL‐5 production by spleen and lymph node cells after stimulated by OVA. Finally, CD40‐silenced DCs facilitated the generation of CD4+CD25+Foxp3+ regulatory T cells. This study is the first to demonstrate a novel allergen‐specific therapy for allergy through DC‐mediated immune modulation following gene silencing of CD40.
Toll‐like receptor mediated activation of ERK1/2 is regulated by GSK3
Authors: Rehani Kunal , Wang Huizhi , Garcia Carlos A. , Martin Michael
Published: June 2021

Toll‐like receptor (TLR) stimulation of innate immune cells can lead to the activation of a diverse array of downstream signal transduction pathways that play critical roles in regulating host inflammatory responses. One of the most highly conserved signaling cascades activated in TLR‐stimulated cells are the mitogen‐associated protein kinases (MAPKs). In the present study, we identify a novel mechanism by which Glycogen‐synthase kinase 3 (GSK3) selectively regulates the activity of the mitogen‐associated protein kinase (MAPK) ERK1/2 in TLR‐stimulated human monocytes. Analysis of upstream kinases within the ERK1/2 pathway demonstrated that GSK3 inhibition resulted in the abrogation of phospho‐rac1 levels at the inhibitory site serine 71 (S71) and increased the ability of rac1 to interact with and activate p21‐activated protein kinase (PAK). The subsequent downstream target of PAK, phospho‐c‐Raf (S338), was highly augmented upon GSK3 inhibition. Inhibition of c‐raf activity abrogated the ability of GSK3 inhibition to increase phospho‐ERK1/2 levels, as well as the direct upstream kinase responsible for phosphorylating ERK1/2, MEK1/2. The capability of GSK3 to differentially regulate pro‐ and anti‐inflammatory cytokines by TLR‐stimulated innate immune cells was shown to be dependent on its ability to regulate ERK1/2 activity. Our findings identify a novel cell‐signaling pathway by which TLR‐mediated activation of ERK1/2 is regulated.
A single‐step enrichment method for generating highly purified mouse NK cell populations by negative selection
Authors: Tran Jimmy , Kung Sam K.P. , Yuan Ning , Milton Graeme T , Wognum Albertus W , Eaves Allen C , Thomas Terry E , Guilbault Benoit G
Published: June 2021

Natural Killer (NK) cells play a critical role in innate immunity and tumor immunology. They are particularly difficult to isolate because they are rare and express several markers found on other cell types. Using column‐free immunomagnetic cell separation technology (EasySep®), we sought to develop a single‐step negative selection method for NK cell isolation, where negative selection enables the isolation of cells with minimal impact on cell function. Unwanted cells were cross‐linked to magnetic particles using a highly optimized cocktail of 10 biotinylated antibodies. This enabled us to achieve NK cell purities of 92.3 ± 2.6% (defined as %CD49b+CD3neg; n=6). The isolated cells retained the expression of several markers associated with an NK phenotype, including NKG2D and LY49C/I/F/H. The purified cells could be readily expanded when cultured in the presence of IL‐2 (> 8‐fold expansion after 5 days, n=3). Cr51 release assays demonstrated the cytotoxic capacity of the isolated cells, as specific lysis was detected at an effector to target ratio of 1:1 and 10:1 for both YAC‐1 cells and Daudi B lymphoma cells, the latter being more resistant to NK‐specific killing as expected (n=3). In conclusion, we present here a novel and unique enrichment method for the isolation of pure NK cells in a single step.

Supported by the Manitoba Health Research Council and The Dean of Medicine Strategic Research Fund.
Novel innate polysaccharide agonists derived from Funtumia elastica tree bark (Yamoa™)
Authors: Hedges Jodi F. , Graff Jill C. , Wilson Suzanne , Freedman Brett , Schepetkin Igor A. , Quinn Mark T. , Jutila Mark A.
Published: June 2021

We have identified a unique polysaccharide innate adjuvant by screening using bovine γδ T cells. The screens were initiated to identify γδ T cell adjuvants, but the identified agonist was much more efficient at priming γδ T cell proliferation when accessory cells were present, suggesting a broader target cell range. The active polysaccharide fraction was isolated from Yamoa™ (ground bark of the Funtumia elastica tree) which is sold as a nutritional supplement. Microarray data from purified bovine γδ T cells and monocytes indicated that Yamoa™ stimulation resulted in a very similar gene expression pattern to that following stimulation with LPS, suggesting similar pathways. However, C3H:HeJ (TLR4−/‐) mice were responsive to intraperitoneal injection of Yamoa™, indicating that despite their almost overlapping gene expression profiles, there is a TLR4 independent element in the Yamoa™‐derived polysaccharides. Specifically, the fraction that induces the most robust γδ T cell priming is TLR4 independent. Injection of Yamoa™ results in neutrophil recruitment and potentially enhances innate immunity in bovine and mouse models of Salmonella enterica serotype Typhimurium. Yamoa™ stimulation of human PBMCs resulted in large increases in GM‐CSF expression, and although changes in GM‐CSF were not detected in sera from Yamoa™ stimulated mice, increases in CXCL1/KC were, suggesting a possible mechanism for the observed neutrophil mobilization and innate protection. Thus, polysaccharide agonists derived from Yamoa™ are novel innate adjuvants with conserved activity and potential application in infectious disease settings.
Tyrosine Phosphorylation of MAL in TLR4 Signaling and Endotoxin Tolerance
Authors: Piao Wenji , Song Chang , Chen Haiyan , Fitzgerald Katherine A. , O’Neill Luke A.J. , Medvedev Andrei E.
Published: June 2021

TLR4 recognition of LPS triggers signalosome assembly among TLR4, sorting (e.g. MyD88‐adapter‐like, Mal) and signaling (e.g. MyD88) adapters and kinases, resulting in activation of transcription factors and production of inflammatory mediators. Here we examined whether Mal tyrosine phosphorylation regulates Mal‐TLR4 interactions and signaling. In contrast to wild‐type (WT) Mal, overexpression of tyrosine‐deficient Mal variants with mutated Y86, Y106, and Y159 tyrosine residues impaired Mal tyrosine phosphorylation, interactions with Bruton's tyrosine kinase (Btk), phosphorylation of p38, and NF‐κB activation. LPS triggered tyrosine phosphorylation of endogenous Mal and initiated Mal‐Btk interactions in 293/TLR4/MD‐2 cells and human monocytes that were suppressed in endotoxin‐tolerant cells. Compared to WT‐Mal, Y86A‐ and Y159A‐ Mal variants exhibited higher interactions with TLR4 and MyD88, while associations with IRAK‐2 and TRAF‐6 were not affected. Overexpression of Y86A, Y159A and Y106A Mal in 293/TLR4/MD‐2 cells exerted dominant‐negative effects on TLR4‐inducible p38 phosphorylation and NF‐κB reporter activation. In contrast, tyrosine‐deficient Mal species did not affect NF‐κB activation when signaling was initiated at the post‐receptor level by overexpression of MyD88, IRAK‐2 or TRAF‐6. Thus, tyrosine phosphorylation of Mal is required for TLR signaling, regulates Mal interactions with TLR4 and receptor signaling, and is inhibited in endotoxin tolerance.

This work was supported by NIH grant AI‐059524 (to A.E.M.)
Inhibitory effect of glycyrrhizin on the production of CCL2 by various sources of immunosuppressive neutrophils (PMN‐II)
Authors: Jones Vickie C , Shigematsu Kenji , Kobayashi Makiko , Sanford Arthur P , Herndon David N , Suzuki Fujio
Published: June 2021

Three subtypes of PMN (PMN‐N, PMN‐I and PMN‐II) have been demonstrated. Among them, PMN‐II have been characterized as cells that participate in the increased susceptibility of burned hosts to bacterial translocation‐associated infectious complications. Since host‐mediated antibacterial effects of glycyrrhizin (GL) were demonstrated in these infections, in this study, the inhibitory effects of GL on the CCL2 production by various sources of PMN‐II were examined. CCL2 has been determined as an effector chemokine on the PMN‐II immunosuppressive activity. PMN‐II were prepared from peripheral blood of burn patients by Ficoll‐Hypaque and dextran sedimentations. Also, PMN‐II were induced from band PMN (isolated peripheral blood PMN from healthy volunteers 3 hrs after having exercised for 1 hr) by stimulation with SAC (0.0075%), GM‐CSF (0.1 ng/ml), VEGF (0.1 ng/ml) or terbutaline (β2‐AR agonist, 10−6 M). All of these PMN preparations were identified as PMN‐II by their abilities to produce CCL2 and IL‐10. In the results, the production of CCL2 by PMN‐II isolated from peripheral blood of severely burned patients was inhibited by 10 μg/ml of GL (Minophagen Pharmaceutical Co., Ltd., Tokyo, Japan). Also, the production of CCL2 by PMN‐II preparations, prepared in vitro from band PMN of exercised volunteers after stimulation with SAC, GM‐CSF, VEGF or terbutaline, was inhibited by GL (10 μg/ml). These results suggest that PMN‐II‐associated deficiency of the host antimicrobial resistance may be effectively intervened by GL.

This study was supported by SHC NA #8840.
Formation and disruption of TRIM19 containing nuclear bodies in neurons during herpes simplex virus in vivo
Authors: Sawtell Nancy M , Thompson Richard L
Published: June 2021

The tripartite motif (TRIM) family is emerging as a novel class of antiviral proteins involved in innate host defense. TRIM19 (promyelocytic leukemia protein, PML) orchestrates the formation of ND10 which are distinct intranuclear substructures. The hypothesis that these bodies are part of the cellular antiviral response arose from the observation that herpes viruses as well as other viruses actively disrupt these bodies. Maul and Everett have demonstrated that the HSV IE protein, ICP0 is necessary and sufficient for disruption of ND10 in cell culture. Whether ND10 form and/or are disrupted in the context of a herpes viral infection in vivo is not known. In this study we have utilized wild type and PML‐/‐ mice and parental and ICP0 null mutant viruses to examine the role of ND10 formation and disruption during HSV infection in vivo. Viral replication, establishment of latency, and viral reactivation in vivo were compared in the presence and absence of ICP0 and in the presence and absence of PML. HSV infection resulted in the global upregulation of ND10 within cells of infected tissues which was not restricted to those cells that were infected. In the absence of ICP0 function, ND10 were not disrupted in infected neurons. These neurons underwent apoptosis prior to infectious virus production. Thus ND10 are broadly upregulated by viral infection in vivo and their disruption is correlated with productive viral replication. NIH RO1AI32121
Tregs can directly suppress macrophage TLR responses
Authors: O’Leary Fionnuala , Tajima Goro , McKenna Ann , MacConmara Malcolm , Delisle Adam , Mannick John , Lederer James
Published: June 2021

Mice lacking CD4 T cells develop a more pro‐inflammatory phenotype when injured. We subsequently showed that CD4+CD25+ regulatory T cells (Tregs) were required to control this injury‐induced change in innate immunity. We now demonstrate that injured Treg‐deficient mice are more susceptible to LPS‐induced shock suggesting that Tregs play an active role in controlling Toll‐like receptor 4 (TLR4) reactivity. Since macrophages are the primary cells responsible for enhanced LPS reactivity after injury, we wished to test whether Tregs might directly control macrophage responses to innate stimuli. To accomplish this, we first developed a macrophage/Treg suppression assay. Macrophages were purified from the spleens and CD4+CD25+ T cells (Tregs) were purified from the lymph nodes of C57BL/6J mice. Cells were then mixed at various Treg:macrophage ratios and stimulated with pure LPS (a TLR4 agonist) or Pam3Cys‐Ser‐(Lys)4 (a TLR2 agonist) and anti‐CD3 antibody. Supernatants were then tested for TNF□, IL‐6, and IL‐10 levels. In some experiments, anti‐IL‐10 or anti‐TGFβ antibodies were tested as blocking reagents. Tregs from burn‐injured versus sham mice were also compared for Treg‐mediated suppression of macrophages. Our results indicate that Tregs can directly suppress TLR4‐ and TLR2‐induced inflammatory cytokine production by macrophages. Moreover, we find that both IL‐10 and TGFβ act synergistically to mediate the suppression of TLR4 and TLR2 responses by macrophages. Finally, we demonstrate that Tregs prepared from injured mice are more potent than sham Tregs at suppressing macrophage TLR2 and TLR4 responses. Taken together, our findings show that Tregs can actively suppress macrophage TLR responses through IL‐10‐and TGFβ‐dependent mechanisms and provide new insight into how Tregs control innate immune responses to injury and inflammation.
TLR4 enhances resolution of lung inflammation by promoting neutrophil apoptosis
Authors: zhao hang , Dedaj rejmon , zhao gao feng , leu shaw wei , shen lian jun , lien egil , fitzgerald Kate , Shiedlin Aviva , quinn deborah ann , hales charles a
Published: June 2021

Acute lung injury (ALI), the called acute respiratory distress syndrome in its most severe (ARDS) clinical manifestation, affects about 150,000 patients per year in the US, with recent mortality rates being > 30%. High concentrations of hyaluronan (HA) are found in bronchoalveolar lavage (BAL) fluid of ARDS patients. Neutrophils appear to play a major role in the development of ALI/ARDS. Here we report that in response to inhaled low molecular weight HA (LMW HA), mice lacking the TLR4‐MyD88 independent signaling pathway showed loss of lung epithelial integrity, diffuse alveolar damage, increased interstitial edema, increased inflammatory cytokines and chemokines, including macrophage inflammatory protein (MIP‐2) (a rodent homolog of IL‐8), interleukin (IL)‐6 and KC compared to wild type. In addition, there were large numbers of activated neutrophils in BAL fluid and lung interstitium in TLR4 knock out mice, indicating TLR4 enhances the resolution of inflammation by promoting apoptosis of neutrophils. Our findings show an unrecognized potential for the treatment of ALI/ARDS disorders.

This work was supported by a training grant from the NIH (HL007874 to H.Z.)
Engagement of TLR2 during priming directs enhanced CD4 T cell expansion and protective capacities compared to other TLR agonists
Authors: Chandran Smita Sarat , Teijaro John R , Fenton Matthew , Farber Donna L
Published: June 2021

Toll‐like receptors (TLR) comprise a family of pattern recognition receptors that recognize pathogen‐associated motifs, and trigger antigen‐presenting cell (APC) activation for priming CD4 T cells. We hypothesized that CD4 T cell responses could be optimized by targeting engagement of specific TLR during priming. We used an in vivo system in which HA‐specific or OVA‐specific CD4 T cells are CFSE labeled, adoptively transferred into BALB/c hosts, and subsequently primed with antigenic peptide in the presence/absence of agonists for TLR2 (Pam3Cys), TLR3 (polyI:C) or TLR4 (lipopolysaccharide). CD4 T cells were harvested 1–2 weeks post‐priming to monitor expansion, functional capacity and protective responses against PR8 Influenza challenge. Transferred CD4 T cells underwent robust antigen‐driven proliferation and expansion when primed with both peptide and TLR agonist compared to peptide alone. Pam3Cys‐mediated priming resulted in significantly expanded populations with high levels of IFN‐γ and IL‐2 production, whereas TLR3 and TLR4 engagement resulted in predominant IFN‐γ production with low IL‐2 production. When challenged with a sub‐lethal dose of Influenza virus, Pam3Cys‐primed CD4 T cells afforded better protection in terms of lung function and viral load compared to TLR3 and TLR4 agonists. Thus targeting TLR2 during early priming leads to enhanced CD4 T cell activation, function and protection.

Supported by NIH grants R21AI065950 and RO1AI50632
Olive baboon θ‐defensins
Authors: Garcia Angie Eva , Selsted Michael
Published: June 2021

θ‐defensins are antimicrobial peptides expressed in leukocytes of Old World monkeys, and they have been implicated as effectors of innate immunity. We detected θ‐defensin expression in the Olive and Gelada baboons, Old World monkeys previously reported to lack θ‐defensin genes. We found that Olive baboons expressed four prepro‐θ‐defensin genes that can, in theory, generate ten unique peptides from alternate pairing of the precursor‐derived nonapeptides. Five of the predicted peptides were purified from baboon leukocytes. Immunohistochemical staining demonstrated that θ‐defensins localize to the cytoplasmic granules of neutrophils and monocytes, and they were also detected in cells of thymus, lung, and spleen. Bactericidal and fungicidal assays, conducted in buffered media, demonstrated that 1–4 μM of each peptide killed > 99.9% of all species tested. The mechanism by which θ‐defensins kill E. coli appears to require permeabilization of the cell envelope. Microbicidal activities were also analyzed in the presence of 154 mM NaCl and 10% serum. Under these conditions, approximately 10‐fold higher peptide concentrations were required for bactericidal activity. Significant differences in target‐specific potency were observed among the baboon θ‐defensins tested, suggesting that θ‐defensin sequences diverged to neutralize a broad range of microbial species.
Crosstalk of the Complement and Coagulation System
Authors: Amara Umme , Albers Sonja , Gebhard Florian , Bruckner Uwe B. , Huber‐Lang Markus
Published: June 2021

Introduction: The complement and coagulation systems are phylogenetically old and important ingredients of innate immune effectors that sense incoming signals of danger. The possible crosstalk and the precise pathways, by which the activation of coagulation triggers the complement cascade and vise versa, remain unclear. Therefore, in the current study the coagulation factors FXIa, FXa, FVIII, FVII, FVIIa, plasminogen, PC, aPC and thrombin were investigated for their potency to in vitro generate C3a and C5a.

Material and Methods: In vitro cleavage experiments with incubation of native C3/C5 ± coagulation factors (FXa, FXIa, FIXa, thrombin) ± various serine protease inhibitors (aprotinin, leupeptine, pefabloc, protease inhibitor cocktail) were performed at 37°C dose‐ and time‐dependently by western blots. The enzymatic activity was assessed by ELISA (C3a Quidel REF A015; C5a, DRG Diagnostic REF EIA 3327).

Results: This study demonstrates bi‐directional interaction of both cascades as the coagulation factors Xa, Xia, IXa and thrombin can dose‐ and time‐dependently cleave the key proteases of the complement system, C3 and C5, and generate biologically active C5a and C3a, resp, as detected by WB, ELISA and chemotactic activity assays of either HMC‐1 cells or neutrophils. The clotting factor FXIa, FXa, IXa and thrombin‐induced C3 and C5 cleavage activity could dose‐dependently be blocked by the serine protease inhibitor leupeptine and by the structure‐dependent inhibitor pefabloc. Furthermore, FXa exhibited maximal enzymatic activity of all measured coagulation factors (FXa<FXIa<thrombin<FIXa<ctrl) in accordance to the Michaelis‐Menten equation.

Conclution: The data provide first evidence of activation of the complement system by various serine proteases of the coagulation system.
Both the lectin and alternative pathways of complement activation play roles in Cryptococcus gattii infection
Authors: Mershon Kileen Louise , Vasuthasawat Alex , Morrison Sherie L. , Beenhouwer David O.
Published: June 2021

Complement is a cascade of serum proteins involved in opsonization, membrane lysis, and anaphylaxis. Complement can be activated through three pathways: classical, alternative, and lectin. It has been shown that pathogenic Cryptococcus species activate the alternative pathway of the complement cascade, while the polysaccharide capsule blocks activation of the classical pathway. While human mannose binding lectin (MBL) has been reported to not bind to Cryptococcus, no published studies have characterized murine MBL binding to Cryptococcus. Using flow cytometry, we analyzed C3 deposition on C. gattii in the presence of serum from C57BL/6J (intact complement pathways), factor B‐deficient (lacking alternative pathway), and MBL‐deficient mice (lacking lectin pathway), and found that complement activation occurred via both the alternative and the lectin pathways. When mice were infected intravenously with C. gattii, C3‐ and factor B‐deficient mice died significantly faster than C57BL/6J mice and showed higher fungal burdens in the lungs. Radiolabeled heat‐killed C. gattii injected intravenously cleared rapidly in C57BL/6J mice. However, in factor B‐deficient and C3‐deficient mice, organisms cleared much less rapidly, indicating a role for complement activation in organism clearance.

NIH R01 AI51415‐5
Targeting phosphoinositide 3‐kinase γ (PI3Kγ) in the treatment of cutaneous leishmaniasis caused by L. mexicana.
Authors: Cummings Hannah Elizabeth , Barbi J. , Keiser T. , Zorko N. , Lu B. , Gerard C. , Ruckle T. , Seveau S. , Rommel C. , Whitacre C. , Satoskar A.
Published: June 2021

Class I PI3Ks are dual‐specificity lipid and protein kinases controlling cell growth, proliferation, survival, adhesion and motility. They include class IA (PI3Kα, PI3Kβ and PI3Kδ) and class IB (PI3Kγ). In this study, we examined the role of PI3Kγ in the development of cutaneous leishmaniasis caused by L. mexicana using a PI3Kγ‐specific inhibitor (PI3Kγinh) as well as PI3Kγ−/‐ mice. We found that daily treatment of L. mexicana‐ infected C57BL/6 mice with the PI3Kγinh inhibited lesion growth and reduced parasite burdens without significant changes in host T cell responses. PI3Kγinh reduced the early adherence of L. mexicana to BMDMs and inhibited uptake in both macrophages and neutrophils in vitro, indicating that PI3Kγ is critical for L. mexicana to enter these cells. PI3Kγ−/‐ mice were resistant to L mexicana infection, developing smaller lesions with fewer parasites than WT mice. PI3Kγ−/‐ had fewer macrophages and neutrophils within the lesions. Parasite uptake was also reduced in PI3Kγ−/‐ BMDMs. These data show that PI3Kγ mediates susceptibility to L. mexicana by 1) promoting the migration of neutrophils and macrophages to the lesion and 2) by facilitating entry of the parasite into these cells, which is critical for parasite survival. These data suggest that PI3Kγ is a therapeutic target for treatment of cutaneous L. mexicana infection.
The CD4+ T cell‐ and SAP‐dependent IgG anti‐polysaccharide response to intact Streptococcus pneumoniae, in contrast to pneumococcal conjugate vaccine, is ICOS‐independent
Authors: Chen Quanyi , Akiba Hisaya , Paton James C. , Cannons Jennifer , Schwartzberg Pamela L. , Snapper Clifford
Published: June 2021

The IgG murine anti‐polysaccharide (PS) and anti‐protein responses to intact Streptococcus pneumoniae (Pn) are CD4+ T cell and B7/CD28 dependent. However, in contrast to the IgG anti‐protein response, the primary IgG anti‐PS response peaks more rapidly, fails to generate memory, and requires a shorter period of CD4+ T cell help and B7‐dependenrt costimulation. Although CD28 is constitutively expressed on naïve CD4+ T cells, ICOS is typically induced during the first 48h following immunization. We now show that ICOS−/− mice immunized with intact Pn elicit a normal IgM and IgG anti‐PS response but a completely defective anti‐protein response, relative to WT mice. Neutralizing anti‐ICOS‐ligand mAb injected at the time of primary, but not secondary, Pn immunization was inhibitory for the anti‐protein response. In distinct contrast, ICOS−/− mice injected with a pneumococcal conjugate vaccine, exhibited both a defective IgG anti‐PS, as well as IgG anti‐protein response. Although the adaptor molecule, SAP is thought to play a key role in T cell‐dependent humoral immune responses through induction of ICOS on CD4+ T cells, SAP−/− mice challenged with intact Pn were highly defective in eliciting the ICOS‐independent IgG anti‐PS response. These data are the first to demonstrate a CD4+ T cell and SAP‐dependent humoral immune response that is ICOS‐independent.

Supported by NIH grant 2R01 AI49192
Effect of Clarithromycin on lung inflammation and alveolar macrophage function in Klebsiella pneumoniae B5055 induced acute lung infection in BALB/c mice
Authors: Kumar Vijay , HARZAI KUSUM , CHHIBBER SANJAY
Published: June 2021

Objective To evaluate the anti‐inflammatory effect of the clarithromycin as well as its effect on macrophage function in Klebsiella pneumoniae B5055 induced acute lung infection in BALB/c mice.

Materials and Methods: Acute lung infection was induced with Klebsiella pneumoniae B5055 by instilling 104 cfu/ml via intranasal route in mice. Group A; control, Group B; clarithromycin (30mg/kg/day/os) treated. The lungs were harvested and homogenized in 1 ml of normal saline and bacterial count, malondialdehyde (MDA), nitric oxide (NO) and myeloperoxidase (MPO) levels. The histopathological examination of the lungs was also done. The lung alveolar macrophage functions (i.e. macrophage spreading phagocytosis) were also assessed.

Results: The clarithromycin treated group showed significant (p<0.05) decrease in the neutrophil influx in to the lungs and other markers of inflammation (MDA, NO and MPO) were also significantly reduced (p<0.05). The clarithromycin treatment also significantly (p<0.05) increased the lung alveolar macrophage spreading and their phagocytic potential.

Conclusion: On this basis we conclude that clarithromycin decreases the lung inflammation induced by K. pneumoniae B5055 with increasing the macrophage activity against Klebsiella pneumoniae B5055 in the lung tissue.
Heat shock enhances innate immune response to PG‐PS and M. tuberculosis in C. elegans
Authors: Kannan Krishnaswamy , Wan Xiaoxiao , Kannan Aarthi , Fang Deyu , Ortmann Robert
Published: June 2021

C. elegans is a soil nematode with a functioning immune system. It shares many similarities with mammals in their immune response to various pathogens and serves as a simple in vivo model to study innate immunity, including responses made via the toll‐like receptor (TLR) pathway. We investigated the expression of nlp‐29, a candidate antimicrobial gene, to study the host immune response to two gram positive bacterial isolates: (1) peptidoglycan‐polysaccharide (PG‐PS, an extract from streptococcal cell wall) and (2) the soluble fraction of pulverized heat‐killed M. tuberculosis strain H37ra (M. tb). Neither antigen is pathogenic to C. elegans and the worms do not exhibit any avoidance behavior.

Our results showed a differential response to PG‐PS and M. tb, both rich in TLR2 agonists. As compared to OP50 E. Coli fed control, PG‐PS treated worms evoked a robust response by both DAF‐16 nuclear translocation and nlp‐29 expression. In comparison, M. tb treated worms exhibited less robust DAF‐16 translocation and no detectable nlp‐29 expression. However, mild heat‐shock at 37°C for an hour enhanced nlp‐29 expression significantly in PG‐PS treated worms and reached detectable levels in the M. tb treated group suggesting a differential response. Our results demonstrate the presence of an inducible innate immune response in C. elegans to PG‐PS and M. tb, both gram positive bacterial preparations rich in TLR2 ligands.
IL‐12 induced STAT‐4 signaling is increased in CD8 T cells from aged mice
Authors: Turner Joanne
Published: June 2021

Old mice express an early resistance to infection with Mycobacterium tuberculosis (M.tb) which is associated with CD8 T cells and IFN‐γ production. CD8 T cells from old mice are also capable of secreting IFN‐γ in an antigen independent manner in response to TH1 cytokines. In this study we sought to determine whether CD8 T cells from old mice were more responsive to IL‐12. Using flow cytometric analysis of phosphorylated STAT4 we found that old mice possessed more CD8 T cells that could respond directly to IL‐12 than young mice. Furthermore, on an individual cell basis, CD8 T cells from old mice could phosphorylate more STAT4 than similar cells from young mice. This increased phosphorylation was independent of STAT4, as STAT4 levels in CD8 T cells were equivalent between young and old mice. These data indicate that CD8 T cells from old mice have enhanced IL‐12 signaling, which ultimately leads to the secretion of more IFN‐γ. The presence of increased levels of IL‐12 and IFN‐γ within the lungs of old mice early after infection with M.tb provides additional evidence that CD8 T cells contribute to the expression of early resistance to infection. This work was supported by R01‐AG21097.
Effect of three supplements with identical micronutrient content on anemia in pregnant Mexican women
Authors: HERNANDEZ AMIRA , Garcia‐Guerra Armando , Dominguez Clara P. , Garcia‐Feregrino Raquel , Neufeld Lynnette M.
Published: June 2021

The objective of this analysis was to assess the impact of three nutritional supplements with identical micronutrient content on the prevalence of anemia (PA) during pregnancy (37 wks) and at 3 m postpartum (pp). Communities (18 per supplement) were randomly assigned to receive a fortified beverage (FB), tablets (TB) or Sprinkles (SK). Pregnant women, beneficiaries of the Oportunidades program were recruited before 25 wk pregnancy. Supplements were delivered daily (6 mo) then weekly and compliance recorded. Appropriate statistical analyses for cluster randomized trials were used. 715 women were recruited (FB n=233; TB n=238; SK n=244). The PA was not different between groups at baseline (FB: 28.8%; TB: 18.1%; SK: 18.5%, p>0.05). At 37 wk pregnancy the PA was not different from that at baseline, nor did it differ between groups (FB: 19.6%, TB: 15%, SK: 17%). At 3 m pp, the PA had decreased in all groups and there was no significant difference in prevalence between them (FB: 24.7%; TB: 14.7%; SK: 15.1%). Given that this study has no true placebo, we do not know to what extent the supplements cured/prevented anemia. The lack of increase in anemia between baseline and 37 wk pregnancy suggests that the supplements may have been equally efficacious to prevent anemia. The Oportunidades program financed this study and holds the rites to the data presented.
An Information‐Motivation‐Behavioral Skills Model for Child Sweetened Beverage Consumption
Authors: Goodell L. Suzanne , Amico K. Rivet , Pierce Michelle B. , Ferris Ann M.
Published: June 2021

As childhood overweight rates increase, a critical need exists to understand parental choices impacting their child's caloric intake. One important and often overlooked contributor to a child's caloric intake is sweetened beverage consumption (CSBC). This study sought to apply a well validated theory of health behavior, the Information‐Motivation‐Behavioral Skills (IMB) model, to CSBC. The proposed IMB‐CSBC model hypothesizes that CSBC‐related information, motivation, and behavioral skills are key determinants of CSBC. We evaluated this model with 198 parents of preschoolers via a self‐report IMB‐CSBC survey and home beverage inventory. Using Structural Equation Modeling, we developed a model accounting for 14% of the variation in CSBC. According to the model (see Figure 1), parents’ information and behavioral skills about CSBC negatively related to CSBC. Behavioral skills also mediated a relationship between information and CSBC, as well as motivation and CSBC. This study provides support for the core IMB‐CSBC model constructs. Using this model, we plan to develop interventions for parents targeting areas such as accurately identifying sweetened and unsweetened beverages, learning how to set and implement limits for child beverage consumption, and reducing the impact of TV on child beverage preference.

Funded by Storrs Agricultural Experiment Station, USDA National Needs Fellowship and UConn FSNE
Caregivers’ attitudes regarding portion size served to Head Start children
Authors: Goodell L. Suzanne , Tsuei Eugenia , Hughes Sheryl O. , Nicklas Theresa A.
Published: June 2021

The purpose of this study was to identify caregivers’ attitudes regarding amounts and types of foods served to Head Start preschoolers using qualitative methods. Researchers conducted 8 focus groups (4 African American; 4 Hispanic) with 33African American and 29 Hispanic Head Start caregivers. Moderator guide questions were based on the Knowledge, Attitudes, and Behavior Model articulated to portion sizes served to Head Start preschoolers. Analysts coded verbatim transcripts for emergent patterns and discrepant data between ethnicities. Caregivers identified child preference, exposure, and pickiness, child age and size, and hunger and the home environment as key influencers on the amounts and types of foods served to Head Start children. Caregivers believed they strongly impact child food consumption through role modeling and verbal encouragement. Using the results from this study, researchers plan to establish how Head Start caregivers’ attitudes impact portion sizes and how portion size impacts consumption. Practitioners should be aware of caregivers’ attitudes regarding their influence on child food consumption and teach these caregivers appropriate behavior modeling and affirmation techniques.

Funded by: #NIH 1 R01 CA107545 and NIH Ruth L. Kirschstein National Research Service Award.
Changes in Consumer Knowledge and Behavioral Intention Toward Genetically Engineered (GE) Foods
Authors: Koutoubi Samer , Heiselmann Philip R. , Murdoch Scott , Sasagawa Masa
Published: June 2021

The main objective of this study is to assess change in the average American consumer's behavioral intention (BI) toward the avoidance of GE foods in the marketplace as a function of change in the consumer's level of knowledge about these foods. Thirty adult mainstream grocery store shoppers participated in this randomized, controlled educational intervention study. Participants completed demographic, knowledge, and BI questionnaires both before and after an educational intervention about GE foods in the marketplace. The questionnaire and educational materials were worded for neutrality – that is, to emphasize neither the pros nor the cons of GE foods. Student's t‐Test and Mann‐Whitney's U test were used and statistical significance defined as p<0.05. The experimental group showed a statistically significant (p<0.001) pre/post increase in knowledge score as compared to the control group. There were no statistically significant pre/post changes in BI scores between the two groups. Within groups, there was a statistically significant (p<0.001) increase in mean knowledge score for the experimental group and a statistically significant (p<0.05) decrease in mean knowledge score for the control group. No statistically significant correlations were found between changes in knowledge and BI scores. The results of this study indicate that neutral information may have little effect on consumer BI toward GE foods.
Comparative study on dietary habits and physical development affected by computer use of elementary school children by gender difference
Authors: Lee Bog‐Hieu , Kim Hye‐Seon , Kim Mi‐Young , Han Chan‐Kyu , Choi Changsun
Published: June 2021

This study was to investigate the elementary school student's dietary habits and physical development relating computer use by gender. Two‐hundred and sixty five elementary school students from Gyeonggi and Seoul were employed in 2006. The study tools were survey questionnaires including general characteristics, dietary habits and anthropometric measurement. A dietary survey was conducted using 24‐hour recall method. All the subjects were using computer at least three to five years. Purpose of computer use was game for boys and internet use for girls(p<0.001). Most used it for one to two hours a day at home during night time. Girls’ dietary habits score(37.5 pts) was better than that of the boys(35.9 pts). Both children had snacking habitually during computer use(p<0.05). The most favorite snack was cookies for boys and beverages for girls. Both consumed considerable instant foods and sugary food, did not enough seaweeds. Boys ate less fruits and consumed more fat food. Children snacking during computer session showed the heavier waist circumference and the higher RI and the higher %RBW compared with those not snacking during computer session(p<0.05). From the findings, computer use by the elementary school children negatively affect their dietary habits and physical development. Also, boys had bad dietary habits and nutitional status. The undesirable dietary habit caused by the use of computer should be corrected through long‐term systematic nutrition education in school, society and at home.