Jianwen Bai

TAT-SNAP-23 treatment inhibits the priming of neutrophil functions contributing to shock and/or sepsis-induced extra-pulmonary acute lung injury

Respiratory burst function of neutrophils is thought to play a pivotal role in the development of pathologies such as indirect (extra-pulmonary) acute lung injury (iALI), as well as sepsis. The current study was conducted to determine the effect of an HIV transactivator of transcription (TAT)-fusion protein containing a soluble N-ethylmaleimide-sensitive factor attachment protein receptor domain from synaptosome-associated protein-23 (SNAP-23) on the shock/sepsis- and sepsis-enhanced neutrophil burst capacity using the clinical relevant two-hit iALI mouse model and the classical cecal ligation and puncture (CLP) septic model. TAT-SNAP-23 significantly decreased the blood neutrophil respiratory burst in vitro, and also in vivo in CLP and hemorrhaged mice. We found that the neutrophil influx to the lung tissue, as measured by myeloperoxidase levels and neutrophil-specific esterase+ cells, was also decreased in the TAT-SNAP-23-treated group. Consistent with this, treatment of TAT-SNAP-23 significantly reduced the disruption of lung tissue architecture and protein concentration of bronchoalveolar lavage fluid in iALI mice compared with vehicle-treated iALI mice. In addition, although TAT-SNAP-23 did not alter the extent of local cytokine/chemokine expression, the in vitro migration capacity of neutrophils was blunted from septic and hemorrhagic mice. These data support our hypothesis that TAT-SNAP-23 reduces neutrophil dysfunction in iALI and sepsis by inhibiting neutrophil respiratory burst.

Programmed cell death receptor ligand 1 modulates the regulatory T cells' capacity to repress shock/sepsis-induced indirect acute lung injury by recruiting phosphatase SRC homology region 2 domain-containing phosphatase 1.

ABSTRACT We recently reported that adoptively transferred (AT) exogenous CD4+CD25+ regulatory T cells (Tregs) to wild-type (WT) mice can directly act to repress shock/sepsis–induced experimental indirect acute lung injury (iALI), and this is mediated in part by programmed cell death receptor 1 (PD-1). In this study, we further determine whether recipient mouse lacking PD-L1, one of the primary ligands for PD-1, contributes to the manipulation of the Tregs’ capacity to repress lung injury. To do this, Tregs isolated from the spleen of WT mice were AT into PD-L1−/− mice subjected to hemorrhagic shock and subsequent to cecal ligation and puncture to induce iALI. Samples were collected for analyses 24 h after cecal ligation and puncture. We found that in PD-L1−/−–recipient mice, AT WT-Tregs lost the ability to reverse the development of iALI seen in WT recipient mice (i.e., no reduction of lung injury indices assessed by histology and vascular leakage, failure to decrease the lung neutrophil influx [myeloperoxidase activity], or the rise in lung apoptosis [caspase 3 activity]). Also, a significant increase in interleukin 1&bgr; (IL-1&bgr;) and keratinocyte-derived chemokine, but no changes in IL-6, IL-10, and IL-17A levels in lung tissues were seen in these mice compared with iALI mice without AT of Tregs. Furthermore, we noted that the lung tissue tyrosine phosphatase Src homology region 2 domain-containing phosphatase 1 (SHP-1), but not SHP-2, was activated with the AT of Tregs in PD-L1−/− iALI mice. Finally, through local depletion of CD4+ T cells or CD25+ (Tregs) in the lung, prior to inducing iALI, we found that SHP-1 activation was associated with the loss of Tregs’ protective effects in vivo. Collectively, our data reveal that PD-L1 is a critical modulator of Tregs’ ability to suppress iALI, and this appears to involve SHP-1 activation.

Active players in resolution of shock/sepsis induced indirect lung injury: immunomodulatory effects of Tregs and PD-1.

The immunomodulatory effects of PD‐1 and CD4+CD25+ Tregs in the resolution of ALI are still poorly understood. Accordingly, 1 million Tregs were isolated from spleens of WT C57BL/6 or PD‐1−/− mice (magnetical bead purification and subsequent labeling with/without Vybrant dye) and then AT into mice subjected to Hem shock during their resuscitation period, which were subsequently subjected to CLP/septic challenge (24 h post‐Hem) to induce iALI. Initially, we demonstrated that Vybrant‐labeled AT Tregs appear in the lungs of iALI mice. Subsequently, we found that AT of WT Tregs induced a significant repression of the indices of lung injury: a reduction of neutrophil influx to the lung tissue and a decrease of lung apoptosis compared with vehicle‐treated iALI mice. In addition, these mice had substantially higher concentrations of BALF and lung‐tissue IL‐10 but significantly decreased levels of lung KC. However, these beneficial effects of the AT of Tregs were lost with the administration of PD‐1−/− mouse Tregs to the recipient WT mice. ALI was exacerbated in these recipient mice receiving AT PD‐1−/− Tregs to the same extent as iALI mice that did not receive Tregs. These data imply that Tregs can act directly to modify the innate immune response induced by experimental iALI, and this is mediated, in part, by PD‐1. Hence, the manipulation of Tregs may represent a plausible target for treating iALI.

Interleukin 2 Gene Polymorphisms Are Associated with Non-Hodgkin Lymphoma

Non-Hodgkin lymphoma (NHL) is the most common hematologic malignancy worldwide. Interleukin-2 (IL-2) plays a key role in the proliferation of T cells and natural killer cells. It has been reported that polymorphisms in the IL-2 gene are associated with various cancers. The aim of this study was to examine the effect of polymorphisms in the IL-2 gene on the development of NHL in the Chinese population. IL-2-330T/G and +114T/G polymorphisms were detected by polymerase chain reaction-restriction fragment length polymorphism in 438 NHL cases and 482 age-matched healthy controls. Data were analyzed using the Chi-square test. Results showed that individuals with -330TG genotype or -330GG genotype had significantly increased susceptibility to NHL (Odds ratio [OR] = 1.40, 95% confidence interval [CI]: 1.05-1.85, p = 0.020 and OR = 2.04, 95%CI: 1.28-3.24, p = 0.002). Meanwhile, the +114T/G polymorphism did not show any correlation with NHL. When analyzing the haplotypes of these two polymorphisms, the prevalence of -330G/+114T haplotype was significantly higher in NHL cases than in controls (OR = 1.45, 95%CI: 1.12-1.88, p = 0.005). These data indicate that IL-2 gene polymorphisms may be new risk factors for NHL.

Early Recruitment of IL-10-Producing B Cells Into Alveoli Improved the Resolution of Acute Lung Injury

Background: Acute lung injury (ALI) is characterized by rapid induction of inflammation at the alveolar-capillary membrane, and immunosuppressive mechanisms were shown to contribute to its resolution. Despite the central role of lymphocytes in initiating and mediating an inflammatory response, their influx dynamics in ALI has not been examined. Methods: We collected mini-BAL samples from the lung of ALI patients over a maximum period of 7 days, and examined the lymphocyte composition. Results: CD3+CD4+IFN-gamma+ Th1 cells were detected early on in all patients examined, while IL-10-producing B cells and CD3+CD4+CD25hiFoxp3+ Treg cells appeared later. Interestingly, IL-10-producing B cells appeared earlier than Tregs in most subjects, which possibly exerted anti-inflammatory function before Tregs. We then found that in patients with earlier recruitment of IL-10-producing B cells, the magnitude of Th1 inflammation decreased significantly over time, which was not observed in patients with later recruitment of IL-10-producing B cells. Furthermore, early IL-10-producing B cell recruiters also had significantly earlier recruitment of Tregs and better survival than late IL-10-producing B cell recruiters. Conclusion: This study provided data on the alveolar infiltration of lymphocytes during ALI, which suggested an inhibitory role of IL-10-producing B cells in ALI and emphasized the importance of controlling inflammation during the initial stage of ALI.

Enhanced Innate Inflammation Induced by Anti-BTLA Antibody in Dual Insult Model of Hemorrhagic Shock/Sepsis.

ABSTRACT Sepsis following hemorrhagic shock is a common clinical condition, in which innate immune system suffers from severe suppression. B and T lymphocyte attenuator (BTLA) is an immune-regulatory coinhibitory receptor expressed not only on adaptive, but also on innate immune cells. Our previous data showed that BTLA gene deficient mice were protected from septic mortality when compared with wild-type control C57BL/6 mice. Here, we extended our study by treating C57BL/6 mice with an anti-BTLA monoclonal antibody (clone 6A6; reported to have the ability to neutralize or agonize/potentiate BTLA signaling) in a mouse model of hemorrhagic shock (Hem) followed by sepsis induced by cecal ligation and puncture (CLP); positing initially that if BTLA engagement was neutralized, like gene deficiency, an anti-BTLA mAb would have the similar effects on the inflammatory response/morbidity in these mice after such insults. Here, we report that BTLA expression is elevated on innate immune cells after Hem/CLP. However, anti-BTLA antibody treatment increased cytokine (TNF-&agr;, IL-12, IL-10)/chemokine (KC, MIP-2, MCP-1) levels and inflammatory cells (neutrophils, macrophages, dendritic cells) recruitment in the peritoneal cavity, which in turn aggravated organ injury and elevated these animals’ mortality in Hem/CLP. When compared with the protective effects of our previous study using BTLA gene deficient mice in a model of lethal septic challenge, we further confirmed BTLAs contribution to enhanced innate cell recruitment, elevated IL-10 levels, and reduced survival, and that engagement of antibody with BTLA potentiates/exacerbates the pathophysiology in Hem/sepsis.

M2a and M2c Macrophage Subsets Ameliorate Inflammation and Fibroproliferation in Acute Lung Injury Through Interleukin 10 Pathway

ABSTRACT The role of M2 macrophages in the resolution and fibroproliferation of acute lung injury (ALI) is poorly understood. In this study, we investigated the effects of two M2 macrophage subtypes, M2a induced by interleukin (IL)-4/IL-13 and M2c induced by IL-10/transforming growth factor -&bgr;, on the pathogenesis of ALI. M2a and M2c were adoptively transferred into lipopolysaccharide-induced ALI mice model. Data showed that Vybrant-labeled macrophages appeared in the lungs of ALI mice. Subsequently, we observed that both subsets significantly reduced lung inflammation and injury including a reduction of neutrophil influx into the lung and an augmentation of apoptosis. Interestingly, M2c macrophages more effectively suppressed indices of lung injury than M2a macrophages. M2c macrophages were also more effective than M2a in reduction of lung fibrosis. In addition, we found that M2c but not M2a macrophages increased IL-10 level in lung tissues of the recipient ALI mice partially mediated by activating the JAK1/STAT3/suppressor of cytokine signaling 3 signaling pathway. After blocking IL-10, these superior effects of M2c over M2a were abolished. These data imply that M2c are more potent than M2a macrophages in protecting against lung injury and subsequent fibrosis due to their ability to produce IL-10. Therefore, reprogramming macrophages to M2c subset may be a novel treatment modality with transitional potential.

Severe Pneumonia Mortality in Elderly Patients Is Associated With Downregulation of Toll-like Receptors 2 and 4 on Monocytes

Background:Elderly patients with pneumonia have a high mortality rate. Since Toll-like receptor (TLR) signaling is involved in the inflammatory response, we conducted a prospective observational case-control study to assess the relationships of TLR2 and TLR4 mortality with elderly patients with pneumonia. Methods:Clinical and laboratory data were collected from these elderly patients with pneumonia (patient group; n = 40) and healthy age-matched subjects (control 1; n = 20). TLR2 and TLR4 expression levels on blood monocytes were examined, and inflammatory cytokine tumor necrosis factor-&agr; (TNF-&agr;), interleukin-1 (IL-1) and interleukin-6 (IL-6) plasma concentrations were analyzed. Results:Increased TLR2 and TLR4 expression levels were detected in elderly patients with severe pneumonia compared with healthy subjects. Furthermore, we observed a positive correlation between TLR2 and TLR4 expression levels and IL-1 and IL-6 levels, and TLR2- and TLR4-positive cell percent expressions correlated with TNF-&agr; levels. By day 28 of observation, the mortality rate of the patient group was 30%. Decreased TLR2 and TLR4 expression was observed in deceased patients compared with survivors. APACHE (Acute Physiology and Chronic Health Evaluation) II scores and CURB-65 scores (Confusion, blood Urea nitrogen, Respiratory rate and low Blood pressure scores) were lower in survivors. Conclusions:These findings demonstrate a previously undocumented association between mortality in elderly patients with severe pneumonia and decreased TLR2 and TLR4 expression. Our results highlight that TLRs can be targeted in the development of improved immune modulation therapies for these patients.

The Association of Monocyte Chemotactic Protein-1 and CC Chemokine Receptor 2 Gene Variants with Chronic Obstructive Pulmonary Disease

Chemokines are potent proinflammatory cytokines that are implicated in numerous inflammatory diseases. Monocyte chemoattractant protein-1 (MCP-1) and its receptor CC chemokine receptor-2 (CCR2) play a major role in the recruitment of inflammatory cells to the lungs of patients with chronic obstructive pulmonary disease (COPD). We investigated a possible association between polymorphisms in MCP-1 and CCR2 genes (MCP-1 -2518 A/G and CCR2 190G/A or V64I) and the development of COPD. Genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism in 386 COPD cases and 398 age-matched healthy controls. Frequency of MCP-1 2518GG genotype for cases and controls was 0.396 and 0.324, respectively; individuals who had the GG genotype had a 1.59-fold increased risk of COPD (p=0.036). Frequency of CCR2 190AA (64I/64I) genotype for cases and controls was 0.285 and 0.21, respectively; subjects carrying the 64I/64I genotype had a 2.04-fold increased risk of COPD compared with the wild-type genotype (p=0.001). When analyzing the allele combination of these two polymorphisms, the combinations MCP-1-A/CCR2-A and MCP-1-G/CCR2-A were detected in significantly higher numbers in COPD cases than in healthy controls (odds ratio [OR]=1.50, 95% confidence interval [CI]: 1.04-2.17, p=0.032; and OR=1.89, 95% CI: 1.38-2.60, p=7.38×10(-5)). These data suggest that MCP-1 -2518 A/G and CCR2 190G/A polymorphisms are new risk factors for COPD.

Upregulation of CD19 + CD24 hi CD38 hi regulatory B cells is associated with a reduced risk of acute lung injury in elderly pneumonia patients

Abstract Acute lung injury (ALI) is a common complication in elderly pneumonia patients who have a rapid progression, and is accompanied by a high mortality rate. Because the treatment options of ALI are limited to supportive care, identifying pneumonia patients who are at higher risk of ALI development is the emphasis of many studies. Here, we approach this problem from an immunological perspective by examining CD19+CD24hiCD38hi B cells, an important participant in acute and chronic inflammation. We find that elderly pneumonia patients have elevated CD19+CD24hiCD38hi B cell frequency compared to healthy individuals. This B cell population may express a higher level of IL-10, which has been was shown to suppress CD4+ T cell-mediated proinflammatory cytokine interferon gamma (IFNg) and tumor necrosis factor alpha (TNFa) production, through an IL-10-dependent mechanism. We also observe that the frequency of CD19+CD24hiCD38hi B cell is positively correlated with the frequency of CD4+CD25+Foxp3+Tregs in peripheral blood. Moreover, consistent with CD19+CD24hiCD38hi B cell’s anti-inflammatory role, we find that pneumonia patients who later developed ALI have reduced level of CD19+CD24hiCD38hi B cells. Together, our results demonstrated that CD19+CD24hiCD38hi B cells in pneumonia patients possess regulatory function in vivo, and are associated with a reduced ALI risk.