An Integrated Plasmo-Photoelectronic Nanostructure Biosensor Detects an Infection Biomarker Accompanying Cell Death in Neutrophils.

Bacterial infections leading to sepsis are a major cause of deaths in the intensive care unit. Unfortunately, no effective methods are available to capture the early onset of infectious sepsis near the patient with both speed and sensitivity required for timely clinical treatment. To fill the gap, the authors develop a highly miniaturized (2.5 × 2.5 µm2 ) plasmo-photoelectronic nanostructure device that detected citrullinated histone H3 (CitH3), a biomarker released to the blood circulatory system by neutrophils. Rapidly detecting CitH3 with high sensitivity has the great potential to prevent infections from developing life-threatening septic shock. To this end, the author's device incorporates structurally engineered arrayed hemispherical gold nanoparticles that are functionalized with high-affinity antibodies. A nanoplasmonic resonance shift induces a photoconduction increase in a few-layer molybdenum disulfide (MoS2 ) channel, and it provides the sensor signal. The device achieves label-free detection of serum CitH3 with a 5-log dynamic range from 10-4 to 101 ng mL and a sample-to-answer time <20 min. Using this biosensor, the authors longitudinally measure the dynamic CitH3 profiles of individual living mice in a sepsis model at high resolution over 12 hours. The developed biosensor may be poised for future translation to personalized management of systemic bacterial infections.

Citrullinated Histone H3 Mediates Sepsis-Induced Lung Injury Through Activating Caspase-1 Dependent Inflammasome Pathway.

Sepsis is a life-threatening organ dysfunction caused by dysregulated host response to infection that often results in acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). An emerging mechanism of sepsis-induced ARDS involves neutrophils/macrophages undergoing cell death, releasing nuclear histones to cause tissue damage that exacerbates pulmonary injury. While published studies focus on unmodified histones, little is known about the role of citrullinated histone H3 (CitH3) in the pathogenesis of sepsis and ALI. In this study, we found that levels of CitH3 were elevated in the patients with sepsis-induced ARDS and correlated to PaO2/FiO2 in septic patients. Systematic administration of CitH3 peptide in mice provoked Caspase-1 activation in the lung tissue and caused ALI. Neutralization of CitH3 with monoclonal antibody improved survival and attenuated ALI in a mouse sepsis model. Furthermore, we demonstrated that CitH3 induces ALI through activating Caspase-1 dependent inflammasome in bone marrow derived macrophages and bone marrow derived dendritic cells. Our study suggests that CitH3 is an important mediator of inflammation and mortality during sepsis-induced ALI.

Cl-Amidine Improves Survival and Attenuates Kidney Injury in a Rabbit Model of Endotoxic Shock.

Objective: Sepsis causes millions of deaths on a global scale annually. Activation of peptidylarginine deiminase (PAD) enzymes in sepsis causes citrullination of histones, which results in neutrophil extracellular trap formation and sepsis progression. This study evaluates pan-PAD inhibitor, Cl-amidine, in a model of lipopolysaccharide (LPS)-induced endotoxic shock in rabbits. We hypothesized that Cl-amidine would improve survival and attenuate kidney injury. Methods: In the survival model, rabbits were injected injected intravenously with 1 mg/kg of LPS, and then randomly assigned either to receive dimethyl sulfoxide (DMSO; 1 mcL/g) or Cl-amidine (10 mg/kg diluted in 1 mcL/g DMSO). They were then monitored for 14 days to evaluate survival. In the non-survival experiment, the same insult and treatment were administered, however; the animals were euthanized 12 hours after LPS injection for kidney harvest. Acute kidney injury (AKI) scoring was performed by a histopathologist who was blinded to the group assignment. Serial blood samples were also collected and compared. Results: Rabbits that received Cl-amidine had a higher survival (72%) compared with the rabbits that received DMSO (14%; p < 0.05). Cl-amidine-treated rabbits had lower (p < 0.05) histopathologic AKI scores, as well as plasma creatinine and blood urea nitrogen (BUN) levels 12 hours after insult. Conclusions: Pan-PAD inhibitor Cl-amidine improves survival and attenuates kidney injury in LPS-induced endotoxic shock in rabbits.

Efficacy and Safety of Ultrasound-Guided Radiofrequency Treatment for Chronic Pain in Patients with Knee Osteoarthritis: A Systematic Review and Meta-Analysis.

Background: Knee osteoarthritis (KOA) is a common degenerative disease associated with joint dysfunction and pain. Ultrasound-guided radiofrequency (RF) may be a promising therapy in the treatment of chronic pain for KOA patients. Objective: To evaluate the efficacy and safety of ultrasound-guided RF treatment for chronic pain in patients with KOA. Design: A systematic review was conducted, and a meta-analysis was carried out when possible. Setting. We examined the studies evaluating the clinical efficiency of ultrasound-guided RF on chronic pain in KOA population. Method: A systematic review for the efficacy and safety of ultrasound-guided RF treatment for pain management of KOA patients was carried out in PubMed, EMBASE, Cochrane Library, Web of Science, Wanfang Data, and China National Knowledge Infrastructure (CNKI) from the date of inception to February 2020, and a meta-analysis was conducted. The primary outcomes of pain intensity (visual analogue scale or numerical rating scale) and knee function [the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC)] were evaluated from baseline to various follow-up times by random-effects model. Heterogeneity was assessed by I 2 statistic and the potential sources of heterogeneity by subgroup and metaregression analyses, respectively. Results: Eight publications with 256 patients were included in the meta-analysis. RF could relieve pain with -4.196 of pooled mean difference and improve knee function by decreasing 23.155 points in WOMAC. Three patients had ecchymosis, two with hypoesthesia and one with numbness after the procedure, and improved within 6 months. Furthermore, study design and treatment target were the sources of heterogeneity by subgroup and metaregression analyses, accounting for 37% and 74% of variances, respectively. Target of genicular nerve achieved better pain relief than intra-articular or sciatic nerve. Sensitivity analysis showed that removal of any single study was unlikely to overturn the findings. Limitations. There were some limitations in the study. Firstly, the small number of relevant studies limited the confidence level of the meta-analysis. Also, the significant heterogeneity may not be explained due to the limited data. Secondly, the direct comparison of two different guidance methods (ultrasound vs. fluoroscopy) for RF therapy is lacking. In addition, the outcomes were blindly assessed in the meta-analysis from all studies according to evaluation of bias, which could affect the reality of the data. Finally, most of the studies only provided short follow-up times, so we could not analyze the long-term effectiveness of ultrasound-guided RF in the treatment of patients with KOA. Conclusions: Ultrasonography is an effective, safe, nonradiative, and easily applicable guidance method for RF in pain relief and functional improvement in KOA patients.

Inhibition of PAD2 Improves Survival in a Mouse Model of Lethal LPS-Induced Endotoxic Shock.

Endotoxemia induced by lipopolysaccharide (LPS) is an extremely severe syndrome identified by global activation of inflammatory responses. Neutrophil extracellular traps (NETs) play an important role in the development of endotoxemia. Histone hypercitrullination catalyzed by peptidylarginine deiminases (PADs) is a key step of NET formation. We have previously demonstrated that simultaneous inhibition of PAD2 and PAD4 with pan-PAD inhibitors can decrease NETosis and improve survival in a mouse model of LPS-induced endotoxic shock. However, the effects of PAD2 specific inhibition during NETosis and endotoxic shock are poorly understood. Therefore, in the present study, we aimed to investigate the effect of the specific PAD2 or PAD4 inhibitor on LPS-induced endotoxic shock in mice. We found that PAD2 inhibition but not PAD4 inhibition improves survival. Also, the levels of proinflammatory cytokines and NETosis were significantly reduced by PAD2 inhibitor. To our knowledge, this study demonstrates for the first time that PAD2 inhibition can reduce NETosis, decrease inflammatory cytokine production, and protect against endotoxin-induced lethality. Our findings provided a novel therapeutic strategy for the treatment of endotoxic shock.

Peptidylarginine deiminase 2 has potential as both a biomarker and therapeutic target of sepsis.

Peptidylarginine deiminases (PADs) are a family of calcium-dependent enzymes that are involved in a variety of human disorders, including cancer and autoimmune diseases. Although targeting PAD4 has shown no benefit in sepsis, the role of PAD2 remains unknown. Here, we report that PAD2 is engaged in sepsis and sepsis-induced acute lung injury in both human patients and mice. Pad2-/- or selective inhibition of PAD2 by a small molecule inhibitor increased survival and improved overall outcomes in mouse models of sepsis. Pad2 deficiency decreased neutrophil extracellular trap (NET) formation. Importantly, Pad2 deficiency inhibited Caspase-11-dependent pyroptosis in vivo and in vitro. Suppression of PAD2 expression reduced inflammation and increased macrophage bactericidal activity. In contrast to Pad2-/-, Pad4 deficiency enhanced activation of Caspase-11-dependent pyroptosis in BM-derived macrophages and displayed no survival improvement in a mouse sepsis model. Collectively, our findings highlight the potential of PAD2 as an indicative marker and therapeutic target for sepsis.

HDAC6 mediates an aggresome-like mechanism for NLRP3 and pyrin inflammasome activation.

Inflammasomes are supramolecular complexes that play key roles in immune surveillance. This is accomplished by the activation of inflammatory caspases, which leads to the proteolytic maturation of interleukin 1ß (IL-1ß) and pyroptosis. Here, we show that nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3)- and pyrin-mediated inflammasome assembly, caspase activation, and IL-1ß conversion occur at the microtubule-organizing center (MTOC). Furthermore, the dynein adapter histone deacetylase 6 (HDAC6) is indispensable for the microtubule transport and assembly of these inflammasomes both in vitro and in mice. Because HDAC6 can transport ubiquitinated pathological aggregates to the MTOC for aggresome formation and autophagosomal degradation, its role in NLRP3 and pyrin inflammasome activation also provides an inherent mechanism for the down-regulation of these inflammasomes by autophagy. This work suggests an unexpected parallel between the formation of physiological and pathological aggregates.

Citrullinated Histone H3 as a Therapeutic Target for Endotoxic Shock in Mice.

Sepsis results in millions of deaths every year, with acute lung injury (ALI) being one of the leading causes of mortality in septic patients. As neutrophil extracellular traps (NETs) are abundant in sepsis, neutralizing components of NETs may be a useful strategy to improve outcomes of sepsis. Citrullinated histone H3 (CitH3) has been recently shown to be involved in the NET formation. In this study, we demonstrate that CitH3 damages human umbilical vein endothelial cells (HUVECs) and potentiates NET formation through a positive feedback mechanism. We developed a novel CitH3 monoclonal antibody to target peptidylarginine deiminase (PAD) 2 and PAD 4 generated CitH3. In a mouse model of lethal lipopolysaccharide (LPS) induced shock, neutralizing CitH3 with the newly developed anti-CitH3 monoclonal antibody attenuates inflammatory responses, ameliorates ALI, and improves survival. Our study suggests that effectively blocking circulating CitH3 might be a potential therapeutic method for the treatment of endotoxemia.

Bone Marrow Mesenchymal Stem Cells-Derived Exosomal MiR-29b-3p Regulates Aging-Associated Insulin Resistance.

Insulin resistance is the major pathological characteristic of type 2 diabetes, and the elderly often develop insulin resistance. However, the deep-seated mechanisms for aging-related insulin resistance remain unclear. Here, we showed that nanosized exosomes released by bone marrow mesenchymal stem cells (BM-MSCs) of aged mice could be taken up by adipocytes, myocytes, and hepatocytes, resulting in insulin resistance both in vivo and in vitro. Using microRNA (miRNA) array assays, we found that the amount of miR-29b-3p was dramatically increased in exosomes released by BM-MSCs of aged mice. Mechanistically, SIRT1 (sirtuin 1) was identified to function as the downstream target of exosomal miR-29b-3p in regulating insulin resistance. Notably, utilizing an aptamer-mediated nanocomplex delivery system that down-regulated the level of miR-29b-3p in BM-MSCs-derived exosomes significantly ameliorated the insulin resistance of aged mice. Meanwhile, BM-MSCs-specific overexpression of miR-29b-3p induced insulin resistance in young mice. Taken together, these findings suggested that BM-MSCs-derived exosomal miR-29b-3p could modulate aging-related insulin resistance, which may serve as a potential therapeutic target for aging-associated insulin resistance.

Inhibition of peptidylarginine deiminase alleviates LPS-induced pulmonary dysfunction and improves survival in a mouse model of lethal endotoxemia.

Immune cell death caused by neutrophil extracellular traps (NETs), referred to as NETosis, can contribute to the pathogenesis of endotoxemia and organ damage. Although the mechanisms by which infection induces NETosis and how that leads to organ dysfunction remain largely unknown, NET formation is often found following citrullination of histone H3 (CitH3) by peptidylarginine deiminase (PAD). We hypothesized that lipopolysaccharide (LPS)-induced activation of PAD and subsequent CitH3-mediated NET formation increases endothelial permeability and pulmonary dysfunction and, therefore, that inhibition of PAD can mitigate damage and improve survival in lethal endotoxemia. Here, we showed that treatment with YW3-56, a PAD2/PAD4 inhibitor, significantly diminished PAD activation, blocked LPS-induced pulmonary vascular leakage, alleviated acute lung injury, and improved survival in a mouse model of lethal LPS-induced endotoxemia. We found CitH3 in the bloodstream 30 min after intraperitoneal injection of LPS (35 mg/kg) into mice. Additionally, CitH3 production was induced in cultured neutrophils exposed to LPS, and NETs derived from these LPS-treated neutrophils increased the permeability of endothelial cells. However, YW3-56 reduced CitH3 production and NET formation by neutrophils following LPS exposure. Moreover, treatment with YW3-56 decreased the levels of circulating CitH3 and abolished neutrophil activation and NET formation in the lungs of mice with endotoxemia. These data suggest a novel mechanism by which PAD-NET-CitH3 can play a pivotal role in pulmonary vascular dysfunction and the pathogenesis of lethal endotoxemia.

Protective Effect of Tubastatin A in CLP-Induced Lethal Sepsis.

We have found earlier that Tubastatin A (TubA), a selective inhibitor of histone deacetylase 6 (HDAC6), improves survival in a mouse model of lethal cecal ligation and puncture (CLP)-induced sepsis. However, the underlying mechanisms have not been fully established. This study sought to test the hypothesis that TubA could affect both lung and splenic functions. C57BL/6J mice were subjected to CLP, and randomized to receive either TubA (70 mg/kg) dissolved in dimethyl sulfoxide (DMSO), or DMSO alone, 1 h following CLP. Sham animals acted as control. Twenty-four hours later, lung tissue was harvested for pathological examination, and splenic tissue was harvested for bacterial colonization. In a parallel study, the spleen was collected 48 h following CLP, and single cell suspension was prepared. Splenocytes then underwent flow cytometry to analyze the immune cell population. RAW264.7 macrophages were treated with lipopolysaccharide (LPS) with or without the presence of TubA (10 µM) at 37 °C for 3 h to assess the effect on macrophage phagocytosis. We found that acute lung injury secondary to lethal sepsis was attenuated by TubA. Treatment with TubA restored the percentage of B lymphocytes, and significantly increased percentages of innate immune cells and macrophages compared to the vehicle-treated CLP group. Moreover, TubA significantly decreased the bacterial load in the spleen, and improved the phagocytic ability of RAW264.7 murine macrophages in vitro. Such findings may help to explain the beneficial effects of TubA treatment in a model of lethal sepsis, as previously reported.

Complete and Partial Aortic Occlusion for the Treatment of Hemorrhagic Shock in Swine.

Hemorrhage remains the leading cause of preventable deaths in trauma. Endovascular management of non-compressible torso hemorrhage has been at the forefront of trauma care in recent years. Since complete aortic occlusion presents serious concerns, the concept of partial aortic occlusion has gained a growing attention. Here, we present a large animal model of hemorrhagic shock to investigate the effects of a novel partial aortic balloon occlusion catheter and compare it with a catheter that works on the principles of complete aortic occlusion. Swine are anesthetized and instrumented in order to conduct controlled fixed-volume hemorrhage, and hemodynamic and physiological parameters are monitored. Following hemorrhage, aortic balloon occlusion catheters are inserted and inflated in the supraceliac aorta for 60 min, during which the animals receive whole-blood resuscitation as 20% of the total blood volume (TBV). Following balloon deflation, the animals are monitored in a critical care setting for 4 h, during which they receive fluid resuscitation and vasopressors as needed. The partial aortic balloon occlusion demonstrated improved distal mean arterial pressures (MAPs) during the balloon inflation, decreased markers of ischemia, and decreased fluid resuscitation and vasopressor use. As swine physiology and homeostatic responses following hemorrhage have been well-documented and are like those in humans, a swine hemorrhagic shock model can be used to test various treatment strategies. In addition to treating hemorrhage, aortic balloon occlusion catheters have become popular for their role in cardiac arrest, cardiac and vascular surgery, and other high-risk elective surgical procedures.

CitH3: a reliable blood biomarker for diagnosis and treatment of endotoxic shock.

Current biomarkers for sepsis are limited by their non-specificity, short half-life, and insensitive response to therapy. Recently, we have demonstrated that citrullinated histone H3(CitH3) is released into the blood from neutrophil extracellular traps(NETs) in response to severe infection, and CitH3 may be a potential biomarker for sepsis. In the present study, we found that NET components were released in mouse models of both lipopolysaccharide(LPS)-induced shock (LPSS) and hemorrhagic shock (HS). To further quantify CitH3 in the NETs, we established a CitH3 specific enzyme-linked immunosorbent assay. Circulating CitH3 was found to be elevated only in LPSS but not in HS. Importantly, blood CitH3 was detected 30 minutes after LPS insult, and remained elevated for 24 hours (period of the highest mortality). Treatment of endotoxic mice with YW3-56, a peptidylarginine deiminase-2/4 inhibitor, significantly diminished levels of CitH3 in the blood. Interleukin-1ß did not respond to LPS early, and interleukin-1ß and interleukin-6 fluctuated although they responded to treatment. Procalcitonin reacted to LPS insult late. Compared to CitH3, these biomarkers were non-specifically induced in LPSS and HS. Collectively, our results demonstrate that YW3-56 protects animals from LPSS, and CitH3 is a reliable biomarker due to its early appearance, specificity, duration, and response to therapeutic intervention.

Fully integrated multi-optoelectronic synthesizer for THz pumping source in wireless communications with rich backup redundancy and wide tuning range.

We report a monolithic photonic integrated circuit (PIC) for THz communication applications. The PIC generates up to 4 optical frequency lines which can be mixed in a separate device to generate THz radiation, and each of the optical lines can be modulated individually to encode data. Physically, the PIC comprises an array of wavelength tunable distributed feedback lasers each with its own electro-absorption modulator. The lasers are designed with a long cavity to operate with a narrow linewidth, typically <4 MHz. The light from the lasers is coupled via an multimode interference (MMI) coupler into a semiconductor optical amplifier (SOA). By appropriate selection and biasing of pairs of lasers, the optical beat signal can be tuned continuously over the range from 0.254 THz to 2.723 THz. The EAM of each channel enables signal leveling balanced between the lasers and realizing data encoding, currently at data rates up to 6.5 Gb/s. The PIC is fabricated using regrowth-free techniques, making it economic for volume applications, such for use in data centers. The PIC also has a degree of redundancy, making it suitable for applications, such as inter-satellite communications, where high reliability is mandatory.

Protective effect of Cl-amidine against CLP-induced lethal septic shock in mice.

Production of innate and adaptive immune cells from hematopoietic stem cells, and maturation of T lymphocytes are effective immune responses to fight severe microbial infection. In sepsis, this emergency myelopoiesis is damaged, leading to failure of bacterial clearance, and excessive stress-induced steroids cause immature T-lymphocyte apoptosis in thymus. We recently found that Cl-amidine, a peptidylarginine deiminase (PAD) inhibitor, improves survival in a mouse model of cecal ligation and puncture (CLP)-induced septic shock. In the present study we investigated how Cl-amidine promotes survival, focusing on protective effects of Cl-amidine on immune response. We confirmed survival-improving effect of Cl-amidine and are the first to explore the role of Cl-amidine in immune response. CLP caused bone marrow (BM) and thymus atrophy, decreased innate immune cells in BM. CLP increased levels of cytokines (IL-1ß, IL-6, and TNF-α) and bacteria load in blood/liver. In primary splenocyte culture, lipopolysaccharide increased TNF-α production. In contrast, Cl-amidine attenuated these CLP and lipopolysaccharide-induced alterations. Moreover, Cl-amidine increased circulating monocytes. Collectively, our results demonstrate Cl-amidine plays protective roles by significantly decreasing BM and thymus atrophy, restoring innate immune cells in BM, increasing blood monocytes and blood/liver bacteria clearance, and attenuating pro-inflammatory cytokine production in a murine model of lethal sepsis.