Inositol hexakisphosphate kinase 1 regulates neutrophil function in innate immunity by inhibiting phosphatidylinositol-(3,4,5)-trisphosphate signaling.

Inositol phosphates are widely produced throughout animal and plant tissues. Diphosphoinositol pentakisphosphate (InsP7) contains an energetic pyrophosphate bond. Here we demonstrate that disruption of inositol hexakisphosphate kinase 1 (InsP6K1), one of the three mammalian inositol hexakisphosphate kinases (InsP6Ks) that convert inositol hexakisphosphate (InsP6) to InsP7, conferred enhanced phosphatidylinositol-(3,4,5)-trisphosphate (PtdIns(3,4,5)P3)-mediated membrane translocation of the pleckstrin homology domain of the kinase Akt and thus augmented downstream PtdIns(3,4,5)P3 signaling in mouse neutrophils. Consequently, these neutrophils had greater phagocytic and bactericidal ability and amplified NADPH oxidase-mediated production of superoxide. These phenotypes were replicated in human primary neutrophils with pharmacologically inhibited InsP6Ks. In contrast, an increase in intracellular InsP7 blocked chemoattractant-elicited translocation of the pleckstrin homology domain to the membrane and substantially suppressed PtdIns(3,4,5)P3-mediated cellular events in neutrophils. Our findings establish a role for InsP7 in signal transduction and provide a mechanism for modulating PtdIns(3,4,5)P3 signaling in neutrophils.

Reactive oxygen species-induced actin glutathionylation controls actin dynamics in neutrophils.

The regulation of actin dynamics is pivotal for cellular processes such as cell adhesion, migration, and phagocytosis and thus is crucial for neutrophils to fulfill their roles in innate immunity. Many factors have been implicated in signal-induced actin polymerization, but the essential nature of the potential negative modulators are still poorly understood. Here we report that NADPH oxidase-dependent physiologically generated reactive oxygen species (ROS) negatively regulate actin polymerization in stimulated neutrophils via driving reversible actin glutathionylation. Disruption of glutaredoxin 1 (Grx1), an enzyme that catalyzes actin deglutathionylation, increased actin glutathionylation, attenuated actin polymerization, and consequently impaired neutrophil polarization, chemotaxis, adhesion, and phagocytosis. Consistently, Grx1-deficient murine neutrophils showed impaired in vivo recruitment to sites of inflammation and reduced bactericidal capability. Together, these results present a physiological role for glutaredoxin and ROS- induced reversible actin glutathionylation in regulation of actin dynamics in neutrophils.

[Valve-sparing Root Reimplantation for Stanford Type A Acute Aortic Dissection Combined with Aortic Root Dilation and Bicuspid Aortic Valve;Report of a Case].

A 45-year-old male developed Stanford type A acute aortic dissection combined with aortic root dilation and congenital bicuspid aortic valve (BAV). He had a Sieveres type 0 BAV, lateral subtype with right and left cusps. Valve-sparing root reimplantation was performed with decalcification of the cusps. Transthoracic echocardiography(TTE) at discharge revealed no aortic regurgitation, and peak velocity of BAV was 2.15 m/second, mean pressure gradient was 9.6 mmHg and aortic valve area was 2.15 cm2. TTE after 6 months revealed only slight elevation of the peak velocity to 2.78 m/second. To perform successful reimplantation in the case of BAV, anatomic orientation of the cusps should be approximately at 180° and the tissue of the cusps should either be normal or have only minor abnormalities. Valve-sparing root reimplantation for BAV needs a careful follow-up for progression of the aortic valve dysfunction.

The Role of BAFF System Molecules in Host Response to Pathogens.

The two ligands B cell-activating factor of the tumor necrosis factor family (BAFF) and a proliferation-inducing ligand (APRIL) and the three receptors BAFF receptor (BAFF-R), transmembrane activator and calcium-modulating cyclophilin ligand interactor (TACI), and B cell maturation antigen (BCMA) are members of the "BAFF system molecules." BAFF system molecules are primarily involved in B cell homeostasis. The relevance of BAFF system molecules in host responses to microbial assaults has been investigated in clinical studies and in mice deficient for each of these molecules. Many microbial products modulate the expression of these molecules. Data from clinical studies suggest a correlation between increased expression levels of BAFF system molecules and elevated B cell responses. Depending on the pathogen, heightened B cell responses may strengthen the host response or promote susceptibility. Whereas pathogen-mediated increases in the expression levels of the ligands and/or the receptors appear to promote microbial clearance, certain pathogens have evolved to ablate B cell responses by suppressing the expression of TACI and/or BAFF-R on B cells. Other than its well-established role in B cell responses, the TACI-mediated activation of macrophages is also implicated in resistance to intracellular pathogens. An improved understanding of the role that BAFF system molecules play in infection may assist in devising novel strategies for vaccine development.

[Valve Replacement for Infective Endocarditis of Pulmonary Valve;Report of a Case].

Generally, infective endocarditis is found at the left side of the heart. The right side infective endocaritis accounts for only 5~10% of all cases of infective endocarditis. The right side infective endocarditis occurs especially among drug users employing intravenous injection. A typical site of infection is the tricupid valve, and isolated pulmonary valve involvement is rare. It is assumed that its rarity is due to the low pressure gradients within the right heart, the low prevalence of valve disease, and the lower oxygen content of the venous blood. We describe a case of isolated pulmonary valve endocarditis requiring valve replacement. Antibiotic therapy was conducted for 4 weeks before surgery and 2 weeks after surgery. Clinical course was favorable and the patent was discharged home 18 days after surgery.

[Quadricuspid Aortic Valve Repair].

A 58-year-old man presented with severe aortic regurgitation(AR) with quadricuspid valve. Intraoperatively, the small accessory cusp was separated from non-coronary cusp( NCC). The NCC and small accessory cusp were sutured to obtain 1 competent cusp, aiming at an effective height of 8 mm. His AR was trivial at the postoperative 7th day.

De novo chemoattractants form supramolecular hydrogels for immunomodulating neutrophils in vivo.

Most immunomodulatory materials (e.g., vaccine adjuvants such as alum) modulate adaptive immunity, and yet little effort has focused on developing materials to regulate innate immunity, which get mentioned only when inflammation affects the biocompatibility of biomaterials. Traditionally considered as short-lived effector cells from innate immunity primarily for the clearance of invading microorganisms without specificity, neutrophils exhibit a key role in launching and shaping the immune response. Here we show that the incorporation of unnatural amino acids into a well-known chemoattractant-N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLF)-offers a facile approach to create a de novo, multifunctional chemoattractant that self-assembles to form supramolecular nanofibrils and hydrogels. This de novo chemoattractant not only exhibits preserved cross-species chemoattractant activity to human and murine neutrophils, but also effectively resists proteolysis. Thus, its hydrogel, in vivo, releases the chemoattractant and attracts neutrophils to the desired location in a sustainable manner. As a novel and general approach to generate a new class of biomaterials for modulating innate immunity, this work offers a prolonged acute inflammation model for developing various new applications.

Effectiveness of intraoperative endoscopic evaluation in aortic valve repair with valve-sparing aortic root replacement: a comparison of short- and mid-term results.

OBJECTIVES: Valve-sparing aortic root replacement requires expertise to predict repair results and prevent secondary aortic clamping for valve repair or replacement secondary to aortic valve insufficiency. Thus, intraoperative evaluation of the aortic valve using diastolic pressure at the aortic root may be helpful. The goal of this retrospective study was to compare the early and mid-term results of aortic valve repair with those of valve-sparing aortic root replacement using intraoperative endoscopic evaluation. METHODS: We included 158 patients who underwent aortic valve repair with valve-sparing aortic root replacement at our hospital between December 2003 and January 2022. The patients were divided into a non-endoscopic evaluation group (group NE, n = 97; mean age 55 years) and an endoscopic evaluation group (group E, n = 61; mean age 51 years). RESULTS: The incidence of a second aortic clamping for aortic valve insufficiency was significantly greater in group NE (17.5%) than in group E (1.6%; P = 0.002). The presence of none or trivial aortic valve insufficiency on transthoracic echocardiography at discharge in group E (87.6%) was significantly lower than in group NE (98.4%; P = 0.017). No significant difference in the cumulative incidence of recurrence of moderate AI (P = 0.47), hospitalization for heart failure (P = 0.84) and reoperation (P = 0.25) between groups NE and E. CONCLUSIONS: Intraoperative endoscopic evaluation during aortic valve repair with valve-sparing aortic root replacement correlated with a lower incidence of second aortic clamping because of aortic valve insufficiency and effective aortic valve insufficiency control.

Sustained antigen delivery improves germinal center reaction and increases antibody responses in neonatal mice.

Neonates and young infants are known to have limited responses to pediatric vaccines due to reduced germinal center formation. Extended vaccine antigen dosing was previously shown to expand germinal center formation and improve humoral responses in adult mice. We report that sustained antigen delivery through sequential dosing overcomes neonatal limitations to form germinal center reactions and improves humoral immunity. Thus, vaccine strategies that extend the release of vaccine antigens may reduce the number of doses, and time needed, to achieve protective immunity in neonates and young infants.

Small-molecule screen identifies reactive oxygen species as key regulators of neutrophil chemotaxis.

Neutrophil chemotaxis plays an essential role in innate immunity, but the underlying cellular mechanism is still not fully characterized. Here, using a small-molecule functional screening, we identified NADPH oxidase-dependent reactive oxygen species as key regulators of neutrophil chemotactic migration. Neutrophils with pharmacologically inhibited oxidase, or isolated from chronic granulomatous disease (CGD) patients and mice, formed more frequent multiple pseudopodia and lost their directionality as they migrated up a chemoattractant concentration gradient. Knocking down NADPH oxidase in differentiated neutrophil-like HL60 cells also led to defective chemotaxis. Consistent with the in vitro results, adoptively transferred CGD murine neutrophils showed impaired in vivo recruitment to sites of inflammation. Together, these results present a physiological role for reactive oxygen species in regulating neutrophil functions and shed light on the pathogenesis of CGD.

B cell receptor-induced IL-10 production from neonatal mouse CD19+CD43- cells depends on STAT5-mediated IL-6 secretion.

Newborns are unable to reach the adult-level humoral immune response partly due to the potent immunoregulatory role of IL-10. Increased IL-10 production by neonatal B cells has been attributed to the larger population of IL-10-producting CD43+ B-1 cells in neonates. Here, we show that neonatal mouse CD43- non-B-1 cells also produce substantial amounts of IL-10 following B cell antigen receptor (BCR) activation. In neonatal mouse CD43- non-B-1 cells, BCR engagement activated STAT5 under the control of phosphorylated forms of signaling molecules Syk, Btk, PKC, FAK, and Rac1. Neonatal STAT5 activation led to IL-6 production, which in turn was responsible for IL-10 production in an autocrine/paracrine fashion through the activation of STAT3. In addition to the increased IL-6 production in response to BCR stimulation, elevated expression of IL-6Rα expression in neonatal B cells rendered them highly susceptible to IL-6-mediated STAT3 phosphorylation and IL-10 production. Finally, IL-10 secreted from neonatal mouse CD43- non-B-1 cells was sufficient to inhibit TNF-α secretion by macrophages. Our results unveil a distinct mechanism of IL-6-dependent IL-10 production in BCR-stimulated neonatal CD19+CD43- B cells.

IL6 suppresses vaccine responses in neonates by enhancing IL2 activity on T follicular helper cells.

The inability of neonates to develop CD4+FoxP3-CXCR5hiPD-1hi T follicular helper (TFH) cells contributes to their weak vaccine responses. In previous studies, we measured diminished IgG responses when IL-6 was co-injected with a pneumococcal conjugate vaccine (PCV) in neonatal mice. This is in sharp contrast to adults, where IL-6 improves vaccine responses by downregulating the expression of IL-2Rß on TFH cells and protecting them from the inhibitory effect of IL-2. In this study, we found that splenic IL-6 levels rapidly increased in both adult and neonatal mice following immunization, but the increase in neonatal mice was significantly more than that of adult mice. Moreover, immunized neonatal TFH cells expressed significantly more IL-2 as well as its receptors, IL-2Rα and IL-2Rß, than the adult cells. Remarkably, IL-6 co-injection with PCV vaccine further increased the production of IL-2 and the expression of its receptors by neonatal TFH cells, whereas excess IL-6 had totally opposite effect in immunized adult mice. Underscoring the role of IL-6 in activating the IL-2 mediated suppression of vaccine responses, immunization of IL-6 knock-out neonates led to improved antibody responses accompanied by expanded TFH cells as well as lower levels of IL-2 and IL-2 receptors on TFH cells. Moreover, CpG containing PCV improved TFH response in neonates by suppressing the expression of IL-2 receptors on TFH cells and inhibiting IL-2 activity. These findings unveil age-specific differences in IL-6 mediated vaccine responses and highlight the need to consider age-related immunobiological attributes in designing vaccines.

Single-stage repair of the right aortic arch with Kommerell diverticulum through the right thoracotomy with SIRC approach.

Kommerell diverticulum (KD) is a congenital vascular disease associated with dilatation at an aberrant subclavian artery's origin. The surgical repair should be considered for the symptomatic patients due to the adjacent organ's compression by the aneurysmal change of KD. An appropriate approach should be selected for the open repair to suit the anatomical type of disease. We reported a 50-year-old female diagnosed with KD undergoing the single-stage open repair through the right thoracotomy with the straight skin incision with rib cross.

[Long-term result of the maze procedure for atrial fibrillation].

Long-term result of the maze operation is unknown. Anticoagulation therapy is controversial even after success of the maze operation. Between 1997 and 2008, 213 patients underwent the maze procedure. Atrial fibrillation (Af) was chronic in 151 patients and paroxysmal in 62. Concomitant mitral valve procedure were performed in 72%. Early mortality was 1.9%. By dismissal electrocardiography 157 patients (74%) wes free from Af. At the last follow-up (mean 38 +/- 30 months), late Af recurrence had occurred in 16 patients. Af recurrence had occurred within 1-year in 15 patients. Risk factors for late Af recurrence was left atrial dimension of 50 mm or greater (P<0.001). There was no difference in 5-year freedom from Af between the Cox maze procedure versions. Anticoagulation for patients with large left atrium is mandatory for 1-year after surgery.

A case of aggressive aortic prosthetic valve endocarditis aggressive caused by Staphylococcus lugdunensis.

BACKGROUND: Staphylococcus lugdunensis is a coagulase-negative Staphylococcus species, which are weak pathogenic bacteria generally. However, the acute and severe pathogenicity of Staphylococcus lugdunensis infective endocarditis may be due to the rapid growth of large vegetation and consequent valve destruction. CASE PRESENTATION: The patient was an 81-year-old male who visited our hospital with chief complaints of low back pain and high fever. Four years before this visit, he had undergone aortic valve replacement for aortic regurgitation. He was found to be hypotensive. Although there is no heart murmur on auscultation and echocardiography revealed negative findings with aortic valve, a blood test showed increases in the white blood cell count and C-reactive protein concentration. On the next day, Gram-positive cocci were detected in a blood culture and echocardiography detected a large vegetation on the prosthetic valve with increased flow velocity. Therefore, he underwent redo aortic valve replacement emergently. Staphylococcus lugdunensis was identified in blood samples and vegetation culture. Consequently, the patient was treated with antibiotics for 5 weeks after the operation and discharged home. CONCLUSIONS: We experienced rapidly progressive prosthetic valve endocarditis caused by Staphylococcus lugdunensis. Hence, Staphylococcus lugdunensis infective endocarditis requires aggressive treatment, and the pathogenicity of this coagulase-negative Staphylococcus with high drug susceptibility should not be underestimated.

Criticality of plasma membrane lipids reflects activation state of macrophage cells.

Signalling is of particular importance in immune cells, and upstream in the signalling pathway many membrane receptors are functional only as complexes, co-locating with particular lipid species. Work over the last 15 years has shown that plasma membrane lipid composition is close to a critical point of phase separation, with evidence that cells adapt their composition in ways that alter the proximity to this thermodynamic point. Macrophage cells are a key component of the innate immune system, are responsive to infections and regulate the local state of inflammation. We investigate changes in the plasma membrane's proximity to the critical point as a response to stimulation by various pro- and anti-inflammatory agents. Pro-inflammatory (interferon γ, Kdo 2-Lipid A, lipopolysaccharide) perturbations induce an increase in the transition temperature of giant plasma membrane vesicles; anti-inflammatory interleukin 4 has the opposite effect. These changes recapitulate complex plasma membrane composition changes, and are consistent with lipid criticality playing a master regulatory role: being closer to critical conditions increases membrane protein activity.

Flow-dependent accumulation of LDL in co-cultures of endothelial and smooth muscle cells in the presence of filtration flow through the cell layer.

To elucidate the roles of blood flow and transmural filtration flow in localized LDL accumulation in vascular walls, we studied the effects of flow velocity on LDL concentration at the cell surface and LDL uptake by co-cultures of vascular endothelial cells (ECs) and smooth muscle cells using a parallel-plate flow cell with or without filtration flow. Co-cultures were prepared on porous membranes. In the presence of filtration flow through the cell layer, the LDL concentration at the cell surface increased when the perfusion velocity was decreased (shear stress was decreased from 1.5 to 0.2 Pa). In the absence of filtration flow, LDL concentration remained unchanged despite changes in flow velocity. LDL uptake by the cells was proportional to its surface concentration that varied inversely with flow velocity. Therefore, in the presence of filtration flow, LDL accumulation was greater under conditions of low shear stress (0.2 Pa) than with high shear stress (1.5 Pa). In contrast, in the absence of filtration flow, LDL uptake was almost proportional to the magnitude of shear stress. These results suggest that shear stress-induced biological responses of ECs and transmural filtration flow, both play important roles in localized LDL accumulation in vascular walls.

IL-6 Impairs Vaccine Responses in Neonatal Mice.

The inability of infants to mount proper follicular helper T (TFH) cell response renders this age group susceptible to infectious diseases. Initial instruction of T cells by antigen presenting cells and subsequent differentiation into TFH cells are controlled by T cell receptor signal strength, co-stimulatory molecules and cytokines such as IL-6 and IL-21. In immunized adults, IL-6 promotes TFH development by increasing the expression of CXCR5 and the TFH master transcription factor, B cell lymphoma 6. Underscoring the importance of IL-6 in TFH generation, we found improved antibody responses accompanied by increased TFH cells and decreased follicular regulatory helper T (TFR) cells, a Foxp3 expressing inhibitory CD4+ T cell occupying the germinal center (GC), when a tetanus toxoid conjugated pneumococcal polysaccharide type 14 vaccine was injected in adult mice together with IL-6. Paradoxically, in neonates IL-6 containing PPS14-TT vaccine suppressed the already impaired TFH development and antibody responses in addition to increasing TFR cell population. Supporting the diminished TFH development, we detected lower frequency of phospho-STAT-3+ TFH in immunized neonatal T cells after IL-6 stimulation than adult cells. Moreover, IL-6 induced more phospho-STAT-3+ TFR in neonatal cells than adult cells. We also measured lower expression of IL-6R on TFH cells and higher expression on TFR cells in neonatal cells than adult cells, a possible explanation for the difference in IL-6 induced signaling in different age groups. Supporting the flow cytometry findings, microscopic examination revealed the localization of Treg cells in the splenic interfollicular niches of immunized adult mice compared to splenic follicles in neonatal mice. In addition to the limitations in the formation of IL-21 producing TFH cells, neonatal mice GC B cells also expressed lower levels of IL-21R in comparison to the adult mice cells. These findings point to diminished IL-6 activity on neonatal TFH cells as an underlying mechanism of the increased TFR: TFH ratio in immunized neonatal mice.

Activation of Toll-like receptors nucleates assembly of the MyDDosome signaling hub.

Infection and tissue damage induces assembly of supramolecular organizing centres (SMOCs)), such as the Toll-like receptor (TLR) MyDDosome, to co-ordinate inflammatory signaling. SMOC assembly is thought to drive digital all-or-none responses, yet TLR activation by diverse microbes induces anything from mild to severe inflammation. Using single-molecule imaging of TLR4-MyDDosome signaling in living macrophages, we find that MyDDosomes assemble within minutes of TLR4 stimulation. TLR4/MD2 activation leads only to formation of TLR4/MD2 heterotetramers, but not oligomers, suggesting a stoichiometric mismatch between activated receptors and MyDDosomes. The strength of TLR4 signalling depends not only on the number and size of MyDDosomes formed but also how quickly these structures assemble. Activated TLR4, therefore, acts transiently nucleating assembly of MyDDosomes, a process that is uncoupled from receptor activation. These data explain how the oncogenic mutation of MyD88 (L265P) assembles MyDDosomes in the absence of receptor activation to cause constitutive activation of pro-survival NF-κB signalling.

Improved arterial wall model by coculturing vascular endothelial and smooth muscle cells.

We have constructed an in vitro arterial wall model by coculturing bovine arterial endothelial cells (ECs) and smooth muscle cells (SMCs). When ECs were seeded directly over SMCs and cocultured in an ordinary culture medium, ECs grew sparsely and did not form a confluent monolayer. Addition of ascorbic acid to the culture medium at concentrations greater than 50 mug/ml increased the production of type IV collagen by the SMCs, and ECs formed a confluent monolayer covering the entire surface of SMCs. Histological studies showed that the thickness of the cell layer composed of ECs and SMCs increased with increasing duration of coculture. This arterial wall model, prepared by our method, may serve as a simple and good in vitro model to study the effects of factors such as biological chemicals and shear stress on cell proliferation and other physiological functions of arterial walls.