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Phagocytosis is a complex process that eliminates microbes and is performed by specialised cells such as macrophages. Toll-like receptor 4 (TLR4) is expressed on the surface of macrophages and recognizes Gram-negative bacteria. Moreover, TLR4 has been suggested to play a role in the phagocytosis of Gram-negative bacteria, but the mechanisms remain unclear. Here we have used primary human macrophages and engineered THP-1 monocytes to show that the TLR4 sorting adapter, TRAM, is instrumental for phagocytosis of Escherichia coli as well as Staphylococcus aureus. We find that TRAM forms a complex with Rab11 family interacting protein 2 (FIP2) that is recruited to the phagocytic cups of E. coli. This promotes activation of the actin-regulatory GTPases Rac1 and Cdc42. Our results show that FIP2 guided TRAM recruitment orchestrates actin remodelling and IRF3 activation, two events that are both required for phagocytosis of Gram-negative bacteria.

Abstract Introduction The treatment of moderate functionalmitral regurgitation (FMR) during coronary artery bypass grafting (CABG) is still debated. Our primary end point was to assess the improvement of “mitral valve reserve” (MVR) after CABG alone as a clinical demonstration of left ventricular (LV) recovery. Materials and methods Between June 2019 and June 2021, we prospectively enrolled 104 consecutive patients undergoing CABG with moderate FMR. Inclusion criteria were inferior-posterior-lateral wall hypokinesia and revascularization of the circumflex or right coronary artery. MVR was calculated as the ratio between anterior and posterior leaflets’ straight length. All patients were followed for 1 year. The improvement of MVR has been considered as a reduction of the ratio between anterior and posterior leaflets straight length. Results Compared to baseline, mean MVR was significantly reduced both at 6 (2.24 ± 0.95 vs. 1,91 ± 0.6; p = 0,047) and 12 months follow-up (2.24 ± 0.95 vs. 1,69 ± 0.49; p = 0,006). Left ventricular (LV) reverse remodeling, meant as improvement of LV ejection fraction and reduction of LV end-systolic volume index and mitral anulus diameter were evaluated at 6 months and 1 year. Mitral regurgitation grade were also significantly reduced at 6 months (p

Complications after device closure of ostium secundum defects are rare but possible. We present a very late erosion of the interatrial septum after a percutaneous closure of an ostium secundum defect. Identification of early clinical and imaging clues associated with this condition is fundamental for a timely diagnosis and treatment. (Level of Difficulty: Intermediate.)

Background and Objectives: Several studies revealed a relation between abnormal cardiac remodeling and glomerular filtration rate (GFR) decline, but there are limited data regarding echocardiographic changes in chronic kidney disease (CKD). This study evaluated the abnormal cardiac structures characterizing patients with CKD, assessing the independent association between echocardiographic parameters and the risk of decline in renal function. Materials and Methods: In total, 160 patients with CKD were studied. All patients underwent an echocardiographic exam and 99mTc-DTPA renal scintigraphy to measure the GFR. After the baseline assessments, patients were followed prospectively for 12 months, or until the endpoint achievement, defined as a worsening in renal function (doubling of baseline serum creatinine, GFR decline ≥25%, the start of dialysis). Results: Patients with GFR values of 34.8 ± 15 mL/min, identifying stages III–IV of CKD, were associated with high levels of left ventricular mass index (LVMi) (101.9 ± 12.2 g/m2), which was related to proteinuria, systolic blood pressure, and pulmonary artery systolic pressure in a multiple regression model. During the observational period, 26% of patients reached the endpoint. Regression analysis revealed LVMi as a predictor of change in renal function after adjusting for kidney and cardiac risk factors. Multiple Cox regression indicated that an increase in LVMi was associated with a 12% increased risk of kidney disease progression (HR: 1.12; 95% CI: 1.04–1.16; p = 0.001). Conclusions: In patients with CKD, high LVMi represents an independent predictor of the progressive decline of the renal function, until the start of renal replacement therapy. Echocardiography can help identify patients at high risk for renal disease worsening in patients with CKD independently of clinical cardiac involvement.

Background and Objectives: Minimally invasive cardiac surgery (MICS) has been developing since 1996. Peripheral cannulation is required to perform MICS, and good venous drainage and a bloodless field are crucial for the success of this procedure. We assessed the benefits of using a virtually wall-less cannula in comparison with the standard thin-wall cannula in clinical practice. Materials and Methods: Between January 2021 and December 2022, we evaluated 65 elective patients, who underwent isolated minimally invasive mitral valve surgery. Both the virtually wall-less and the thin-wall cannulas were placed through a surgical cut-down. Patients’ characteristics at baseline were similar in the two groups, except for the body surface area (BSA), which was greater in the virtually wall-less group compared to the thin-wall one. In the standard group, the size of the cannula was chosen depending on the patient’s BSA, and the choice of the Smartcannula was based on their height. Results: There were no significant differences between the two groups in terms of negative pressure applied, target flow achieved, hemolysis, the need for blood transfusion, and the post-operative increases in liver and renal enzymes. However, in all the patients, the estimated target flow was achieved, thereby showing the better hemodynamic performance of the virtually wall-less cannula, since, in this group, the patients’ BSA was significantly greater compared to the thin-wall group. Ultimately, the mean cross-clamp time, as an indirect index of the effectiveness of the venous drainage, is shorter in the virtually wall-less group compared with the thin-wall group. Conclusions: The virtually wall-less cannula should be preferred in minimally invasive mitral valve surgery due to its superior performance in terms of venous drainage compared with the standard thin-wall cannula.

Intraoperative assessment of graft patency is pivotal for successful coronary revascularization. In the present study we aimed to propose a new, easy to perform tool to assess anastomotic quality intraoperatively, and to investigate its potential reliability in predicting early graft failure. Intraoperative graft flowmetry of 63 consecutive patients undergoing CABG were prospectively collected. Transit time flowmetry and its derivatives were recorded. Coronary resistances were calculated according to Hagen–Poiseuille equation both during cardioplegic arrest and after withdrawal from cardiopulmonary bypass. Angiographic evidence of graft occlusion at follow-up was cross-checked with intraoperative recordings. After a mean follow-up of 10.4 ± 6.0 months, 22 grafts were studied, and occlusion was documented in five (22.7%). Occluded grafts showed lower flows and higher resistances recorded during aortic cross-clamping. Coronary resistances, recorded during aortic cross-clamping, greater than 2.0 mmHg/mL/min, showed a sensitivity of 80% and a specificity of 100% to predict graft failure. We propose the routine recording of coronary resistances during aortic cross-clamping as an additional tool to overcome the acknowledged limitation of TTF to predict graft occlusion at 1 year.

We explore here the potential of a newly described technology, which is named PROFILER and is based on next generation sequencing of gene-specific lambda phage-displayed libraries, to rapidly and accurately map monoclonal antibody (mAb) epitopes. For this purpose, we used a novel mAb (designated 31E10/E7) directed against Neisserial Heparin-Binding Antigen (NHBA), a component of the anti-group B meningococcus Bexsero® vaccine. An NHBA phage-displayed library was affinity-selected with mAb 31E10/E7, followed by massive sequencing of the inserts present in antibody-selected phage pools. Insert analysis identified an amino acid stretch (D91-A128) in the N-terminal domain, which was shared by all of the mAb-enriched fragments. Moreover, a recombinant fragment encompassing this sequence could recapitulate the immunoreactivity of the entire NHBA molecule against mAb 31E10/E7. These results were confirmed using a panel of overlapping recombinant fragments derived from the NHBA vaccine variant and a set of chemically synthetized peptides covering the 10 most frequent antigenic variants. Furthermore, hydrogen-deuterium exchange mass-spectrometry analysis of the NHBA-mAb 31E10/E7 complex was also compatible with mapping of the epitope to the D91-A128 region. Collectively, these results indicate that the PROFILER technology can reliably identify epitope-containing antigenic fragments and requires considerably less work, time and reagents than other epitope mapping methods.

Previous studies have shown that the pro-inflammatory cytokine IL-1β has a crucial role in host defenses against group B streptococcus (GBS), a frequent human pathogen, by recruiting neutrophils to infection sites. We examined here the cell types and mechanisms involved in IL-1β production during infection. Using a GBS-induced peritonitis model in mice, we first found that a large proportion of exudate cells contain intracellular IL-1β by immunofluorescence. Of the IL-1β positive cells, 82 and 7% were neutrophils and macrophages, respectively, suggesting that the former cell type might significantly contribute to IL-1β production. Accordingly, depletion of neutrophils with anti-Ly6G antibodies resulted in a significant reduction in the levels of IL-1β, but not of TNF-α or IL-6. We next found that neutrophils are capable of releasing mature IL-1β and TNF-α directly in response to in vitro stimulation with GBS. The production of pro-IL-1β and TNF-α in these cells required the Toll-like receptor (TLR) adaptor MyD88 and the chaperone protein UNC93B1, which is involved in mobilization of a subfamily of TLRs to the endosomes. Moreover, pro-IL-1β processing and IL-1β release was triggered by GBS hemolysin and required components of the canonical inflammasome, including caspase-1, ASC and NLRP3. Collectively our findings indicate that neutrophils make a significant contribution to IL-1β production during GBS infection, thereby amplifying their own recruitment. These cells directly recognize GBS by means of endosomal TLRs and cytosolic sensors, leading to activation of the caspase-1 inflammasome.