Probing mechanical interaction of immune receptors and cytoskeleton by membrane nanotube extraction.

The role of force application in immune cell recognition is now well established, the force being transmitted between the actin cytoskeleton to the anchoring ligands through receptors such as integrins. In this chain, the mechanics of the cytoskeleton to receptor link, though clearly crucial, remains poorly understood. To probe this link, we combine mechanical extraction of membrane tubes from T cells using optical tweezers, and fitting of the resulting force curves with a viscoelastic model taking into account the cell and relevant molecules. We solicit this link using four different antibodies against various membrane bound receptors: antiCD3 to target the T Cell Receptor (TCR) complex, antiCD45 for the long sugar CD45, and two clones of antiCD11 targeting open or closed conformation of LFA1 integrins. Upon disruption of the cytoskeleton, the stiffness of the link changes for two of the receptors, exposing the existence of a receptor to cytoskeleton link-namely TCR-complex and open LFA1, and does not change for the other two where a weaker link was expected. Our integrated approach allows us to probe, for the first time, the mechanics of the intracellular receptor-cytoskeleton link in immune cells.

A dynamic causal modeling of the second outbreak of COVID-19 in Italy.

While the vaccination campaign against COVID-19 is having its positive impact, we retrospectively analyze the causal impact of some decisions made by the Italian government on the second outbreak of the SARS-CoV-2 pandemic in Italy, when no vaccine was available. First, we analyze the causal impact of reopenings after the first lockdown in 2020. In addition, we also analyze the impact of reopening schools in September 2020. Our results provide an unprecedented opportunity to evaluate the causal relationship between the relaxation of restrictions and the transmission in the community of a highly contagious respiratory virus that causes severe illness in the absence of prophylactic vaccination programs. We present a purely data-analytic approach based on a Bayesian methodology and discuss possible interpretations of the results obtained and implications for policy makers.

A text data mining approach to the study of emotions triggered by new advertising formats during the COVID-19 pandemic.

Under the influence of the health emergency triggered by the COVID-19 pandemic, many brands changed their communication strategy and included more or less explicit references to the principles of solidarity and fraternity in their TV commercials to boost the confidence and hope of Italian families during the lockdown. The traditional attitudes of the advertising format, which focused on product characteristics, were relegated to the background in order to reinforce the "brand image" through words, signs, hashtags and music that spread empathetic messages to all those who needed to regain hope and trust in a time of extreme emotional fragility. The objective of this paper is to identify the emotions and brand awareness during the lockdown using text mining techniques by measuring customer sentiment expressed on the Twitter social network. Our proposal starts from an unstructured corpus of 20,982 tweets processed with text data mining techniques to identify patterns and trends in people's posts related to specific hashtags and TV ads produced during the COVID-19 pandemic. The innovations in the brand's advertising among consumers seem to have triggered some sense of appreciation and gratitude, as well as a strong sense of belonging that was not present before, as the TV ads were perceived as a disruptive element in consumers' tweets. Although this effect is clearly documented, in this paper we demonstrate its transitory nature, in the sense that the frequency of occurrence of terms associated with an emotional dimension peaks during the weeks of lockdown, and then gradually decreases.

Keratocytes migrate against flow with a roly-poly-like mechanism.

While cell migration can be directed by various mechanical cues such as force, deformation, stiffness, or flow, the associated mechanisms and functions may remain elusive. Single cell migration against flow, repeatedly reported with leukocytes, is arguably considered as active and mediated by integrin mechanotransduction, or passive and determined by a mechanical bias. Here, we reveal a phenotype of flow mechanotaxis with fish epithelial keratocytes that orient upstream or downstream at shear stresses around tens of dyn cm-2. We show that each cell has an intrinsic orientation that results from the mechanical interaction of flow with its morphology. The bulbous trailing edge of a keratocyte generates a hydrodynamical torque under flow that stabilizes an upstream orientation, just as the heavy lower edge of a roly-poly toy generates a gravitational torque that stabilizes an upright position. In turn, the wide and flat leading edge of keratocytes destabilizes upstream orientation, allowing the existence of two distinct phenotypes. To formalize these observations, we propose a simple mechanical model that considers keratocyte morphology as a hemisphere preceded by a wide thin sheet. Our findings show that this model can recapitulate the phase diagram of single cell orientation under flow without adjustable parameters. From a larger perspective, this passive mechanism of keratocytes flow mechanotaxis implies a potential absence of physiological function and evolution-driven process.

The Prognostic Role of ST2L and sST2 in Patients Who Underwent Carotid Plaque Endarterectomy: A Five-Year Follow-Up Study.

Soluble suppressor of tumorigenicity (sST)-2 plasma concentration is related to atherosclerosis. The aim of this study was to assess the prognostic impact of sST2 and its membrane-associated form (ST2L) in patients with carotid atherosclerotic plaque who underwent endarterectomy (CEA). Eighty-two consecutive patients (age range: 48−86 years) who underwent CEA were enrolled. Anthropometric, clinical, instrumental, and laboratory evaluations were gathered. Thirty-seven (45%) patients were symptomatic of cerebrovascular diseases. Patients underwent a five-year follow-up. Phone calls and the analysis of national and regional databases were performed in order to evaluate the occurrence of the primary outcome (all-cause mortality). The population was divided according to survival status. Statins were administered in 81% and 87.5% of survivors and non-survivors, respectively. sST2 levels were higher in non-survivors than in survivors (117.0 ± 103.9 vs. 38.0 ± 30.0 ng/mL, p < 0.001) and in symptomatic individuals, compared with asymptomatic (80.3 ± 92.1 ng/mL vs. 45.4 ± 41.4 ng/mL, p = 0.02). ROC curve analysis identified sST2 cut-off: >98.44 ng/mL as the best predictor for mortality. At the one-year follow-up, the survival rate decreased up to 20% in patients with sST2 higher than the cut-off value. A multivariate regression analysis revealed that only sST2 (HR: 1.012, 95% CI: 1.008−1.016, p < 0.0001) and triglycerides plasma levels (HR: 1.008, 95% CI: 1.002−1.015, p = 0.0135) remained significantly associated with all-cause mortality. ST2L was not associated with all-cause mortality risk. sST2 may act as an independent prognostic determinant of all-cause mortality and symptomatic cerebrovascular diseases in patients with carotid atherosclerotic plaque who underwent CEA.

Mitochondrial DNA diversity of the Sardinian local cattle stock.

The aim of this research was to characterize the genetic diversity of the Sarda (Sa, n = 131), Sardo Bruna (SB, n = 44) and Sardo Modicana (SM, n = 26) cattle breeds, reared in the island of Sardinia (Italy). A portion of the mitochondrial DNA hypervariable region was sequenced, in order to identify a potential signature of African introgression. The FST coefficients among populations ranged between 0.056 for Sa vs SB and 0.167 for SB vs SM. AMOVA analysis indicated there was a significant differentiation of the three breeds, although most of diversity was gathered at the within-breed level. The Median Joining Network of the Sardinian sequences showed a potential founder effect signature. A MJ network including Sardinian cattle plus African, Italian, Iberian and Asian sequences, revealed the presence of haplogroup T3, already detected in Sa cattle, and the presence of Hg T1 and Hg T1'2'3, in Sa and SB. The presence of a private haplotype belonging to haplogroup T1, which is characteristic of African taurine breeds, may be due to the introgression of Sardinian breeds with African cattle, either directly (most probable source: North African cattle) or indirectly (through a Mediterranean intermediary already introgressed with African blood).

Influence of systemic manifestations of inflammatory bowel diseases on endothelial function and cardiovascular risk.

BACKGROUND: Inflammatory bowel diseases (IBD) may be complicated by extraintestinal manifestations (EIM). Both conditions may be implicated in the overall increase of cardiovascular (CV) risk profile of the patients. The study aimed to assess CV risk in IBD patients with EIMs in relation to the stages of both diseases. METHODS: A total of 70 (38 men, mean age 51.7±12.4 years) patients with IBD and 22 controls (12 men, mean age 49.2±13.6 years) were enrolled. All patients and controls were screened for extraintestinal manifestations and underwent physical and anthropometric examinations, standard laboratory investigations, ultrasound evaluation of carotid arteries and flow-mediated vasodilatation (FMD). Patients were divided into four groups in relation to their active or remission stage of disease: 1) IBD+ EIM+; 2) IBD+ EIM-; 3) IBD- EIM+; and 4) IBD- EIM-. RESULTS: The groups were homogenous according to their clinical characteristics. Patients with both IBD and EIM in active phase showed significantly lower values in FMD than controls (P=0.024). Carotid intima-media thickness values (cIMT) were similar among groups. Patients with active phases of IBD and/or EIM showed statistically significant lower values in FMD measurements (P=0.0008 and P=0.0011, respectively). Multivariate regression did not reveal any independent predictors for FMD values. CONCLUSIONS: The active phase of IBD or EIM or both may promote endothelial dysfunction in patients, thus increasing their CV risk profile. Patients in remission phase showed endothelial function similar at controls.

Human neutrophils swim and phagocytise bacteria.

BACKGROUND INFORMATION: Leukocytes migrate in an amoeboid fashion while patrolling our organism in the search for infection or tissue damage. Their capacity to migrate has been proven integrin independent, however, non-specific adhesion or confinement remain a requisite in current models of cell migration. This idea has been challenged twice within the last decade with human neutrophils and effector T lymphocytes, which were shown to migrate in free suspension, a phenomenon termed swimming. While the relevance of leukocyte swimming in vivo remains under judgment, a growing amount of clinical evidence demonstrates that leukocytes are indeed found in liquid-filled body cavities, occasionally with phagocyted pathogens, such as in the amniotic fluid, the cerebrospinal fluid (CSF), or the eye vitreous and aqueous humor. RESULTS: We studied in vitro swimming of primary human neutrophils in the presence of live bacteria, in 2 and 3 dimensions. We show that swimming neutrophils perform phagocytosis of bacteria in suspension. By micropatterning live bacteria on a substrate with an optical technique, we further prove that they use chemotaxis to swim towards their targets. Moreover, we provide evidence that neutrophil navigation can alternate between adherent and non-adherent modes. CONCLUSIONS: Our results suggest that human neutrophils do not rely on adhesion to carry out their functions, supporting a versatile phagocytic function adaptable to the various environmental conditions encountered in vivo, as already suggested by clinical data. SIGNIFICANCE: We verified a claim stated 10 years ago and never reproduced, on the capacity of human neutrophils to swim and perform swimming chemotaxis. We further extended those results to prove that swimming neutrophils can phagocytise bacteria, disregarding adhesion nor confinement as a requisite for accomplishing their function, which differs with current paradigms of leukocyte migration.

The Diagnosis of Acute Myocarditis in Emergency (DAME) score: improving diagnostics within the emergency department.

PURPOSE: . The final diagnosis of myocarditis is challenging. The aim of our study was to provide the D.A.M.E. (Diagnosis of Acute Myocarditis in Emergency) Score for the fast identification of patients suffering from myocarditis at Emergency Department (ED). METHODS: . This was a multicenter, retrospective study involving three centers. All medical records from January 2010 to December 2014 reporting a final discharge diagnosis of myocarditis were considered. One hundred-four patients (mean age: 40.2±16.5 years) were enrolled. Clinical, biochemical and instrumental data were gathered. Data were analysed by means of logistic regression model and factorial analysis. A validation cohort from a fourth center was enrolled. RESULTS: . The final determinants of the DAME score were six: fever, chest pain, erythrocyte sedimentation rate (ESR) > 20 mm/h, C-reactive protein (hs-CRP) >3 mg/L, troponin serum levels >3 ng/L, and left ventricle ejection fraction < 50%. All of them received a specified score ranging from 0 to 4. A score > 4 was related to 75% probability of myocarditis; a final score ranging between 1 and 4 was related to 57% probability of myocarditis. ROC curve on the validation cohort (289 patients, 27 with myocarditis) demonstrated the best cut-off to be 7: AUC 0.958 (p< 0.001), sensibility: 100%, specificity: 85.11%, PPV: 40.9%, NPV: 100% (LR+: 6.72; LR-: 0.00). Logistic regression analysis revealed Odds Ratio equal to 2.83 (95% CI 1.90 - 4.20, p < 0.0001). CONCLUSIONS: . DAME score can offer a reliable tool in ED setting for the evaluation of patients suffering from suspected myocarditis.

Rate-dependent force-extension models for single-molecule force spectroscopy experiments.

Single-molecule force spectroscopy techniques allow for the measurement of several static and dynamic features of macromolecules of biological origin. In particular, atomic force microscopy, used with a variable pulling rate, provides valuable information on the folding/unfolding dynamics of proteins. We propose here two different models able to describe the out-of-equilibrium statistical mechanics of a chain composed of bistable units. These latter represent the protein domains, which can be either folded or unfolded. Both models are based on the Langevin approach and their implementation allows for investigating the effect of the pulling rate and of the device intrinsic elasticity on the chain unfolding response. The theoretical results (both analytical and numerical) have been compared with experimental data concerning the unfolding of the titin and filamin proteins, eventually obtaining a good agreement over a large range of the pulling rates.

SNARE machinery is optimized for ultrafast fusion.

SNARE proteins zipper to form complexes (SNAREpins) that power vesicle fusion with target membranes in a variety of biological processes. A single SNAREpin takes about 1 s to fuse two bilayers, yet a handful can ensure release of neurotransmitters from synaptic vesicles much faster: in a 10th of a millisecond. We propose that, similar to the case of muscle myosins, the ultrafast fusion results from cooperative action of many SNAREpins. The coupling originates from mechanical interactions induced by confining scaffolds. Each SNAREpin is known to have enough energy to overcome the fusion barrier of 25-[Formula: see text]; however, the fusion barrier only becomes relevant when the SNAREpins are nearly completely zippered, and from this state, each SNAREpin can deliver only a small fraction of this energy as mechanical work. Therefore, they have to act cooperatively, and we show that at least three of them are needed to ensure fusion in less than a millisecond. However, to reach the prefusion state collectively, starting from the experimentally observed half-zippered metastable state, the SNAREpins have to mechanically synchronize, which takes more time as the number of SNAREpins increases. Incorporating this somewhat counterintuitive idea in a simple coarse-grained model results in the prediction that there should be an optimum number of SNAREpins for submillisecond fusion: three to six over a wide range of parameters. Interestingly, in situ cryoelectron microscope tomography has very recently shown that exactly six SNAREpins participate in the fusion of each synaptic vesicle. This number is in the range predicted by our theory.

Surgical and pharmacological reassignment: influence on transsexual cardiovascular risk profile.

BACKGROUND/AIM: To evaluate and stratify early cardiovascular risk of transsexuals who underwent pharmacological and/or surgical gender reassignment. METHODS: Fifty-six transsexuals were divided into two groups: group 1 - underwent gonadectomy (orchiectomy for transwomen and hystero-annessiectomy for transmen); group 2 - hormone replacement therapy alone. All participants underwent carotid artery intima-media thickness (C-IMT) and flow-mediated vasodilation (FMD) of brachial artery evaluations. RESULTS: FMD was lower in patients who had undergone gonadectomy compared with non-surgically treated patients (Group 1: 5.711 vs Group 2: 7.339, P < 0.0001). Mean C-IMT was higher in group 1 than group 2 (group 1: 0.733 vs group 2: 0.582). The duration of hormone therapy correlates positively with mean C-IMT (B = 0.001) and negatively with FMD (%) (B = - 0.007). CONCLUSIONS: Cardiovascular risk, which is expressed in terms of endothelial (FMD) and morphological (C-IMT) dysfunction, increases in subjects undergoing gonadectomy compared with those receiving cross-sex reassignment therapy alone.

Neuro-Psychological Pattern in Patients Suffering from Primitive Dilated Cardiomyopathy with Impairment in Executive Function.

According to the American Heart Association (AHA), primitive dilated cardiomyopathy (PDCM) is a "progressive dilation of the left or both ventricles and a depressed contractility in the absence of abnormal load conditions". It evolves in progressive heart-failure. The term "cardiogenic dementia" expresses the intimate connection between heart diseases and cognitive functions. The association between PDCM and the neuropsychological functions is unclear: the main pathophysiological hypotheses are cerebral hypoperfusion and cardiogenic emboli. The aim of this study is to evaluate the impact that the PDCM has on neuropsychological decline and to detect early echocardiographic markers of cognitive impairment. We enrolled 235 patients: 168 suffering from PDCM as sample group and 67 suffering from hypertensive dilated cardiomyopathy (HTCM) as control group. They underwent a cardiology examination and a neuropsychological assessment. A p<0.05 was considered significant. The two groups showed no differences in risk factors, demographic and cardiovascular parameters (except for dimensions of aortic root, left atrium and ventricle which appeared greater in PDCM and left ventricle ejection fraction that appeared lower in PDCM). Among administered neuropsychological tests, only the Stroop Test (which explores executive and attentive functions) appeared significantly lower in PDCM (p = 0.029). Moreover left ventricle end-diastolic diameter was inversely related to the Stroop Test Score (r= -0.32). PDCM doesn't appear to be at the basis of a generalized cognitive and neuropsychological decline. Only the executive functions seem impaired in PDCM. Left ventricle dilation seems to be associated to attentive and executive functions decline.

ST2L Transmembrane Receptor Expression: An Immunochemical Study on Endarterectomy Samples.

BACKGROUND: ST2 (suppression of tumorigenity) has been described as a receptor for the interleukin-33, a member of the IL-1 family of cytokines. It is associated to coronary artery disease, all-causes mortality and cardiovascular mortality. AIMS: The present study was designed to assess the immunohistochemical expression of the ST2 receptor (ST2L/Il-1R) in atherosclerotic plaques of formalin fixed paraffin-embedded internal carotid arteries of patients with and without cerebro-vascular symptoms. METHODS AND RESULTS: The study involved 41 cases (23 asymptomatic and 18 symptomatic). All the clinical and morphological parameters examined were uniformly distributed between the two groups, with a mild predominance of degree of calcification in asymptomatic cases (p = 0.01). ST2L expression was found to be more evident as a membrane pattern in macrophages when observing carotid atherosclerotic plaques of symptomatic patients, rather than in asymptomatic patients' plaques (77.7% vs 39.1%; p = 0.015), and its expression was particularly remarkable in VI type plaque (AHA). Significantly, ST2L was marked by the endothelium of neoangiogenetic vessels on the shoulder region of the plaque, but not (apart from a few cases) in the endothelium covering the residual lumen of the vessel. CONCLUSIONS: The ST2L immunohistochemical expression was for the first time investigated in a large number of human carotid atherosclerotic plaques, as for its pattern of distribution in the different plaque cell populations. Furthermore, ST2L was particularly remarkable on macrophages, as a membrane pattern, of symptomatic patients' plaque. Considering our data, we hypothesize that ST2L/IL33 axis could drive the mechanism of plaque development and eventually rupture.

Perioperative hemodynamic goal-directed therapy and mortality: a systematic review and meta-analysis with meta-regression.

INTRODUCTION: Recent data found that perioperative goal directed therapy (GDT) was effective only in higher control mortality rates (>20%) with a relatively high heterogeneity that limited the strength of evidence. The aim of the present meta-analysis was to clearly understand which high risk patients may benefit of GDT. EVIDENCE ACQUISITION: Systematic review of randomized controlled trials with meta-analyses, including a meta-regression technique. MEDLINE, EMBASE, and The Cochrane Library databases were searched (1980-January 2015). Trials enrolling adult surgical patients and comparing the effects of GDT versus standard hemodynamic therapy were considered. The primary outcome measure was mortality. Data synthesis was obtained by using Odds Ratio (OR) with 95% confidence interval (CI) by random-effects model. EVIDENCE SYNTHESIS: Fifty eight studies met the inclusion criteria (8171 participants). Pooled OR for mortality was 0.70 (95% CI 0.56-0.88, P=0.002, no statistical heterogeneity). GDT significantly reduced mortality when it is >10% in control group (OR 0.43, 95% CI 0.30-0.61, P<0.00001). The meta-regression model showed that the cut off of 10% of mortality rate in control group significantly differentiates 43 studies from the other 15, with a regression coefficient b of -0.033 and a P value of 0.0001. The significant effect of GDT was driven by high risk of bias studies (OR 0.48, 95% CI 0.34-0.67, P<0.0001). CONCLUSIONS: The present meta-analysis, adopting the meta-regression technique, suggests that GDT significantly reduces mortality even when the event control rate is >10%.

Real-time mechanical characterization of DNA degradation under therapeutic X-rays and its theoretical modeling.

The killing of tumor cells by ionizing radiation beams in cancer radiotherapy is currently based on a rather empirical understanding of the basic mechanisms and effectiveness of DNA damage by radiation. By contrast, the mechanical behaviour of DNA encompassing sequence sensitivity and elastic transitions to plastic responses is much better understood. A novel approach is proposed here based on a micromechanical Silicon Nanotweezers device. This instrument allows the detailed biomechanical characterization of a DNA bundle exposed to an ionizing radiation beam delivered here by a therapeutic linear particle accelerator (LINAC). The micromechanical device endures the harsh environment of radiation beams and still retains molecular-level detection accuracy. In this study, the first real-time observation of DNA damage by ionizing radiation is demonstrated. The DNA bundle degradation is detected by the micromechanical device as a reduction of the bundle stiffness, and a theoretical model provides an interpretation of the results. These first real-time observations pave the way for both fundamental and clinical studies of DNA degradation mechanisms under ionizing radiation for improved tumor treatment.

Stochastic mechanical degradation of multi-cracked fiber bundles with elastic and viscous interactions.

The mechanics of fiber bundles has been largely investigated in order to understand their complex failure modes. Under a mechanical load, the fibers fail progressively while the load is redistributed among the unbroken fibers. The classical fiber bundle model captures the most important features of this rupture process. On the other hand, the homogenization techniques are able to evaluate the stiffness degradation of bulk solids with a given population of cracks. However, these approaches are inadequate to determine the effective response of a degraded bundle where breaks are induced by non-mechanical actions. Here, we propose a method to analyze the behavior of a fiber bundle, undergoing a random distribution of breaks, by considering the intrinsic response of the fibers and the visco-elastic interactions among them. We obtain analytical solutions for simple configurations, while the most general cases are studied by Monte Carlo simulations. We find that the degradation of the effective bundle stiffness can be described by two scaling regimes: a first exponential regime for a low density of breaks, followed by a power-law regime at increasingly higher break density. For both regimes, we find analytical effective expressions described by specific scaling exponents.

Scaling shift in multicracked fiber bundles.

Bundles of fibers, wires, or filaments are ubiquitous structures in both natural and artificial materials. We investigate the bundle degradation induced by an external damaging action through a theoretical model describing an assembly of parallel fibers, progressively damaged by a random population of cracks. Fibers in our model interact by means of a lateral linear coupling, thus retaining structural integrity even after substantial damage. Monte Carlo simulations of the Young's modulus degradation for increasing crack density demonstrate a remarkable scaling shift between an exponential and a power-law regime. Analytical solutions of the model confirm this behavior, and provide a thorough understanding of the underlying physics.