A Multiscale In Silico Investigation of the Mechanics and Dynamics of Ex Vivo Coagulation

Theories describing the coagulation cascade have been around for decades and have greatly expanded in functional detail over the past several years. However, there still exists a void in the literature on the quantification of the microscale contribution of individual blood cell mechanics on the macroscale behavior of blood clots. This is due, in part, to the fact that the trans-scale relationships between blood components are not fully understood. In this work, we aim to bridge the gap between the known cell-scale phenomena of coagulation, specifically platelet and fibrin interactions, and the measurable mechanics and dynamics of whole blood clots. The developed multiscale model consists of two main components: (1) a phenomenological model of activated platelet adhesion and contraction and (2) a mechanistic model of fibrin viscoelasticity and strain-hardening extensibility. The components of the multiscale model include stand-alone discrete element method (DEM)-based cell-scale models for the primary components of blood. The unique mechanical and dynamical behaviors observed experimentally in single-platelet and single-fiber studies can be captured by this technique due to the inclusion of phenomenological force models, namely piecewise linear functions for the adhesion exhibited by activated platelets and Hill functions for the nonlinear elastic modulus of fibrin. The isolated platelet adhesion and fibrin extension models were developed and calibrated separately before they were combined to study the emergent behavior of platelet and fibrin assemblies. The platelet and fibrin compositions were varied between simulations to assess the morphological and mechanistic differences of in silico-formed aggregates. Applying the model within a dynamic framework was also used to obtain a macroscale metric of in silico aggregate behavior that is comparable to one from existing clinical whole-blood diagnostic devices like the thromboelastogram, or TEG. Specifically, we can quantify the platelet contribution to the strength of platelet and fibrin in silico aggregates. We observed a nonlinear relationship between platelet concentration and platelet contribution that corroborates experimental studies. The culmination of the modeling efforts from this dissertation is a tool that can be used and expanded to better understand the mechanistic detail of platelet and fibrin contributions during coagulation

Volume regulation by Amphiuma red blood cells. The membrane potential and its implications regarding the nature of the ion-flux pathways

After osmotic perturbation, the red blood cells of Amphiuma exhibited a volume-regulatory response that returned cell volume back to or toward control values. After osmotic swelling, cell-volume regulation (regulatory volume decrease; RVD) resulted from net cellular loss of K, Cl, and osmotically obliged H2O. In contrast, the volume-regulatory response to osmotic shrinkage (regulatory volume increase; RVI) was characterized by net cellular uptake of Na, Cl, and H2O. The net K and Na fluxes characteristic of RVD and RVI are increased by 1-2 orders of magnitude above those observed in studies of volume-static control cells. The cell membrane potential of volume-regulating and volume-static cells was measured by impalement with glass microelectrodes. The information gained from the electrical and ion-flux studies led to the conclusion that the ion fluxes responsible for cell-volume regulation proceed via electrically silent pathways. Furthermore, it was observed that Na fluxes during RVI were profoundly sensitive to medium [HCO3] and that during RVI the medium becomes more acid, whereas alkaline shifts in the suspension medium accompany RVD. The experimental observations are explained by a model featuring obligatorily coupled alkali metal-H and Cl-HCO3 exchangers. The anion- and cation-exchange pathways are separate and distinct yet functionally coupled via the net flux of H. As a result of the operation of such pathways, net alkali metal, Cl, and H2O fluxes proceed in the same direction, whereas H and HCO3 fluxes are cyclic. Data also are presented that suggest that the ion-flux pathways responsible for cell-volume regulation are not activated by changes in cell volume per se but by some event associated with osmotic perturbation, such as changes in intracellular pH

Cluster of legionnaires’ disease in an Italian prison

Background: Legionella pneumophila (Lp) is the most common etiologic agent causing Legionnaires’ Disease (LD). Water systems offer the best growth conditions for Lp and support its spread by producing aerosols. From 2015 to 2017, the Regional Reference Laboratory of Clinical and Environmental Surveillance of Legionellosis of Palermo monitored the presence of Lp in nine prisons in Western Sicily. During this investigation, we compared Lp isolates from environmental samples in a prison located in Palermo with isolates from two prisoners in the same prison. Methods: We collected 93 water samples from nine Sicilian prisons and the bronchoalveolar lavages (BALs) of two prisoners considered cases of LD. These samples were processed following the procedures described in the Italian Guidelines for the Prevention and Control of Legionellosis of 2015. Then, genotyping was performed on 19 Lp colonies (17 from water samples and 2 from clinical samples) using the Sequence-Based Typing (SBT) method, according to European Study Group for Legionella Infections (ESGLI) protocols. Results: Lp serogroup (sg) 6 was the most prevalent serogroup isolated from the prisons analyzed (40%), followed by Lp sg 1 (16%). Most of all, in four penitentiary institutions, we detected a high concentration of Lp >104 Colony Forming Unit/Liter (CFU/L). The environmental molecular investigation found the following Sequence Types (STs) in Lp sg 6: ST 93, ST 292, ST 461, ST 728, ST 1317 and ST 1362, while most of the isolates in sg 1 belonged to ST 1. We also found a new ST that has since been assigned the number 2451 in the ESGLI-SBT database. From the several Lp sg 1 colonies isolated from the two BALs, we identified ST 2451. Conclusions: In this article, we described the results obtained from environmental and epidemiological investigations of Lp isolated from prisons in Western Sicily. Furthermore, we reported the first cluster of Legionnaires’ in an Italian prison and the molecular typing of Lp sg 1 from one prison’s water system and two BALs, identified the source of the contamination, and discovered a new ST

Transitions of protein traffic from cardiac ER to junctional SR

The junctional sarcoplasmic reticulum (jSR) is an important and unique ER subdomain in the adult myocyte that concentrates resident proteins to regulate Ca(2+) release. To investigate cellular mechanisms for sorting and trafficking proteins to jSR, we overexpressed canine forms of junctin (JCT) or triadin (TRD) in adult rat cardiomyocytes. Protein accumulation over time was visualized by confocal fluorescence microscopy using species-specific antibodies. Newly synthesized JCTdog and TRDdog appeared by 12-24h as bright fluorescent puncta close to the nuclear surface, decreasing in intensity with increasing radial distance. With increasing time (24-48h), fluorescent puncta appeared at further radial distances from the nuclear surface, eventually populating jSR similar to steady-state patterns. CSQ2-DsRed, a form of CSQ that polymerizes ectopically in rough ER, prevented anterograde traffic of newly made TRDdog and JCTdog, demonstrating common pathways of intracellular trafficking as well as in situ binding to CSQ2 in juxtanuclear rough ER. Reversal of CSQ-DsRed interactions occurred when a form of TRDdog was used in which CSQ2-binding sites are removed ((del)TRD). With increasing levels of expression, CSQ2-DsRed revealed a novel smooth ER network that surrounds nuclei and connects the nuclear axis. TRDdog was retained in smooth ER by binding to CSQ2-DsRed, but escaped to populate jSR puncta. TRDdog and (del)TRD were therefore able to elucidate areas of ER-SR transition. High levels of CSQ2-DsRed in the ER led to loss of jSR puncta labeling, suggesting a plasticity of ER-SR transition sites. We propose a model of ER and SR protein traffic along microtubules, with prominent transverse/radial ER trafficking of JCT and TRD along Z-lines to populate jSR, and an abundant longitudinal/axial smooth ER between and encircling myonuclei, from which jSR proteins traffic

Assessment of the CPL-0015 isolate as a vaccine strain for the control of canine parvovirus in Cuba

The safety and protective efficacy of the CPL-0015 Cuban isolate of canine parvovirus type 2 (CPV-2) were evaluated for its possible use as a vaccine strain candidate. The study included a total of 23 healthy Beagle dogs of both sexes, aged 84 days and without specific maternal antibodies against canine parvovirus. Safety was analysed by comparing clinicopathological values, food consumption, body weight, rectal temperature and white blood cell counts for 14 consecutive days between control dogs (n=5) and dogs subcutaneously injected (n=10) with 2 mL (equivalent to two doses) of the CPL-0015 strain with an antigenic titer of 106.0 infectious dose50 in cell culture/mL. The protective effectiveness was determined by measuring and comparing anti-CPV-2 IgG levels and clinical signs during 56 experimental days between control dogs (n=2) and dogs inoculated (n=6) with double doses of 1 mL each, separated by a 21-day interval. All animals were challenged orally on day 35 with the virulent strain Cornell-780916 (105.0 infective dose50 in cell culture/mL). The results showed that the CPL-0015 strain did not negatively impact the physiological condition of the exposed animals. The inoculated and challenged animals showed not only significantly increased levels of anti-CPV-2 IgG (P<0.05) when compared to days 0, 35, and the control group animals but also had 100% survival without clinical signs of the disease, unlike the control group. It is concluded that CPL-0015 is safe and provides effective protection against homologous virulent strains

A NEW CASE OF LOUSE-BORNE RELAPSING FEVER IN SICILY: CASE REPORT AND MINI REVIEW

Body lice transport B. recurrentis from man to man and humans are the only host. The presence of lice in Italy and an increasing number of cases in migrants can contribute to the onset of autochthonous cases. In this paper, we report a new case of Louse-borne Relapsing Fever (LBRF) diagnosed among migrants in Sicily exactly one year after the first case was recorded. We reviewed all cases reported in Europe from February 2016 until now. Our study identified two new cases of LBRF in migrants arrived in Europe: one who came from Somalia and one from Mali. Here we report data on a new case in Sicily. The number of migrants and refugees to transit in Sicily has increased, and this has led to the introduction of infectious diseases. Therefore, in our opinion it is essential to upgrade control of the sanitation conditions of migrants

Trastornos mentales y consumo de drogas en mujeres embarazadas con alto riesgo obstétrico

Introducción: El consumo de sustancias psicoactivas en el embarazo puede desencadenar patologías como cuadro psiquiatrico agudo, síndrome de abstinencia y algunos riesgos relacionados con las conductas asociadas al consumo. Objetivo: Describir la prevalencia de trastornos mentales y consumo de drogas en mujeres embarazadas con alto riesgo obstétrico. Materiales y métodos: Estudio observacional, descriptivo transversal en 220 mujeres embarazadas con alto riesgo obstétrico. Se calcularon prevalencias de trastornos mentales y de consumo de drogas. Resultados: El trastorno mental con mayor prevalencia fue el episodio depresivo mayor (10,0%). Las sustancias con mayor prevalencia de consumo en el último año fueron: alcohol (58,3%), bebidas energéticas (36,4%), cigarrillo (10,9%) y marihuana (6,4%). Conclusiones: Es necesario incluir en los protocolos de atención de la mujer gestante, la exploración de estos aspectos, incluido el tamizaje de drogas en orina, dadas las implicaciones que para la salud pública tiene el consumo de sustancias psicoactivas y la presencia de trastornos mentales durante el embarazo.   &nbsp

Socio-Demographic Characteristics and Sexual Behavioral Factors of Patients with Sexually Transmitted Infections Attending a Hospital in Southern Italy

Sexually transmitted infections (STIs) are a serious global health problem. In Italy, data describing the vulnerability to STIs of specific sexual minorities and the influence of sociodemographic and behavioral determinants are limited, as most infections are not subject to mandatory notification. This retrospective study describes the sociodemographic profile and main sexual behaviors of patients attending a hospital in Palermo (Sicily, Italy) from January 2018 to March 2019 as predictors of STI risk. Patients were divided in subgroups: men-who-have-sex-with-men (MSM), men-who-have-sex-with-women (MSW), bisexual men and females. Data were obtained through an anonymous questionnaire. Patients were tested for chlamydia, syphilis, Mycoplasma genitalium infection, genital herpes and HPV infection. A total of 294 subjects with STIs (male/female ratio about 2:1) were screened. Of the total sample, 79.6% of patients were Italian. MSM accounted for 34.3%, MSW for 29.6%, bisexual men for 5.8% and females for 30.3%. A total of 44.5% of patients had a high education level, 42.5% reported irregular use of condoms, 20.7% reported having had 5-10 partners in the six months prior to the visit and 32.9% were HIV-positive. HPV infection and syphilis were the most prevalent STIs. Conclusions: The most common profile of patients attending our clinic was that of an adult, Italian man with a high level of education, poor use of condoms and a high number of partners. MSM had the highest sex-behavior-related risk for STIs. In addition, our results suggest that all STD teams need to implement counselling topics and recommendations to share with patients, as well as tips on how to approach sexual health education/counselling, thereby promoting patient-centered approaches and educational programs