Derivation and Validation of a Biomarker-Based Clinical Algorithm to Rule Out Sepsis From Noninfectious Systemic Inflammatory Response Syndrome at Emergency Department Admission: A Multicenter Prospective Study.

OBJECTIVES: To derive and validate a predictive algorithm integrating a nomogram-based prediction of the pretest probability of infection with a panel of serum biomarkers, which could robustly differentiate sepsis/septic shock from noninfectious systemic inflammatory response syndrome. DESIGN: Multicenter prospective study. SETTING: At emergency department admission in five University hospitals. PATIENTS: Nine-hundred forty-seven adults in inception cohort and 185 adults in validation cohort. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: A nomogram, including age, Sequential Organ Failure Assessment score, recent antimicrobial therapy, hyperthermia, leukocytosis, and high C-reactive protein values, was built in order to take data from 716 infected patients and 120 patients with noninfectious systemic inflammatory response syndrome to predict pretest probability of infection. Then, the best combination of procalcitonin, soluble phospholipase A2 group IIA, presepsin, soluble interleukin-2 receptor α, and soluble triggering receptor expressed on myeloid cell-1 was applied in order to categorize patients as "likely" or "unlikely" to be infected. The predictive algorithm required only procalcitonin backed up with soluble phospholipase A2 group IIA determined in 29% of the patients to rule out sepsis/septic shock with a negative predictive value of 93%. In a validation cohort of 158 patients, predictive algorithm reached 100% of negative predictive value requiring biomarker measurements in 18% of the population. CONCLUSIONS: We have developed and validated a high-performing, reproducible, and parsimonious algorithm to assist emergency department physicians in distinguishing sepsis/septic shock from noninfectious systemic inflammatory response syndrome.

Multiple sites of vascular dilation or aneurysmal disease and matrix metalloproteinase genetic variants in patients with abdominal aortic aneurysm.

OBJECTIVE: The objective of this study was to assess whether functional genetic polymorphisms of matrix metalloproteinases (MMPs) 1, 3, 9, and 12 are associated with arterial enlargements or aneurysms of the thoracic aorta or popliteal arteries in patients with abdominal aortic aneurysm (AAA). METHODS: The associations between MMP1 (-1607 G in/del, rs1799750), MMP3 (-1171 A in/del rs35068180), MMP9 (13-26 CA repeats around -90, rs2234681, rs917576, rs917577), and MMP12 (G/T missense variation, rs652438) polymorphisms and enlargements or aneurysms of the thoracic aorta and popliteal arteries were tested in 169 consecutive AAA patients. RESULTS: Thoracic aorta enlargement or aneurysm (TE/A; maximum diameter, >35 mm) was detected in 34 patients (20.1% prevalence). MMP9 rs2234681 microsatellite was the only genetic determinant of TE/A in AAA patients (P = .003), followed by hypercholesterolemia and antiplatelet use. Carriers of both alleles with ≥22 CA repeats had a 5.9 (95% confidence interval, 1.9-18.6; P < .0001) increased odds of TE/A, and a score considering all three variables showed 98% negative predictive value and 30% positive predictive value for thoracic aortic aneurysm detection. Eighty-two popliteal artery enlargements or aneurysms (diameter >10 mm) occurred in 55 patients (33.1% prevalence). Carriers of MMP12 rs652438 C allele showed an 18% (P = .006) increased diameter in popliteal arteries and a 2.8 (95% confidence interval, 1.3-6; P = .008) increased odds of popliteal artery enlargement or aneurysm compared with TT genotype. CONCLUSIONS: Among patients with AAA, carriers of homozygous ≥22 CA repeats in MMP9 rs12234681 and of C allele in MMP12 rs652438 have a substantial risk of carrying thoracic and popliteal enlargements, respectively.

Procalcitonin in early rule-in/rule-out of sepsis in SIRS patients admitted to a medical ward.

BACKGROUND: A relevant amount of patients with clinical suspect of sepsis is admitted and treated in medical wards (MW). These patients have a better prognosis but are older and with more comorbidities compared to those admitted to intensive care units (ICU). Procalcitonin (PCT) is extensively used in emergency departments for the diagnosis of sepsis, but its accuracy in the setting of a MW has not been thoroughly investigated. Predicted low PCT levels also call for the comparison of immunomagnetic-chemiluminescent (L-PCT) and time-resolved amplified cryptate emission (TRACE, K-PCT) technologies, in PCT determination. METHODS: In 80 patients with systemic inflammatory response syndrome (SIRS) diagnostic criteria and suspect of sepsis newly admitted to a MW, PCT was determined with L- and K-PCT method. RESULTS: Sixty patients were diagnosed as sepsis (20 microbiologically and 40 clinically proven) and 20 with non-infective SIRS. The sepsis group had significantly higher levels of both PCTs, with no differences between the clinically and microbiologically proven subgroups. The areas under ROC curves for L- and K-PCT were 0.72 and 0.78 (p<0.001 for each), respectively. Based on MW customized cut-off values of 0.150 (L-PCT) and 0.143 ng/mL (K-PCT), overall accuracies were 66.8 (95% CI 58.7-78.9) and 78.2% (69.8-87.2), respectively, compared to the 55% (44.2-66) of 0.5 ng/mL canonical cut-off. Neither PCT-L nor -K held prognostic value on survival. CONCLUSIONS: In MW patients, customized PCT cut-off levels provide better accuracy than customary levels adopted from ICU, and TRACE technology seems to offer a wider analysis range.

Lipoprotein(a) as a Risk Factor for Cardiovascular Diseases: Pathophysiology and Treatment Perspectives.

Cardiovascular disease (CVD) is still a leading cause of morbidity and mortality, despite all the progress achieved as regards to both prevention and treatment. Having high levels of lipoprotein(a) [Lp(a)] is a risk factor for cardiovascular disease that operates independently. It can increase the risk of developing cardiovascular disease even when LDL cholesterol (LDL-C) levels are within the recommended range, which is referred to as residual cardiovascular risk. Lp(a) is an LDL-like particle present in human plasma, in which a large plasminogen-like glycoprotein, apolipoprotein(a) [Apo(a)], is covalently bound to Apo B100 via one disulfide bridge. Apo(a) contains one plasminogen-like kringle V structure, a variable number of plasminogen-like kringle IV structures (types 1-10), and one inactive protease region. There is a large inter-individual variation of plasma concentrations of Lp(a), mainly ascribable to genetic variants in the Lp(a) gene: in the general po-pulation, Lp(a) levels can range from <1 mg/dL to >1000 mg/dL. Concentrations also vary between different ethnicities. Lp(a) has been established as one of the risk factors that play an important role in the development of atherosclerotic plaque. Indeed, high concentrations of Lp(a) have been related to a greater risk of ischemic CVD, aortic valve stenosis, and heart failure. The threshold value has been set at 50 mg/dL, but the risk may increase already at levels above 30 mg/dL. Although there is a well-established and strong link between high Lp(a) levels and coronary as well as cerebrovascular disease, the evidence regarding incident peripheral arterial disease and carotid atherosclerosis is not as conclusive. Because lifestyle changes and standard lipid-lowering treatments, such as statins, niacin, and cholesteryl ester transfer protein inhibitors, are not highly effective in reducing Lp(a) levels, there is increased interest in developing new drugs that can address this issue. PCSK9 inhibitors seem to be capable of reducing Lp(a) levels by 25-30%. Mipomersen decreases Lp(a) levels by 25-40%, but its use is burdened with important side effects. At the current time, the most effective and tolerated treatment for patients with a high Lp(a) plasma level is apheresis, while antisense oligonucleotides, small interfering RNAs, and microRNAs, which reduce Lp(a) levels by targeting RNA molecules and regulating gene expression as well as protein production levels, are the most widely explored and promising perspectives. The aim of this review is to provide an update on the current state of the art with regard to Lp(a) pathophysiological mechanisms, focusing on the most effective strategies for lowering Lp(a), including new emerging alternative therapies. The purpose of this manuscript is to improve the management of hyperlipoproteinemia(a) in order to achieve better control of the residual cardiovascular risk, which remains unacceptably high.

Epidemiology of atherosclerotic cardiovascular disease in polygenic hypercholesterolemia with or without high lipoprotein(a) levels.

Background and aims: Epidemiology of atherosclerotic cardiovascular disease might be different in patients with polygenic hypercholesterolemia plus high levels (≥30 mg/dl) of Lp(a) (H-Lpa) than in those with polygenic hypercholesterolemia alone (H-LDL). We compared the incidence of peripheral artery disease (PAD), coronary artery disease (CAD), and cerebrovascular disease (CVD) in patients with H-Lpa and in those with H-LDL. Methods: Retrospective analysis of demographics, risk factors, vascular events, therapy, and lipid profile in outpatient clinical data. Inclusion criteria was adult age, diagnosis of polygenic hypercholesterolemia, and both indication and availability for Lp(a) measurement. Results: Medical records of 258 patients with H-Lpa and 290 H-LDL were reviewed for occurrence of vascular events. The median duration of follow-up was 10 years (IQR 3-16). In spite of a similar reduction of LDL cholesterol, vascular events occurred more frequently, and approximately 7 years earlier (P = 0.024) in patients with H-Lpa than in H-LDL (HR 1.96 1.21-3.17, P = 0.006). The difference was around 10 years for acute events (TIA, Stroke, acute coronary events) and one year for chronic ones (P = 0.023 and 0.525, respectively). Occurrence of acute CAD was higher in H-Lpa men (HR 3.1, 95% CI 1.2-7.9, P = 0.007) while, among women, PAD was observed exclusively in H-Lpa subjects with smoking habits (P = 0.009). Conclusions: Patients with high Lp(a) levels suffer from a larger and earlier burden of the disease compared to those with polygenic hypercholesterolemia alone. These patients are at higher risk of CAD if they are men, and of PAD if they are women.

Oral anticoagulation and VKORC1 polymorphism in patients with a mechanical heart prosthesis: a 6-year follow-up.

Therapy with Vitamin K antagonists (VKA) effectively reduces the thrombosis risk in many clinical conditions. Genetic variants of vitamin K epoxide reductase (VKORC-1) are associated with increased VKA effect and bleeding risk. It is unknown whether these variants could also affect the long-term outcome in patients with high-dosage oral anticoagulation and/or more difficult adherence to the therapeutic INR range. Hundred and twenty-four patients with mechanical heart valve replacement assuming VKA were genotyped for VKORC-1 -1639G>A (Rs9923231) polymorphism. Hemorrhage, venous thrombosis and atherothrombotic events were retrospectively assessed for a 6-year period. Furthermore, stability of their INR in relationship with the VKORC-1 genotype was investigated day-by-day for 3 months. No differences were observed in hemorrhage and venous thrombosis events according to rs 9923231. GG genotype carriers (n = 41) had no atherothrombotic events, while 4 strokes, 4 TIA and 3 AMI were diagnosed in A carriers (n = 83; P = 0.0008). During the daily observation period, A allele carriers had lower VKA requirements (4.7, 3.7, 2.2 mg/day for GG/GA/AA genotype respectively; P = 0.00001), higher mean INR (2.7, 2.8, 2.9; P = 0.05) and a higher number of examinations above the therapeutic range than GG carriers (17 % vs. 0 % in GG genotype, P = 0.036). Conversely, patients with GG genotype had a more stable dosage of VKA (P = 0.006) and a higher percentage of examinations under the therapeutic range (51, 43 and 36 % in GG, GA and AA genotype, respectively, P = 0.040). In patients with high dosage VKA, VKORC-1 polymorphism is associated to a different warfarin dosage, anticoagulation level, time spent outside the therapeutic range and, in the long-term, a different incidence of atherothrombotic events.

Remodeling of abdominal aortic aneurysm sac following endovascular aortic repair: association with clinical, surgical, and genetic factors.

After successful endovascular aortic repair (EVAR), abdominal aortic aneurysms (AAA) sac will undergo negative remodeling (i.e., shrinkage) as a measure of successful exclusion. Determinants of shrinkage after EVAR are not fully known. In 84 post-EVAR patients, time course of AAA diameter after repair and occurrence of endoleaks (ELs) have been correlated with clinical history, medications, anthropometric data, vascular anatomy, and matrix metalloprotease (MMP) genetic variants (namely MMP-1 rs1799750, MMP-3 rs35068180, MMP-9 rs2234681, rs917576, rs917577, MMP-12 rs652438, and TIMP1 rs4898). During follow-up, 41 ELs were detected in 37 patients (44%, 10.4 events/100 pt./y), accounting for AAA dilation or reduced shrinkage (P < .001). High-flow ELs (type 1 and/or 3) occurrence was associated with warfarin use, MMP9 rs17577 polymorphism, and unfavorable anatomy, while low-flow type 2 ELs occurred more often in TIMP1 rs4898 non-T carriers. In EL-free patients, AAA diameter decreased for the first three years, (-4, -3 and - 2 mm/year respectively) and remained stable thereafter. Shrinkage between two measurements (n = 120) was associated with smaller AAA diameter at the baseline, peripheral arterial disease (PAD), patients' older age at intervention, and G-/G- genotype in MMP1 rs1799750 (binary logistic regression, P = .0001). Aneurysmal sac shrinking occurs for few years after EVAR, only in patients without EL, and is related to older age, PAD, smaller aneurysm size and putative lower MMP1 expression while EL occurrence prevents such a remodeling and is mainly related to local-acting factors like unfavorable anatomy, anticoagulation, and MMP9 and TIMP1 genetic polymorphisms.

Ruling Out Coronavirus Disease 2019 in Patients with Pneumonia: The Role of Blood Cell Count and Lung Ultrasound.

Coronavirus disease 2019 (COVID-19) is characterized by a distinctive blood leucocyte pattern and B-lines on lung ultrasound (LUS) as marker of alveolar-interstitial syndrome. We aimed to evaluate the accuracy of blood leucocyte count alone or in combination with LUS for COVID-19 diagnosis. We retrospectively enrolled consecutive patients diagnosed with community acquired pneumonia (CAP) at hospital admission to derive and validate cutoff values for blood cell count that could be predictive of COVID-19 before confirmation by the nucleic acid amplification test (NAAT). Cutoff values, generated and confirmed in inception (41/115, positive/negative patients) and validation (100/180, positive/negative patients) cohorts, were ≤17 and ≤10 cells/mm3 for basophils and eosinophils, respectively. Basophils and/or eosinophils below cutoff were associated with sensitivity of 98% (95%CI, 94-100) and negative likelihood ratio of 0.04 (95%CI, 0.01-0.11). In a subgroup of 265 subjects, the sensitivity of B-line on LUS was 15% lower (p < 0.001) than that of basophils and/or eosinophils below cutoff. The combination of B-lines with basophils and eosinophils below cutoff was associated with a moderate increase of the positive likelihood ratio: 5.0 (95%CI, 3.2-7.7). In conclusion, basophil and eosinophil counts above the generated cutoff virtually rule out COVID-19 in patients with CAP. Our findings can help optimize patient triage pending the NAAT results.

Aquaporin-8-facilitated mitochondrial ammonia transport.

Aquaporin-8 (AQP8) is a membrane channel permeable to water and ammonia. As AQP8 is expressed in the inner mitochondrial membrane of several mammalian tissues, we studied the effect of the AQP8 expression on the mitochondrial transport of ammonia. Recombinant rat AQP8 was expressed in the yeast Saccharomyces cerevisiae. The presence of AQP8 in the inner membrane of yeast mitochondria was demonstrated by subcellular fractionation and immunoblotting analysis. The ammonia transport was determined in isolated mitochondria by stopped flow light scattering using formamide as ammonia analog. We found that the presence of AQP8 increased by threefold mitochondrial formamide transport. AQP8-facilitated mitochondrial formamide transport in rat native tissue was confirmed in liver (a mitochondrial AQP8-expressing tissue) vs. brain (a mitochondrial AQP8 non-expressing tissue). Comparative studies indicated that the AQP8-mediated mitochondrial movement of formamide was markedly higher than that of water. Together, our data suggest that ammonia diffusional transport is a major function for mitochondrial AQP8.

Epidemiological and clinical features of rotavirus among children younger than 5 years of age hospitalized with acute gastroenteritis in Northern Italy.

BACKGROUND: Rotavirus is the major cause of acute gastroenteritis and severe dehydrating diarrhea in young children. METHODS: To estimate the proportion of hospital admissions for rotavirus acute gastroenteritis and identify the circulating G and P genotypes among children under five years of age, we conducted a prospective observational study from January to December 2008, recruiting children consecutively admitted to six hospitals in Milan and nearby towns in northern Italy. Typing was done on stool samples by reverse transcriptase polymerase chain reaction amplification. RESULTS: Of the 521 stool samples from children with acute gastroenteritis, 34.9% (95%CI, 30.8 to 39.2%) were rotavirus-positive. Two thirds (67.6%) were under two years of age, and 13.2% were under six months. The predominant G type was G1 (40.7%), followed by G9 (22.5%), G2 (13.2%), G3 (5.5%), G4 (3.8%) and G10 (1.6%). Twenty-one (11.7%) mixed-G infections were identified: G1+G10 (8.8%); G1+G9 (1.6%); and G2+G10 (1.2%). Only P[8] (67.6%) and P[4] (12.6%) types were P genotyped. The predominant single G/P combination was G1P[8] (39.7%), followed by G9P[8] (25.3%), G2P[4] (14.3%), and G3P[8] (4.1%). All G-mixed types combined with P[8]. CONCLUSIONS: These findings show an high prevalence of rotavirus infections among children admitted to hospital for acute gastroenteritis caused by different rotavirus strains circulating in the area studied.

Screening Readthrough Compounds to Suppress Nonsense Mutations: Possible Application to ß-Thalassemia.

Several types of thalassemia (including ß039-thalassemia) are caused by nonsense mutations in genes controlling globin production, leading to premature translation termination and mRNA destabilization mediated by the nonsense mediated mRNA decay. Drugs (for instance, aminoglycosides) can be designed to suppress premature translation termination by inducing readthrough (or nonsense suppression) at the premature termination codon. These findings have introduced new hopes for the development of a pharmacologic approach to cure this genetic disease. In the present review, we first summarize the principle and current status of the chemical relief for the expression of functional proteins from genes otherwise unfruitful for the presence of nonsense mutations. Second, we compare data available on readthrough molecules for ß0-thalassemia. The examples reported in the review strongly suggest that ribosomal readthrough should be considered as a therapeutic approach for the treatment of ß0-thalassemia caused by nonsense mutations. Concluding, the discovery of molecules, exhibiting the property of inducing ß-globin, such as readthrough compounds, is of great interest and represents a hope for several patients, whose survival will depend on the possible use of drugs rendering blood transfusion and chelation therapy unnecessary.

Short term effects of doxycycline on matrix metalloproteinases 2 and 9.

PURPOSE: To investigate the short term effects of Doxycycline on MMP-2 and MMP-9. METHODS: Short term effects of Doxycycline (100 mg B.I.D.) on plasma levels of MMP-2 and MMP-9 were investigated in 20 healthy subjects; the effects of Doxy, Acetylsalicylic acid, Nitrates, and Enalapril on MMP-9 release from were assessed in isolated polymorphonuclear cells. RESULTS: In plasma, MMP-9 activity was reduced (-22%, 95% CI -32/-11; P = 0.002) starting at 12 h after doxy; in vitro, MMP-9 released from stimulated neutrophils was reduced by Doxy (-28%, 95% CI -43/-14; P = 0.001), inhibiting degranulation, and by nitrates (-52%, 95% CI -76/-28 P = 0.005), increasing three times both pro- and active-MMP-9 bound to neutrophils (P = 0.007 and 0.040, respectively). CONCLUSIONS: Doxy decreases MMP-9 plasma levels by around 20%, within the first 12 h. The mechanism leading to such reduction seems due to the inhibition of PMN degranulation.

Alcohol reduces MMP-2 in humans and isolated smooth muscle cells.

Alcoholic beverages are known to exert a protective effect on atherosclerotic disease. This study aimed to assess the in vivo and in vitro effects of alcohol on matrix metalloproteinase (MMP) -2 and -9, known to determine atherosclerosis progression. Eighteen healthy volunteers, regular drinkers (two standard alcohol servings/day, on average) at first examination (baseline) were asked to abstain from any alcoholic beverage for one week (abstention), and then to assume two standard alcohol servings of beer daily for 1 week (re-exposure). Activity of MMP-2 and -9, total antioxidant activity (AOA), glutathione (GSH) plasma levels were carried out at baseline, at the end of abstention, and after 1 week of re-exposure. To validate the in vivo results, MMP-2 activity and expression, AOA, and GSH, were determined in human smooth muscle cells treated for 96 h with increasing concentrations (12.5-100 mM) of ethanol. MMP-2, but not MMP-9 plasma activity was higher at abstention than at baseline or re-exposure (P<.001 and P< or =.005, respectively). Changes in AOA and GSH throughout the study were not significant. No correlation was found between MMPs and antioxidant activity. In vitro, ethanol at 25 mM reduced by around 10% MMP-2 activity (P=.003) in smooth muscle cells, whereas MMP-2 expression, AOA, and GSH were unaffected. Alcohol reduces MMP-2 plasma activity in healthy humans and in isolated vascular smooth muscle cells. This in vitro reduction is unrelated to MMP-2 expression in vascular cells or to antioxidant levels changes.

MMP-9 microsatellite polymorphism and susceptibility to carotid arteries atherosclerosis.

OBJECTIVE: The aims of this study were to compare a microsatellite polymorphism (PM) of matrix metalloproteinase (MMP)-9 in patients with carotid atherosclerosis and control population, and to assess the relationship between this PM and plaque structure. METHODS AND RESULTS: One hundred fifty patients referring to vascular diagnostic centers for suspected carotid atherosclerosis (at ultrasound examination: 110 positive, 40 negative) and controls (n=110) have been genotyped for MMP-9 PM. After controlling for risk factors, allelic and genotype frequencies were significantly different among the groups, with significant prevalence of long microsatellites in patients with carotid atherosclerosis. Long microsatellites (settled as 22 to 27 repeats) were associated with carotid atherosclerosis (odds ratio [OR], 5.2; 95% confidence interval [CI], 2.9 to 9.2), compared with controls; an independent case control study on patients with coronary atherosclerosis confirmed such result. Binary logistic regression showed that hypertension, long microsatellites in MMP-9 PM and smoking habits were variables accounting for the difference between ultrasound-positive patients and controls. Long microsatellites were also associated to plaques with thin fibrous cap and echolucent core (OR, 13.1; 95% CI, 1.6 to 100). These alleles were slightly more represented in female patients (chi2 test=0.019; OR, 2.7; 95% CI, 1.2 to 6) but not associated with other risk factors. Plasma MMP-9 levels were related neither to MMP-9 PM nor to plaque type, and were related to gender and extension of atherosclerosis in carotid arteries. CONCLUSIONS: The number of repeats (> or =22 CA) in the microsatellite of MMP-9 promoter, but not MMP-9 plasma levels, is associated to carotid atherosclerosis and particularly to plaques with a thin fibrous cap.

Chemical-Induced Read-Through at Premature Termination Codons Determined by a Rapid Dual-Fluorescence System Based on S. cerevisiae.

Nonsense mutations generate in-frame stop codons in mRNA leading to a premature arrest of translation. Functional consequences of premature termination codons (PTCs) include the synthesis of truncated proteins with loss of protein function causing severe inherited or acquired diseases. A therapeutic approach has been recently developed that is based on the use of chemical agents with the ability to suppress PTCs (read-through) restoring the synthesis of a functional full-length protein. Research interest for compounds able to induce read-through requires an efficient high throughput large scale screening system. We present a rapid, sensitive and quantitative method based on a dual-fluorescence reporter expressed in the yeast Saccharomyces cerevisiae to monitor and quantitate read-through at PTCs. We have shown that our novel system works equally well in detecting read-through at all three PTCs UGA, UAG and UAA.

MMP-9 microsatellite polymorphism: association with the progression of intima-media thickening and constrictive remodeling of carotid atherosclerotic plaques.

Intima-media thickening (IMT) of carotid arteries and constrictive remodeling (CR) of atherosclerotic plaques are vascular pathologic characteristics that precede the onset of vascular events. SMC migration and proliferation are linked both to IMT and CR and are matrix metalloproteinase 9 (MMP-9) dependent. A genetic polymorphism (PM) of MMP-9, a CA (13-27) microsatellite in the promoter region, which accounts for differential expression of MMP-9, could be linked to progression of IMT and CR. Progression of IMT and CR of plaques in carotid arteries were studied in 55 consecutive patients with a 12-18 months follow-up. All patients were genotyped for MMP-9 PM. A positive linear relationship between the number of repeats and the progression of IMT (P=0.028) as well as of CR (P=0.018) was found. The analogous relationship was obtained when only the allele with longer microsatellite was considered. Carriers of more than 20 repeats in one allele showed faster both IMT growth (P=0.045) and stenosis progressions of plaques (P=0.019). In multivariate analysis, age, dyslipidemia, and MMP-9 PM were determinants of IMT progression, while MMP-9 PM was the only one of CR. In conclusion, the high number of CA repeats in MMP-9 promoter is positively correlated with faster IMT and CR progression.

MMP-9 microsatellite polymorphism and multiple sclerosis.

A polymorphism (PM) in the microsatellite of the promoter region of matrix metalloproteinase 9 (MMP-9), modulating its expression, could play a role in susceptibility to multiple sclerosis (MS). MMP-9 PM was determined in 95 patients with MS (MS Group) and 95 age- and sex-matched controls (Control Group). Comparison of allelic frequencies showed that a higher number of CA repeats characterized the MS group (P<0.0001) and prevalence of carriers of > or =22 CA repeats was higher in the MS than in the Control Group (OR 3.4, 95% CI: 1.7-6.8, P<0.0001). An earlier age at disease onset was a characteristic of patients with >22 CA repeats (33+/-10 vs. 28+/-10, P=0.027). No differences were found in the main MRI parameters.

Invasive filamentous fungus infection with secondary cerebral vasculitis in a patient with no obvious immune suppression.

Invasive mold infections represent an emerging and important diagnostic challenge, especially in immunocompetent patients when microscopy and cultures of the biological fluids remain negative. A central nervous system localization is not common and the clinical presentation is aspecific.

Heterogeneous models for an early discrimination between sepsis and non-infective SIRS in medical ward patients: a pilot study.

The objective of the study was to determine the accuracy of phospholipase A2 group II (PLA2-II), interferon-gamma-inducible protein 10 (IP-10), angiopoietin-2 (Ang-2), and procalcitonin (PCT) plasma levels in early ruling in/out of sepsis among systemic inflammatory response syndrome (SIRS) patients. Biomarker levels were determined in 80 SIRS patients during the first 4 h of admission to the medical ward. The final diagnosis of sepsis or non-infective SIRS was issued according to good clinical practice. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) for sepsis diagnosis were assessed. The optimal biomarker combinations with clinical variables were investigated by logistic regression and decision tree (CART). PLA2-II, IP-10 and PCT, but not Ang-2, were significantly higher in septic (n = 60) than in non-infective SIRS (n = 20) patients (P ≤ 0.001, 0.027, and 0.002, respectively). PLA2-II PPV and NPV were 88 and 86%, respectively. The corresponding figures were 100 and 31% for IP-10, and 93 and 35% for PCT. Binary logistic regression model had 100% PPV and NPV, while manual and software-generated CART reached an overall accuracy of 95 and 98%, respectively, both with 100% NPV. PLA2-II and IP-10 associated with clinical variables in regression or decision tree heterogeneous models may be valuable biomarkers for sepsis diagnosis in SIRS patients admitted to medical ward (MW). Further studies are needed to introduce them into clinical practice.

Preparative scale production and functional reconstitution of a human aquaglyceroporin (AQP3) using a cell free expression system.

Understanding the selectivity of aquaporin (AQP) membrane channels and exploiting their biotechnological potential will require structural and functional studies of wild type and modified proteins; however, expression systems have not previously yielded AQPs in the necessary milligrams quantities. Cell free (CF) systems have emerged in recent years as fast, efficient and versatile technologies for the production of high quality membrane proteins. Here, we establish a convenient method to synthesize large amounts of functional human aquaglyceroporin 3 protein (AQP3), an AQP of physiological relevance conducting glycerol and some small neutral solutes besides water. Milligram amounts of AQP3 were produced as a histidine-tagged protein (hAQP3-6His) in an Escherichia coli extract-based CF system in the presence of the non-ionic detergent Brij-98. The recombinant AQP3 was purified by affinity chromatography, incorporated into liposomes and evaluated functionally by stopped-flow light scattering. Correct protein folding was indicated by the high glycerol and water permeability exhibited by the hAQP3-6His proteoliposomes as compared to empty control liposomes. Functionality of hAQP3-6His was further confirmed by the strong inhibition of the glycerol and water permeability by phloretin and HgCl2, respectively, two blockers of AQP3. Fast and convenient CF production of functional AQP3 may serve as basis for further structural/functional assessment of aquaglyceroporins and help boosting the AQP-based biomimetic technologies.