Low-Chloride Diet Prevents the Development of Arterial Hypertension and Protects Kidney Function in Angiotensin II-Infused Mice.

INTRODUCTION: A comprehensive pathophysiological mechanism to explain the relationship between high-salt intake and hypertension remains undefined. Evidence suggests that chloride, as the accompanying anion of sodium in dietary salt, is necessary to develop hypertension. We evaluated whether reducing dietary Cl- while keeping a standard Na+ intake modified blood pressure, cardiac hypertrophy, renal function, and vascular contractility after angiotensin II (AngII) infusion. METHODS: C56BL/6J mice fed with standard Cl- diet or a low-Cl- diet (equimolar substitution of Cl- by a mixture of Na+ salts, both diets with standard Na+ content) received AngII (infusion of 1.5 mg/kg/day) or vehicle for 14 days. We measured systolic blood pressure (SBP), glomerular filtration rate (GFR), natriuretic response to acute saline load, and contractility of aortic rings from mice infused with vehicle and AngII, in standard and low-Cl- diet. RESULTS: The mice fed the standard diet presented increased SBP and cardiac hypertrophy after AngII infusion. In contrast, low-Cl- diet prevented the increase of SBP and cardiac hypertrophy. AngII-infused mice fed a standard diet presented hampered natriuretic response to saline load, meanwhile the low-Cl- diet preserved natriuretic response in AngII-infused mice, without change in GFR. Aortic rings from mice fed with standard diet or low-Cl- diet and infused with AngII presented a similar contractile response. CONCLUSION: We conclude that the reduction in dietary Cl- as the accompanying anion of sodium in salt is protective from AngII pro-hypertensive actions due to a beneficial effect on kidney function and preserved natriuresis.

Seroprevalence of human leptospirosis in a rural community from Tandil, Argentina. Assessment of risk factors and spatial analysis.

Leptospirosis is a neglected zoonosis that is widely distributed in the world. Although it is endemic in Argentina, prevalence remains unknown. The aims of the study were: (i) to determine the prevalence of leptospirosis in humans from a rural community in Tandil Argentina, (ii) to identify infecting Leptospira spp. serogroups, (iii) to identify factors associated with the infection, (iv) to estimate the population attributable fraction (PAF) of the risk factors and (v) to determine the spatial patterns of disease presentation and related risk factors. Blood samples from 202 participants were collected. A survey was conducted to obtain clinical and epidemiological data. Serological testing was performed by the microscopic agglutination test (MAT). Univariate and multivariate methods were applied to evaluate associations. Spatial clusters were investigated for seroprevalence and risk factors. Antibodies were found in 32.2% of participants (95% CI: 25.8-39.1). The most prevalent serogroup was Hebdomadis followed by Sejroe; Icterohaemorrhagiae; Tarassovi and Canicola. Living at lower altitudes (OR: 13.04; 95% CI: 2.60-65.32); not having access to water supply network (OR: 2.95; 95% CI: 1.30-6.69); living close to flooded streets (OR: 2.94; 95% CI: 1.14-7.69) and practicing water sports (OR: 3.12; 95% CI: 1.12-8.33) were associated with seropositivity. Factors related with housing characteristics, services and infrastructure had the higher PAF (from 17% to 81%). A spatial cluster with higher rates of positivity and of the main risk factors was determined. This work contributes useful data for specific preventive measures that should be implemented for the control of the disease.

Fetal cardiac dysfunction in pregnancies affected by intrahepatic cholestasis of pregnancy: A cohort study.

AIM: To analyze the presence of fetal myocardial dysfunction in intrahepatic cholestasis of pregnancy (ICP) at diagnosis. METHODS: This prospective cohort study included 49 pregnant participants with ICP at diagnosis and 49 nonaffected controls from a single public hospital. ICP was diagnosed based on clinical symptoms after excluding other causes of pruritus and presence of autoimmune diseases. Total bile acids were not obtained in this cohort. ICP pregnancies were assessed with a functional echocardiography at diagnosis including PR-interval, isovolumetric contraction time (ICT), ejection time (ET), and isovolumetric relaxation time (IRT) for electrical, systolic, and diastolic function, respectively. Controls were assessed at recruitment. Perinatal outcomes were obtained from delivery books. The main outcome was the presence of PR-interval prolongation or first-degree fetal heart block, and echographic signs of diastolic and systolic dysfunction. RESULTS: Compared to controls, ICP were above upper limit in conjugated bilirubin (2.0% vs. 20.4%; p = 0.008), aspartate aminotransferase (2.0% vs. 24.5%; p = 0.002), and alanine aminotransferase (4.1% vs. 28.6%; p = 0.002). ICP was associated with a higher PR-interval (130 ± 12 ms vs. 121 ± 6 ms; p < 0.0001) with five first-degree fetal heart blocks. IRT was significantly higher in ICP (42 ± 6 ms vs. 37 ± 5 ms; p = 0.0001), with no differences in ICT and ET. PR-interval trend was only positively correlated with IRT in ICP pregnancies (p = 0.04 and p = 0.34, in ICP and controls, respectively). CONCLUSIONS: Our study demonstrates that fetuses affected by maternal ICP are associated with electrical and diastolic myocardial dysfunction. More studies focused on antenatal and postnatal functional echocardiography are necessary to validate our results and consider these markers in the clinical management of ICP pregnancies.

SPARC inhibition accelerates NAFLD-associated hepatocellular carcinoma development by dysregulating hepatic lipid metabolism.

BACKGROUND AND AIMS: Non-alcoholic fatty liver (NAFLD) and its more serious form non-alcoholic steatohepatitis increase risk of hepatocellular carcinoma (HCC). Lipid metabolic alterations and its role in HCC development remain unclear. SPARC (Secreted Protein, Acidic and Rich in Cysteine) is involved in lipid metabolism, NAFLD and diabetes, but the effects on hepatic lipid metabolism and HCC development is unknown. The aim of this study was to evaluate the role of SPARC in HCC development in the context of NAFLD. METHODS: Primary hepatocyte cultures from knockout (SPARC-/- ) or wild-type (SPARC+/+ ) mice, and HepG2 cells were used to assess the effects of free fatty acids on lipid accumulation, expression of lipogenic genes and de novo triglyceride (TG) synthesis. A NAFLD-HCC model was stabilized on SPARC-/- or SPARC+/+ mice. Correlations among SPARC, lipid metabolism-related gene expression patterns and clinical prognosis were studied using HCC gene expression dataset. RESULTS: SPARC-/- mice increases hepatic lipid deposits over time. Hepatocytes from SPARC-/- mice or inhibition of SPARC by an antisense adenovirus in HepG2 cells resulted in increased TG deposit, expression of lipid-related genes and nuclear translocation of SREBP1c. Human HCC database analysis revealed that SPARC negatively correlated with genes involved in lipid metabolism, and with poor survival. In NAFLD-HCC murine model, the absence of SPARC accelerates HCC development. RNA-seq study revealed that pathways related to lipid metabolism, cellular detoxification and proliferation were upregulated in SPARC-/- tumour-bearing mice. CONCLUSIONS: The absence of SPARC is associated with an altered hepatic lipid metabolism, and an accelerated NAFLD-related HCC development.

Human umbilical cord perivascular cells-derived extracellular vesicles mediate the transfer of IGF-I to the liver and ameliorate hepatic fibrogenesis in mice.

Extracellular vesicles (EVs) can mediate mesenchymal stromal cells (MSCs) paracrine effects. We aimed to evaluate the therapeutic potential of human umbilical cord perivascular cells (HUCPVCs) engineered to produce Insulin Growth Factor like-I (IGF-I) in experimental liver fibrosis and the role of EVs in this effect. HUCPVCs were engineered to produce human IGF-I (AdhIGF-I) or green fluorescence protein (AdGFP) using adenoviruses, and EVs were isolated from their conditioned medium (CM). In vitro effects of CM and EVs on hepatic stellate cells and hepatic macrophages were studied. Cells or EVs-based treatments were evaluated in thioacetamide-induced liver fibrosis in mice. The application of AdhIGF-I-HUCPVCs resulted in a further amelioration of liver fibrosis when compared to AdGFP-HUCPVCs and saline. Similarly, treatment with AdhIGF-I-HUCPVCs-derived EVs resulted in a reduction of collagen deposition and gene expression of the fibrogenic related molecules TGF-ß1, α-SMA, and COL1A2. In vitro incubation of hepatic stellate cells with EVs-AdhIGF-I-HUCPVCs significantly reduced activation of fibrogenic cells. In addition, EVs-AdhIGF-I-HUCPVCs trigger hepatic macrophages to switch their phenotype towards anti-inflammatory phagocytes, which might be involved in the antifibrotic effect. Consistently, high levels of IGF-I were observed within EVs-AdhIGF-I-HUCPVCs but not in controls EVs. Our results showed that hIGF-I carrying EVs could mediate the paracrine mechanism by which AdhIGF-I-HUCPVCs reduce liver fibrosis.

Testing of In Vitro Susceptibility of Neisseria gonorrhoeae to Azithromycin: Comparison of Disk Diffusion and Reference Agar Dilution Methods.

Azithromycin in combination with ceftriaxone is recommended as the first-line treatment for uncomplicated gonorrhea in many countries. Therefore, monitoring of azithromycin susceptibility of Neisseria gonorrhoeae isolates is essential. In 2019, the Clinical and Laboratory Standards Institute (CLSI) listed the MIC breakpoint for a susceptible-only category to azithromycin, but breakpoints for disk diffusion are not yet available. In this study, we evaluated the usefulness of disk diffusion for testing the susceptibility of N. gonorrhoeae isolates to azithromycin. A total of 189 clinical isolates susceptible and nonsusceptible to azithromycin were used. Agar dilution MICs were correlated with inhibition zone diameters of azithromycin disks (15-µg) manufactured by BBL and Oxoid. In addition, an interlaboratory study involving two clinical microbiology laboratories was conducted. There was a strong correlation between disk diffusion and agar dilution for BBL disks (r = -0.74; P < 0.001) and Oxoid disks (r = -0.75; P < 0.001). Using a zone diameter breakpoint of ≥27 mm (susceptible) and ≤26 mm (nonsusceptible) yielded good separation between susceptible and nonsusceptible isolates and the least number of discrepancies. Compared to agar dilution, disk diffusion showed high agreement and kappa values of 95.2% and 0.899 (P < 0.001) for BBL disks and 96.8% and 0.933 (P < 0.001) for Oxoid disks, respectively. Major and very major discrepancies were observed in isolates with azithromycin MICs (1 and 2 µg/ml, respectively) near to the breakpoint. These data illustrate that disk diffusion could be a reliable method in clinical laboratories to test susceptibility to azithromycin in N. gonorrhoeae isolates.

Effects of Campylobacter fetus on bull sperm quality.

Campylobacter fetus is a gram-negative, motile, spiral or S-shaped bacterium, which induces campylobacteriosis. This disease causes decrease productivity of cattle. Although considerable research has been done on the role of C. fetus on female fertility, little is known about the effect on bulls. The aim of this work was to evaluate the effect of C. fetus subsp. fetus (Cff) and C. fetus subsp. venerealis (Cfv) on bull sperm quality. Samples of frozen semen (n = 29 straws) were each distributed into three groups: two of them incubated with the microorganism (Cff, Cfv) and a control group. The proportions of live spermatozoa, with functional membrane and true acrosomal reaction in control group were significantly (P < 0.01) greater than those observed in Cff and Cfv groups. However, no significant differences (P > 0.05) were found in sperm chromatin structure among treatments. In adhesion assay, proportions of spermatozoa with adhered Campylobacter were similar for both subspecies. Results confirm that Cff and Cfv have the same ability to bind in an irreversible way to bull spermatozoa and to affect sperm quality. It is proposed that adherence could be considered as the main cause of sperm alterations, and also an important step of pathogenesis and venereal transmission.

Uso de mioinositol más Bifidobacterium lactis y Lactobacillus rhamnosus para la prevención de diabetes mellitus gestacional en mujeres mexicanas.

AIM: To compare the incidence of gestational diabetes mellitus (GDM) in women with three or more risk factor to developing GDM supplemented with myo-inositol plus probiotics versus women care without supplementation. METHODS: Retrospective cohort study, group 1, women with supplementation (myo-inositol 2g plus Bifidobacterium lactis and Lactobacillus rhamnosus 5x108 UFC, twice per day, from 12-14 to 28 weeks of gestation; group 2, women with prenatal care without supplementation, matched by age and body mass index (BMI). The primary outcome was the incidence of GDM using the International Association of Diabetes and Pregnancy Study Groups criteria. RESULTS: Group 1 n=48, group 2 n=96. There were no significant baseline differences between groups in age, BMI and number of risk factors. The incidence of GDM in group 1 was n=14 (29.2%), and for group 2 n=46 (47.9%); RR: 0.61 (95% CI: 0.37-0.99; p = 0.03). CONCLUSIONS: Supplementation from 12-14 weeks of gestation with myo-inositol plus probiotics decrease the incidence of GDM in Mexican women.

OBJETIVO: Comparar la incidencia de diabetes mellitus gestacional (DMG) en mujeres con tres o más factores de riesgo para desarrollar DMG suplementadas con mioinositol más probióticos versus mujeres sin suplementación.­. MATERIAL Y ­MÉTODOS: Estudio de cohorte retrospectivo, grupo 1, mujeres con suplementación (mioinositol 2 g más Bifidobacterium lactis y Lactobacillus rhamnosus 5x108unidades formadoras de colonias, dos veces al día, de las 12-14 hasta las 28 semanas de gestación); grupo 2, mujeres con control prenatal habitual sin suplementación, pareadas por edad e índice de masa corporal (IMC). El resultado primario fue la incidencia de DMG utilizando los criterios de la Asociación Internacional de Grupos de Estudio de Diabetes y Embarazo. RESULTADOS: Grupo 1, n = 48, y grupo 2 n = 96. No hubo diferencias significativas en características basales como edad, IMC, y numero de factores de riesgo entre los grupos. La incidencia de DMG en el grupo 1 fue n = 14 (29.2%) y en el grupo 2 n = 46 (47.9%); RR: 0.61 (IC 95%: 0.37-0.99; p = 0.03). CONCLUSIONES: La suplementación desde las 12-14 semanas de gestación con mioinositol más probióticos disminuye la incidencia de DMG en mujeres mexicanas.

4Mu Decreases CD47 Expression on Hepatic Cancer Stem Cells and Primes a Potent Antitumor T Cell Response Induced by Interleukin-12.

The tumor microenvironment (TME) represents a complex interplay between different cellular components, including tumor cells and cancer stem cells (CSCs), with the associated stroma; such interaction promotes tumor immune escape and sustains tumor growth. Several experimental approaches for cancer therapy are focused on TME remodeling, resulting in increased antitumor effects. We previously demonstrated that the hyaluronan synthesis inhibitor 4-methylumbelliferone (4Mu) decreases liver fibrosis and induces antitumor activity in hepatocellular carcinoma (HCC). In this work, 4Mu, in combination with an adenovirus encoding interleukin-12 genes (AdIL-12), elicited a potent antitumor effect and significantly prolonged animal survival (p < 0.05) in an orthotopic HCC model established in fibrotic livers. In assessing the presence of CSCs, we found reduced mRNA levels of CD133+, CD90+, EpCAM+, CD44+, and CD13+ CSC markers within HCC tumors (p < 0.01). Additionally, 4Mu downregulated the expression of the CSC marker CD47+ on HCC cells, promoted phagocytosis by antigen-presenting cells, and, combined with Ad-IL12, elicited a potent cytotoxic-specific T cell response. Finally, animal survival was increased when CD133low HCC cells, generated upon 4Mu treatment, were injected in a metastatic HCC model. In conclusion, the combined strategy ameliorates HCC aggressiveness by targeting CSCs and as a result of the induction of anticancer immunity.

Herpud1 impacts insulin-dependent glucose uptake in skeletal muscle cells by controlling the Ca2+-calcineurin-Akt axis.

Skeletal muscle plays a central role in insulin-controlled glucose homeostasis. The molecular mechanisms related to insulin resistance in this tissue are incompletely understood. Herpud1 is an endoplasmic reticulum membrane protein that maintains intracellular Ca2+ homeostasis under stress conditions. It has recently been reported that Herpud1-knockout mice display intolerance to a glucose load without showing altered insulin secretion. The functions of Herpud1 in skeletal muscle also remain unknown. Based on these findings, we propose that Herpud1 is necessary for insulin-dependent glucose disposal in skeletal muscle. Here we show that Herpud1 silencing decreased insulin-dependent glucose uptake, GLUT4 translocation to the plasma membrane, and Akt Ser473 phosphorylation in cultured L6 myotubes. A decrease in insulin-induced Akt Ser473 phosphorylation was observed in soleus but not in extensor digitorum longus muscle samples from Herpud1-knockout mice. Herpud1 knockdown increased the IP3R-dependent cytosolic Ca2+ response and the activity of Ca2+-dependent serine/threonine phosphatase calcineurin in L6 cells. Calcineurin decreased insulin-dependent Akt phosphorylation and glucose uptake. Moreover, calcineurin inhibition restored the insulin response in Herpud1-depleted L6 cells. Based on these findings, we conclude that Herpud1 is necessary for adequate insulin-induced glucose uptake due to its role in Ca2+/calcineurin regulation in L6 myotubes.

The interplay between local immune response and Epstein-Barr virus-infected tonsillar cells could lead to viral infection control.

Epstein Barr virus (EBV) gains access to the host through tonsillar crypts. Our aim was to characterize microenvironment composition around EBV+ cells in tonsils from pediatric carriers, to disclose its role on viral pathogenesis. LMP1 expression, assessed by immunohistochemistry (IHC), was used to discriminate EBV + and - zones in 41 tonsil biopsies. Three regions were defined: Subepithelial (SE), interfollicular (IF) and germinal center (GC). CD8, GrB, CD68, IL10, Foxp3, PD1, CD56 and CD4 markers were evaluated by IHC; positive cells/100 total cells were counted. CD8+, GrB+, CD68+ and IL10+ cells were prevalent in EBV+ zones at the SE region (p < 0.0001, p = 0.03, p = 0.002 and p = 0.002 respectively, Wilcoxon test). CD4+ and CD68+ cell count were higher in EBV + GC (p = 0.01 and p = 0.0002 respectively, Wilcoxon test). Increment of CD8, GrB and CD68 at the SE region could indicate a specific response that may be due to local homing at viral entry, which could be counterbalanced by IL10, an immunosuppressive cytokine. Additionally, it could be hypothesized that CD4 augment at the GC may be involved in the EBV-induced B-cell growth control at this region, in which macrophages could also participate.

A neuropsychological study of older adult first-time sex offenders.

BACKGROUND/AIMS: The issue of whether sex offenders have cognitive deficits remains controversial. The objective of this study was to compare the neuropsychological function of older adult first time sex-offenders (FTSO), who had not previously been charged with a sexual offence prior to the age of 50, to historical long-term sex offenders (HSO) and non-sex offenders (NSO). The hypotheses were (a) that FTSO would demonstrate greater deficits in executive function, decision-making, and memory compared to non-sex offenders; and (b) the HSOs would present similar neuropsychological deficits to non-sex offenders. METHOD: A battery of neuropsychological measures was administered to 100 participants comprising 32 FTSOs, 36 HSOs, and 32 NSOs. RESULTS: Both FTSOs and HSOs showed significant impairment on tests of executive function (including verbal fluency, trail-making, and the Hayling test of response inhibition) as well as on tests of verbal and verbal memory compared to NSOs; however, there was no difference between the two sex offender groups. CONCLUSIONS: Older adult sex offenders, overall, demonstrated poorer neuropsychological performance than older adult non-sex offenders did, although there was no difference between older first-time and historical offenders. Cognitive deficits may increase the risk of sexual offending due to impaired capacity in self-regulation, planning, judgment, and inhibition. A proportion of older adult sex offenders may be harboring acquired frontal lobe pathology.

Reversal of the Hartmann's procedure: A comparative study of laparoscopic versus open surgery.

BACKGROUND: The Hartmann's operation, although less frequently performed today, is still used when initial colonic anastomosis is too risky in the short term. However, the subsequent procedure to restore gastrointestinal continuity is associated with significant morbidity and mortality. PATIENTS AND METHODS: The review of an institutional review board (IRB)-approved prospectively maintained database provided data on the Hartmann's reversal procedure performed by either laparoscopic or open technique at our institution. The data collected included: demographic data, operative approach, conversion for laparoscopic cases and perioperative morbidity and mortality. RESULTS: Over a 14-year period from January 1997 to August 2011, 74 Hartmann's reversal procedures were performed (laparoscopic surgery-49, open surgery-25). The average age was 55 years for the laparoscopic and 57 years for the open surgery group, respectively. Male patients represent 61% of both groups. There was no significant difference in operative time between the two groups (149 min vs 151 min; P = 0.95), and there was a tendency to lower morbidity (3/49-7.3% vs 4/25-16%; P = 0.24) in the laparoscopic surgery group. In the laparoscopic group, eight patients (16.3%) were converted to open surgery, mostly due to severe adhesions. The length of hospital stay was significantly shorter for the laparoscopic group (5 days vs 7 days; P = 0.44). CONCLUSIONS: The Hartmann's reversal procedure can be safely performed in the majority of the cases using a laparoscopic approach with a low morbidity rate and achieving a shorter hospital stay.

Tumor Microenvironment Remodeling by 4-Methylumbelliferone Boosts the Antitumor Effect of Combined Immunotherapy in Murine Colorectal Carcinoma.

We have previously demonstrated that a low dose of cyclophosphamide (Cy) combined with gene therapy of interleukin-12 (AdIL-12) has a synergistic, although limited, antitumoral effect in mice with colorectal carcinoma. The main mechanism involved in the efficacy of Cy+AdIL-12 was the induction of a specific immune response mediated by cytotoxic T lymphocytes. Our current aims were to evaluate the effects of 4-methylumbelliferone (4Mu), a selective inhibitor of hyaluronan (HA) synthesis, on tumor microenvironment (TME) and to investigate how 4Mu affects the therapeutic efficacy of Cy+AdIL-12. The results showed that 4Mu significantly reduced the amount of tumoral HA leading to a significant decrease in tumor interstitial pressure (TIP). As a consequence, tumor perfusion was improved allowing an increased adenoviral transgene expression. In addition, treatment with 4Mu boosted the number of cytotoxic T lymphocytes that reach the tumor after adoptive transfer resulting in a potent inhibition of tumor growth. Importantly, we observed complete tumor regression in 75% of mice when 4Mu was administrated in combination with Cy+AdIL-12. The triple combination 4Mu+Cy+AdIL-12 also induced a shift toward antiangiogenic factors production in tumor milieu. Our results showed that TME remodeling is an interesting strategy to increase the efficacy of anticancer immunotherapies based on gene and/or cell therapy.

Increased PR Interval in Fetuses of Patients with Intrahepatic Cholestasis of Pregnancy.

OBJECTIVE: To evaluate the fetal mechanical PR interval in fetuses from pregnancies with intrahepatic cholestasis of pregnancy (ICP). METHODS: A case-control study was conducted in the Maternal-Fetal Medicine Unit at Hospital Carlos Van Buren between 2011 and 2013. Fetal echocardiography was performed in patients with ICP and normal pregnancies. Demographic and clinical characteristics were compared using the Mann-Whitney U test for continuous variables. A p value <0.05 was considered significant. RESULTS: 51 patients with ICP were compared with 51 unaffected pregnancies. There were no significant differences in neither demographic nor clinical characteristics between the two groups. The fetal PR interval was significantly longer in the ICP group when compared to the control group (134.6 ± 12 vs. 121.4 ± 10 ms, p < 0.001). Moreover, four fetuses from the ICP group had a mechanical PR interval >150 ms, which is compatible with a first-degree atrioventricular block. Two fetuses were identified in the neonatal period and were transferred to pediatric cardiology for follow-up, with a normal mechanical PR after the first month of life. CONCLUSIONS: We demonstrated that the fetal cardiac conduction system is altered in fetuses of patients with ICP. Further research is necessary to determine whether this alteration is related to stillbirths seen in ICP.

Epstein-Barr virus load in transplant patients: Early detection of post-transplant lymphoproliferative disorders.

High levels of circulating EBV load are used as a marker of post-transplant lymphoproliferative disorders (PTLD). There is no consensus regarding the threshold level indicative of an increase in peripheral EBV DNA. The aim of the study was to clinically validate a developed EBV quantification assay for early PTLD detection. Transversal study: paired peripheral blood mononuclear cells (PBMC), plasma and oropharyngeal lymphoid tissue (OLT) from children undergoing a solid organ transplant with (n=58) and without (n=47) PTLD. Retrospective follow-up: 71 paired PBMC and plasma from recipients with (n=6) and without (n=6) PTLD history. EBV load was determined by real-time PCR. The diagnostic ability to detect all PTLD (categories 1-4), advanced PTLD (categories 2-4) or neoplastic PTLD (categories 3 and 4) was estimated by analyzing the test performance at different cut-off values or with a load variation greater than 0.5log units. The higher diagnostic performance for identifying all, advanced or neoplastic PTLD, was achieved with cut-off values of 1.08; 1.60 and 2.47log EBVgEq/10(5) PBMC or 2.30; 2.60; 4.47loggEq/10(5) OLT cells, respectively. EBV DNA detection in plasma showed high specificity but low (all categories) or high (advanced/neoplastic categories) sensitivity for PTLD identification. Diagnostic performance was greater when: (1) a load variation in PBMC or plasma was identified; (2) combining the measure of EBV load in PBMC and plasma. The best diagnostic ability to identify early PTLD stages was achieved by monitoring EBV load in PBMC and plasma simultaneously; an algorithm was proposed.

Action of anti-M3muscarinic acetylcholine receptor IgG of primary Sjögren's syndrome on the enzymatic antioxidant system in rat submandibular gland.

BACKGROUND: We demonstrate that serum immunoglobulin G (IgG) directed against glandular M3 muscarinic acetylcholine receptors (M3mAChR) and pilocarpine triggers the increment of superoxide dismutase (SOD) and catalase (CAT) and the production of nitric oxide (NO) and prostaglandin E2(PGE2). METHODS: Enzyme-linked immunosorbent assay (ELISA) was performed in the presence of the human M2mAChR synthetic peptide as antigen to detect in serum of pSS patients the autoantibodies. Further, SOD and CAT specific activity and NO were determined chemically in the presence of anti-M3mAChR IgG and pilocarpine. The level of PGE2generation in the presence of autoantibody and pilocarpine was determined by ELISA. RESULTS: An association between anti-M2mAChR autoantibodies and pilocarpine given the increment of the specific activity of SOD and CAT in the serum of pSS patients and in the rat submandibular gland was observed. As a result of this action, M3synthetic peptide and atropine abrogated the stimulatory action. The L-type calcium channel, calcium/calmodulin complex and COX-2 inhibitors selectively blocked the increment of the specific activity of SOD and CAT in the rat submandibular gland. An increased production of NO and PGE2by the cholinergic autoantibody and pilocarpine was also detected. CONCLUSION: On the basis of these results, the increment of the specific activity of SOD and CAT in pSS patients as compared to control healthy individuals may be seen as a defensive reaction to the increment of the amount of ROS, which becoming uncontrollable, leads to irreversible cellular and tissue damage.

Effect of Campylobacter fetus on in vitro fertilization and embryonic development of preimplantation bovine embryos.

Campylobacter fetus is an important veterinary pathogen that causes campylobacteriosis. This disease causes decreased productivity of cattle by inducing reproductive losses. Although several virulence factors have been recognized in C. fetus, including a cytolethal distending toxin (CDT), the exact mechanism responsible for embryonic death remains unknown. The aim of this work was to evaluate the effect of C. fetus subsp. fetus (Cff) and C. fetus subsp. venerealis (Cfv), and their toxin activity on the in vitro fertilization of bovine ova and early embryonic development. Two different experiments were performed. In Experiment 1, a total of 1524 cumulus-oocyte complexes (COCs) were inseminated, distributed into three groups: two of them infected with the microorganism (Cff, Cfv) and a control group. Percentages COCs cleaved were similar among groups (p = 0.1243); however, the embryonic development rate (blastocyst at day 7) in the control group was greater (p < 0.001) than those obtained in Cff and Cfv groups. In experiment 2, a total of 746 COCs were inseminated, divided into three groups: two of them treated with the bacterial-free culture filtrates to test toxin activity (Cff-CDT, Cfv-CDT) and a control group. Both cleavage and embryonic development rates were greater (p < 0.001) in the control group than those obtained in Cff-CDT and Cfv-CDT groups. This study provides evidence that both subspecies of C. fetus do not interfere with fertilization but do affect in vitro embryonic development. It is the first report on the biological effect of the CDT on bovine embryonic development.

Development and standardization of an enzyme-linked inmunosorbent for the detection of orthohantavirus infection in Argentina based on its bacterial-expressed nucleocapside protein.

We conducted a development and standardization of an IgG ELISA assay for serological detection of human orthohantavirus infections using the recombinant antigen rLECH13 produced in bacterial and derived from the LECHV. The evaluation and standardization were carried out by analyzing serum samples from a total of 50 patients with confirmed Hantavirus Pulmonary Syndrome (HPS) diagnosis through the reference technique, 50 negative sera, and 53 patients with other medical conditions. The data from the assay analysis showed a diagnostic sensitivity value of 95% and a diagnostic specificity of 80%. The high sensitivity of this novel assay leads us to conclude that rLECH13 is a feasible option for use in the immunodiagnostic of orthohantavirus infection. Additionally, it is crucial to have an antigen that can be produced under conditions that do not require highly complex laboratories. Furthermore, the new assay is cost-effective, reproducible, and demonstrates excellent performance.

Comparative evaluation of lateral flow assays to diagnose chronic Trypanosoma cruzi infection in Bolivia.

Bolivia has the highest incidence of Chagas disease (CD) worldwide. Caused by the parasite Trypanasoma cruzi, CD is generally a chronic condition. Diagnosis is logistically and financially challenging, requiring at least two different laboratory-based serological tests. Many CD cases are missed; in Bolivia it is estimated just 6% of individuals chronically infected with T. cruzi get diagnosed. Achieving control on the way to elimination of CD requires a radical simplification of the current CD testing pathways, to overcome the barriers to accessing CD treatment. We aimed to generate unbiased performance data of lateral flow assays (LFAs) for T. cruzi infection in Bolivia, to evaluate their usefulness for improving T. cruzi diagnosis rates in a precise and efficient manner. This retrospective, laboratory-based, diagnostic evaluation study sought to estimate the sensitivity/specificity of 10 commercially available LFAs for T. cruzi, using the current CD diagnostic algorithm employed in Bolivia as the reference test method. All tests were blinded at the study site and performed by three operators. In total, 470 serum samples were tested, including 221 and 249 characterized as CD-positive/-negative, respectively. The LFAs were scored according to their relative importance using a decision-tree-based algorithm, with the mean decrease in Gini index as the scoring metric. The estimates of sensitivities ranged from 62.2-97.7% (95% confidence interval (CI) lower bound 55.0-94.7%); for specificities the range was 78.6-100% (95% CI lower bound 72.0-97.5%); 5/10 and 6/10 tests had sensitivity >90% and specificity >95%, respectively. Four LFAs showed high values of both sensitivity (93-95%) and specificity (97-99%). The agreement between 6 LFAs and the reference tests was almost perfect (Kappa 0.83-0.94). Most LFAs evaluated thus showed performances comparable with current laboratory-based diagnostic methods.