Human vascular endothelial cells with extended life spans: in vitro cell response,protein expression,and angiogenesis

An in vitro angiogenesis system was designed for screening angiogenic agonists and antagonists. In order to obtain large quantities of cells and reproducibility, human endothelial cells with extended life spans were developed by retroviral transfection. The resulting cells grown in a serum-free medium containing endothelial cell growth supplement (ECGS) have a telomerase activity, extended life spans of at least 21 passages, and an endothelial cell phenotype (diI-acetylated-LDL upake, factor VIII-related antigen, VEGFR-1 and R-2, and tissue-type plasminogen activator (tPA)) that resembled that of unaltered primary endothelial cells. Exceptions were (i) a higher expression of tPA, and (ii) a non-significant growth response to FGF-2 or VEGF stimulation. Within three-dimensional fibrin gels, specific cell clones rapidly formed tubular structures in a more reproducible manner than those observed with low-passage primary cells. Tube formation by primary endothelial cells and those with extended life spans was dependent upon FGF-2 and ECGS, respectively. Both cell types produced FGF-2 and VEGF cytokines. Increasing doses of suramin significantly decreased the size of microvessels formed by both cell lines. These functional results indicate that a vascular matrix system containing human cells with extended life spans can be successfully utilized as an in vitro assay for antiangiogenic compounds.     

How to improve screening in first-degree relatives of patients with premature coronary heart disease.

BACKGROUND: Guidelines on the prevention of cardiovascular disease recommend screening in close relatives of patients with premature coronary heart disease (CHD). This family history puts them at increased risk for CHD, independent of other major risk factors, but screening for CHD risk factors in these relatives is not widely practiced in Europe. This demonstration project examined how to improve screening of close relatives of patients with premature CHD in daily practice. METHODS: A controlled study design was used. Four hospitals were compared in a pre-test as to the actual screening of relatives of patients with premature CHD. Then they were arranged in pairs and randomly assigned to the Usual care (U) or Intervention group (I). An information and health education program--involving patients, relatives and family doctors--was developed in I to improve screening by the family doctor. RESULTS: The pre-test confirmed that screening of relatives of patients with premature CHD is poorly practiced in the four regions; no significant differences between I and U were observed. The screening of relatives during the study period reached 63.9% in I compared to 25.4% in U. This difference between I and U was present in siblings and offspring. The cardiovascular risk profile of the relatives of I was not optimal and needed improvement. CONCLUSION: Screening of first-degree relatives of patients with premature CHD can be significantly improved through a health education program. This is the first and necessary step to improve the management of risk factors in these people, who are at increased risk for CHD.     

COMMUNITY ACTIVISTS’ COMPETENCE: THE CONTRIBUTING FACTORS

This study examined the contribution of the personal and social resources to community activists' competence. The research population included 163 activists who engage in volunteer activity in traditional communities. The findings revealed that the activists' gender, supervision by community‐organizers, sense of mastery, sense of belonging to the community, citizen participation, representation, and perceptions of leadership all contributed significantly to the activists' competence. A comprehensive analysis of the findings is presented, as well as practical recommendations for community organization. The recommendations highlight the importance of professional supervision for community activists, which aims to develop perceptions of community leadership, community belonging, citizen participation, and representation in order to enhance the success of community activity. Finally, the examination of personal and social resources that contribute to activists' competence can facilitate identification of potential community activists, in addition to shedding light on the content that professional supervisors should incorporate in their work with activists.     

Evaluation of Bactec Mycosis IC/F and Plus Aerobic/F Blood Culture Bottles for Detection of Candida in the Presence of Antifungal Agents

Clinical practice guidelines recommend performing follow-up cultures for patients with candidemia in order to determine the time when Candida is cleared from the bloodstream. Since this requires culturing blood samples from patients undergoing antifungal treatment, we evaluated two blood culture bottles (the Bactec Mycosis IC/F [MICF], specifically adapted to the growth of fungi, and the Bactec Plus Aerobic/F [PAF], containing resins to inactivate anti-infective agents) for their effectiveness in detecting Candida albicans and Candida glabrata when seeded in concentrations of 1 CFU/ml and 10 CFU/ml, respectively, together with human whole blood and various antifungal agents in therapeutic peak serum concentrations (C_max). Significant differences between the MICF and PAF vials for the detection of Candida spp. were found when inoculated with caspofungin (0/12 versus 8/12) (P < 0.001) or amphotericin B (3/12 versus 12/12) (P < 0.001). Inoculation of fluconazole or voriconazole did not influence the effectiveness of detection in the MICF and PAF bottles (P = 1.0). Neither the MICF nor the PAF bottles detected Candida spp. reliably when seeded together with anidulafungin (1/12 versus 1/12) (P = 1.0) or micafungin (0/12 versus 1/12) (P = 1.0). The times to positivity of both bottles were significantly prolonged when antifungal agents were added compared to those of controls without antimycotic drugs (P < 0.001). Overall, the results of this in vitro study indicate that the PAF bottles detected Candida spp. more reliably than the MICF bottles when supplemented with certain antifungal agents. Consequently, clinical studies should evaluate whether this holds true when blood cultures from patients undergoing antifungal treatment are performed.     

Neural modelling of the semantic predictability gain under challenging listening conditions

When speech intelligibility is reduced, listeners exploit constraints posed by semantic context to facilitate comprehension. The left angular gyrus (AG) has been argued to drive this semantic predictability gain. Taking a network perspective, we ask how the connectivity within language‐specific and domain‐general networks flexibly adapts to the predictability and intelligibility of speech. During continuous functional magnetic resonance imaging (fMRI), participants repeated sentences, which varied in semantic predictability of the final word and in acoustic intelligibility. At the neural level, highly predictable sentences led to stronger activation of left‐hemispheric semantic regions including subregions of the AG (PGa, PGp) and posterior middle temporal gyrus when speech became more intelligible. The behavioural predictability gain of single participants mapped onto the same regions but was complemented by increased activity in frontal and medial regions. Effective connectivity from PGa to PGp increased for more intelligible sentences. In contrast, inhibitory influence from pre‐supplementary motor area to left insula was strongest when predictability and intelligibility of sentences were either lowest or highest. This interactive effect was negatively correlated with the behavioural predictability gain. Together, these results suggest that successful comprehension in noisy listening conditions relies on an interplay of semantic regions and concurrent inhibition of cognitive control regions when semantic cues are available.     

Rare Pathogenic Variants in Mitochondrial and Inflammation-Associated Genes May Lead to Inflammatory Cardiomyopathy in Chagas Disease

Abstract

Cardiomyopathies are an important cause of heart failure and sudden cardiac death. Little is known about the role of rare genetic variants in inflammatory cardiomyopathy. Chronic Chagas disease cardiomyopathy (CCC) is an inflammatory cardiomyopathy prevalent in Latin America, developing in 30% of the 6 million patients chronically infected by the protozoan Trypanosoma cruzi, while 60% remain free of heart disease (asymptomatic (ASY)). The cytokine interferon-γ and mitochondrial dysfunction are known to play a major pathogenetic role. Chagas disease provides a unique model to probe for genetic variants involved in inflammatory cardiomyopathy.

Methods

We used whole exome sequencing to study nuclear families containing multiple cases of Chagas disease. We searched for rare pathogenic variants shared by all family members with CCC but absent in infected ASY siblings and in unrelated ASY.

Results

We identified heterozygous, pathogenic variants linked to CCC in all tested families on 22 distinct genes, from which 20 were mitochondrial or inflammation-related – most of the latter involved in proinflammatory cytokine production. Significantly, incubation with IFN-γ on a human cardiomyocyte line treated with an inhibitor of dihydroorotate dehydrogenase brequinar (enzyme showing a loss-of-function variant in one family) markedly reduced mitochondrial membrane potential (ΔψM), indicating mitochondrial dysfunction.

Conclusion

Mitochondrial dysfunction and inflammation may be genetically determined in CCC, driven by rare genetic variants. We hypothesize that CCC-linked genetic variants increase mitochondrial susceptibility to IFN-γ-induced damage in the myocardium, leading to the cardiomyopathy phenotype in Chagas disease. This mechanism may also be operative in other inflammatory cardiomyopathies.

     

A novel temporal‐predominant neuro‐astroglial tauopathy associated with TMEM106B gene polymorphism in FTLD/ALS‐TDP

Polymorphisms in TMEM106B, a gene on chromosome 7p21.3 involved in lysosomal trafficking, correlates to worse neuropathological, and clinical outcomes in frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS) with TDP‐43 inclusions. In a small cohort of C9orf72 expansion carriers, we previously found an atypical, neuroglial tauopathy in cases harboring a TMEM106B rs1990622 A/A genotype. To test whether TMEM106B genotype affects the risk of developing atypical tauopathy under a recessive genotype model (presence versus absence of two major alleles: A/A vs. A/G and G/G). We characterized the atypical tauopathy neuropathologically and determined its frequency by TMEM106B rs1990622 genotypes in 90 postmortem cases with a primary diagnosis of FTLD/ALS‐TDP [mean age at death 65.5 years (±8.1), 40% female]. We investigated the effect of this new atypical tauopathy on demographics and clinical and neuropsychological metrics. We also genotyped TMEM106B in an independent series with phenotypically similar cases. Sixteen cases (16/90, 17.7 %) showed the temporal‐predominant neuro‐astroglial tauopathy, and 93.7% of them carried an A/A genotype (vs. ~35% in a population cohort). The odds ratio of FTLD/ALS‐TDP individuals with the A/A genotype showing neuro‐astroglial tauopathy was 13.9. Individuals with this tauopathy were older at onset (p = 0.01). The validation cohort had a similarly high proportion of rs1990622 A/A genotype. TDP‐43 and tau changes co‐occur in a subset of neurons. Our data add to the growing body of evidence that TMEM106B polymorphisms may modulate neurodegeneration. A distinctive medial temporal predominant, 4‐repeat, neuro‐astroglial tauopathy strongly correlates to TMEM106B A/A genotype in FTLD/ALS‐TDP cases.     

Disturbed gastric and small bowel transit in severe idiopathic constipation

Many patients with severe idiopathic constipation complain of upper gastrointestinal symptoms, and these often persist after subtotal colectomy. To determine if there is a disturbance of upper gastrointestinal motility in this condition, we have studied gastric emptying for solids (¹¹¹In-containing pancake) and liquids (^99mTc-containing orange, juice) for a longer period after a meal (6 hr) than in previously reported gastric emptying studies. Small bowel transit for solids was also measured. All patients had emptied their colon the day before the study. Twelve women (mean age 36 years) with a bowel frequency of less than once per week, proven slow intestinal transit, and a normal diameter colon were studied. Twelve healthy controls (eight female and four male, mean age 33) were also studied. As a group the constipated patients demonstrated no statistically significant delay in emptying during the first 3 hr, although the emptying rate for three of 12 individuals fell outside the normal range. However, at 6 hr after ingestion of the meal, six of 10 patients had residual gastric contents greater than normal-up to 48% solid residue (median: 11% for patients and 0% for controls,P<0.01) and 40% of liquid (median 9% vs 0%P<0.01). Three of four patients with upper gastrointestinal symptoms 6 hr after the meal had gastric retention of solids markedly outside the normal range (48%, 32%, and 16%; normal<4%). Small bowel transit time was assessed as the time for the solid phase to pass from the duodenum to the cecum; the constipated patients demonstrated delayed transit (median: 75 vs 55 min,P<0.01). Effectiveness of small bowel transit was assessed by the proportion of solids in the cecum at the time the stomach had emptied 50% of the solid meal; this was reduced in the patients (median: 6 vs 18%,P<0.01). All patients with normal gastric emptying had normal small bowel transit, and all those with delayed gastric emptying had prolonged small bowel transit. Colonic transit of the radioisotope was slow in all patients (head of the radioisotope column, cecum to stool, median: 96 vs 31 hr,P<0.01). Many patients with severe idiopathic constipation have a disturbance of gastric and small bowel transit that may be related to symptoms and that have implications for treatment.