Expressional changes in cerebrovascular receptors after experimental transient forebrain ischemia

Background

Global ischemic stroke is one of the most prominent consequences of cardiac arrest, since the diminished blood flow to the brain results in cell damage and sometimes permanently impaired neurological function. The post-arrest period is often characterised by cerebral hypoperfusion due to subacute hemodynamic disturbances, the pathophysiology of which are poorly understood. In two other types of stroke, focal ischemic stroke and subarachnoid hemorrhage, it has earlier been demonstrated that the expression of certain vasoconstrictor receptors is increased in cerebral arteries several days after the insult, a phenomenon that leads to increased contraction of cerebral arteries, reduced perfusion of the affected area and worsened ischemic damage. Based on these findings, the aim of the present study was to investigate if transient global cerebral ischemia is associated with upregulation of vasoconstrictive endothelin and 5-hydroxytryptamine receptors in cerebral arteries. Experimental transient forebrain ischemia of varying durations was induced in male wistar rats, followed by reperfusion for 48 hours. Neurological function was assessed daily by three different tests and cerebrovascular expression and contractile function of endothelin and 5-hydroxytryptamine receptors were evaluated by wire myography, immunohistochemistry and western blotting.

Results

Transient forebrain ischemia induced neurological deficits as well as functional upregulation of vasoconstrictive ET_B and 5-HT_1B receptors in cerebral arteries supplying mid- and forebrain regions. No receptor upregulation was seen in arteries supplying the hindbrain. Immunohistochemical stainings and western blotting demonstrated expressional upregulation of these receptor subtypes in the mid- and forebrain arteries and confirmed that the receptors were located in the smooth muscle layer of the cerebral arteries.

Conclusions

This study reveals a new pathophysiological aspect of global ischemic stroke, namely expressional upregulation of vasoconstrictor receptors in cerebral arteries two days after the insult, which might contribute to cerebral hypoperfusion and delayed neuronal damage after cardiac arrest.     

A Prospective Population Study of Resting Heart Rate and Peak Oxygen Uptake (the HUNT Study,Norway)

Objectives

We assessed the prospective association of resting heart rate (RHR) at baseline with peak oxygen uptake (VO_2peak) 23 years later, and evaluated whether physical activity (PA) could modify this association.

Background

Both RHR and VO_2peak are strong and independent predictors of cardiovascular morbidity and mortality. However, the association of RHR with VO_2peak and modifying effect of PA have not been prospectively assessed in population studies.

Methods

In 807 men and 810 women free from cardiovascular disease both at baseline (1984–86) and follow-up 23 years later, RHR was recorded at both occasions, and VO_2peak was measured by ergospirometry at follow-up. We used Generalized Linear Models to assess the association of baseline RHR with VO_2peak, and to study combined effects of RHR and self-reported PA on later VO_2peak.

Results

There was an inverse association of RHR at baseline with VO_2peak (p<0.01). Men and women with baseline RHR greater than 80 bpm had 4.6 mL·kg⁻¹·min⁻¹ (95% confidence interval [CI], 2.8 to 6.3) and 1.4 mL·kg⁻¹·min⁻¹ (95% CI, −0.4 to 3.1) lower VO_2peak at follow-up compared with men and women with RHR below 60 bpm at baseline. We found a linear association of change in RHR with VO_2peak (p = 0.03), suggesting that a decrease in RHR over time is likely to be beneficial for cardiovascular fitness. Participants with low RHR and high PA at baseline had higher VO_2peak than inactive people with relatively high RHR. However, among participants with relatively high RHR and high PA at baseline, VO_2peak was similar to inactive people with relatively low RHR.

Conclusion

RHR is an important predictor of VO_2peak, and serial assessments of RHR may provide useful and inexpensive information on cardiovascular fitness. The results suggest that high levels of PA may compensate for the lower VO_2peak associated with a high RHR.     

Hepcidin-25 in chronic hemodialysis patients is related to residual kidney function and not to treatment with erythropoiesis stimulating agents

Hepcidin-25, the bioactive form of hepcidin, is a key regulator of iron homeostasis as it induces internalization and degradation of ferroportin, a cellular iron exporter on enterocytes, macrophages and hepatocytes. Hepcidin levels are increased in chronic hemodialysis (HD) patients, but as of yet, limited information on factors associated with hepcidin-25 in these patients is available. In the current cross-sectional study, potential patient-, laboratory- and treatment-related determinants of serum hepcidin-20 and -25, were assessed in a large cohort of stable, prevalent HD patients. Baseline data from 405 patients (62% male; age 63.7 ± 13.9 [mean SD]) enrolled in the CONvective TRAnsport STudy (CONTRAST; NCT00205556) were studied. Predialysis hepcidin concentrations were measured centrally with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Patient-, laboratory- and treatment related characteristics were entered in a backward multivariable linear regression model. Hepcidin-25 levels were independently and positively associated with ferritin (p<0.001), hsCRP (p<0.001) and the presence of diabetes (p = 0.02) and inversely with the estimated glomerular filtration rate (p = 0.01), absolute reticulocyte count (p = 0.02) and soluble transferrin receptor (p<0.001). Men had lower hepcidin-25 levels as compared to women (p = 0.03). Hepcidin-25 was not associated with the maintenance dose of erythropoiesis stimulating agents (ESA) or iron therapy. In conclusion, in the currently studied cohort of chronic HD patients, hepcidin-25 was a marker for iron stores and erythropoiesis and was associated with inflammation. Furthermore, hepcidin-25 levels were influenced by residual kidney function. Hepcidin-25 did not reflect ESA or iron dose in chronic stable HD patients on maintenance therapy. These results suggest that hepcidin is involved in the pathophysiological pathway of renal anemia and iron availability in these patients, but challenges its function as a clinical parameter for ESA resistance.     

Cellular effects of HER3-specific affibody molecules

Recent studies have led to the recognition of the epidermal growth factor receptor HER3 as a key player in cancer, and consequently this receptor has gained increased interest as a target for cancer therapy. We have previously generated several Affibody molecules with subnanomolar affinity for the HER3 receptor. Here, we investigate the effects of two of these HER3-specific Affibody molecules, Z05416 and Z05417, on different HER3-overexpressing cancer cell lines. Using flow cytometry and confocal microscopy, the Affibody molecules were shown to bind to HER3 on three different cell lines. Furthermore, the receptor binding of the natural ligand heregulin (HRG) was blocked by addition of Affibody molecules. In addition, both molecules suppressed HRG-induced HER3 and HER2 phosphorylation in MCF-7 cells, as well as HER3 phosphorylation in constantly HER2-activated SKBR-3 cells. Importantly, Western blot analysis also revealed that HRG-induced downstream signalling through the Ras-MAPK pathway as well as the PI3K-Akt pathway was blocked by the Affibody molecules. Finally, in an in vitro proliferation assay, the two Affibody molecules demonstrated complete inhibition of HRG-induced cancer cell growth. Taken together, our findings demonstrate that Z05416 and Z05417 exert an anti-proliferative effect on two breast cancer cell lines by inhibiting HRG-induced phosphorylation of HER3, suggesting that the Affibody molecules are promising candidates for future HER3-targeted cancer therapy.     

Job strain and tobacco smoking: an individual-participant data meta-analysis of 166,130 adults in 15 European studies

Background

Tobacco smoking is a major contributor to the public health burden and healthcare costs worldwide, but the determinants of smoking behaviours are poorly understood. We conducted a large individual-participant meta-analysis to examine the extent to which work-related stress, operationalised as job strain, is associated with tobacco smoking in working adults.

Methodology and Principal Findings

We analysed cross-sectional data from 15 European studies comprising 166 130 participants. Longitudinal data from six studies were used. Job strain and smoking were self-reported. Smoking was harmonised into three categories never, ex- and current. We modelled the cross-sectional associations using logistic regression and the results pooled in random effects meta-analyses. Mixed effects logistic regression was used to examine longitudinal associations. Of the 166 130 participants, 17% reported job strain, 42% were never smokers, 33% ex-smokers and 25% current smokers. In the analyses of the cross-sectional data, current smokers had higher odds of job strain than never-smokers (age, sex and socioeconomic position-adjusted odds ratio: 1.11, 95% confidence interval: 1.03, 1.18). Current smokers with job strain smoked, on average, three cigarettes per week more than current smokers without job strain. In the analyses of longitudinal data (1 to 9 years of follow-up), there was no clear evidence for longitudinal associations between job strain and taking up or quitting smoking.

Conclusions

Our findings show that smokers are slightly more likely than non-smokers to report work-related stress. In addition, smokers who reported work stress smoked, on average, slightly more cigarettes than stress-free smokers.     

Pharmacological undertreatment of coronary risk factors in patients with psoriasis: observational study of the Danish nationwide registries

Background

Patients with psoriasis have increased prevalence of coronary risk factors and limited recent results have suggested that these risk factors are undertreated in patients with psoriasis. This may contribute to the increased risk of cardiovascular diseases observed in patients with psoriasis.

Objective

To examine the pharmacological treatment of coronary risk factors in patients with severe psoriasis treated with biologic agents in a real-world setting.

Methods and Findings

Medical history of patients with severe psoriasis treated with biologic agents in the time period 2007–09 was retrieved from a Danish nationwide registry (DERMBIO). Individual-level linkage of nationwide administrative registries of hospitalizations, concomitant medications, and socioeconomic status was performed to gain insights into the use of pharmacological treatment. A total of 693 patients (mean age 46.1±12.7 years, 65.7% male) with severe psoriasis treated with biologic agents were identified. Hypertension, hypercholesterolemia, and diabetes mellitus were identified in 16.6%, 9.2%, and 6.7% of cases, respectively. Patients with severe psoriasis were significantly less likely to receive cardiovascular pharmacotherapy compared to age, sex, and coronary risk factor matched controls. In psoriatic patients with hypertension 27.7% received no antihypertensive pharmacotherapy. Patients with dyslipidemia received cholesterol-lowering medications in 55.8% of cases and patients with diabetes mellitus received angiotensin converting enzyme inhibitors/angiotensin II receptor blockers and cholesterol-lowering medications in 42.1% and 23.7% of cases, respectively. Similar results were found for the subset of patients with >1 coronary risk factor and for high risk patients with established atherosclerotic disease.

Conclusion

This nationwide study of patients with severe psoriasis demonstrated substantial undertreatment of coronary risk factors. Increased focus on identifying cardiovascular risk factors and initiation of preventive cardiovascular pharmacotherapy in patients with psoriasis is warranted.     

The Density of Knobs on Plasmodium falciparum-Infected Erythrocytes Depends on Developmental Age and Varies among Isolates

Background

The virulence of Plasmodium falciparum malaria is related to the parasite’s ability to evade host immunity through clonal antigenic variation and tissue-specific adhesion of infected erythrocytes (IEs). The P. falciparum erythrocyte membrane protein 1 (PfEMP1) family expressed on dome-shaped protrusions called knobs on the IE surface is central to both. Differences in receptor specificity and affinity of expressed PfEMP1 are important for IE adhesiveness, but it is not known whether differences in the number and size of the knobs on which the PfEMP1 proteins are expressed also play a role. Therefore, the aim of this study was to provide detailed information on isolate- and time-dependent differences in knob size and density.

Methodology/Principal Findings

We used atomic force microscopy to characterize knobs on the surface of P. falciparum-infected erythrocytes. Fourteen ex vivo isolates from Ghanaian children with malaria and 10 P. falciparum isolates selected in vitro for expression of a particular PfEMP1 protein (VAR2CSA) were examined. Knob density increased from ∼20 h to ∼35 h post-invasion, with significant variation among isolates. The knob density ex vivo, which was about five-fold higher than following long-term in vitro culture, started to decline within a few months of culture. Although knob diameter and height varied among isolates, we did not observe significant time-dependent variation in these dimensions.

Conclusions/Significance

The density of knobs on the P. falciparum-IE surface depends on time since invasion, but is also determined by the infecting isolate in a time-independent manner. This is the first study to quantitatively evaluate knob densities and dimensions on different P. falciparum isolates, to examine ex vivo isolates from humans, and to compare ex vivo and long-term in vitro-cultured isolates. Our findings contribute to the understanding of the interaction between P. falciparum parasites and the infected host.     

The Peripheral Binding of 14-3-3γ to Membranes Involves Isoform-Specific Histidine Residues

Mammalian 14-3-3 protein scaffolds include seven conserved isoforms that bind numerous phosphorylated protein partners and regulate many cellular processes. Some 14-3-3-isoforms, notably γ, have elevated affinity for membranes, which might contribute to modulate the subcellular localization of the partners and substantiate the importance of investigating molecular mechanisms of membrane interaction. By applying surface plasmon resonance we here show that the binding to phospholipid bilayers is stimulated when 14-3-3γ is complexed with its partner, a peptide corresponding to the Ser19-phosphorylated N-terminal region of tyrosine hydroxylase. Moreover, membrane interaction is dependent on salts of kosmotropic ions, which also stabilize 14-3-3γ. Electrostatic analysis of available crystal structures of γ and of the non-membrane-binding ζ-isoform, complemented with molecular dynamics simulations, indicate that the electrostatic potential distribution of phosphopeptide-bound 14-3-3γ is optimal for interaction with the membrane through amphipathic helices at the N-terminal dimerization region. In addition, His158, and especially His195, both specific to 14-3-3γ and located at the convex lateral side, appeared to be pivotal for the ligand induced membrane interaction, as corroborated by site-directed mutagenesis. The participation of these histidine residues might be associated to their increased protonation upon membrane binding. Overall, these results reveal membrane-targeting motifs and give insights on mechanisms that furnish the 14-3-3γ scaffold with the capacity for tuned shuffling from soluble to membrane-bound states.     

Persurf,a New Method to Improve Surfactant Delivery: A Study in Surfactant Depleted Rats

Purpose

Exogenous surfactant is not very effective in adults with ARDS, since surfactant does not reach atelectatic alveoli. Perfluorocarbons (PFC) can recruit atelectatic areas but do not replace impaired endogenous surfactant. A surfactant-PFC-mixture could combine benefits of both therapies. The aim of the proof-of-principal-study was to produce a PFC-in-surfactant emulsion (Persurf) and to test in surfactant depleted Wistar rats whether Persurf achieves I.) a more homogenous pulmonary distribution and II.) a more homogenous recruitment of alveoli when compared with surfactant or PFC alone.

Methods

Three different PFC were mixed with surfactant and phospholipid concentration in the emulsion was measured. After surfactant depletion, animals either received 30 ml/kg of PF5080, 100 mg/kg of stained (green dye) Curosurf™ or 30 ml/kg of Persurf. Lungs were fixated after 1 hour of ventilation and alveolar aeration and surfactant distribution was estimated by a stereological approach.

Results

Persurf contained 3 mg/ml phospholipids and was stable for more than 48 hours. Persurf-administration improved oxygenation. Histological evaluation revealed a more homogenous surfactant distribution and alveolar inflation when compared with surfactant treated animals.

Conclusions

In surfactant depleted rats administration of PFC-in-surfactant emulsion leads to a more homogenous distribution and aeration of the lung than surfactant alone.