Serial isoelectric focusing as an effective and economic way to obtain maximal resolution and high-throughput in 2D-based comparative proteomics of scarce samples: proof-of-principle

In 2D-based comparative proteomics of scarce samples, such as limited patient material, established methods for prefractionation and subsequent use of different narrow range IPG strips to increase overall resolution are difficult to apply. Also, a high number of samples, a prerequisite for drawing meaningful conclusions when pathological and control samples are considered, will increase the associated amount of work almost exponentially. Here, we introduce a novel, effective, and economic method designed to obtain maximum 2D resolution while maintaining the high throughput necessary to perform large-scale comparative proteomics studies. The method is based on connecting different IPG strips serially head-to-tail so that a complete line of different IPG strips with sequential pH regions can be focused in the same experiment. We show that when 3 IPG strips (covering together the pH range of 3-11) are connected head-to-tail an optimal resolution is achieved along the whole pH range. Sample consumption, time required, and associated costs are reduced by almost 70%, and the workload is reduced significantly.     

A meta-analysis of seaweed impacts on seagrasses: generalities and knowledge gaps

Seagrasses are important habitat-formers and ecosystem engineers that are under threat from bloom-forming seaweeds. These seaweeds have been suggested to outcompete the seagrasses, particularly when facilitated by eutrophication, causing regime shifts where green meadows and clear waters are replaced with unstable sediments, turbid waters, hypoxia, and poor habitat conditions for fishes and invertebrates. Understanding the situations under which seaweeds impact seagrasses on local patch scales can help proactive management and prevent losses at greater scales. Here, we provide a quantitative review of available published manipulative experiments (all conducted at the patch-scale), to test which attributes of seaweeds and seagrasses (e.g., their abundances, sizes, morphology, taxonomy, attachment type, or origin) influence impacts. Weighted and unweighted meta-analyses (Hedges d metric) of 59 experiments showed generally high variability in attribute-impact relationships. Our main significant findings were that (a) abundant seaweeds had stronger negative impacts on seagrasses than sparse seaweeds, (b) unattached and epiphytic seaweeds had stronger impacts than 'rooted' seaweeds, and (c) small seagrass species were more susceptible than larger species. Findings (a) and (c) were rather intuitive. It was more surprising that 'rooted' seaweeds had comparatively small impacts, particularly given that this category included the infamous invasive Caulerpa species. This result may reflect that seaweed biomass and/or shading and metabolic by-products like anoxia and sulphides could be lower for rooted seaweeds. In conclusion, our results represent simple and robust first-order generalities about seaweed impacts on seagrasses. This review also documented a limited number of primary studies. We therefore identified major knowledge gaps that need to be addressed before general predictive models on seaweed-seagrass interactions can be build, in order to effectively protect seagrass habitats from detrimental competition from seaweeds.     

Correlation analysis of clinical parameters with epigenetic modifications in the DUX4 promoter in FSHD

The aim of our study was to identify relationships between epigenetic parameters correlating with a relaxed chromatin state of the DUX4 promoter region and clinical severity as measured by a clinical severity score or muscle pathologic changes in D4Z4 contraction-dependent (FSHD1) and –independent (FSHD2) facioscapulohumeral muscular dystrophy patients. Twenty primary fibroblast (5 control, 10 FSHD1 and 5 FSHD2) and 26 primary myoblast (9 control, 12 FSHD1 and 5 FSHD2) cultures originating from patients with FSHD and controls were analyzed. Histone modification levels were determined by chromatin immunoprecipitation. We examined correlations between the chromatin compaction score (ChCS) defined by the H3K9me3:H3K4me2 ratio and an age corrected clinical severity score (CSS) or muscle pathology score (MPS). Possible relationships were investigated using linear regression analysis and significance was tested by Pearson’s product-moment coefficient.   We found a significant difference of the ChCS between controls and patients with FSHD1 and between controls and patients with FSHD2. Tissue specific differences in ChCS were also observed. We also found a near-significant relationship between ChCS and the age corrected CSS in fibroblasts but not in myoblasts. Surprisingly, we found a strong correlation between the MPS of the vastus lateralis and the CSS. Our results confirm the D4Z4 chromatin relaxation previously shown to be associated with FSHD in a small number of samples. A possible relationship between clinical and epigenetic parameters could be established in patient fibroblasts, but not in myoblasts. The strong correlation between the MPS of the vastus lateralis and the CSS suggests that this muscle can be used to study for surrogate markers of overall disease severity.     

Cytomegalovirus seropositivity is associated with glucose regulation in the oldest old. Results from the Leiden 85-plus Study

ABSTRACT: BACKGROUND: Cytomegalovirus (CMV) infection has been reported to contribute to the pathogenesis of type 1 diabetes and post-transplantation diabetes. However, CMV infection has not been evaluated as a possible risk factor for type 2 diabetes. Our aim was to investigate potential associations between CMV seropositivity, CMV IgG antibody level and glucose regulation in the oldest old. RESULTS: CMV seropositive subjects were more likely to have type 2 diabetes (17.2% vs 7.9%, p = 0.016), had a higher level of HbA1c (p = 0.014) and higher non-fasting glucose (p = 0.024) in the oldest olds. These associations remained significant after adjustment for possible confounders. CMV IgG antibody level was not significantly associated with glucose regulation (all p > 0.05). CONCLUSIONS: In the oldest old, CMV seropositivity is significantly associated with various indicators of glucose regulation. This finding suggests that CMV infection might be a risk factor for the development of type 2 diabetes in the elderly.     

Clinical features and predictors for disease natural progression in adults with Pompe disease: a nationwide prospective observational study

Background

Due partly to physicians’ unawareness, many adults with Pompe disease are diagnosed with great delay. Besides, it is not well known which factors influence the rate of disease progression, and thus disease outcome. We delineated the specific clinical features of Pompe disease in adults, and mapped out the distribution and severity of muscle weakness, and the sequence of involvement of the individual muscle groups. Furthermore, we defined the natural disease course and identified prognostic factors for disease progression.

Methods

We conducted a single-center, prospective, observational study. Muscle strength (manual muscle testing, and hand-held dynamometry), muscle function (quick motor function test), and pulmonary function (forced vital capacity in sitting and supine positions) were assessed every 3–6 months and analyzed using repeated-measures ANOVA.

Results

Between October 2004 and August 2009, 94 patients aged between 25 and 75 years were included in the study. Although skeletal muscle weakness was typically distributed in a limb-girdle pattern, many patients had unfamiliar features such as ptosis (23%), bulbar weakness (28%), and scapular winging (33%). During follow-up (average 1.6 years, range 0.5-4.2 years), skeletal muscle strength deteriorated significantly (mean declines of ?1.3% point/year for manual muscle testing and of ?2.6% points/year for hand-held dynamometry; both p<0.001). Longer disease duration (>15 years) and pulmonary involvement (forced vital capacity in sitting position <80%) at study entry predicted faster decline. On average, forced vital capacity in supine position deteriorated by 1.3% points per year (p=0.02). Decline in pulmonary function was consistent across subgroups. Ten percent of patients declined unexpectedly fast.

Conclusions

Recognizing patterns of common and less familiar characteristics in adults with Pompe disease facilitates timely diagnosis. Longer disease duration and reduced pulmonary function stand out as predictors of rapid disease progression, and aid in deciding whether to initiate enzyme replacement therapy, or when.

     

Neuromelanin can protect against iron-mediated oxidative damage in system modeling iron overload of brain aging and Parkinson's disease

In Parkinson's disease (PD), dopamine neurons containing neuromelanin selectively degenerate. Neuromelanin binds iron and accumulates in aging. Iron accumulates in reactive form during aging, PD, and is involved in neurodegeneration. It is not clear how the interaction of neuromelanin and iron can be protective or toxic by modulating redox processes. Here, we investigated the interaction of neuromelanin from human substantia nigra with iron in the presence of ascorbic acid, dopamine, and hydrogen peroxide. We observed that neuromelanin blocks hydroxyl radical production by Fenton's reaction, in a dose-dependent manner. Neuromelanin also inhibited the iron-mediated oxidation of ascorbic acid, thus sparing this major antioxidant molecule in brain. The protective effect of neuromelanin on ascorbate oxidation occurs even in conditions of iron overload into neuromelanin. The blockade of iron into a stable iron–neuromelanin complex prevents dopamine oxidation, inhibiting the formation of neurotoxic dopamine quinones. The above processes occur intraneuronally in aging and PD, thus showing that neuromelanin is neuroprotective. The iron–neuromelanin complex is completely decomposed by hydrogen peroxide and its degradation rate increases with the amount of iron bound to neuromelanin. This occurs in PD when extraneuronal iron–neuromelanin is phagocytosed by microglia and iron–neuromelanin degradation releases reactive/toxic iron.     

Phenotypic variability of the cat eye syndrome. Case report and review of the literature.

We present a male infant with preauricular skin tags and pits, downslanting palpebral fissures, hypertelorism, ectopic anus, hypospadias, and hypoplastic left heart syndrome. The clinical features in our patient show phenotypic overlap with the cat eye syndrome, as illustrated by the review of 105 reported cases. Cytogenetic analysis revealed a supernumerary marker chromosome, which was identified by microdissection and fluorescence in situ hybridization as an isodicentric chromosome 22(pter --> q11.2::q11.2 --> pter). It was proved with probes specific for the cat eye syndrome critical region that this region was present in quadruplicate in the propositus. We conclude that CES is characterized by large phenotypic variability, ranging from near normal to severe malformations, as reflected in the neurodevelopmental outcome. Preauricular skin tags and/or pits are the most consistent features, and suggest the presence of a supernumerary bisatellited marker chromosome 22 derived from duplication of the CES critical region.