Thalidomide induces limb defects by preventing angiogenic outgrowth during early limb formation.

Thalidomide is a potent teratogen that induces a range of birth defects, most commonly of the developing limbs. The mechanisms underpinning the teratogenic effects of thalidomide are unclear. Here we demonstrate that loss of immature blood vessels is the primary cause of thalidomide-induced teratogenesis and provide an explanation for its action at the cell biological level. Antiangiogenic but not antiinflammatory metabolites/analogues of thalidomide induce chick limb defects. Both in vitro and in vivo, outgrowth and remodeling of more mature blood vessels is blocked temporarily, whereas newly formed, rapidly developing, angiogenic vessels are lost. Such vessel loss occurs upstream of changes in limb morphogenesis and gene expression and, depending on the timing of drug application, results in either embryonic death or developmental defects. These results explain both the timing and relative tissue specificity of thalidomide embryopathy and have significant implications for its use as a therapeutic agent.

Zebrafish notch signalling pathway mutants exhibit trunk vessel patterning anomalies that are secondary to somite misregulation.

The Notch signalling pathway mutants, after-eight (aei), beamter (bea), and deadly-seven (des) have previously been used to study somitogenesis and neurogenesis. Notch signalling has also been shown to have roles in vascular development. However, vascular development in each of these three Notch mutants has not been described, and so their potential usefulness for further understanding the role of Notch signalling in angiogenesis is unknown. Here we demonstrate each of the mutants also exhibit vascular defects in inter-somitic vessel (ISV) positioning and patterning. Ectopic filopodia were also observed on the ISVs of the mutants. Ectopic filopodia are not due to loss of dll4. Somite expression of known vascular guidance cues, efnb2, sema3a2, and plexinD1 are disrupted, suggesting that the ISV vascular phenotype is due to disruption of these cues.

Approaches and challenges in the diagnosis and management of secondary progressive multiple sclerosis: A Central Eastern European perspective from healthcare professionals.

Secondary progressive multiple sclerosis (SPMS) is a debilitating condition characterized by gradual worsening after an initial relapsing disease course. Despite the recent advances in our understanding of the disease, the diagnosis and treatment of SPMS continue to be challenging in routine clinical practice. The aim of this review article is to present the views of leading MS experts on the challenges in the diagnosis and management of SPMS and clinicians' perspectives in Central and Eastern Europe. This article also provides recommendations of MS experts to improve the situation with diagnosis and management of SPMS. Many countries within Central and Eastern Europe have high prevalence of MS (>100 per 100,000 population). Consistent with the global trend, in the absence of reliable tests or biomarkers, SPMS at early stage remains undiagnosed. Due to diagnostic uncertainty and lack of a universally accepted disease definition, clinicians rely more on retrospective analysis of the clinical symptoms to confirm the diagnosis. With the lack of awareness and poor understanding of the timing of the onset of SPMS, clinicians may tend to direct attention to relapses than the symptoms of progression, which leads to underestimation of SPMS. Although several predictors of progression to SPMS have been identified, their predictive value is highly variable. Therefore, defining the transitioning period as a separate stage of MS is essential. According to experts' opinion, frequent follow-up of patients and periodic assessment of progression are recommended for the timely identification of patients transitioning from RRMS to SPMS. MSProDiscuss Tool is an example of a quick assessment tool for identifying patients progressing from RRMS to SPMS. MS progression is usually assessed by changes in Expanded Disability Status Scale (EDSS) scores. As EDSS scores tend to fluctuate when measured in the short term (3-6 months), a longer period (≥12 months) may be needed to confirm the progression. Assessment of cognitive function is also important for evaluating secondary progression. Compartmentalization of inflammation within the central nervous system is an important reason behind the limited success of disease-modifying therapies (DMTs) for treating SPMS. Most of the DMTs fail to cross the blood-brain barrier; only 38% of the tested DMTs achieved their primary endpoint in SPMS. In Europe, siponimod is the first oral treatment for adults with active SPMS. Particularly, in Central and Eastern Europe, patients with SPMS are still being prescribed less efficacious DMTs and interferons. The absence of alternative treatments in SPMS supports the use of new products (siponimod and others); however the decision to initiate siponimod therapy in more severe patients (EDSS score of 7 or higher) should be individualized in consultation with the payers. The focus should be on early treatment initiation to delay disease progression.

Functional consequences of cysteine modification in the ligand binding sites of peroxisome proliferator activated receptors by GW9662.

In the course of a high throughput screen to search for ligands of peroxisome proliferator activated receptor-gamma (PPARgamma), we identified GW9662 using a competition binding assay against the human ligand binding domain. GW9662 had nanomolar IC(50) versus PPARgamma and was 10- and 600-fold less potent in binding experiments using PPARalpha and PPARdelta, respectively. Pretreatment of all three PPARs with GW9662 resulted in the irreversible loss of ligand binding as assessed by scintillation proximity assay. Incubation of PPAR with GW9662 resulted in a change in the absorbance spectra of the receptors consistent with covalent modification. Mass spectrometric analysis of the PPARgamma ligand binding domain treated with GW9662 established Cys(285) as the site of covalent modification. This cysteine is conserved among all three PPARs. In cell-based reporter assays, GW9662 was a potent and selective antagonist of full-length PPARgamma. The functional activity of GW9662 as an antagonist of PPARgamma was confirmed in an assay of adipocyte differentiation. GW9662 showed essentially no effect on transcription when tested using both full-length PPARdelta and PPARalpha. Time-resolved fluorescence assays of ligand-modulated receptor heterodimerization, coactivator binding, and corepressor binding were consistent with the effects observed in the reporter gene assays. Control activators increased PPAR:RXR heterodimer formation and coactivator binding to both PPARgamma and PPARdelta. Corepressor binding was decreased. In the case of PPARalpha, GW9662 treatment did not significantly increase heterodimerization and coactivator binding or decrease corepressor binding. The experimental data indicate that GW9662 modification of each of the three PPARs results in different functional consequences. The selective and irreversible nature of GW9662 treatment, and the observation that activity is maintained in cell culture experiments, suggests that this compound may be a useful tool for elucidation of the role of PPARgamma in biological processes.