Polar body diagnosis for hemophilia a using multiplex PCR for linked polymorphic markers.

Preimplantation genetic diagnosis (PGD) is usually performed on blastomeres. In Germany, the only possibility to perform PGD is by analysis of polar bodies. We performed PGD using polar bodies in a woman who is a carrier of hemophilia A. Multiplex PCR followed by nested fluorescent PCR for five linked polymorphic markers was established. From 11 analyzed polar bodies, only 1 showed alleles linked to the mutation. The corresponding oocyte was transferred and no pregnancy was established. As seen in other investigations, the rate of heterozygous first polar bodies is surprisingly high.     

Developmental origin of smooth muscle cells in the descending aorta in mice

Aortic smooth muscle cells (SMCs) have been proposed to derive from lateral plate mesoderm. It has further been suggested that induction of SMC differentiation is confined to the ventral side of the aorta, and that SMCs later migrate to the dorsal side. In this study, we investigate the origin of SMCs in the descending aorta using recombination-based lineage tracing in mice. Hoxb6-cre transgenic mice were crossed with Rosa 26 reporter mice to track cells of lateral plate mesoderm origin. The contribution of lateral plate mesoderm to SMCs in the descending aorta was determined at different stages of development. SMC differentiation was induced in lateral plate mesoderm-derived cells on the ventral side of the aorta at embryonic day (E) 9.0-9.5, as indicated by expression of the SMC-specific reporter gene SM22alpha-lacZ. There was, however, no migration of SMCs from the ventral to the dorsal side of the vessel. Moreover, the lateral plate mesoderm-derived cells in the ventral wall of the aorta were replaced by somite-derived cells at E10.5, as indicated by reporter gene expression in Meox1-cre/Rosa 26 double transgenic mice. Examination of reporter gene expression in adult aortas from Hoxb6-cre/Rosa 26 and Meox1-cre/Rosa 26 double transgenic mice suggested that all SMCs in the adult descending aorta derive from the somites, whereas no contribution was recorded from lateral plate mesoderm.     

Chiisanoside is not absorbed but inhibits oil absorption in the small intestine of rodents

Chiisanoside is the main component of Acanthopanax sessiliflorus leaves. Simultaneous administration of chiisanoside resulted in a decrease in the plasma TG level and increase of undigested TG in the intestinal lumen after oil gavage to mice. This suggests that chiisanoside has the potential to prevent obesity as a lipase inhibitor which suppresses fat absorption in vivo.     

Novel insights into systemic autoimmune rheumatic diseases using shared molecular signatures and an integrative analysis

We undertook this study to identify DNA methylation signatures of three systemic autoimmune rheumatic diseases (SARDs), namely rheumatoid arthritis, systemic lupus erythematosus, and systemic sclerosis, compared to healthy controls. Using a careful design to minimize confounding, we restricted our study to subjects with incident disease and performed our analyses on purified CD4⁺ T cells, key effector cells in SARD. We identified differentially methylated (using the Illumina Infinium HumanMethylation450 BeadChip array) and expressed (using the Illumina TruSeq stranded RNA-seq protocol) sites between cases and controls, and investigated the biological significance of this SARD signature using gene annotation databases. We recruited 13 seropositive rheumatoid arthritis, 19 systemic sclerosis, 12 systemic lupus erythematosus subjects, and 8 healthy controls. We identified 33 genes that were both differentially methylated and expressed (26 over- and 7 under-expressed) in SARD cases versus controls. The most highly overexpressed gene was CD1C (log fold change in expression = 1.85, adjusted P value = 0.009). In functional analysis (Ingenuity Pathway Analysis), the top network identified was lipid metabolism, molecular transport, small molecule biochemistry. The top canonical pathways included the mitochondrial L-carnitine shuttle pathway (P = 5E-03) and PTEN signaling (P = 8E-03). The top upstream regulator was HNF4A (P = 3E-05). This novel SARD signature contributes to ongoing work to further our understanding of the molecular mechanisms underlying SARD and provides novel targets of interest.     

Chemical Variability of Ivoirian Xylopia rubescens Leaf Oil

Forty‐two essential oil samples were isolated from leaves of Xylopia rubescens harvested in three forests of Southern Ivory Coast. All the samples have been submitted to GC‐FID and the retention indices (RIs) of individual components have been measured on two capillary columns of different polarity. In addition, 20 oil samples, selected on the basis of their chromatographic profile, were also analyzed by ¹³C‐NMR and 24 components (78.0 – 92.4% of the whole compositions) have been identified. The content of the main components varied drastically from sample to sample: furanoguaia‐1,4‐diene (5.7 – 54.1%), furanoguaia‐1,3‐diene (1.1 – 10.5%), (8Z,11Z,14Z)‐heptadeca‐8,11,14‐trien‐2‐one (4.3 – 16.0%), and (E)‐β‐caryophyllene (1.7 – 17.3%). Hierarchical cluster and principal components analysis of the 42 oil compositions allowed the distinction of two well‐differentiated groups of unequal importance within the oil samples. Oil samples of the main group (Group II) contained mainly furanoguaia‐1,4‐diene (mean [M] = 43.1%; standard deviation [SD] = 3.2%) while furanoguaia‐1,3‐diene (M = 8.4%; SD = 0.9%) and (8Z,11Z,14Z)‐heptadeca‐8,11,14‐trien‐2‐one (M = 7.1%; SD = 1.5%) were present at appreciable contents. The composition of Group I was dominated by furanoguaia‐1,4‐diene (M = 17.0%; SD = 8.5%), (8Z,11Z,14Z)‐heptadeca‐8,11,14‐trien‐2‐one (M = 10.2%; SD = 2.4%) and (E)‐β‐caryophyllene (M = 9.5%; SD = 5.3%).     

CAK-independent Activation of CDK6 by a Viral Cyclin

In normal cells, activation of cyclin-dependent kinases (cdks) requires binding to a cyclin and phosphorylation by the cdk-activating kinase (CAK). The Kaposi's sarcoma-associated herpesvirus encodes a protein with similarity to D-type cyclins. This KSHV-cyclin activates CDK6, alters its substrate specificity, and renders CDK6 insensitive to inhibition by the cdk inhibitor p16(INK4a). Here we investigate the regulation of the CDK6/KSHV-cyclin kinase with the use of purified proteins and a cell-based assay. We find that KSHV-cyclin can activate CDK6 independent of phosphorylation by CAK in vitro. In addition, CAK phosphorylation decreased the p16(INK4a) sensitivity of CDK6/KSHV-cyclin complexes. In cells, expression of CDK6 or to a lesser degree of a nonphosphorylatable CDK6(T177A) together with KSHV-cyclin induced apoptosis, indicating that CDK6 activation by KSHV-cyclin can proceed in the absence of phosphorylation by CAK in vivo. Coexpression of p16 partially protected cells from cell death. p16 and KSHV-cyclin can form a ternary complex with CDK6 that can be detected by binding assays as well as by conformational changes in CDK6. The Kaposi's sarcoma-associated herpesvirus has adopted a clever strategy to render cell cycle progression independent of mitogenic signals, cdk inhibition, or phosphorylation by CAK.     

Structural and conformational modifications of α-MSH/ACTH4–10 provide melanotropin analogues with highly potent behavioral activities

Previous studies have identified the (4–10) heptapeptide sequence as the central core of α-MSH/ACTH peptides required for mediation of important biological activities. In the present study, the structure-activity relationships of Nle⁴-substituted and -bridged cyclic α-MSH analogues, which were previously shown to exhibit a wide range of melanotropic potencies from weak agonism to super potency, were examined for grooming behavioral activity in the rat following intracerebroventricular injections. The results showed that stepwise C-terminal elongation of the linear Nle⁴-substituted Ac-α-MSH_4–10-NH₂ increased grooming potencies of the peptides in a manner similar to their actions on melanocytes. The most interesting finding was the observation that cyclization of the inactive linear “central (4–10) core” of α-MSH (Ac-α-MSH_4–10) to form Ac-[ ]-α-MSH_4–10-NH₂ resulted in a super potent agonist in the grooming assay. However, while cyclization of the (4–10) heptapeptide produced potent agonists on grooming behavior, the structure-activity relationships were different than the frog skin bioassay. These findings support the hypothesis that appropriate structural and confirmational modifications of α-MSH-related peptides can produce profound effects on the bioactivities of the peptides, and suggest that different structural-conformational requirements exist for α-MSH interactions with its various receptors.