Contribution of iron and protein contents from rat brain subcellular fractions to MR phase imaging.

PURPOSE: Investigation of magnetic susceptibility and chemical exchange as sources of MRI phase contrast between gray and white matter resulting from protein and iron content from subcellular fractions. METHODS: This study analyzes the iron and macromolecule content of different subcellular fractions from rat brain and their relation to the water-resonance frequency by NMR spectroscopy. Additionally, the contributions of susceptibility and exchange were determined with different NMR reference substances. RESULTS: Only weak correlations between iron (r = 0.4318, P = 0.76) or protein content (r = 0.4704, P = 0.70) and frequency shift were observed. After membrane depletion, the correlation for iron increased to r = -0.9006 (P = 0.0009), whereas the shift relative to protein content increased much less (r = -0.4982, P = 0.64). Exchange-driven frequency shifts were 1.283 ppb/(mg/ml) for myelin and 0.775 ppb/(mg/ml) for synaptosomes; susceptibility-driven shifts were -1.209 ppb/(mg/ml) for myelin and -0.368 ppb/(mg/ml) for synaptosomes. The ratios between susceptibility and exchange differ significantly from simple protein solutions. CONCLUSIONS: As a result of counteracting susceptibility and exchange and increased relative shifts in membrane-depleted fractions, we conclude that tissue microstructure accounts more for the in vivo phase contrast than in the situation of homogenized tissue. Thus, membranes may generate much of the in vivo MR phase contrast resulting from anisotropy. Magn Reson Med 77:2028-2039, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Isolation of cDNAs for two closely related members of the axolotl Wnt family, Awnt-5A and Awnt-5B, and analysis of their expression during development.

To characterize molecular interactions between cells in the early amphibian embryo, we have isolated cDNAs for two members of the axolotl (Ambystoma mexicanum) Wnt family, Awnt-5A and Awnt-5B. The encoded proteins share 83% amino acid identity. Using a reverse transcription-polymerase chain reaction (RT-PCR) assay, we find that Awnt-5A transcripts are abundant in the blastula until gastrulation, barely detectable during gastrulation, and increase again during neurulation. They are detected throughout the remaining development and in hatched larvae. In contrast, transcripts for Awnt-5B are undetectable in the blastula. They appear with gastrulation, are present throughout neurulation and organogenesis, and decrease to barely detectable levels in hatched larvae. PCR reactions performed using cDNA library-phage DNA templates derived from whole neurulae versus embryos with the neuroectoderm removed suggest that, in the neurula, Awnt-5A transcripts are present in neuroectodermal as well as non-neuroectodermal tissues while Awnt-5B mRNAs are predominantly localized in the neuroectoderm. To localize Awnt-5A expression in embryos before gastrulation, early gastrulae were dissected by cutting along the animal-vegetal and future dorso-ventral axes and analyzed by RT-PCR. At this early stage, Awnt-5A transcripts appear to be predominantly localized in the dorso-vegetal region of the embryo. These results suggest that the two closely related Awnt-5 genes participate in different morphogenetic processes during early axolotl development.

Evidence for the stability of positional markers in the goldfish tectum.

Positional markers in the tectum, which are thought to guide growing axons to their target sites, have been proposed to be induced by axons, to be only transiently associated with the tectal cells, and then lost after long-term denervation periods (Schmidt: J. Comp. Neurol. 177:279-300, '78). To further investigate this concept, retinal axons were induced to regenerate into ipsilateral tecta which had been deprived of their retinal afferents for shorter (0-4 months) and longer periods (4-8 months). The paths of HRP-labeled regenerating axons of known retinal origin were traced and used as an operational test to decide whether the axons might navigate under the influence of positional markers. Two different kinds of experiments were performed: 1. The axons from a subpopulation of all ganglion cells in the retina were labeled by applying a small crystal of HRP at defined retinal regions. Independent of the denervation period of the tectum, the labeled regenerating axons traveled in abnormal but nonrandom routes. In early regeneration stages, axons exhibited signs of exploratory growth. They extended branches equipped with growth cones and filopodia into various regions of the tectum. In late regeneration stages, the axons lost these branches, exhibited U-turns and bends, and ended in terminal arbors in the retinotopic target region. These findings suggest that the axons travel under the influence of tectal positional markers and that these markers are not transient. 2. Axons from a surgically created temporal hemiretina were labeled by application of HRP to the optic nerve to test whether the temporal axons might expand into the caudal tectum in long-term-denervated tecta. The HRP-labeled axons coursed over rostral and midtectal regions. Instead of invading the caudal tectum they bent and terminated in the rostral tectal half. These results add further support for the conclusion that the path of regenerating retinal axons is governed by long-lasting positional markers.

Production and characterization of polyclonal antibodies against Awnt-1, the axolotl homologue of the proto-oncogene product Wnt-1.

The proto-oncogene Wnt-1 (int-1) is activated by mouse mammary tumor virus retroviral insertion and contributes to the formation of mammary gland tumors in mice. During early development, it is expressed in small groups of cells in the developing central nervous system of fish, amphibians and mice. In the Ambystoma mexicanum (axolotl) embryo, the expression profile of Wnt-1 is biphasic. We have raised polyclonal antibodies against a synthetic peptide corresponding to a surface-exposed epitope of Awnt-1. The antibodies react with Awnt-1 synthesized in bacteria as determined by Western blot analysis. They immunoprecipitate the 40 kD Awnt-1 polypeptide translated in vitro from its corresponding cDNA and immunoprecipitation can be efficiently blocked by the immunizing peptide. These data show that the polyclonal anti-peptide antibodies specifically recognize Awnt-1 in its denatured and native conformation.

Molecular analysis of the Wnt-1 proto-oncogene in Ambystoma mexicanum (axolotl) embryos.

To analyze Wnt-1 expression during neurulation in urodele embryos, we have isolated a Wnt-1 cDNA clone, Awnt-1, from an Ambystoma mexicanum (axolotl) neurula-stage cDNA library. Awnt-1 codes for a protein of 369 amino acids rich in cysteine residues, is preceded by a hydrophobic leader peptide sequence and contains four possible sites for N-linked glycosylation. The temporal expression profile of Awnt-1 was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Awnt-1 expression in the axolotl embryo is biphasic. Awnt-1 transcripts are found in early blastulae until gastrulation, are barely detectable during gastrulation, and are present again from neurulation until late embryogenesis. Transcripts are present before the midblastula transition, indicating that they might be of maternal origin. To localize Awnt-1 expression in embryos during the first phase of expression, early gastrulae were dissected by cutting along the animal-vegetal and future dorso-ventral axes and analyzed by RT-PCR. At the early gastrula stage Awnt-1 transcripts appear to be located in the future ventral region of the embryo. Hatching larvae no longer express Awnt-1. PCR reactions performed using cDNA library-phage DNA templates derived from whole neurulae versus embryos with the neuroectoderm removed suggest that, in the neurula, Awnt-1 transcripts are located in the neuroectoderm. This suggest that, as is the case for Wnt-1 in other vertebrates, Awnt-1 may be involved in neurogenesis. These results suggest that Wnt-1 has earlier roles in development than has been considered until now.

[Patient with left-sided pleural effusion and large structures of soft-tissue-density in the mediastinum]. / Patient mit Pleuraerguss links und grossen weichteildichten Strukturen im Mediastinum. Pankreatitis mit Pleuritis und mediastinaler Beteiligung.

A patient was admitted to the hospital to clarifying a left-sided pleural effusion. Typical cytological findings in pleural effusion did not leave a doubt about being present a pancreatitis in conformity with clearly increased levels of lipase and amylase in the effusion fluid. The CT of the upper abdomen showed signs of a exsudative pancreatitis. In the chest CT as well as in transesophageal ultrasound evaluation large structures of soft-tissue-density together with areas of encapsulated fluid were seen in the dorsal mediastinum reaching down to the diaphragm and abdomen. Just with subsequent diet and strict termination of any alcohol consumption the pathological findings recovered completely. Due to the expansive involvement of the pleural, mediastinal and abdominal compartments this is an extraordinary case of acute pancreatitis.

Early assessment of outcome by ST-segment analysis after thrombolytic therapy in acute myocardial infarction.

As an early marker of outcome, the sum of ST-segment elevation resolution between the electrocardiogram before and 3 hours after initiation of thrombolysis was investigated in 77 patients with acute myocardial infarction. Prospectively, three groups were defined according to complete (> or = 70%, n = 34), partial (< 70% to > or = 30%, n = 26), or no (< 30%, n = 17) ST resolution. There were considerable differences in the enzyme-determined infarct size (alpha-hydroxybutyrate dehydrogenase release for complete, partial, and no ST resolution: 529 +/- 397 IU/L, 689 +/- 484 IU/L, and 1293 +/- 742 IU/L, respectively; p = 0.0001) and the angiographic left ventricular function 1 week later (ejection fraction 58% +/- 10%, 53% +/- 13%, and 43% +/- 12%, respectively, p < 0.01; regional dyssynergic area 24 +/- 19, 39 +/- 23, and 50 +/- 21 U2, respectively, p < 0.01). Early reperfusion as assessed by creatine kinase release measured in 15-minute intervals was 90%, 65%, and 18%, respectively (p = 0.0001). Differences in degrees of ST-elevation resolution at 3 hours may help facilitate timely screening of patients for appropriate therapeutic intervention. Patients with complete ST resolution may be considered for early discharge, and patients with no ST resolution may be candidates for an early invasive approach or additional thrombolytic therapy.

Antiplasmodial activity of mono- and dimeric carbazoles.

A series of mono- and dimeric natural and structurally modified carbazoles has been tested for activity against Plasmodium falciparum. One of the monomers, the synthetic compound 1,4-diacetoxy-3-methylcarbazole (17), displays the highest activity, with an IC50 value of 1.79 micrograms/ml. It is distinctly more active than the as yet best natural compound, 1-hydroxy-3-methylcarbazole (4) (4).