Influence of contrast ultrasonography with perflutren lipid microspheres on microvessel injury.

Microbubbles have been reported to enhance ultrasound (US)-related side effects in animal systems. The present study investigated the influence of contrast ultrasonography (US) with perflutren lipid microspheres, a recently developed second-generation contrast agent, on microvessels. Rat mesentery was exposed to 1.8-MHz pulsed US with intravenous injection of perflutren (0.1 or 1.0 ml/kg) or Levovist (300 mg/kg), and the microvessel bleeding and endothelial cell injury was examined. Impaired endothelial cells were identified by the fluorescence of propidium iodide. Microvessel bleeding was examined also in the rat myocardium. The interaction between 0.1 ml/kg of perflutren and US exposure did not cause microvessel bleeding, and did not increase endothelial cell injury compared with the sham operation, unless frequent, strong US exposure occurred. When the dose was increased to 1.0 ml/kg, the combination of perflutren and US exposure resulted in capillary bleeding and increased endothelial cell injury in capillaries and venules (p<0.01). However, the incidence of microvessel bleeding and endothelial cell injury did not exceed that with Levovist microbubbles. In the myocardium, microvessel bleeding was not observed under any conditions. In conclusion, perflutren lipid microspheres enhanced US-related microvessel injury as with other contrast agents at the dose of 1.0 ml/kg, but not with 0.1 ml/kg and the appropriate US setting.     

Hyperthymic temperament and brightness preference in healthy subjects: Further evidence for involvement of left inferior orbitofrontal cortex in hyperthymic temperament

Background

Hyperthymic temperament has been generally accepted as one of premorbid temperament of bipolar disorders. Although several studies indicate that subjects with hyperthymic temperament receive more illuminance, our recent study suggests that the threshold of brightness and darkness judgment is not different between more and less hyperthymic subjects, and that hyperthymic temperament may be associated with left inferior orbitofrontal cortex, which has been reported to be associated with bipolar disorder. Therefore, at the next stage, it can be hypothesized that hyperthymic subjects may prefer brightness (i.e., heliotropism) and thereby seek illuminance, and that percent signal changes of left inferior orbitofrontal cortex during the preference task may be associated with hyperthymic temperament scores.

Methods

We compared brightness preference and un-preference between more and less hyperthymic subjects, and investigated percent signal changes of left inferior orbitofrontal cortex during brightness preference judgment, brightness un-preference judgment, and control task by using functional Magnetic Resonance Imaging (fMRI).

Results

There were significant differences in brightness preference judgment and un-preference judgment, showing that more hyperthymic subjects preferred brighter illuminace levels and un-preferred darker illuminance levels than less hyperthymic subjects. Moreover, fMRI signal changes of left inferior orbitofrontal cortex was significantly and negatively associated with hyperthymic temperament scores.

Limitations

It is unknown why left but not right inferior orbitofrontal cortex was associated with hyperthymic temperament scores.

Conclusions

The present findings suggest that more hyperthymic subjects may prefer brightness and un-prefer darkness than less hyperthymic subjects (i.e., heliotropism), and reconfirm that hyperthymic temperament may be associated with left inferior orbitofrontal cortex, which have been reported to be associated with bipolar disorders.     

Reduction of toxicity in brachytherapy using a new technique

PurposeThe purpose of the study was to elucidate the usefulness of a dose evaluation method for reducing late genitourinary (GU) toxicity in high-dose-rate brachytherapy (HDR-BT) of prostate cancer.Methods and MaterialsGU toxicity was scored in accordance with the Common Terminology Criteria for Adverse Events version 4.0. The prostatic urethra was divided into three segments (base = B, midgland = M, apex = A), which were subclassified into seven subgroups (B, M, A, BM, BA, MA, BMA) using a D_10% color map of the urethra. Significance testing was conducted on urethral D_0.1% and D_10% among the seven subgroups. Grade < 2 GU toxicity was also implemented.ResultsData of 174 patients with localized prostate cancer treated with HDR-BT combined with external beam radiotherapy between November 2011 and July 2014 were analyzed retrospectively. Median age was 74 (53–84) years, and median followup period was 44 (6–69) months. The number of Grade < 2 and Grade ≥ 2 toxicity was significantly different in the M subgroup than in the other subgroups (p < 0.05), suggesting increased radioresistance in the midgland urethra.ConclusionsA high-dose-area evaluation method using a urethral D_10% color map may be helpful in reducing late GU toxicity in HDR-BT for prostate cancer.     

A collision tumor of solitary fibrous tumor/hemangiopericytoma and meningioma: A case report with literature review

An intracranial collision tumor is a rare lesion composed of two histologically different neoplasms in the same anatomic location. Even more rare is the collision tumor of a solitary fibrous tumor/hemangiopericytoma (SFT/HPC) and meningioma. The patient was a 46-year-old woman who had a 40 × 35 × 30-mm mass in the vermis of the cerebellum. Histologically, the mass consisted of two different components. One component showed the morphology of meningioma (World Health Organization (WHO) grade I), and the other component exhibited small round cell proliferation with hypercellular density, which was revealed to be SFT/HPC (WHO grade III) based on STAT6 immunohistochemistry. STAT6 showed completely different immunohistochemistry results in these two components (nuclear-negative in meningioma and nuclear-positive in SFT/HPC). Since these two neoplasms are associated with different prognoses, they should be distinguished from each other. When meningioma and an SFT/HPC-like lesion are identified morphologically, it is important to recognize the presence of such a collision tumor composed of meningioma and SFT/HPC, and identify the SFT/HPC component by employing STAT6 immunohistochemistry.     

Pentraxin 3 as a new biomarker of peritoneal injury in peritoneal dialysis patients

It is well known that bioincompatible peritoneal dialysate plays a central role in the development of peritoneal fibrosis. Peritoneal inflammation continues even after the cessation of peritoneal dialysate stimulation. It is important to establish the definition of persistent inflammation in the peritoneal cavity at the cessation of peritoneal dialysis (PD). The objective of the present study was to determine whether pentraxin 3 (PTX3) in peritoneal effluent (PE) may be a new biomarker in PD patients. Serum, PE, and peritoneal specimens were obtained from 50 patients with end-stage kidney disease at Juntendo University Hospital. Samples of 19 patients were obtained at the initiation of PD and those of 31 patients at the cessation of PD. PTX3, high-sensitivity CRP, and MMP-2 and IL-6 were analyzed. An immunohistological examination using an anti-PTX3 antibody was performed. Expressions of PTX3 were observed in endothelial cells, fibroblasts, and mesothelial cells in the peritoneum. The PTX3 level in PE at the cessation of PD was significantly higher than that at the initiation of PD. Effluent PTX3 levels in patients with a history of peritonitis or a PD duration of more than 8 years were significantly higher than those in patients without peritonitis or patients with a PD duration of <8 years. The PTX3 level was significantly correlated with MMP-2 and IL-6 levels in PE, as well as the thickness of the submesothelial compact zone and the vasculopathy. It appears that PTX3 may be a new biomarker of peritoneal inflammation and progressive fibrosis.     

Stereoselective synthesis of Gly-Gly-type (E)-methylalkene and (Z)-chloroalkene dipeptide isosteres and their application to 14-mer RGG peptidomimetics

Stereoselective and efficient synthesis of Gly-Gly-type (E)-methylalkene and (Z)-chloroalkene dipeptide isosteres is realized by organocuprate-mediated single electron transfer reduction. The synthetic isosteres can be used in Fmoc-based solid phase peptide synthesis, resulting in the preparation of the 14-mer RGG peptidomimetics containing an (E)-methylalkene or a (Z)-chloroalkene unit.

An efficient synthesis of Gly-Gly-type (E)-methylalkene and (Z)-chloroalkene dipeptide isosteres is realized by organocuprate-mediated single electron transfer reduction.

Glycylglycine (Gly-Gly) is the smallest dipeptide and has been synthesized by many approaches in the last 100 years.¹ Due to its versatile nature and ready availability, glycylglycine has been used as a chemical probe² and buffer³ in biochemical studies and also as a reagent which can enhance the solubility of overexpressed proteins.⁴ In addition to the utility of Gly-Gly itself, oligoglycine (oligo-Gly) motifs are notable for being more flexible and less-functionalized than any combination of other amino acids. These features are useful in bioconjugation strategies which link multiple biomolecules without interfering with the function of each biomolecule, allowing synthesis of bioconjugated artificial molecules including dimeric, multi-domain, and fusion proteins.⁵ The flexibility of oligo-Gly enables the formation of unusual secondary structures of peptides and proteins.⁶ Thus, the oligo-Gly motif can be found in the biologically important peptides and proteins such as Met/Leu-enkephalin ( and ), the C-terminus of ubiquitin , ctenidin,⁷ shepherin I,⁸ and DNA/RNA-binding proteins with repeated sequences related to the various physiological processes via protein–protein and protein–nucleic acids interactions.⁹ These proteins relate with gene expression, DNA damage signal and apoptosis, however, the detail effects of Gly-Gly with steric and electronic factors to these functions are unknown. Given the importance of oligo-Gly in various fields, non-hydrolyzable peptidomimetics of oligo-Gly could be attractive building blocks for the synthesis of novel bioconjugated molecules and complex peptidomimetics with improved chemical stability and functionality. For example, even the Gly-Gly dipeptide mimic with the tetra-substituted alkene unit replacing the Gly-Gly peptide bond has been shown to promote the β-hairpin formation, and is thus the smallest peptidomimetic that is known to control a peptide structure.¹⁰ There are two reports of the synthesis of Gly-Gly-type fluoroalkene dipeptide isosteres.¹¹ However, the poor synthetic access to such molecules has hindered their application to the peptidomimetics. Our long-standing interest in the drug discovery with amide-to-alkene isosteric switching prompted this investigation into the stereoselective synthesis of Gly-Gly-type (E)-methylalkene and (Z)-chloroalkene dipeptide isosteres.In this report, we describe the beginning of our oligo-Gly-based peptidomimetic study with the stereoselective synthesis of Gly-Gly-type (E)-methylalkene and (Z)-chloroalkene dipeptide isosteres and their use in Fmoc-based solid phase peptide synthesis (SPPS). In the first application of isosteres of this type to access complex peptidomimetics, we synthesize 14-mer RGG peptidomimetics containing (E)-methylalkene or (Z)-chloroalkene unit as surrogates for a Gly-Gly peptide bond. These two isosteres were selected because the potentials of both isosteres have not been fully uncovered, though there are several promising examples.^12,13 Since the carbonyl oxygen equivalents of those isosteres are similar in their size¹⁴ but differ in their electronic properties,¹⁵ comparative studies of these isosteres would have the advantage of exploring the role of peptide bonds in terms of their steric and electronic natures. This work also uncovered the unique ability of the Gly-Gly-type (Z)-chloroalkene isostere to induce β-turn structures in the almost unfoled peptides (Fig. 1).Open in a separate windowFig. 1Gly-Gly peptide and its alkene-type peptidomimetics.The main challenges in this study include the stereoselective formation of the (E)-methylalkene and (Z)-chloroalkene moieties as the surrogates of the Gly-Gly trans-peptide bond, together with the control of the olefine isomerization under the condition of the isostere synthesis and Fmoc-based SPPS. There are several synthetic approaches toward tri-substituted alkene-type isosteres, including the Overman rearrangement,¹³ the S_N2′-type opening of alkenylaziridines,^12a Cu-mediated CF₃-coupling of vinyl iodide,¹⁶ and cross-couplings of vinyl stannane.¹⁷ Organocuprate-mediated reactions of α,β-unsaturated carbonyl compounds with γ-leaving group(s) are also powerful methods to produce tri-substituted alkene-type isosteres, and are particularly suitable for the stereoselective synthesis of (Z)-fluoroalkene¹⁸ and (Z)-chloroalkene isosteres.¹⁹The preparation of Gly-Gly-type alkene dipeptide isosteres was facilitated by the organocuprate-mediated SET reduction that was used in the stereoselective formation of the (E)-methylalkene and (Z)-chloroalkene moieties. Scheme 1 shows the synthesis of (E)-methylalkene isosteres (5). A Witting reaction of methacrolein (1) with ethyl (triphenylphosphoranylidene)acetate followed by epoxidation with m-CPBA afforded the alkenyl oxirane (2) which, treated with Gillman reagent (n-Bu₂CuLi), produced the allylic alcohol (3) with high E-selectivity. A Mitsunobu reaction of 3 with Ns(Boc)NH and deprotection provided the Gly-Gly-type (E)-methylalkene dipeptide isostere 5 that can be applied to the Fmoc-based SPPS. All reactions leading to the synthesis of 5 were performed on a gram scale.Open in a separate windowScheme 1Synthesis of Gly-Gly-type (E)-methylalkene dipeptide isosteres (5).Synthesis of chloroalkene isostere (14) is shown in Scheme 2. Based on previous results for the successful, efficient synthesis of Val-Xaa-type chloroalkene isosteres,^19c we assumed that a similar protocol would allow for the synthesis of 14. However, our attempts revealed that the Gly substrate used has different reactivity and selectivity compared to the other substrates, particularly in the SET reduction step. Consequently, Gly-specific reaction conditions are necessary. The nucleophilic addition of the lithium enolate of methyl dichloroacetate to the N-sulfinylaldimine (7), prepared from (±)-tert-butylsulfinamide (6) and paraformaldehyde and m-CPBA oxidation of 6 gave the N-tert-butylsulfonyl (Bus)-protected α,α-dichloro-β-amino ester (8). Precise control of the amount of DIBAL-H at low temperatures enables the partial reduction of 8, and this is followed by a Horner–Wadsworth–Emmons reaction to produce the corresponding (E)-enoate (9). Initial efforts to apply our established conditions for SET reduction with Me₂CuLi identified the poor Z-selectivity of the reaction and also its propensity to form the α-methylated side products 11 and 12 (Open in a separate windowScheme 2Synthesis of Gly-Gly-type (Z)-chloroalkene dipeptide isosteres (14).Reactivity of (E)-enoate (9) with organocuprates^a