Zinc and manganese homeostasis closely interact in mammalian cells.

Understanding the homeostatic interactions among essential trace metals is important for explaining their roles in cellular systems. Recent studies in vertebrates suggest that cellular Mn metabolism is related to Zn metabolism in multifarious cellular processes. However, the underlying mechanism remains unclear. In this study, we examined the changes in the expression of proteins involved in cellular Zn and/or Mn homeostatic control and measured the Mn as well as Zn contents and Zn enzyme activities to elucidate the effects of Mn and Zn homeostasis on each other. Mn treatment decreased the expression of the Zn homeostatic proteins metallothionein (MT) and ZNT1 and reduced Zn enzyme activities, which were attributed to the decreased Zn content. Moreover, loss of Mn efflux transport protein decreased MT and ZNT1 expression and Zn enzyme activity without changing extracellular Mn content. This reduction was not observed when supplementing with the same Cu concentrations and in cells lacking Cu efflux proteins. Furthermore, cellular Zn homeostasis was oppositely regulated in cells expressing Zn and Mn importer ZIP8, depending on whether Zn or Mn concentration was elevated in the extracellular milieu. Our results provide novel insights into the intricate interactions between Mn and Zn homeostasis in mammalian cells and facilitate our understanding of the physiopathology of Mn, which may lead to the development of treatment strategies for Mn-related diseases in the future.

Protein kinase Cγ in cerebellar Purkinje cells regulates Ca2+-activated large-conductance K+ channels and motor coordination.

The cerebellum, the site where protein kinase C (PKC) was first discovered, contains the highest amount of PKC in the central nervous system, with PKCγ being the major isoform. Systemic PKCγ-knockout (KO) mice showed impaired motor coordination and deficient pruning of surplus climbing fibers (CFs) from developing cerebellar Purkinje cells (PCs). However, the physiological significance of PKCγ in the mature cerebellum and the cause of motor incoordination remain unknown. Using adeno-associated virus vectors targeting PCs, we showed that impaired motor coordination was restored by re-expression of PKCγ in mature PKCγ-KO mouse PCs in a kinase activity-dependent manner, while normal motor coordination in mature Prkcgfl/fl mice was impaired by the Cre-dependent removal of PKCγ from PCs. Notably, the rescue or removal of PKCγ from mature PKCγ-KO or Prkcgfl/fl mice, respectively, did not affect the CF innervation profile of PCs, suggesting the presence of a mechanism distinct from multiple CF innervation of PCs for the motor defects in PKCγ-deficient mice. We found marked potentiation of Ca2+-activated large-conductance K+ (BK) channel currents in PKCγ-deficient mice, as compared to wild-type mice, which decreased the membrane resistance, resulting in attenuation of the electrical signal during the propagation and significant alterations of the complex spike waveform. These changes in PKCγ-deficient mice were restored by the rescue of PKCγ or pharmacological suppression of BK channels. Our results suggest that PKCγ is a critical regulator that negatively modulates BK currents in PCs, which significantly influences PC output from the cerebellar cortex and, eventually, motor coordination.

Production and synthesis of a novel 191Pt-labeled platinum complex and evaluation of its biodistribution in healthy mice.

We previously reported that our sugar-conjugated platinum complex (cis-dichloro [(2-fluoro-α-d-glucopylanosidyl) propane-1,3-diamine] platinum: FGC-Pt) has low toxicity and tumor growth inhibitory effect comparable to that of cisplatin. We focused on radioactive Pt isotopes in order to analyze the kinetics of FGC-Pt using gamma-ray imaging techniques, assuming that FGC-Pt could be used for chemotherapy in the future. Therefore, in this study, we aimed to develop a non-invasive method to analyze the biodistribution of FGC-Pt using 191Pt-labeled FGC-Pt ([191Pt]FGC-Pt). 191Pt was produced via the (n,2n) reaction induced by accelerator neutrons. [191Pt]FGC-Pt was prepared using two different methods. In the first method, [191Pt]FGC-Pt (method A) was obtained through the accelerator neutron irradiation of FGC-Pt. In the second method, [191Pt]FGC-Pt (method B) was synthesized using [191Pt]K2PtCl4, which was obtained by the accelerator neutron irradiation of K2PtCl4. Highly purified [191Pt]FGC-Pt was obtained using the latter method, which suggests that the synthetic method using a 191Pt-labeled platinum reagent is suitable for the radioactivation of platinum complexes. We also aimed to investigate whether a significant correlation existed between the biodistribution of FGC-Pt and [191Pt]FGC-Pt in healthy mice 24 h after tail vein administration. FGC-Pt and [191Pt]FGC-Pt were similarly distributed in healthy mice, with a higher accumulation in the liver and kidney 24 h post injection. In addition, a significant correlation (p < 0.05, r = 0.92) between the 191Pt radioactivity concentration (%ID/g (gamma counter)) and platinum concentration (%ID/g (ICP-MS)) was observed in 13 organs. These results suggest that 191Pt-labeled compounds, synthesized using radioactive platinum reagents, can be used to confirm the biodistribution of platinum compounds. Our study on the biodistribution of [191Pt]FGC-Pt is expected to contribute to the development of novel platinum-based drugs in the future.

Copper-mediated radioiodination and radiobromination via aryl boronic precursor and its application to 125I/77Br-labeled prostate-specific membrane antigen imaging probes.

Prostate-specific membrane antigen (PSMA), expressed in prostate cancer cells, is being investigated extensively worldwide as a target for imaging and therapy of prostate cancer. Various radioiodinated PSMA imaging probes have been developed, and their structure has a peptidomimetic urea-based skeleton as a pharmacophore. For direct radioiodination of molecules containing these peptidomimetic structures, prior studies performed radioiododestannylation or electrophilic radioiodination of tyrosine residues. However, although these radiolabeling methods are frequently used, there are some issues with precursor toxicity and by-product production. Therefore, it is required to investigate a radiolabeling method that can be used for the radiosynthesis of radioiodinated PSMA imaging probes with urea-based peptidomimetic structures. We recently reported that copper-mediated radioiodination via a boronic precursor is an effective method for directly labeling a peptide. This radiohalogenation method was expected to be an effective method for radiosynthesis of PSMA imaging probes with a peptidomimetic structure. In this study, to confirm that this labeling method applies to the synthesis of the PSMA imaging probe, we synthesized PSMA imaging probes labeled with 125I and 77Br ([125I]mIB-PS and [77Br]mBrB-PS) using a copper-mediated radiohalogenation via common boronic precursors and investigated optimal boronic precursor and labeling conditions. As a result, the radiochemical yields of [125I]mIB-PS and [77Br]mBrB-PS were improved to > 93% at room temperature by optimizing the structure of the boronic precursor. We demonstrate that copper-mediated nucleophilic radiochemistry using a boronic precursor is a promising radiosynthetic method of PSMA imaging probes. Although we focused on the synthesis of PSMA imaging probes, the results in this study will also be useful for the synthesis of various radioiodine or radiobromine-labeled bioactive molecules.

Copper-mediated radioiodination reaction through aryl boronic acid or ester precursor and its application to direct radiolabeling of a cyclic peptide.

A copper-mediated radioiodination using aryl boronic precursors is attracting attention as a solution to oxidative iododestannylation and nickel-mediated radioiodination drawbacks. The copper-mediated radiolabeling method allows radioiodination at room temperature with stable aryl boronic precursors without preparing complex starting materials or reagents and can be performed in a reaction vessel exposed to air. This method has good potential in radiochemistry; however, studies on the scope of copper-mediated radioiodination through boronic precursors are insufficient. In particular, few reports have demonstrated the effect of protecting groups on radiolabeling efficiency. Therefore, the effect of the protecting group of aryl boronic acids on the copper-mediated radioiodination was investigated. In addition, this method, which does not require heating, is expected to be useful for direct radiolabeling of peptides. Thus, we attempted direct radioiodination of c(RGDyk) as an example. The resulting radioiodination method was well tolerated in various substrates and was unaffected by the pinacol ester-type protecting group. Also, c(RGDyk) was labeled with 125 I via copper-mediated radioiodination using an aryl boronic acid precursor. The reaction time and yield were improved, compared with the indirect method. Furthermore, the large difference in polarity between the boronic acid precursor and the radiolabeled compound facilitated purification.

Synthesis of 18F-labeled streptozotocin derivatives and an in-vivo kinetics study using positron emission tomography.

Glucose transporter 2 (GLUT2) is involved in glucose uptake by hepatocytes, pancreatic beta cells, and absorptive cells in the intestine and proximal tubules in the kidney. Pancreatic GLUT2 also plays an important role in the mechanism of glucose-stimulated insulin secretion. In this study, novel Fluorine-18-labeled streptozotocin (STZ) derivatives were synthesized to serve as glycoside analogs for in-vivo GLUT2 imaging. Fluorine was introduced to hexyl groups at the 3'-positions of the compounds, and we aimed to synthesize compounds that were more stable than STZ. The nitroso derivatives exhibited relatively good stability during purification and purity analysis after radiosynthesis. We then evaluated the compounds in PET imaging and ex-vivo biodistribution studies. We observed high levels of radioactivity in the liver and kidney, which indicated accumulation in these organs within 5 min of administration. In contrast, the denitroso derivatives accumulated only in the kidney and bladder shortly after administration. Compounds with nitroso groups are thus expected to accumulate in GLUT2-expressing organs, and the presence of a nitroso group is essential for in-vivo GLUT2 imaging.

Bright and dark sides of exercise effects on biological responses such as energy metabolism and renal function in rats with renal failure and fructose-induced glucose intolerance.

In the present study, we investigated the beneficial and risky effects of exercise intended to prevent or treat lifestyle-related diseases on insulin sensitivity, lactic acid utilization, lipid metabolism, hepatic and renal oxidative stress, hepatic selenoprotein P and renal function in obese and glucose-intolerant rats with renal failure. We fed normal rats a 20% casein diet while the glucose-intolerant, obese rats received a high-fructose diet, and after then rats received single injection of vancomycin at a dose of 400 mg/kg for constructing the duplicative state of renal failure and diabetes mellitus. They were forced to run for 1 h/day, 6 days/week, for 10 weeks. Exercise reduced visceral fat and ameliorated insulin sensitivity in the high-fructose group, improved lactic acid usage efficiency, however, increased hepatic oxidative stress and complicated renal dysfunction in the normal and high-fructose fed groups with renal failure. Additionally, exercise upregulated hepatic selenoprotein P expression and enhanced renal antioxidative system in both groups. It is concluded that strictly controlled exercise conditions must be adapted to patient health states especially in view of kidney protection, and supplemental therapy is also recommended in parallel with exercise, using nutrients and vitamins for kidney protection.

Effects of exercise on biological trace element concentrations and selenoprotein P expression in rats with fructose-induced glucose intolerance.

In the present study, we investigated the effects of exercise intended to prevent or treat lifestyle-related diseases on the glucose tolerance, insulin level, lactic acid utilization, muscle glycogen synthesis, hepatic and renal oxidative stress, hepatic selenoprotein P and biological trace element levels in organs of obese, glucose-intolerant rats. We fed normal, healthy rats a 20% casein diet while the glucose-intolerant, obese rats received a high-fructose diet. They were forced to run for one hour per day, six days per week, for ten weeks. Exercise reduced visceral fat and ameliorated glucose tolerance in the high-fructose group, lowered blood lactic acid levels, improved lactic acid usage efficiency, and increased oxidative stress and hepatic levels of Mn, Fe, Cu, and Zn in the normal and high-fructose groups. Additionally, exercise significantly upregulated hepatic selenoprotein P expression in both groups, however, its effect was remarkable in healthy group. On the other hand, muscle glycogen synthesis was not markedly enhanced in high-fructose-diet rats but in normal-diet rats in response to exercise. It is concluded that exercise conditions rather than exercise load must be customized and optimized for each health and disease states in advance before starting exercise training intended to prevent or treat lifestyle-related diseases.

Total Synthesis of Clavilactones.

Clavilactones A, B, and D are epidermal growth factor receptor tyrosine kinase inhibitors that were isolated from cultures of the fungus Clitocybe clavipes. Here, we report full details of the total synthesis of these clavilactones. A key feature of our synthetic approach is a ring-opening/ring-closing metathesis strategy that allows the concise transformation of a cyclobutenecarboxylate into a γ-butenolide. Coupled with enantioselective Ti/BINOL-catalyzed alkynylation of a multisubstituted benzaldehyde and ring-closing metathesis of a diene-bearing silylene acetal to construct the 10-membered carbocycle, this strategy enabled the total synthesis of the natural enantiomers (+)-clavilactone A and (-)-clavilactone B. In addition, the correct structure of clavilactone D was determined by the synthesis of two newly proposed structures. This research resulted in the asymmetric synthesis of the revised (+)-clavilactone D.

Anti-hyperglycemic Effect of Long-Term Bis(hinokitiolato)zinc Complex ([Zn(hkt)2]) Ingestion on Insulin Resistance and Pancreatic Islet Cells Protection in Type 2 Diabetic KK-Ay Mice.

Zinc (Zn) is a trace element with anti-diabetes mellitus (anti-DM) effects. Zn complexes exhibit stronger insulin-like activity than Zn ions. Bis(hinokitiolato)zinc complex ([Zn(hkt)2]) was recently reported to be a potent anti-DM candidate. We examined the effects of [Zn(hkt)2] on insulin resistance and pancreatic islet cells through in vivo long-term ingestion studies. In an in vivo study, we performed 4-month long-term [Zn(hkt)2] administration experiments in KK-Ay mice as a type 2 DM animal model. Ingestion of [Zn(hkt)2] resulted in lower blood glucose levels compared with the non-treated KK-Ay mice (control group). Additionally, [Zn(hkt)2] treatment decreased plasma insulin concentration compared with that of the non-treated KK-Ay group. [Zn(hkt)2] treatment resulted in a significant suppression of islet cell enlargement and a significantly decreased number of insulin-positive cells compared with the non-treated KK-Ay control group. The [Zn(hkt)2] treatment group showed the increasing tendency in the amount of Zn levels in peripheral organs; liver, muscle, adipose, and pancreas, compared with the non-treated KK-Ay control group. However, the Zn level in the pancreas of the [Zn(hkt)2] treatment group did not show the significant increase compared with the non-treated KK-Ay control group. This accumulation of Zn in pancreas suggested that [Zn(hkt)2] mainly effects on the peripheral tissue, and [Zn(hkt)2] has the less effect on the pancreas directly. Thus, we concluded that [Zn(hkt)2] exerted the main effect on peripheral organs by ameliorating insulin resistance.

Investigation of Biodistribution and Speciation Changes of Orally Administered Dual Radiolabeled Complex, Bis(5-chloro-7-[131I]iodo-8-quinolinolato)[65Zn]zinc.

Many zinc (Zn) complexes have been developed as promising oral antidiabetic agents. In vitro assays using adipocytes have demonstrated that the coordination structures of Zn complexes affect the uptake of Zn into cells and have insulinomimetic activities, for which moderate stability of Zn complexes is vital. The complexation of Zn plays a major role improving its bioavailability. However, investigation of the speciation changes of Zn complexes after oral administration is lacking. A dual radiolabeling approach was applied in order to investigate the speciation of bis(5-chloro-7-iodo-8-quinolinolato)zinc complex [Zn(Cq)2], which exhibits the antidiabetic activity in diabetic mice. In the present study, 65Zn- and 131I-labeled [Zn(Cq)2] were synthesized, and their biodistribution were analyzed after an oral administration using both invasive conventional assays and noninvasive gamma-ray emission imaging (GREI), a novel nuclear medicine imaging modality that enables analysis of multiple radionuclides simultaneously. The GREI experiments visualized the behavior of 65Zn and [131I]Cq from the stomach to large intestine and through the small intestine; most of the administered Zn was transported together with clioquinol (5-chloro-7-iodo-8-quinolinol) (Cq). Higher accumulation of 65Zn for [Zn(Cq)2] than ZnCl2 suggests that the Zn associated with Cq was highly absorbed by the intestinal tract. In particular, the molar ratio of administered iodine to Zn decreased during the distribution processes, indicating the dissociation of most [Zn(Cq)2] complexes. In conclusion, the present study successfully evaluated the speciation changes of orally administered [Zn(Cq)2] using the dual radiolabeling method.

Anti-Diabetic Effect of Organo-Chalcogen (Sulfur and Selenium) Zinc Complexes with Hydroxy-Pyrone Derivatives on Leptin-Deficient Type 2 Diabetes Model ob/ob Mice.

Since the discovery of the anti-diabetic effects of zinc (Zn) complex, we synthesized several Zn complexes and evaluated their effects using the KKAy type 2 diabetes mouse model. Recently, we demonstrated that organo-chalcogen (sulfur and selenium) Zn complexes elicit strong anti-diabetic effects. In this study, we treated leptin-deficient ob/ob mice with organo-chalcogen Zn complexes, and evaluated the resulting anti-diabetic effects in a mouse model of diabetes arising from pathogenic mechanisms different from those in KKAy mice. C57BL/6J ob/ob mice orally received either bis(3-hydroxy-2-methyl-4(H)-pyran-4-thiono)Zn, [Zn(hmpt)2] or bis(3-hydroxy-2-methyl-4(H)-pyran-4-seleno)Zn, [Zn(hmps)2], daily for 28 days. Both Zn complexes elicited potent blood glucose-lowering effects and improved HbA1c values. Moreover, glucose intolerance improved as evidenced by the oral glucose tolerance test, and fasting plasma insulin levels decreased in both types of Zn complex-treated mice. Zn concentrations in the liver and pancreas of [Zn(hmpt)2]-treated mice and in the pancreas of [Zn(hmps)2]-treated mice were increased, respectively. The results suggest that the present Zn complexes mainly exerted an anti-diabetic effect in the liver or pancreas. This study is the first to demonstrate that potent Zn complexes elicit anti-diabetic effects in not only KKAy but also ob/ob mice via a normalizing effect on insulin secretion and fasting blood glucose levels.

Transport of Azithromycin into Extravascular Space in Rats.

Recent clinical trials showed a prolonged retention of subinhibitory concentrations of unbound azithromycin in the interstitial fluid of soft tissues despite the fact that azithromycin is extensively distributed in tissues. In these clinical trials, interstitial fluid samples were obtained by using the microdialysis method, and it was established that drug concentrations represent protein-unbound drug concentrations. The present study was designed to measure total azithromycin concentrations in the interstitial fluid of the skin of rats by directly collecting interstitial fluid samples from a pore formed on the skin by a dissolving microneedle array. The total azithromycin concentrations in interstitial fluid of the skin were about 4 to 5 times higher than those in plasma throughout the experimental period, and stasis of the azithromycin concentration in interstitial fluid was observed when the concentration of azithromycin in plasma was at the lower limit of quantification. In addition, the skin/plasma concentration ratio transiently increased after dosing (from 4.3 to 83.1). Our results suggest that azithromycin was trapped inside white blood cells and/or phagocytic cells in not only blood but also interstitial fluid, resulting in a high total azithromycin concentration and the retention of its antimicrobial activity at the primary infection site. The stasis of azithromycin in interstitial fluid and skin would lead to long-lasting pharmacological effects (including those against skin infection) at concentrations exceeding the MIC.

Contrasting intra- and extracellular distribution of catalytic ferrous iron in ovalbumin-induced peritonitis.

Iron is an essential nutrient for every type of life on earth. However, excess iron is cytotoxic and can lead to an increased cancer risk in humans. Catalytic ferrous iron [Fe(II)] is an initiator of the Fenton reaction, which causes oxidative stress by generating hydroxyl radicals. Recently, it became possible to localize catalytic Fe(II) in situ with a turn-on fluorescent probe, RhoNox-1. Here, we screened each organ/cell of rats to globally evaluate the distribution of catalytic Fe(II) and found that eosinophils showed the highest abundance. In various cells, lysosomes were the major organelle, sharing ∼40-80% of RhoNox-1 fluorescence. We then used an ovalbumin-induced allergic peritonitis model to study the dynamics of catalytic Fe(II). Peritoneal lavage revealed that the total iron contents per cell were significantly decreased, whereas an increase in the number of inflammatory cells (macrophages, neutrophils, eosinophils and lymphocytes) resulted in an increased total iron content of the peritoneal inflammatory cells. Notably, macrophages, eosinophils and neutrophils exhibited significantly increased catalytic Fe(II) with increased DMT1 expression and decreased ferritin expression, though catalytic Fe(II) was significantly decreased in the peritoneal lavage fluid. In conclusion, catalytic Fe(II) in situ more directly reflects cellular activity and the accompanying pathology than total iron does.

Oral treatment with a zinc complex of acetylsalicylic acid prevents diabetic cardiomyopathy in a rat model of type-2 diabetes: activation of the Akt pathway.

BACKGROUND: Type-2 diabetics have an increased risk of cardiomyopathy, and heart failure is a major cause of death among these patients. Growing evidence indicates that proinflammatory cytokines may induce the development of insulin resistance, and that anti-inflammatory medications may reverse this process. We investigated the effects of the oral administration of zinc and acetylsalicylic acid, in the form of bis(aspirinato)zinc(II)-complex Zn(ASA)2, on different aspects of cardiac damage in Zucker diabetic fatty (ZDF) rats, an experimental model of type-2 diabetic cardiomyopathy. METHODS: Nondiabetic control (ZL) and ZDF rats were treated orally with vehicle or Zn(ASA)2 for 24 days. At the age of 29-30 weeks, the electrical activities, left-ventricular functional parameters and left-ventricular wall thicknesses were assessed. Nitrotyrosine immunohistochemistry, TUNEL-assay, and hematoxylin-eosin staining were performed. The protein expression of the insulin-receptor and PI3K/AKT pathway were quantified by Western blot. RESULTS: Zn(ASA)2-treatment significantly decreased plasma glucose concentration in ZDF rats (39.0 ± 3.6 vs 49.4 ± 2.8 mM, P < 0.05) while serum insulin-levels were similar among the groups. Data from cardiac catheterization showed that Zn(ASA)2 normalized the increased left-ventricular diastolic stiffness (end-diastolic pressure-volume relationship: 0.064 ± 0.008 vs 0.084 ± 0.014 mmHg/µl; end-diastolic pressure: 6.5 ± 0.6 vs 7.9 ± 0.7 mmHg, P < 0.05). Furthermore, ECG-recordings revealed a restoration of prolonged QT-intervals (63 ± 3 vs 83 ± 4 ms, P < 0.05) with Zn(ASA)2. Left-ventricular wall thickness, assessed by echocardiography, did not differ among the groups. However histological examination revealed an increase in the cardiomyocytes' transverse cross-section area in ZDF compared to the ZL rats, which was significantly decreased after Zn(ASA)2-treatment. Additionally, a significant fibrotic remodeling was observed in the diabetic rats compared to ZL rats, and Zn(ASA)2-administered ZDF rats showed a similar collagen content as ZL animals. In diabetic hearts Zn(ASA)2 significantly decreased DNA-fragmentation, and nitro-oxidative stress, and up-regulated myocardial phosphorylated-AKT/AKT protein expression. Zn(ASA)2 reduced cardiomyocyte death in a cellular model of oxidative stress. Zn(ASA)2 had no effects on altered myocardial CD36, GLUT-4, and PI3K protein expression. CONCLUSIONS: We demonstrated that treatment of type-2 diabetic rats with Zn(ASA)2 reduced plasma glucose-levels and prevented diabetic cardiomyopathy. The increased myocardial AKT activation could, in part, help to explain the cardioprotective effects of Zn(ASA)2. The oral administration of Zn(ASA)2 may have therapeutic potential, aiming to prevent/treat cardiac complications in type-2 diabetic patients.

Bis(hinokitiolato)zinc complex ([Zn(hkt)2]) activates Akt/protein kinase B independent of insulin signal transduction.

Since many Zn complexes have been developed to enhance the insulin-like activity and increase the exposure and residence of Zn in the animal body, these complexes are recognized as one of the new candidates with action mechanism different from existing anti-diabetic drugs. However, the molecular mechanism by which Zn complexes exert an anti-DM effect is unknown. Therefore, we evaluated the activity of Zn complexes, especially related to the phosphorylation of insulin signaling pathway components. We focused on the insulin-like effects of the bis(hinokitiolato)zinc complex, [Zn(hkt)2], using 3T3-L1 adipocytes. [Zn(hkt)2] was taken up by cells and induced Akt phosphorylation in a time-dependent manner. Additionally, it showed inhibitory activity against PTP1B and PTEN, which are major negative regulators of insulin signaling. It did not promote the phosphorylation of IR (insulin receptor)-ß or IRS (insulin receptor substrate)-1 by itself, but in combination with insulin, it enhanced the phosphorylation of IRß. We conclude that [Zn(hkt)2] has effects on the proteins of insulin signaling pathway without insulin receptor mediation, and [Zn(hkt)2] promotes insulin function and shows the anti-DM effects. Thus, [Zn(hkt)2] may be the basis for improved DM treatments.

Fecal Excretion of Orally Administered Collagen-Like Peptides in Rats: Contribution of the Triple-Helical Conformation to Their Stability.

Orally ingested peptides are generally digested in the gastrointestinal (GI) tract and absorbed in the form of oligopeptides. We previously reported that intravenously administered collagen-like triple-helical peptides circulated in the bloodstream and were excreted in their intact forms in urine nearly quantitatively. In the present study, we investigated the fates of orally administered collagen-like peptides in rats. (Pro-Hyp-Gly)10 (Hyp: 4-hydroxyproline), which formed a stable triple-helical structure, was stable in the GI tract, and 72.3±13.0% of the peptide was excreted in the feces. Its recovery ratio was similar to that of all-D-(Pro-Pro-Gly)10 (75.1±15.7%), the indigestible control. In contrast, (Pro-Hyp-Gly)5 and (Pro-Pro-Gly)10, the random coil conformations of which were dominant at body temperature, were not detected in fecal samples, indicating that they were digested by proteases. The high stability of the triple-helical conformation in mammalian bodies suggests the potential use of collagen-like peptides as novel scaffolds of peptide drugs.

Anti-Oxidative Activity of Mytiloxanthin, a Metabolite of Fucoxanthin in Shellfish and Tunicates.

Anti-oxidative activities of mytiloxanthin, a metabolite of fucoxanthin in shellfish and tunicates, were investigated. Mytiloxanthin showed almost the same activities for quenching singlet oxygen and the inhibition of lipid peroxidation as those of astaxanthin, which is a well-known singlet oxygen quencher. Furthermore, mytiloxanthin showed excellent scavenging activity for hydroxyl radicals and this activity was markedly higher than that of astaxanthin.

Preparation of chiral ligands connected with quaternary ammonium group for recyclable catalytic asymmetric transfer hydrogenation in ionic liquid.

Reuse of chiral ruthenium catalyst in catalytic asymmetric transfer hydrogenation (CATH) has attracted attention from economic and environmental viewpoints, and reactions using ionic liquids (ILs) as solvent are recognized as one of the most useful methods for reuse of the catalyst. We synthesized (1S,2S)-N-(p-toluenesulfonyl)-1,2-diphenylethylenediamine (TsDPEN) derivatives with various ionic moieties, and investigated the effect of their structure with respect to catalytic ability and recyclability in CATH with ILs. Ligand 3a having an imidazolium group showed the best results, and significant differences were observed depending on the structure of the ionic moiety or the length of the alkyl chain connecting the ligand site and the ionic moiety. Among various prochiral ketones used as substrates at various cycles, 3a showed a relatively good result.

Bis (aspirinato) zinc (II) complex successfully inhibits carotid arterial neointima formation after balloon-injury in rats.

PURPOSE: Neointima formation following angioplasty is a serious consequence of endothelial damage in arteries. Inflammatory mediators and lack of endothelial regulatory mechanisms lead to migration and proliferation of smooth-muscle cells and thus to restenosis. This study examines the effect of the novel bis (aspirinato) zinc (II) complex on neointima formation in a rat model of carotid balloon-injury. METHODS: Rats underwent balloon-injury of the right common carotid artery, then received PEG400 vehicle (untreated-group), acetylsalicylic-acid (ASA-group), zinc-chloride (Zn-group) and bis (aspirinato) zinc (II) complex (Zn(ASA) 2-group) orally for 18 consecutive days. From harvested carotid arteries, histology, immunohistochemistry and mRNA expression analysis were performed. RESULTS: Compared to the untreated-group, Zn (ASA) 2-treatment significantly lowered stenosis ratio (54.0 ± 5.8% to 25.5 ± 3.9%) and reduced neointima/media ratio (1.5 ± 0.2 to 0.5 ± 0.1). Significantly higher alpha smooth muscle actin mRNA and protein expression were measured after Zn (ASA)2 and Zn-treatment in comparison with the untreated and ASA-groups while the expression of matrix-metalloproteinase-9 was significantly higher in these groups compared to Zn (ASA)2. The presence of collagen in media was significantly decreased in all treated groups. mRNA expressions of nuclear factor kappa-b, transforming growth-factor-ß and proliferating cell nuclear antigen were significantly down-regulated, whereas a20 was up-regulated by Zn (ASA)2 treatment compared to the untreated and ASA-groups. CONCLUSION: This study proves the effectivity of the novel bis (aspirinato) zinc complex in reducing neointima formation and restenosis after balloon-injury and supports the hypothesis that inhibition of smooth-muscle transformation/proliferation plays a key role in the prevention of restenosis.