Effect of RU 486 on luteal function in the early pregnant rat

A dose of 30 mg RU 486/kg, an antiprogesterone, was administered to pregnant rats on Day 2 (Group 1) or Day 4 (Group 2) of pregnancy. RU 486 significantly changed serum progesterone and oestradiol concentrations and luteal 3 beta-HSD and 20 alpha-HSD activities in Group 1, and implantation was significantly inhibited. The luteal 3 beta-HSD activity in Group 2 rats on Day 6 was significantly (P less than 0.01) lower than the control value (7.5 +/- 0.6 and 10.1 +/- 0.6 mU/mg protein respectively). This decline in the 3 beta-HSD activity was followed by a marked decrease in the serum progesterone concentration, resulting in a significant decrease of the progesterone/oestradiol ratio and implantation was completely inhibited. The 20 alpha-HSD activity, which could not be detected on Day 6 in the control rats, was twice as great in Group 2 than in Group 1 rats (17.5 +/- 1.2 and 7.4 +/- 3.1 mU/mg protein respectively). Ultrastructural examination of corpora lutea of Group 2 rats confirmed luteolysis. These results suggest that RU 486 has a luteolytic effect and its anti-implantation effect is concomitant with luteolysis of the corpora lutea of pregnancy.     

Genetic polymorphism of alpha-2-HS-glycoprotein: four new alleles and allele frequencies in Japanese

alpha 2-HS-glycoprotein (AHSG) phenotyping was done in 655 Japanese from the Goto Islands, western Japan, using isoelectric focusing followed by immunoblotting. Four new AHSG alleles were encountered, AHSG*G1-G4, whose genetic transmissions were established in family studies. The allele frequencies were: AHSG*1 = 0.7221; AHSG*2 = 0.2748, and AHSG*G1-G4 = 0.0008, respectively.     

Polyamines in bacteriophage R17 and its RNA.

Bacteriophage R17 and its RNA were found to contain significant amounts of spermidine but not of putrescine. When isolated at 0.01 M KCl, up to 1,000 molecules of spermidine were associated with the virion. The phage RNA isolated with phenol plus sodium lauryl sulfate contained approximately 70 to 90 molecules of spermidine. The association appeared to be ionic because the bound spermidine could be dissociated by KCl, MgCl2, or both. Effects of polyamines on in vitro translation were studied using both poly(U) and phage R17-RNA as mRNA. Addition of spermidine to the system at suboptimal concentrations of Mg2+ resulted in marked stimulations of the rate of protein synthesis. Putrescine alone had no effect but stimulated the incorporation in the presence of suboptimal concentrations of spermidine plus Mg2+. The isolated amino acid-incorporating system contained suboptimal soluble and bound polyamines. A comparison of incorporation was made in this system using R17-RNA with and without bound spermidine. No effects of these bound cations were detected on the rate or extent of incorporation of valine. The ratio of incorporation of histidine (present in non-coat proteins) to valine (total protein) revealed little difference as a functions of cation in the system or a function of the spermidine present in R17-RNA.     

A role of the Duffy antigen for the maintenance of plasma chemokine concentrations

We examined plasma chemokine concentrations and chemokine clearance rates in Duffy antigen knockout mice. The plasma concentrations of eotaxin and MCP-1 in Duffy antigen knockout mice were less than one-third of those in wild-type mice. When eotaxin or hMGSA was intravenously injected, the chemokine disappeared more rapidly from the plasma of Duffy antigen knockout mice than from the plasma of wild-type mice. The half-lives of hIP-10 and interferon-gamma, which do not have an affinity for the Duffy antigen, in plasma were indistinguishable between Duffy antigen knockout mice and wild-type mice. These results suggest that the Duffy antigen delays the disappearance of chemokines from the plasma, resulting in the maintenance of plasma chemokine concentrations.     

Toxic effects of mercury(II), cadmium(II) and lead(II) porphyrins on Trypanosoma brucei brucei growth

The effects of free mercury(II), cadmium(II) and lead(II) ions and their metalloporphyrin-derivatives on Trypanosoma brucei brucei growth in culture were studied. All experiments were conducted in the dark. IC(50) values on growth obtained in 24-h time-course experiments were 1.5 x 10(-7), 2.4 x 10(-6), 4.4 x 10(-6) and 2.6 x 10(-5) M for mercury(II) porphyrin, cadmium(II) porphyrin, lead(II) porphyrin and free base porphyrin, respectively. While the IC50 values for Hg2+, Cd2+ and Pb2+ were 3.6 x 10(-6), 1.5 x 10(-5) and 1.6 x 10(-5) M, respectively. These results clearly indicate that the toxicity of the metalloporphyrin complexes of mercury(II), cadmium(II) and lead(II) to T. b. brucei parasites was much higher compared to their free metal ions and free base porphyrin at low concentrations. It was also observed after 8 h incubation that the metalloporphyrins were effective in inhibiting the division of the parasites at concentrations >1.25 x 10(-7) M for mercury(II) porphyrin, concentrations >1.2 x 10(-6) M for cadmium(II) and lead(II) porphyrins and at concentrations >3.6 x 10(-6) M for Hg2+ ion. These observations were not detected in samples treated with the free metal ions and the free base porphyrin at the same concentrations. Interestingly, trypanosomes treated with metalloporphyrin complexes displayed different morphological features from those cells treated with free base porphyrin or metal ions. The chemotherapeutic potential of the metalloporphyrins of H2TMPyP for treatment of African trypanosomiasis is discussed.     

Diurnal rhythms of retinal phospholipid synthetic enzymes are retained but their activities are decreased in rats under alpha-linolenic acid deficiency

Rats fed a safflower oil (alpha-linolenic acid (ALNA)-deficient) diet over the course of two generations had significantly decreased docosahexaenoic acid (22:6n-3) and increased docosapentaenoic acid (22:5n-6) contents in the major retinal phospholipids such as phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylserine (PS) when compared with those fed a perilla oil (ALNA-sufficient) diet, but the compositions of phosphatidylinositol acyl chains were relatively unaffected. The contents of individual phospholipids in the retina were essentially the same for the two dietary groups. The activities of the rate-limiting enzymes in the de novo synthesis of PC and PE, CTP:phosphocholine cytidylyltransferase (CT), and CTP:phosphoethanolamine cytidylyltransferase (ET), respectively, were measured in the retinas excised at 5:00, 9:00, 13:00, and 17:00 h from rats adapted to a 24-h cycle with lights on from 7:00 to 19:00 h. Both enzymes exhibited significant diurnal rhythms with the lowest activities at 5:00 h and gradually increasing activities following exposure of the rats to light; the maximum activities were at 13:00 h for CT and 17:00 h for ET. The diurnal rhythms were not significantly affected by the above-mentioned diets. However, both enzyme activities at each collection time point were significantly lower in the safflower oil group than in the perilla oil group. These results suggest that retinal phospholipid turnover associated with shedding, phagocytosis, and resynthesis of the rod outer segments is limited by ALNA deficiency.     

Seizure Outcomes and Predictors of Recurrent Post-Stroke Seizure: A Retrospective Observational Cohort Study

Background

Seizure is a common complication after stroke (termed “post-stroke seizure,” PSS). Although many studies have assessed outcomes and risk factors of PSS, no reliable predictors are currently available to determine PSS recurrence. We compared baseline clinical characteristics and post-stroke treatment regimens between recurrent and non-recurrent PSS patients to identify factors predictive of recurrence.

Methods

Consecutive PSS patients admitted to our stroke center between January 2011 and July 2013 were monitored until February 2014 (median 357 days; IQR, 160–552) and retrospectively evaluated for baseline clinical characteristics and PSS recurrence. Cumulative recurrence rates at 90, 180, and 360 days post-stroke were estimated by Kaplan—Meier analysis. Independent predictors of recurrent PSS were identified by Cox proportional-hazards analysis.

Results

A total of 104 patients (71 men; mean age, 72.1 ± 11.2 years) were analyzed. PSS recurred in 31 patients (30%) during the follow-up. Factors significantly associated with PSS recurrence by log-rank analysis included previous PSS, valproic acid (VPA) monotherapy, polytherapy with antiepileptic drugs (AEDs), frontal cortical lesion, and higher modified Rankin Scale score at discharge (all p < 0.05). Independent predictors of recurrent PSS were age <74 years (HR 2.38, 95% CI 1.02–5.90), VPA monotherapy (HR 3.86, 95% CI 1.30–12.62), and convulsions on admission (HR 3.87, 95% CI 1.35–12.76).

Conclusions

Approximately one-third of PSS patients experienced seizure recurrence within one year. The predictors of recurrent PSS were younger age, presence of convulsions and VPA monotherapy. Our findings should be interpreted cautiously in countries where monotherapy with second-generation AEDs has been approved because this study was conducted while second-generation AEDs had not been officially approved for monotherapy in Japan.     

Involvement of regulatory T cells in the experimental autoimmune encephalomyelitis-preventive effect of dendritic cells expressing myelin oligodendrocyte glycoprotein plus TRAIL

We previously reported the protection from myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) by the adoptive transfer of genetically modified embryonic stem cell-derived dendritic cells (ES-DC) presenting MOG peptide in the context of MHC class II molecules and simultaneously expressing TRAIL (ES-DC-TRAIL/MOG). In the present study, we found the severity of EAE induced by another myelin autoantigen, myelin basic protein, was also decreased after treatment with ES-DC-TRAIL/MOG. This preventive effect diminished, if the function of CD4(+)CD25(+) regulatory T cells (Treg) was abrogated by the injection of anti-CD25 mAb into mice before treatment with ES-DC-TRAIL/MOG. The adoptive transfer of CD4(+)CD25(+) T cells from ES-DC-TRAIL/MOG-treated mice protected the recipient mice from MOG- or myelin basic protein-induced EAE. The number of Foxp3(+) cells increased in the spinal cords of mice treated with ES-DC-TRAIL/MOG. In vitro experiments showed that TRAIL expressed in genetically modified ES-DC and also in LPS-stimulated splenic macrophages had a capacity to augment the proliferation of CD4(+)CD25(+) T cells. These results suggest that the prevention of EAE by treatment with ES-DC-TRAIL/MOG is mediated, at least in part, by MOG-reactive CD4(+)CD25(+) Treg propagated by ES-DC-TRAIL/MOG. For the treatment of organ-specific autoimmune diseases, induction of Treg reactive to the organ-specific autoantigens by the transfer of DC-presenting Ags and simultaneously overexpressing TRAIL therefore appears to be a promising strategy.     

Mutational analysis of conserved outer sphere arginine residues of chalcone synthase

Chalcone synthase (CHS), a key enzyme in flavonoid biosynthesis, catalyses sequential decarboxylative condensations of p-coumaroyl-CoA with three malonyl-CoA molecules and cyclizes the resulting tetraketide intermediate to produce chalcone. Phenylglyoxal, an Arg selective reagent, was found to inactivate the enzyme, although no Arg is found at the active site. Conserved, non-active site Arg residues of CHS were individually mutated and the results were discussed in the context of the 3D structure of CHS. Arg199 and Arg350 were shown to provide important interactions to maintain the structural integrity and foldability of the enzyme. Arg68, Arg172 and Arg328 interact with highly conserved Gln33/Phe215, Glu380 and Asp311/Glu314, respectively, thus helping position the catalytic Cys-His-Asn triad and the (372)GFGPG loop in correct topology at the active site. In particular, a mutation of Arg172 resulted in selective impairment in the cyclization activities of CHS and stilbene synthase, a related enzyme that catalyses a different cyclization of the same tetraketide intermediate. These Arg residues and their interactions are well conserved in other enzymes of the CHS superfamily, suggesting that they may serve similar functions in other enzymes. Mutations of Arg68 and Arg328 had been found in mutant plants that showed impaired CHS activity.