Social memory,amnesia, and autism: Brain oxytocin secretion is regulated by NAD+ metabolites and single nucleotide polymorphisms of CD38

Previously, we demonstrated that CD38, a transmembrane protein with ADP-ribosyl cyclase activity, plays a critical role in mouse social behavior by regulating the release of oxytocin (OXT), which is essential for mutual recognition. When CD38 was disrupted, social amnesia was observed in Cd38 knockout mice. The autism spectrum disorders (ASDs), characterized by defects in reciprocal social interaction and communication, occur either sporadically or in a familial pattern. However, the etiology of ASDs remains largely unknown. Therefore, the theoretical basis for pharmacological treatments has not been established. Hence, there is a rationale for investigating single nucleotide polymorphisms (SNPs) in the human CD38 gene in ASD subjects. We found several SNPs in this gene. The SNP rs3796863 (C > A) was associated with high-functioning autism (HFA) in American samples from the Autism Gene Resource Exchange. Although this finding was partially confirmed in low-functioning autism subjects in Israel, it has not been replicated in Japanese HFA subjects. The second SNP of interest, rs1800561 (4693C > T), leads to the substitution of an arginine (R) at codon 140 by tryptophan (W; R140W) in CD38. This mutation was found in four probands of ASD and in family members of three pedigrees with variable levels of ASD or ASD traits. The plasma levels of OXT in ASD subjects with the R140W allele were lower than those in ASD subjects lacking this allele. The OXT levels were unchanged in healthy subjects with or without this mutation. One proband with the R140W allele receiving intranasal OXT for approximately 3 years showed improvement in areas of social approach, eye contact and communication behaviors, emotion, irritability, and aggression. Five other ASD subjects with mental deficits received nasal OXT for various periods; three subjects showed improved symptoms, while two showed little or no effect. These results suggest that SNPs in CD38 may be possible risk factors for ASD by abrogating OXT function and that some ASD subjects can be treated with OXT in preliminary clinical trials.     

Complement C1q activates canonical Wnt signaling and promotes aging-related phenotypes

Wnt signaling plays critical roles in development of various organs and pathogenesis of many diseases, and augmented Wnt signaling has recently been implicated in mammalian aging and aging-related phenotypes. We here report that complement C1q activates canonical Wnt signaling and promotes aging-associated decline in tissue regeneration. Serum C1q concentration is increased with aging, and Wnt signaling activity is augmented during aging in the serum and in multiple tissues of wild-type mice, but not in those of C1qa-deficient mice. C1q activates canonical Wnt signaling by binding to Frizzled receptors and subsequently inducing C1s-dependent cleavage of the ectodomain of Wnt coreceptor low-density lipoprotein receptor-related protein 6. Skeletal muscle regeneration in young mice is inhibited by exogenous C1q treatment, whereas aging-associated impairment of muscle regeneration is restored by C1s inhibition or C1qa gene disruption. Our findings therefore suggest the unexpected role of complement C1q in Wnt signal transduction and modulation of mammalian aging.     

An enzyme cycling method for measurement of allantoin in human serum

Current conventional measurement of allantoin levels in human serum uses an HPLC method. However, performing this assay is time-consuming and sample-intensive, and it requires expensive equipment. We have developed a novel enzyme cycling method for measuring allantoin concentrations in human serum. In the first step, serum allantoin is converted to allantoate by the action of allantoinase (EC 3.5.2.5), and endogenous ammonia is simultaneously removed by the action of glutamine synthetase II (EC 6.3.1.2). In the second step, l-methionine sulfoximine is used to inhibit glutamine synthetase II, and ammonia is liberated from allantoate by the activity of allantoate amidohydrolase (EC 3.5.3.9). In the final step, the ammonia is then converted to NAD by NAD synthetase (EC 6.3.1.5). Subsequent action of glucose dehydrogenase (EC 1.1.1.47) and diaphorase (EC 1.6.99.2) in the presence of glucose and 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium (WST-1) acts to cycle the formed NAD between its oxidized and reduced forms, resulting in the production of WST-1 formazan, which is monitored at 450 nm. The assay standard curve is linear from 0 to 70 μM allantoin. The level of allantoin in healthy subjects was measured to be 8.2 ± 3.1 μM (n = 30).     

Physicochemical properties and in vitro intestinal permeability properties and intestinal cell toxicity of silica particles,performed in simulated gastrointestinal fluids

Background

Amorphous silica particles with the primary dimensions of a few tens of nm, have been widely applied as additives in various fields including medicine and food. Especially, they have been widely applied in powders for making tablets and to coat tablets. However, their behavior and biological effects in the gastrointestinal tracts associated with oral administration remains unknown.

Methods

Amorphous silica particles with diameters of 50, 100, and 200 nm were incubated in the fasted-state and fed-state simulated gastric and intestinal fluids. The sizes, intracellular transport into Caco-2 cells (model cells for intestinal absorption), the Caco-2 monolayer membrane permeability, and the cytotoxicity against Caco-2 cells were then evaluated for the silica particles.

Results

Silica particles agglomerated in fed-state simultaneous intestinal fluids. The agglomeration and increased particles size inhibited the particles' absorption into the Caco-2 cells or particles' transport through the Caco-2 cells. The in vitro cytotoxicity of silica particles was not observed when the average size was larger than 100 nm, independent of the fluid and the concentration.

Conclusion

Our study indicated the effect of diet on the agglomeration of silica particles. The sizes of silica particles affected the particles' absorption into or transport through the Caco-2 cells, and cytotoxicity in vitro, depending on the various biological fluids.

General significance

The findings obtained from our study may offer valuable information to evaluate the behavior of silica particles in the gastrointestinal tracts or safety of medicines or foods containing these materials as additives.     

Differential Regulation of Cellular Senescence and Differentiation by Prolyl Isomerase Pin1 in Cardiac Progenitor Cells

Autologous c-kit⁺ cardiac progenitor cells (CPCs) are currently used in the clinic to treat heart disease. CPC-based regeneration may be further augmented by better understanding molecular mechanisms of endogenous cardiac repair and enhancement of pro-survival signaling pathways that antagonize senescence while also increasing differentiation. The prolyl isomerase Pin1 regulates multiple signaling cascades by modulating protein folding and thereby activity and stability of phosphoproteins. In this study, we examine the heretofore unexplored role of Pin1 in CPCs. Pin1 is expressed in CPCs in vitro and in vivo and is associated with increased proliferation. Pin1 is required for cell cycle progression and loss of Pin1 causes cell cycle arrest in the G₁ phase in CPCs, concomitantly associated with decreased expression of Cyclins D and B and increased expression of cell cycle inhibitors p53 and retinoblastoma (Rb). Pin1 deletion increases cellular senescence but not differentiation or cell death of CPCs. Pin1 is required for endogenous CPC response as Pin1 knock-out mice have a reduced number of proliferating CPCs after ischemic challenge. Pin1 overexpression also impairs proliferation and causes G₂/M phase cell cycle arrest with concurrent down-regulation of Cyclin B, p53, and Rb. Additionally, Pin1 overexpression inhibits replicative senescence, increases differentiation, and inhibits cell death of CPCs, indicating that cell cycle arrest caused by Pin1 overexpression is a consequence of differentiation and not senescence or cell death. In conclusion, Pin1 has pleiotropic roles in CPCs and may be a molecular target to promote survival, enhance repair, improve differentiation, and antagonize senescence.     

Studies on turfgrass snow mold caused byTyphula ishikariensis. II. Microscopical observation of infected bentgrass leaves

Light and transmission electron microscopy revealed thatTyphula ishikariensis penetrated into bentgrass leaves either through cuticles or stomata either by single hyphae or infection cushions formed on host surfaces. Time course study on infected leaves showed that penetration through stomatal subsidiary cells and their adjacent cells seemed to occur earlier than that through epidermal cells located farther from stomata. More than 30% of epidermal cells were infected by 10 days after inoculation. When hyphae penetrated through an intact cuticle of epidermal cells, they seemed to dissolve host cell walls enzymatically at penetration sites. Physical pressure also seemed to be involved in penetration.     

Genetic Polymorphism of Cat (Felis catus) Plasma Orosomucoid

Genetic polymorphism of orosomucoid (ORM) was observed in 22 breeds of cats (Felis catus) using isoelectric focusing (pH 4.0–6.5) of desialylated plasmas followed by immunoblotting with rabbit antiserum to human ORM. From a total of 943 plasma samples examined, 15 phenotypes were identified and family studies demonstrated an inheritance of five codominant alleles, ORM^A, ORM^B, ORM^C, ORM^D, and ORM^E, at a single locus.     

Regulation of reductive production of succinate under anaerobic conditions in baker's yeast

When baker's yeast grown aerobically on ethanol as a carbon source was anaerobically cultured in a medium containing glucose, the activity of a cytoplasmic fumarate reductase irreversibly catalyzing the conversion of fumarate to succinate increased, reaching about 3 times the original activity after 12 h, while the activity of succinate dehydrogenase was almost lost after 10 h. These results indicate that the citrate cycle is partially modified to become a reductive pathway leading to succinate during the anaerobic cultivation. In non-proliferating cells grown anaerobically on glucose, the rates of accumulating succinate and pyruvate were decreased and increased, respectively, with increasing concentrations of L-aspartate or NH4Cl in the medium containing glucose as a substrate. These changes were accompanied with increase in the cellular content of aspartate, an inhibitor of pyruvate carboxylase that is involved in supplying the intermediates of the citrate cycle, and pyruvate, a substrate of the enzyme. The aminotransferase inhibitor, aminooxyacetate, prevented the changes in succinate accumulation and cellular aspartate following the addition of NH4Cl. The addition of L-glutamate caused a marked increase in the rate of succinate accumulation without changing the cellular content of aspartate. Neither L-glutamate nor L-aspartate had the ability to produce succinate. The rate of glucose consumption was not changed upon adding these nitrogen compounds. Similar findings were also observed in experiments using proliferating cells. This report presents evidence that in cells containing a large amount of the fumarate reductase, the production of succinate from glucose is regulated by the cellular level of aspartate through the pyruvate carboxylase reaction and that glutamate regulates the succinate production by a mechanism distinct from that involved in the regulation by L-aspartate.     

Isolation and characterization of intraspecific cybrids : Effect of mitochondrial DNA on their cellular properties

Cybrid clones were obtained by fusing whole cells of rat glioma C6BU-1, resistant to 5-bromodeoxyuridine (BrdU), with cytoplasts of embryonic rat 3Y1CAP cells, resistant to chloramphenicol (CAP), in selective medium with BrdU and CAP. The clones resistant to BrdU and CAP were confirmed to be cybrids by chromosome and mtDNA analyses. More than half the mtDNA of all the cybrid clones was from the 3Y1CAP cells. After cultivation of a cybrid clone Y22 for 3 months in the absence of CAP, subclones were isolated. One subclone Y22-22 contained predominantly mitochondrial DNA (mtDNA) from the 3Y1CAP cells. Using this subclone, the effects of the mitochondrial genome on cellular properties were examined. The growth patterns, expression of glioma-specific beta-adrenergic receptor, and composition of the major proteins of C6BU-1 cells were not affected by transmitted mtDNA from the 3Y1CAP cells. This procedure for isolating cells containing predominantly foreign mtDNA will be useful in studies on the interaction between genomes of the mitochondria and nucleus.     

Design and synthesis of estrogen receptor degradation inducer based on a protein knockdown strategy

We designed and synthesized estrogen receptor (ER) degradation inducers 5, 6, and 7, which crosslink the ER and the cellular inhibitor of apoptosis protein 1 (cIAP1). Compounds 5, 6, and 7 induced cIAP1-mediated ubiquitylation of ERα resulting in its proteasomal degradation.