Learning-Dependent Gene Expression of CREB1 Isoforms in the Molluscan Brain

Cyclic AMP-responsive element binding protein1 (CREB1) has multiple functions in gene regulation. Various studies have reported that CREB1-dependent gene induction is necessary for memory formation and long-lasting behavioral changes in both vertebrates and invertebrates. In the present study, we characterized Lymnaea CREB1 (LymCREB1) mRNA isoforms of spliced variants in the central nervous system (CNS) of the pond snail Lymnaea stagnalis. Among these spliced variants, the three isoforms that code a whole LymCREB1 protein are considered to be the activators for gene regulation. The other four isoforms, which code truncated LymCREB1 proteins with no kinase inducible domain, are the repressors. For a better understanding of the possible roles of different LymCREB1 isoforms, the expression level of these isoform mRNAs was investigated by a real-time quantitative RT-PCR method. Further, we examined the changes in gene expression for all the isoforms in the CNS after conditioned taste aversion (CTA) learning or backward conditioning as a control. The results showed that CTA learning increased LymCREB1 gene expression, but it did not change the activator/repressor ratio. Our findings showed that the repressor isoforms, as well as the activator ones, are expressed in large amounts in the CNS, and the gene expression of CREB1 isoforms appeared to be specific for the given stimulus. This was the first quantitative analysis of the expression patterns of CREB1 isoforms at the mRNA level and their association with learning behavior

Monoamines, Insulin and the Roles They Play in Associative Learning in Pond Snails

Molluscan gastropods have long been used for studying the cellular and molecular mechanisms underlying learning and memory. One such gastropod, the pond snail Lymnaea stagnalis, exhibits long-term memory (LTM) following both classical and operant conditioning. Using Lymnaea, we have successfully elucidated cellular mechanisms of learning and memory utilizing an aversive classical conditioning procedure, conditioned taste aversion (CTA). Here, we present the behavioral changes following CTA training and show that the memory score depends on the duration of food deprivation. Then, we describe the relationship between the memory scores and the monoamine contents of the central nervous system (CNS). A comparison of learning capability in two different strains of Lymnaea, as well as the filial 1 (F1) cross from the two strains, presents how the memory scores are correlated in these populations with monoamine contents. Overall, when the memory scores are better, the monoamine contents of the CNS are lower. We also found that as the insulin content of the CNS decreases so does the monoamine contents which are correlated with higher memory scores. The present review deepens the relationship between monoamine and insulin contents with the memory score

The activity of isolated neurons and the modulatory state of an isolated nervous system represent a recent behavioural state

Behavioural/motivational state is known to influence nearly all aspects of physiology and behaviour. The cellular basis of behavioural state control is only partially understood. Our investigation, performed on the pond snail Lymnaea stagnalis whose nervous system is useful for work on completely isolated neurons, provided several results related to this problem. First, we demonstrated that the behavioural state can produce long-term changes in individual neurons that persist even after neuron isolation from the nervous system. Specifically, we found that pedal serotonergic neurons that control locomotion show higher activity and lower membrane potential after being isolated from the nervous systems of hungry animals. Second, we showed that the modulatory state (the chemical neuroactive microenvironment of the central ganglia) changes in accordance with the nutritional state of an animal and produces predicted changes in single isolated locomotor neurons. Third, we report that observed hunger-induced effects can be explained by the increased synthesis of serotonin in pedal serotonergic neurons, which has an impact on the electrical activity of isolated serotonergic neurons and the intensity of extrasynaptic serotonin release from the pedal ganglia

A unique profile of insulin antibody titer in islet‐transplanted patients

Insulin antibodies (IAs) can cause glycemic variability. Islet transplantation (ITx) is a treatment for insulin-deficient diabetes that aims to establish on-target glycemic control in the absence of hypoglycemia. To date, there has not been a detailed case study of the association between ITx and IA levels. In this study, we identified a unique profile of IA titers, which differed from glutamic acid decarboxylase antibody titers, in four ITx patients. IA levels decreased with intensified immunosuppressive therapy, whereas glutamic acid decarboxylase antibodies increased transiently after ITx. These data suggest the possibility that IAs, unlike other islet autoantibodies, were eliminated due to immunosuppression after transplantation therapy. The disappearance of IAs, as well as the restoration of regulated insulin secretion after ITx, might have a positive effect on glycemic control in recipients with diabetes. Furthermore, this unique feature is suggestive of immunological pathogenesis and has implications for the treatment of IA-causing disease conditions

Cord Blood Transplantation from Unrelated Donors for Children with Acute Lymphoblastic Leukemia in Japan: The Impact of Methotrexate on Clinical Outcomes

Cord blood transplantation (CBT) from an unrelated donor is recognized as one of the major treatment modalities in allogeneic stem cell transplantation (SCT) for children with hematologic malignancies. We analyzed the clinical outcomes of CBT for children with acute lymphoblastic leukemia (ALL) in Japan and identified the risk factors for the transplant outcomes. From 1997 to 2006, 332 children with ALL underwent CBT from unrelated donors, 270 of which had no prior transplant. Their disease statuses at transplant were first complete remission (CR) (n = 120), second CR (n = 71), and more advanced stages (n = 75). As preconditioning for SCT, total body irradiation (TBI) was given to 194 patients and, for the prophylaxis of graft-versus-host disease (GVHD), methotrexate (MTX) was given to 159 patients. The cumulative incidents of neutrophil and platelet recovery (>20 K) were 88.5% and 78.4%, respectively. The incidents of grade II-IV, III-IV acute GVHD (aGVHD), and chronic GVHD (cGVHD) were 45.6%, 20.4%, and 19.2%, respectively, and treatment-related mortality was 22.6%. The 5-year event-free survival (EFS) and overall survival (OS) at CR1, CR2, and advanced status were 47.4%, 45.5%, 15.0%, and 63.7%, 59.7%, and 20.7%, respectively. Multivariate analysis revealed that MTX with calcineurin inhibitor (CNI) was associated with decreased incidence of grade II-IV GVHD (CNI alone: hazard ratio [HR] = 1.74, 95% confidence interval [CI] = 1.06-2.83, P = .027; CNI + prednisolone (PSL), HR = 1.61, 95% CI = 1.03-2.50, P = .036), III-IV aGVHD (CNI alone: HR = 3.02, 95% CI = 1.55-5.91, P = 0.001; CNI + PSL, HR = 1.89, 95% CI = 0.93-3.83, P = .078), or cGVHD (CNI alone: HR = 1.78, 95% CI = 0.83-3.82, P = .143; CNI + PSL, HR = 2.44, 95% CI = 1.24-4.82, P = .01), compared with CNI alone or CNI + PSL. At an advanced stage of disease, GVHD prophylaxis with MTX + CNI is associated with improved OS compared with CNI alone (CNI alone: HR = 3.20, 95% CI = 1.43-7.15, P = .005; CNI + PSL, HR = 1.47, CI = 0.67-3.20, P = .332). Our retrospective study showed that CBT for children with ALL is feasible and GVHD prophylaxis with MTX + CNI is associated with significant favorable outcomes in prevention of aGVHD and cGVHD as well as survival advantage in advanced cases

The impact of human leukocyte antigen mismatch on recipient outcomes in living‐donor liver transplantation

Donor–recipient human leukocyte antigen (HLA) compatibility has not been considered to significantly affect liver transplantation (LT) outcomes; however, its significance in living-donor LT (LDLT), which is mostly performed between blood relatives, remains unclear. This retrospective cohort study included 1954 LDLTs at our institution (1990–2020). The primary and secondary endpoints were recipient survival and the incidence of T cell–mediated rejection (TCMR) after LDLT, respectively, according to the number of HLA mismatches at all five loci: HLA-A, HLA-B, HLA-C, HLA-DR, and HLA-DQ. Subgroup analyses were also performed in between-siblings that characteristically have widely distributed 0–10 HLA mismatches. A total of 1304 cases of primary LDLTs were finally enrolled, including 631 adults (recipient age at LT ≥18 years) and 673 children (<18 years). In adult-to-adult LDLT, the more HLA mismatches at each locus, the significantly worse the recipient survival was (p = 0.03, 0.01, 0.03, 0.001, and <0.001 for HLA-A, HLA-B, HLA-C, HLA-DR, and HLA-DQ, respectively). This trend was more pronounced when multiple loci were combined (all p < 0.001 for A + B + DR, A + B + C, DR + DQ, and A + B + C + DR + DQ). Notably, a total of three or more HLA-B + DR mismatches was an independent risk factor for both TCMR (hazard ratio [HR] 2.66, 95% confidence interval [CI] 1.21–5.87; p = 0.02) and recipient survival (HR 2.44, 95% CI 1.11–5.35; p = 0.03) in between-siblings. By contrast, HLA mismatch did not affect pediatric LDLT outcomes at any locus or in any combinations; however, it should be noted that all donor–recipient relationships are parent-to-child that characteristically possesses one or less HLA mismatch at each locus and maximally five or less mismatches in total. In conclusion, HLA mismatch significantly affects not only TCMR development but also recipient survival in adult LDLT, but not in children

キョウイン ノ ICT キキ リヨウ シエン ノ タメ ノ ドウガ コウシュウ キョウザイ ノ サクセイ ト ヒョウカ

研究報

Highly Sensitive Determination of Hydrogen Peroxide and Glucose by Fluorescence Correlation Spectroscopy

BACKGROUND: Because H(2)O(2) is generated by various oxidase-catalyzed reactions, a highly sensitive determination method of H(2)O(2) is applicable to measurements of low levels of various oxidases and their substrates such as glucose, lactate, glutamate, urate, xanthine, choline, cholesterol and NADPH. We propose herein a new, highly sensitive method for the measurement of H(2)O(2) and glucose using fluorescence correlation spectroscopy (FCS). METHODOLOGY/PRINCIPAL FINDINGS: FCS has the advantage of allowing us to determine the number of fluorescent molecules. FCS measures the fluctuations in fluorescence intensity caused by fluorescent probe movement in a small light cavity with a defined volume generated by confocal illumination. We thus developed a highly sensitive determination system of H(2)O(2) by FCS, where horseradish peroxidase (HRP) catalyzes the formation of a covalent bond between fluorescent molecules and proteins in the presence of H(2)O(2). Our developed system gave a linear calibration curve for H(2)O(2) in the range of 28 to 300 nM with the detection limit of 8 nM. In addition, by coupling with glucose oxidase (GOD)-catalyzed reaction, the method allows to measure glucose in the range of 80 nM to 1.5 µM with detection limit of 24 nM. The method was applicable to the assay of glucose in blood plasma. The mean concentration of glucose in normal human blood plasma was determined to be 4.9 mM. CONCLUSIONS/SIGNIFICANCE: In comparison with commercial available methods, the detection limit and the minimum value of determination for glucose are at least 2 orders of magnitude more sensitive in our system. Such a highly sensitive method leads the fact that only a very small amount of plasma (20 nL) is needed for the determination of glucose concentration in blood plasma