Deficit of tRNA(Lys) modification by Cdkal1 causes the development of type 2 diabetes in mice

The worldwide prevalence of type 2 diabetes (T2D), which is caused by a combination of environmental and genetic factors, is increasing. With regard to genetic factors, variations in the gene encoding Cdk5 regulatory associated protein 1-like 1 (Cdkal1) have been associated with an impaired insulin response and increased risk of T2D across different ethnic populations, but the molecular function of this protein has not been characterized. Here, we show that Cdkal1 is a mammalian methylthiotransferase that biosynthesizes 2-methylthio-N6-threonylcarbamoyladenosine (ms2t6A) in tRNA(Lys)(UUU) and that it is required for the accurate translation of AAA and AAG codons. Mice with pancreatic β cell-specific KO of Cdkal1 (referred to herein as β cell KO mice) showed pancreatic islet hypertrophy, a decrease in insulin secretion, and impaired blood glucose control. In Cdkal1-deficient β cells, misreading of Lys codon in proinsulin occurred, resulting in a reduction of glucose-stimulated proinsulin synthesis. Moreover, expression of ER stress-related genes was upregulated in these cells, and abnormally structured ER was observed. Further, the β cell KO mice were hypersensitive to high fat diet-induced ER stress. These findings suggest that glucose-stimulated translation of proinsulin may require fully modified tRNA(Lys)(UUU), which could potentially explain the molecular pathogenesis of T2D in patients carrying cdkal1 risk alleles.     

Selective inhibitors of methionyl-tRNA synthetase have potent activity against Trypanosoma brucei Infection in Mice

Human African trypanosomiasis continues to be an important public health threat in extensive regions of sub-Saharan Africa. Treatment options for infected patients are unsatisfactory due to toxicity, difficult administration regimes, and poor efficacy of available drugs. The aminoacyl-tRNA synthetases were selected as attractive drug targets due to their essential roles in protein synthesis and cell survival. Comparative sequence analysis disclosed differences between the trypanosome and mammalian methionyl-tRNA synthetases (MetRSs) that suggested opportunities for selective inhibition using drug-like molecules. Experiments using RNA interference on the single MetRS of Trypanosoma brucei demonstrated that this gene product was essential for normal cell growth. Small molecules (diaryl diamines) similar to those shown to have potent activity on prokaryotic MetRS enzymes were synthesized and observed to have inhibitory activity on the T. brucei MetRS (50% inhibitory concentration, <50 nM) and on bloodstream forms of T. brucei cultures (50% effective concentration, as low as 4 nM). Twenty-one compounds had a close correlation between enzyme binding/inhibition and T. brucei growth inhibition, indicating that they were likely to be acting on the intended target. The compounds had minimal effects on mammalian cell growth at 20 μM, demonstrating a wide therapeutic index. The most potent compound was tested in the murine model of trypanosomiasis and demonstrated profound parasite suppression and delayed mortality. A homology model of the T. brucei MetRS based on other MetRS structures was used to model binding of the lead diaryl diamine compounds. Future studies will focus on improving the pharmacological properties of the MetRS inhibitors.     

Otx2 binding to perineuronal nets persistently regulates plasticity in the mature visual cortex

Specific transfer of (orthodenticle homeobox 2) Otx2 homeoprotein into GABAergic interneurons expressing parvalbumin (PV) is necessary and sufficient to open, then close, a critical period (CP) of plasticity in the developing mouse visual cortex. The accumulation of endogenous Otx2 in PV cells suggests the presence of specific Otx2 binding sites. Here, we find that perineuronal nets (PNNs) on the surfaces of PV cells permit the specific, constitutive capture of Otx2. We identify a 15 aa domain containing an arginine-lysine doublet (RK peptide) within Otx2, bearing prototypic traits of a glycosaminoglycan (GAG) binding sequence that mediates Otx2 binding to PNNs, and specifically to chondroitin sulfate D and E, with high affinity. Accordingly, PNN hydrolysis by chondroitinase ABC reduces the amount of endogenous Otx2 in PV cells. Direct infusion of RK peptide similarly disrupts endogenous Otx2 localization to PV cells, reduces PV and PNN expression, and reopens plasticity in adult mice. The closure of one eye during this transient window reduces cortical acuity and is specific to the RK motif, as an Alanine-Alanine variant or a scrambled peptide fails to reactivate plasticity. Conversely, this transient reopening of plasticity in the adult restores binocular vision in amblyopic mice. Thus, one function of PNNs is to facilitate the persistent internalization of Otx2 by PV cells to maintain CP closure. The pharmacological use of the Otx2 GAG binding domain offers a novel, potent therapeutic tool with which to restore cortical plasticity in the mature brain.     

Development of an experimental model of cholestasis induced by hypoxic/ischemic damage to the bile duct and liver tissues in infantile rats

Background

We aimed to develop experimental models of hypoxia/ischemia-induced cholestasis using neonatal and infantile rats.

Methods

Hypoxia/ischemia was induced in the bile duct (BD) by injecting prostaglandin (PG) at birth and/or by coagulation of the hepatic artery (CHA) at about 3?weeks after birth. The rats were divided into 6 groups: control; PG-injected; sham-operated with or without PG; CHA; and CHA?+?PG. CHA was also performed in adult rats. Liver specimens and blood samples were obtained at 5?weeks after birth, and immunohistochemical and biochemical examinations were performed.

Results

(1) BD proliferation with fibrosis (BDPF) was found in the intrahepatic portal tract in the CHA and CHA?+?PG groups. Low-grade BDPF was observed in the PG group. (2) Cyst formation in the extrahepatic BD (EBD) was observed in the porta hepatis of some rats in the CHA and CHA?+?PG groups. In these groups, the number of peribiliary vascular plexuses (PVPs) decreased. BD proliferation and infiltration of inflammatory cells were observed in the EBD wall in the CHA?+?PG group. (3) Ki-67 was expressed in BD and EBD cells in the CHA?+?PG group. (4) BDPF was not detected in adult rats with CHA. (5) Serum liver function tests indicated obstructive changes in the EBD in the CHA and CHA?+?PG groups.

Conclusion

Reduced blood flow in the EBD during infancy induced BDPF and obstructive changes in the EBD, which may, along with immature PVP and inflammatory changes in the EBD, contribute to hypoxia/ischemia of the EBD.     

Evaluation of valproate effects on acylcarnitine in epileptic children by LC-MS/MS

Background: Valproate (VPA) is a simple fatty acid and a substrate for the fatty acid β-oxidation pathway. Previous data suggested that the toxicity of VPA may be provoked by carnitine deficiency and the inhibition of mitochondrial β-oxidation. Objective: The aim of the present study was to elucidate the effect of VPA treatment on carnitine and isomer-differentiated acylcarnitine disposition, and determined the relationships between acylcarnitines and blood VPA levels in long-term treated patients with VPA and/or other antiepileptic drugs. Methods: Serum samples were obtained from children aged 1-15 years old treated for at least 6 months with VPA alone (n = 28) or VPA combined with other anticonvulsants (n = 23) and untreated controls (n = 23). Serum acylcarnitines were separated from their isomers and quantified using high-performance liquid chromatography-tandem mass spectrometry. Results: We found higher 3-hydroxyisovalerylcarnitine levels and trace amounts of valproylcarnitine in both VPA monotherapy and polytherapy patients. Other acylcarnitines, hexanoylcarnitine, C12, C14:1-carnitines and the ratio of long-chain acylcarnitine to free carnitine were also higher in VPA polytherapy individuals than in controls. VPA monotherapy does not result in decreases in free carnitine or in the accumulation of long-chain acylcarnitines. Blood VPA concentrations correlated positively with hexanoylcarnitine, C12, C14:1, C16:1, C18:1-carnitines in all VPA-treated children (n = 51). Conclusion: Long-term VPA treatment in pediatric patients could affect some specific acylcarnitines, which is enhanced by the concomitant use of other anticonvulsants, and the formation of valproylcarnitine alone seems insufficient to develop severe carnitine deficiency at therapeutic doses of VPA.     

Development of a multi-dimensional scale for PDD and ADHD

A novel assessment scale, the multi-dimensional scale for pervasive developmental disorder (PDD) and attention-deficit/hyperactivity disorder (ADHD) (MSPA), is reported. Existing assessment scales are intended to establish each diagnosis. However, the diagnosis by itself does not always capture individual characteristics or indicate the level of support required, since inter-individual differences are substantial and co-morbidity is common. The MSPA consists of 14 domains and each domain is rated by a nine-point quantitative scale. The clinical and behavioral features are projected onto a radar-chart, which facilitates understanding of the disorders both by the patients themselves and by those in their surroundings. We assessed 179 patients and analyzed features by six diagnostic subgroups, which showed relationships between features and diagnoses. The inter-rater reliability was satisfactory.     

Neurosarcoidosis presenting as spontaneously remitting hypersomnia

Journal of Neurology -     

Alcohol intake and 19-year mortality in diabetic men: NIPPON DATA80

Although moderate alcohol intake in diabetic Caucasians is associated with a reduction in coronary heart disease mortality, no study in Japanese with diabetes has examined the association between alcohol intake and mortality outcomes. We analyzed the relationship between alcohol intake and all-cause and cause-specific mortality using the database from NIPPON DATA80. At the baseline in 1980, data on history, lifestyle, and physical examinations were collected on study participants aged 30 years and older from randomly selected areas in Japan. After excluding participants with comorbidities, we followed 4,018 male participants (3,614 nondiabetics, 195 with impaired glucose tolerance and 209 diabetic) for 19 years. During the 19 years of follow-up, there were 990 deaths; 328 were from cardiovascular disease and 157 from all-heart diseases. With the never-drinking category serving as a reference, the Cox multivariate-adjusted hazard ratios for non-daily and daily drinkers for cardiovascular mortality were 0.43 (95% confidence intervals: 0.19-0.95) and 0.45 (0.25-0.80), respectively, and 0.33 (0.12-0.91) and 0.31 (0.15-0.67) for all-heart disease mortality in the combined impaired glucose tolerance and diabetic Japanese men. Alcohol drinking in men with glucose intolerance was associated with a significant reduction in cardiovascular and all-heart disease mortality as seen in the general population in Japan.     

An overview and prospect of demented outpatient care in a department of geriatric medicine: report based on statistical review of the memory clinic at the geriatric outpatient unit of the Nagoya University Hospital

AIM: Although there are many reports regarding the status of memory clinics in Japan, most are from the clinical departments of psychiatry or neurology, and there are few from the geriatric outpatient clinics. This study aimed to review the status of the memory clinic at the geriatric outpatient unit of a university hospital and also to compare the results with other reports. METHODS: Patient records of the memory clinic at the geriatric outpatient unit of the Nagoya University Hospital between January 2000 and June 2006, which included clinical information and neuropsychological profiles were extensively reviewed for statistical analyses. Of the patients who first visited the memory clinic between January 2004 and June 2006, prior written consent are obtained from 232 outpatients, among which 223 individuals who had intact sets of data were subjected to detailed analyses. RESULTS: During the period investigated, we had a total of 778 visits by 577 patients. The characteristics of patients were: age: 74.5+/-8.3 years; MMSE: 23.8+/-4.7; education year: 10.7+/-2.9. Clinical profiles of the patients who visited during the most recent 2.5 years were as follow: cognitively normal, 8.1%; dementia of Alzheimer's type, 45.3%; vascular type, 5.4%; mixed type, 2.2%; frontotemporal dementia, 3.1%; mild cognitive impairment, 15.7%, and others. CONCLUSION: Compared with previous reports from other institutions, we observed that the visitors to our geriatric memory clinic had a relatively higher educational background with earlier stages of dementing disorders, which also included pre-clinical cognitive impairment.