The Agatston score, commonly used to quantify coronary artery calcification (CAC), is determined by the plaque area and density. Despite an excellent predictability of the Agatston score for cardiovascular events, the density of CAC has never been studied in patients with pre-dialysis chronic kidney disease (CKD). This study aimed to analyze the CAC density and its association with serum mineral levels in CKD.

PSD-95 associated PSD proteins play a critical role in regulating the density and activity of glutamate receptors. Numerous previous studies have shown an association between the genes that encode these proteins and schizophrenia (SZ) and autism spectrum disorders (ASD), which share a substantial portion of genetic risks. We sequenced the protein-encoding regions of DLG1, DLG2, DLG4, DLGAP1, DLGAP2, and SynGAP in 562 cases (370 SZ and 192 ASD patients) on the Ion PGM platform. We detected 26 rare (minor allele frequency <1%), non-synonymous mutations, and conducted silico functional analysis and pedigree analysis when possible. Three variants, G344R in DLG1, G241S in DLG4, and R604C in DLGAP2, were selected for association analysis in an independent sample set of 1315 SZ patients, 382 ASD patients, and 1793 healthy controls. Neither DLG4-G241S nor DLGAP2-R604C was detected in any samples in case or control sets, whereas one additional SZ patient was found that carried DLG1-G344R. Our results suggest that rare missense mutations in the candidate PSD genes may increase susceptibility to SZ and/or ASD. These findings may strengthen the theory that rare, non-synonymous variants confer substantial genetic risks for these disorders.

Malocclusion induced by raising the bite causes chronic stress. Chronic stress leads to increased plasma corticosterone levels and impaired hippocampal function due to impaired neurogenesis or increased apoptosis in the hippocampus. The present study aimed to clarify the mechanisms underlying the impaired hippocampal function induced by the bite-raised condition in aged senescence-accelerated mouse prone 8 (SAMP8).

Protein phosphatase 4 catalytic subunit (PP4c) is a PP2A-related protein serine/threonine phosphatase with important functions in a variety of cellular processes, including microtubule (MT) growth/organization, apoptosis, and tumor necrosis factor signaling. In this study, we report that NDEL1 is a substrate of PP4c, and PP4c selectively dephosphorylates NDEL1 at Cdk1 sites. We also demonstrate that PP4c negatively regulates Cdk1 activity at the centrosome. Targeted disruption of PP4c reveals disorganization of MTs and disorganized MT array. Loss of PP4c leads to an unscheduled activation of Cdk1 in interphase, which results in the abnormal phosphorylation of NDEL1. In addition, abnormal NDEL1 phosphorylation facilitates excessive recruitment of katanin p60 to the centrosome, suggesting that MT defects may be attributed to katanin p60 in excess. Inhibition of Cdk1, NDEL1, or katanin p60 rescues the defective MT organization caused by PP4 inhibition. Our work uncovers a unique regulatory mechanism of MT organization by PP4c through its targets Cdk1 and NDEL1 via regulation of katanin p60 distribution.

This study aims to elucidate age-related intrinsic brain volume changes over the adult lifespan using an unbiased data-driven structural brain parcellation. Anatomical brain images from a cohort of 293 healthy volunteers ranging in age from 21 to 86 years were analyzed using independent component analysis (ICA). ICA-based parcellation identified 192 component images, of which 174 (90.6%) showed a significant negative correlation with age and with some components being more vulnerable to aging effects than others. Seven components demonstrated a convex slope with aging; 3 components had an inverted U-shaped trajectory, and 4 had a U-shaped trajectory. Linear combination of 86 components provided reliable prediction of chronological age with a mean absolute prediction error of approximately 7.2 years. Structural co-variation analysis showed strong interhemispheric, short-distance positive correlations and long-distance, inter-lobar negative correlations. Estimated network measures either exhibited a U- or an inverted U-shaped relationship with age, with the vertex occurring at approximately 45-50 years. Overall, these findings could contribute to our knowledge about healthy brain aging and could help provide a framework to distinguish the normal aging processes from that associated with age-related neurodegenerative diseases.

The chromosome 22q11.2 deletion is an extremely high risk genetic factor for various neuropsychiatric disorders; however, the 22q11.2 deletion-related brain pathology in humans at the cellular and molecular levels remains unclear.

The estimation of functional connectivity (FC) measures using resting state functional MRI (fMRI) is often affected by head motion during functional imaging scans. Head motion is more common in the elderly than in young participants and could therefore affect the evaluation of age-related changes in brain networks. Thus, this study aimed to investigate the influence of head motion in FC estimation when evaluating age-related changes in brain networks.

Leptospirosis is a public health challenge in Sabah State of Malaysian Borneo. Rapid urbanization, rural-to-urban migration, and undocumented immigration in Sabah have increased the pressure on the urban garbage disposal system. Rodents and other small animals thrive under these conditions. We hypothesized that urban sanitation workers would be at risk of developing leptospirosis. In total, 303 urban sanitation workers with a mean age of 42.6 years were enrolled in this study. The serum samples collected from these workers were subjected to the microscopic agglutination test (MAT), PCR and nucleotide sequencing of the amplicons to confirm the presence of Leptospira. The phylogenetic analysis using the neighbor joining method was performed to assess whether they were pathogenic. In this study 43.8% (133/303) of the samples were MAT-seropositive and among them, 29 (21.8%) were positive by PCR. Nucleotide sequencing of the amplicons confirmed the presence of Leptospira. Phylogenetic analysis showed that our strains belonged to the pathogenic group of Leptospira. A high proportion of urban sanitation workers were seropositive for leptospirosis, and a considerable number were PCR positive for Leptospira, thereby indicating asymptomatic infections. Further research is needed to confirm whether this is a transient phenomenon or antibiotic therapy is required.

HIF-1α, an essential transcription factor under hypoxic condition, is indispensable for chondrocytes during skeletal development but its expression and roles in articular chondrocytes are yet to be revealed. We examined HIF-1α protein expression and the hypoxic condition during mouse osteoarthritis (OA) development using state of the art hypoxic probes and found that its expression decreased as OA progressed, coinciding with the change in hypoxic conditions in articular cartilage. Gain- and loss-of-function of HIF-1α in cell culture experiments showed that HIF-1α suppressed catabolic genes such as Mmp13 and Hif2a. We confirmed these anticatabolic effects by measuring glycosaminoglycan release from wild type and conditional knock-out mice femoral heads cultured ex vivo. We went on to surgically induce OA in mice with chondrocyte-specific deletion of Hif1a and found that the development of OA was exacerbated. Increased expression of catabolic factors and activation of NF-κB signalling was clearly evident in the knock-out mice. By microarray analysis, C1qtnf3 was identified as a downstream molecule of HIF-1α, and experiments showed it exerted anti-catabolic effects through suppression of NF-κB. We conclude that HIF-1α has an anti-catabolic function in the maintenance of articular cartilage through suppression of NF-κB signalling.

The 22q11.2 deletion syndrome (22q11.2DS) is associated with an increased risk for psychiatric disorders. Although most of the 22q11.2DS patients have a 3.0-Mb deletion, existing mouse models only mimic a minor mutation of 22q11.2DS, a 1.5-Mb deletion. The role of the genes existing outside the 1.5-Mb deletion in psychiatric symptoms of 22q11.2DS is unclear. In this study, we generated a mouse model that reproduced the 3.0-Mb deletion of the 22q11.2DS (Del(3.0 Mb)/ +) using the CRISPR/Cas9 system. Ethological and physiological phenotypes of adult male mutants were comprehensively evaluated by visual-evoked potentials, circadian behavioral rhythm, and a series of behavioral tests, such as measurement of locomotor activity, prepulse inhibition, fear-conditioning memory, and visual discrimination learning. As a result, Del(3.0 Mb)/ + mice showed reduction of auditory prepulse inhibition and attenuated cue-dependent fear memory, which is consistent with the phenotypes of existing 22q11.2DS models. In addition, Del(3.0 Mb)/ + mice displayed an impaired early visual processing that is commonly seen in patients with schizophrenia. Meanwhile, unlike the existing models, Del(3.0 Mb)/ + mice exhibited hypoactivity over several behavioral tests, possibly reflecting the fatigability of 22q11.2DS patients. Lastly, Del(3.0 Mb)/ + mice displayed a faster adaptation to experimental jet lag as compared with wild-type mice. Our results support the validity of Del(3.0 Mb)/ + mice as a schizophrenia animal model and suggest that our mouse model is a useful resource to understand pathogenic mechanisms of schizophrenia and other psychiatric disorders associated with 22q11.2DS.

Rotavirus infection is a dilemma for developing countries, including Malaysia. Although commercial rotavirus vaccines are available, these are not included in Malaysia's national immunization program. A scarcity of data about rotavirus genotype distribution could be partially to blame for this policy decision, because there are no data for rotavirus genotype distribution in Malaysia over the past 20 years. From January 2018 to March 2019, we conducted a study to elucidate the rotavirus burden and genotype distribution in the Kota Kinabalu and Kunak districts of the state of Sabah. Stool specimens were collected from children under 5 years of age, and rotavirus antigen in these samples was detected using commercially available kit. Electropherotypes were determined by polyacrylamide gel electrophoresis of genomic RNA. G and P genotypes were determined by RT-PCR using type specific primers. The nucleotide sequence of the amplicons was determined by Sanger sequencing and phylogenetic analysis was performed by neighbor-joining method. Rotavirus was identified in 43 (15.1%) children with watery diarrhea. The male:female ratio (1.9:1) of the rotavirus-infected children clearly showed that it affected predominantly boys, and children 12-23 months of age. The genotypes identified were G3P[8] (74% n = 31), followed by G1P[8] (14% n = 6), G12P[6](7% n = 3), G8P[8](3% n = 1), and GxP[8] (3% n = 1). The predominant rotavirus circulating among the children was the equine-like G3P[8] (59.5% n = 25) with a short electropherotype. Eleven electropherotypes were identified among 34 strains, indicating substantial diversity among the circulating strains. The circulating genotypes were also phylogenetically diverse and related to strains from several different countries. The antigenic epitopes present on VP7 and VP4 of Sabahan G3 and equine-like G3 differed considerably from that of the RotaTeq vaccine strain. Our results also indicate that considerable genetic exchange is occurring in Sabahan strains. Sabah is home to a number of different ethnic groups, some of which culturally are in close contact with animals, which might contribute to the evolution of diverse rotavirus strains. Sabah is also a popular tourist destination, and a large number of tourists from different countries possibly contributes to the diversity of circulating rotavirus genotypes. Considering all these factors which are contributing rotavirus genotype diversity, continuous surveillance of rotavirus strains is of utmost importance to monitor the pre- and post-vaccination efficacy of rotavirus vaccines in Sabah.

Hand, foot, and mouth disease (HFMD) is endemic in Malaysia, with the number of cases increasing. Sabah has experienced several HFMD outbreaks, but information on the epidemiology and molecular characteristics of responsible viruses is scarce. In this study, data of 17,574 reports of HFMD cases in Sabah from 2015 to 2019 were extracted from a public health disease surveillance system and analyzed. Twenty-one swab samples from 13 children were collected from Beaufort, Sabah, during an outbreak in August 2018 for detection and serotyping of causative viruses by semi-nested reverse transcription-polymerase chain reaction (snRT-PCR) of the VP4-VP2 region and consensus degenerate hybrid oligonucleotide primer PCR of the VP1 region, respectively. Nucleotide sequencing and phylogenetic analysis were conducted by the neighbor-joining method. The average annual incidence of HFMD was 94.3 per 100,000 people, with the greatest yearly increase between 2017 and 2018. Swabs from six children were tested positive for enterovirus, of which five were positive for CVA16 and one for EV71. All CVA16 strains belonged to sub-genotype B1a, and the EV71 strain belonged to sub-genotype B5. Phylogenetic analyses indicate that enterovirus genotype shift might be responsible for the increasing trend of HFMD in Sabah, however, further study is needed.

The Reelin gene (RELN) encodes a large extracellular protein, which has multiple roles in brain development and adult brain function. It activates a series of neuronal signal transduction pathways in the adult brain that function in synaptic plasticity, dendritic morphology, and cognitive function. To further investigate the roles of Reln in brain function, we generated a mouse line using the C57BL/6 J strain with the specific Reln deletion identified from a Japanese patient with schizophrenia (Reln-del mice). These mice exhibited abnormal sociality, but the pathophysiological significance of the Reln deletion for higher brain functions, such as learning and behavioral flexibility remains unclear. In this study, cognitive function in Reln-del mice was assessed using touchscreen-based visual discrimination (VD) and reversal learning (RL) tasks. Reln-del mice showed normal learning in the simple VD task, but the learning was delayed in the complex VD task as compared to their wild-type (WT) littermates. In the RL task, sessions were divided into early perseverative phase (sessions with <50% correct) and later learning phase (sessions with ≥50% correct). Reln-del mice showed normal perseveration but impaired relearning ability in both simple RL and complex RL task as compared to WT mice. These results suggest that Reln-del mice have impaired learning ability, but the behavioral flexibility is unaffected. Overall, the observed behavioral abnormalities in Reln-del mice suggest that this mouse model is a useful preclinical tool for investigating the neurobiological mechanism underlying cognitive impairments in schizophrenia and a therapeutic strategy.

Rotaviruses are major causative agents of acute diarrhea in children under 5 years of age in Malaysia. However, a rotavirus vaccine has not been included in the national vaccination program. To date, only two studies have been carried out in the state of Sabah, Malaysia, although children in this state are at risk of diarrheal diseases. Previous studies showed that 16%-17% of cases of diarrhea were caused by rotaviruses and that equine-like G3 rotavirus strains are predominant. Because the prevalence of rotaviruses and their genotype distribution vary over time, this study was conducted at four government healthcare facilities from September 2019 through February 2020. Our study revealed that the proportion of rotavirus diarrhea increased significantly to 37.2% (51/137) after the emergence of the G9P[8] genotype in replacement of the G12P[8] genotype. Although equine-like G3P[8] strains remain the predominant rotaviruses circulating among children, the Sabahan G9P[8] strain belonged to lineage VI and was phylogenetically related to strains from other countries. A comparison of the Sabahan G9 strains with the G9 vaccine strains used in the RotaSiil and Rotavac vaccines revealed several mismatches in neutralizing epitopes, indicating that these vaccines might not be effective in Sabahan children. However, a vaccine trial may be necessary to understand the precise effects of vaccination.

22q11.2 deletion syndrome (22q11.2DS) is associated with a high risk of developing various psychiatric and developmental disorders, including schizophrenia and early-onset Parkinson's disease. Recently, a mouse model of this disease, Del(3.0Mb)/+, mimicking the 3.0 Mb deletion which is most frequently found in patients with 22q11.2DS, was generated. The behavior of this mouse model was extensively studied and several abnormalities related to the symptoms of 22q11.2DS were found. However, the histological features of their brains have been little addressed. Here we describe the cytoarchitectures of the brains of Del(3.0Mb)/+ mice. First, we investigated the overall histology of the embryonic and adult cerebral cortices, but they were indistinguishable from the wild type. However, the morphologies of individual neurons were slightly but significantly changed from the wild type counterparts in a region-specific manner. The dendritic branches and/or dendritic spine densities of neurons in the medial prefrontal cortex, nucleus accumbens, and primary somatosensory cortex were reduced. We also observed reduced axon innervation of dopaminergic neurons into the prefrontal cortex. Given these affected neurons function together as the dopamine system to control animal behaviors, the impairment we observed may explain a part of the abnormal behaviors of Del(3.0Mb)/+ mice and the psychiatric symptoms of 22q11.2DS.

Although rabies is endemic in Laos, genetic characterization of the viruses in this country is limited. There are growing concerns that development in the region may have increased transport of dog through Laos for regional dog meat consumption, and that this may cause spillover of the viruses from dogs brought here from other countries. This study was therefore undertaken to evaluate the current rabies situation and the genetic characteristics of rabies viruses currently circulating in Laos.

Transcription factors SOX9, SOX5 and SOX6 are indispensable for generation and differentiation of chondrocytes. However, molecular mechanisms to induce the SOX genes are poorly understood. To address this issue, we previously determined the human embryonic enhancer of SOX6 by 5'RACE analysis, and identified the 46-bp core enhancer region (CES6). We initially performed yeast one-hybrid assay for screening other chondrogenic factors using CES6 as bait, and identified a zinc finger protein ZNF449. ZNF449 and Zfp449, a counterpart in mouse, transactivated enhancers or promoters of SOX6, SOX9 and COL2A1. Zfp449 was expressed in mesenchyme-derived tissues including cartilage, calvaria, muscle and tendon, as well as in other tissues including brain, lung and kidney. In limb cartilage of mouse embryo, Zfp449 protein was abundantly located in periarticular chondrocytes, and decreased in accordance with the differentiation. Zfp449 protein was also detected in articular cartilage of an adult mouse. During chondrogenic differentiation of human mesenchymal stem cells, ZNF449 was increased at an early stage, and its overexpression enhanced SOX9 and SOX6 only at the initial stage of the differentiation. We further generated Zfp449 knockout mice to examine the in vivo roles; however, no obvious abnormality was observed in skeletal development or articular cartilage homeostasis. ZNF449 may regulate chondrogenic differentiation from mesenchymal progenitor cells, although the underlying mechanisms are still unknown.

Vitamin D hydroxylated at carbon 25 (25(OH)D) is generally recognized as a precursor of active vitamin D. Despite its low affinity for the vitamin D receptor (VDR), both deficient and excessive 25(OH)D levels are associated with poor clinical outcomes. Here we studied direct effects of 25(OH)D3 on the kidney using 25(OH)D-1α-hydroxylase (CYP27B1) knockout mice. The effects of 25(OH)D3 on unilateral ureteral obstruction were analyzed as proximal tubular cells and macrophages are two major cell types that take up 25(OH)D and contribute to the pathogenesis of kidney injury. Excess 25(OH)D3 in obstructed mice worsened oxidative stress and tubulointerstitial fibrosis, whereas moderate levels of 25(OH)D3 had no effects. The exacerbating effects of excess 25(OH)D3 were abolished in CYP27B1/VDR double-knockout mice and in macrophage-depleted CYP27B1 knockout mice. Excess 25(OH)D3 upregulated both M1 marker (TNF-α) and M2 marker (TGF-β1) levels of kidney-infiltrating macrophages. In vitro analyses verified that excess 25(OH)D3 directly upregulated TNF-α and TGF-β1 in cultured macrophages but not in tubular cells. TNF-α and 25(OH)D3 cooperatively induced oxidative stress by upregulating iNOS in tubular cells. Aggravated tubulointerstitial fibrosis in mice with excess 25(OH)D3 indicated that macrophage-derived TGF-β1 also had a key role in the pathogenesis of surplus 25(OH)D3. Thus, excess 25(OH)D3 worsens tubulointerstitial injury by modulating macrophage phenotype.

A novel organic cation transporter OCTN2 is indispensable for carnitine transport across plasma membrane and subsequent fatty acid metabolism in the mitochondria. Here, we report a novel splice variant of OCTN2 (OCTN2VT), in which a 72-base-pair sequence located in the first intron of OCTN2 gene was spliced between exons 1 and 2 of OCTN2, causing the insertion of 24 amino acids in the first extracellular loop of OCTN2. Despite the similarity between OCTN2 and OCTN2VT regarding primary structure and tissue distribution, their biochemical characteristics were significantly different. OCTN2 was expressed on the plasma membrane with robust N-glycosylation, whereas OCTN2VT was retained in the endoplasmic reticulum (ER) with poor N-glycosylation. In addition, the retention in the ER caused no carnitine uptake into the cells. These results demonstrate that the biochemical and functional characteristics of OCTN2VT are distinct from OCTN2 due to the insertion of 24 amino acids in the first extracellular loop.

Healthy aging is associated with structural and functional changes in the brain even in individuals who are free of neurodegenerative diseases. Using resting state functional magnetic resonance imaging data from a carefully selected cohort of participants, we examined cross sectional changes in the functional organization of several large-scale brain networks over the adult lifespan and its potential association with general cognitive performance. Converging results from multiple analyses at the voxel, node, and network levels showed widespread reorganization of functional brain networks with increasing age. Specifically, the primary processing (visual and sensorimotor) and visuospatial (dorsal attention) networks showed diminished network integrity, while the so-called core neurocognitive (executive control, salience, and default mode) and basal ganglia networks exhibited relatively preserved between-network connections. The visuospatial and precuneus networks also showed significantly more widespread increased connectivity with other networks. Graph analysis suggested that this reorganization progressed towards a more integrated network topology. General cognitive performance, assessed by Addenbrooke's Cognitive Examination-Revised total score, was positively correlated with between-network connectivity among the core neurocognitive and basal ganglia networks and the integrity of the primary processing and visuospatial networks. Mediation analyses further indicated that the observed association between aging and relative decline in cognitive performance could be mediated by changes in relevant functional connectivity measures. Overall, these findings provided further evidence supporting widespread age-related brain network reorganization and its potential association with general cognitive performance during healthy aging.