Plasma ANGPTL8 Levels and Risk for Secondary Cardiovascular Events in Japanese Patients With Stable Coronary Artery Disease Receiving Statin Therapy.

BACKGROUND: The ability to predict secondary cardiovascular events could improve health of patients undergoing statin treatment. Circulating ANGPTL8 (angiopoietin-like protein 8) levels, which positively correlate with proatherosclerotic lipid profiles, activate the pivotal proatherosclerotic factor ANGPTL3. Here, we assessed potential association between circulating ANGPTL8 levels and risk of secondary cardiovascular events in statin-treated patients. METHODS: We conducted a biomarker study with a case-cohort design, using samples from a 2018 randomized control trial known as randomized evaluation of high-dose (4 mg/day) or low-dose (1 mg/day) lipid-lowering therapy with pitavastatin in coronary artery disease (REAL-CAD [Randomized Evaluation of Aggressive or Moderate Lipid-Lowering Therapy With Pitavastatin in Coronary Artery Disease])." From that study's full analysis set (n=12 413), we selected 2250 patients with stable coronary artery disease (582 with the primary outcome, 1745 randomly chosen, and 77 overlapping subjects). A composite end point including cardiovascular-related death, nonfatal myocardial infarction, nonfatal ischemic stroke, or unstable angina requiring emergent admission was set as a primary end point. Circulating ANGPTL8 levels were measured at baseline and 6 months after randomization. RESULTS: Over a 6-month period, ANGPTL8 level changes significantly decreased in the high-dose pitavastatin group, which showed 19% risk reduction of secondary cardiovascular events compared with the low-dose group in the REAL-CAD [Randomized Evaluation of Aggressive or Moderate Lipid-Lowering Therapy With Pitavastatin in Coronary Artery Disease] study. In the highest quartiles, relative increases in ANGPTL8 levels were significantly associated with increased risk for secondary cardiovascular events, after adjustment for several cardiovascular disease risk factors and pitavastatin treatment (hazard ratio in Q4, 1.67 [95% CI, 1.17-2.39). Subgroup analyses showed relatively strong relationships between relative ANGPTL8 increases and secondary cardiovascular events in the high-dose pitavastatin group (hazard ratio in Q4, 2.07 [95% CI, 1.21-3.55]) and in the low ANGPTL8 group at baseline (166

Increased numbers of pre-operative circulating monocytes predict risk of developing cardiac surgery-associated acute kidney injury in conditions requiring cardio pulmonary bypass.

BACKGROUND: Evaluating patients' risk for acute kidney injury (AKI) is crucial for positive outcomes following cardiac surgery. Our aims were first to select candidate risk factors from pre- or intra-operative real-world parameters collected from routine medical care and then evaluate potential associations between those parameters and risk of onset of post-operative cardiac surgery-associated AKI (CSA-AKI). METHOD: We conducted two cohort studies in Japan. The first was a single-center prospective cohort study (n = 145) to assess potential association between 115 clinical parameters collected from routine medical care and CSA-AKI (≥ Stage1) risk in the population of patients undergoing cardiac surgery involving cardiopulmonary bypass (CPB). To select candidate risk factors, we employed random forest analysis and applied survival analyses to evaluate association strength. In a second retrospective cohort study, we targeted patients undergoing cardiac surgery with CPB (n = 619) and evaluated potential positive associations between CSA-AKI incidence and risk factors suggested by the first cohort study. RESULTS: Variable selection analysis revealed that parameters in clinical categories such as circulating inflammatory cells, CPB-related parameters, ventilation, or aging were potential CSA-AKI risk factors. Survival analyses revealed that increased counts of pre-operative circulating monocytes and neutrophils were associated with CSA-AKI incidence. Finally, in the second cohort study, we found that increased pre-operative circulating monocyte counts were associated with increased CSA-AKI incidence. CONCLUSIONS: Circulating monocyte counts in the pre-operative state are associated with increased risk of CSA-AKI development. This finding may be useful in stratifying patients for risk of developing CSA-AKI in routine clinical practice.

Complex and dynamic expression of cadherins in the embryonic marmoset cerebral cortex.

Cadherin is a cell adhesion molecule widely expressed in the nervous system. Previously, we analyzed the expression of nine classic cadherins (Cdh4, Cdh6, Cdh7, Cdh8, Cdh9, Cdh10, Cdh11, Cdh12, and Cdh20) and T-cadherin (Cdh13) in the developing postnatal common marmoset (Callithrix jacchus) brain, and found differential expressions between mice and marmosets. In this study, to explore primate-specific cadherin expression at the embryonic stage, we extensively analyzed the expression of these cadherins in the developing embryonic marmoset brain. Each cadherin showed differential spatial and temporal expression and exhibited temporally complicated expression. Furthermore, the expression of some cadherins differed from that in rodent brains, even at the embryonic stage. These results suggest the possibility that the differential expressions of diverse cadherins are involved in primate specific cortical development, from the prenatal to postnatal period.

Periostin, a neurite outgrowth-promoting factor, is expressed at high levels in the primate cerebral cortex.

Periostin (POSTN or osteoblast specific factor) is an extracellular matrix protein originally identified as a protein highly expressed in osteoblasts. Recently, periostin has been reported to function in axon regeneration and neuroprotection. In the present study, we focused on periostin function in cortical evolution. We performed a comparative gene expression analysis of periostin between rodents (mice) and primates (marmosets and macaques). Periostin was expressed at higher levels in the primate cerebral cortex compared to the mouse cerebral cortex. Furthermore, we performed overexpression experiments of periostin in vivo and in vitro. Periostin exhibited neurite outgrowth activity in cortical neurons. These results suggested the possibility that prolonged and increased periostin expression in the primate cerebral cortex enhances the cortical plasticity of the mammalian cerebral cortex.

Identification of tool use acquisition-associated genes in the primate neocortex.

Japanese macaques are able to learn how to use rakes to take food after only a few weeks of training. Since tool-use training induced rapid morphological changes in some restricted brain areas, this system will be a good model for studying the neural basis of plasticity in human brains. To examine the mechanisms of tool-use associated brain expansion on the molecular and cellular level, here, we performed comprehensive analysis of gene expressions with microarray. We identified various transcripts showing differential expression between trained and untrained monkeys in the region around the lateral and intraparietal sulci. Among candidates, we focused on genes related to synapse formation and function. Using quantitative reverse transcription-polymerase chain reaction and histochemical analysis, we confirmed at least three genes (ADAM19, SPON2, and WIF1) with statistically different expression levels in neurons and glial cells. Comparative analysis revealed that tool use-associated genes were more obviously expressed in macaque monkeys than marmosets or mice. Thus, our findings suggest that cognitive tasks induce structural changes in the neocortex via gene expression, and that learning-associated genes innately differ with relation to learning ability.

Complementary and dynamic type II cadherin expression associated with development of the primate visual system.

The middle temporal visual area (MT, also known as V5) is a visual association area that is particularly evolved in the primate brain. The MT receives input from the primary visual area (V1), constitutes part of the dorsal visual pathway, and plays an essential role in processing motion. Connections between the MT and V1 in the primate brain are formed after birth, and are related to the maturation of visual system. However, it remains to be determined what molecular mechanisms control the formation and maturation of the visual system. Cadherins are transmembrane proteins, originally isolated as cell adhesion molecules, which have multiple roles in synapse formation and function. To investigate potential involvement of cadherins in development of the primate visual system, we examined type II cadherin expression (cadherin-6, -8, -12) in cortical and thalamic visual areas of pre- and postnatal brains of the common marmoset (Callithrix jacchus). In the prenatal brain, cadherin-6 was dominantly expressed in the pulvino-MT pathway whereas cadherin-8 was dominant in the lateral geniculate nucleus (LGN)-V1 pathway. During postnatal development, there was a downregulation of cadherin-6 and upregulation of cadherin-8 expression in the MT. The timing of this cadherin exchange preceded the development of V1-MT connections. Our results suggest the possibility that changes in cadherin expression are involved in the development of the primate visual system, and that a switch in cadherin expression may be a general mechanism to control neural plasticity of highly cognitive abilities.

Efficacy and safety in mice of repeated, lifelong administration of an ANGPTL3 vaccine.

Previously, we reported that an ANGPTL3 vaccine is a hopeful therapeutic option against dyslipidemia. In our current study, we assess durability and booster effects of that vaccine over a period representing a mouse's lifespan. The vaccine remained effective for over one year, and booster vaccination maintained suppression of circulating triglyceride levels thereafter without major adverse effects on lungs, kidneys, or liver, suggesting vaccine efficacy and safety.

Comparative analysis of mineralocorticoid receptor expression among vocal learners (Bengalese finch and budgerigar) and non-vocal learners (quail and ring dove) has implications for the evolution of avian vocal learning.

Mineralocorticoid receptor is the receptor for corticosteroids such as corticosterone or aldosterone. Previously, we found that mineralocorticoid receptor was highly expressed in song nuclei of a songbird, Bengalese finch (Lonchura striata var. domestica). Here, to examine the relationship between mineralocorticoid receptor expression and avian vocal learning, we analyzed mineralocorticoid receptor expression in the developing brain of another vocal learner, budgerigar (Melopsittacus undulatus) and non-vocal learners, quail (Coturnix japonica) and ring dove (Streptopelia capicola). Mineralocorticoid receptor showed vocal control area-related expressions in budgerigars as Bengalese finches, whereas no such mineralocorticoid receptor expressions were seen in the telencephalon of non-vocal learners. Thus, these results suggest the possibility that mineralocorticoid receptor plays a role in vocal development of parrots as songbirds and that the acquisition of mineralocorticoid receptor expression is involved in the evolution of avian vocal learning.

RGM and its receptor neogenin regulate neuronal survival.

Repulsive guidance molecule (RGM) is an axon guidance protein that repels retinal axons upon activation of the neogenin receptor. To understand the functions of RGM-neogenin complexes in vivo, we used gene transfer technology to perturb their expression in the developing neural tube of chick embryos. Surprisingly, neogenin over-expression or RGM down-expression in the neural tube induces apoptosis. Neogenin pro-apoptotic activity in immortalized neuronal cells and in the neural tube is associated with the cleavage of its cytoplasmic domain by caspases. Thus neogenin is a dependence receptor inducing cell death in the absence of RGM, whereas the presence of RGM inhibits this effect.

Vaccine targeting ANGPTL3 ameliorates dyslipidemia and associated diseases in mouse models of obese dyslipidemia and familial hypercholesterolemia.

Dyslipidemia is a risk factor for cardiovascular disease (CVD), a major cause of death worldwide. Angiopoietin-like protein 3 (ANGPTL3), recognized as a new therapeutic target for dyslipidemia, regulates the metabolism of low-density lipoprotein-cholesterol (LDL-C) and triglycerides. Here, we design 3 epitopes (E1-E3) for use in development of a peptide vaccine targeting ANGPTL3 and estimate effects of each on obesity-associated dyslipidemia in B6.Cg-Lepob /J (ob/ob) mice. Vaccination with the E3 (32EPKSRFAMLD41) peptide significantly reduces circulating levels of triglycerides, LDL-C, and small dense (sd)-LDL-C in ob/ob mice and decreases obese-induced fatty liver. Moreover, E3 vaccination does not induce cytotoxicity in ob/ob mice. Interestingly, the effect of E3 vaccination on dyslipidemia attenuates development of atherosclerosis in B6.KOR/StmSlc-Apoeshl mice fed a high-cholesterol diet, which represent a model of severe familial hypercholesterolemia (FH) caused by ApoE loss of function. Taken together, ANGPTL3 vaccination could be an effective therapeutic strategy against dyslipidemia and associated diseases.

Vocal control area-related expression of neuropilin-1, plexin-A4, and the ligand semaphorin-3A has implications for the evolution of the avian vocal system.

The avian vocal system is a good model for exploring the molecular basis of neural circuit evolution related to behavioral diversity. Previously, we conducted a comparative gene expression analysis among two different families of vocal learner, the Bengalese finch (Lonchura striata var. domestica), a songbird, and the budgerigar (Melopsittacus undulatus), a parrot; and a non-learner, the quail (Coturnix coturnix), to identify various axon guidance molecules such as cadherin and neuropilin-1 as vocal control area-related genes. Here, we continue with this study and examine the expression of neuropilin and related genes in these species in more detail. We found that neuropilin-1 and its coreceptor, plexin-A4, were expressed in several vocal control areas in both Bengalese finch and budgerigar brains. In addition, semaphorin-3A, the ligand of neuropilin-1, expression was not detected in vocal control areas in both species. Furthermore, there was some similar gene expression in the quail brain. These results suggest the possibility that a change in the expression of a combination of semaphorin/neuropilin/plexin was involved in the acquisition of vocal learning ability during evolution.

Evolution and diversity in avian vocal system: an Evo-Devo model from the morphological and behavioral perspectives.

Birds use various vocalizations to mark their territory and attract mates. Three groups of birds (songbirds, parrots, and hummingbirds) learn their vocalizations through imitation. In the brain of such vocal learners, there is a neural network called the song system specialized for vocal learning and production. In contrast, birds such as chickens and pigeons do not have such a neural network and can only produce innate sounds. Since each avian species shows distinct, genetically inherited vocal learning abilities that are related to its morphology, the avian vocal system is a good model for studying the evolution of functional neural circuits. Nevertheless, studies on avian vocalization from an evolutionary developmental-biological (Evo-Devo) perspective are scant. In the present review, we summarize the results of songbird studies and our recent work that used the Evo-Devo approach to understand the evolution of the avian vocal system.

Intraretinal RGMa is involved in retino-tectal mapping.

The repulsive guidance molecule (RGMa) is involved in controlling the topography of retinal ganglion cell axons along the anterioposterior axis of the tectum. Here, we generated a new RGMa-monoclonal antibody and show that it is expressed in the developing retina, suggesting that it may regulate retinal axon pathfinding. We tested this hypothesis by using in ovo electroporation to either overexpress or downregulate RGMa in the eye. Anterograde labeling of retinal axons entering the optic tecta revealed abnormal phenotypes when RGMa expression is perturbed. These included the absence of terminal zone, the premature stalling of arborization of fibers, overshooting of terminal zone, aberrant axonal turns in the optic tectum and abnormal projections into deeper tectal layers. Moreover, RGMa overexpression frequently leads to intraretinal pathfinding errors. Thus, these data suggest that RGMa expression on retinal axons is a major determinant of topographic targeting in the retino-tectal projection and in the retina.

The slit receptor Rig-1/Robo3 controls midline crossing by hindbrain precerebellar neurons and axons.

During development, precerebellar neurons migrate dorsoventrally from the rhombic lip to the floor plate. Some of these neurons cross the midline while others stop. We have identified a role for the slit receptor Rig-1/Robo3 in directing this process. During their tangential migration, neurons of all major hindbrain precerebellar nuclei express high levels of Rig-1 mRNA. Rig-1 expression is rapidly downregulated as their leading process crosses the floor plate. Interestingly, most precerebellar nuclei do not develop normally in Rig-1-deficient mice, as they fail to cross the midline. In addition, inferior olivary neurons, which normally send axons into the contralateral cerebellum, project ipsilaterally in Rig-1 mutant mice. Similarly, neurons of the lateral reticular nucleus and basilar pons are unable to migrate across the floor plate and instead remain ipsilateral. These results demonstrate that Rig-1 controls the ability of both precerebellar neuron cell bodies and their axons to cross the midline.

Vocal area-related expression of the androgen receptor in the budgerigar (Melopsittacus undulatus) brain.

The androgen receptor is a steroid hormone receptor widely expressed in the vocal control nuclei in songbirds. Here, we analysed androgen receptor expression in the brains of juvenile and adult budgerigars. With a species-specific probe for budgerigar androgen receptor mRNA, we found that the androgen receptor was expressed in the vocal areas, such as the central nucleus of the lateral nidopallium, the anterior arcopallium, the oval nucleus of the mesopallium, the oval nucleus of the anterior nidopallium and the tracheosyringeal hypoglossal nucleus. With the present data, together with previous reports, it turned out that the androgen receptor expression in telencephalic vocal control areas is similar amongst three groups of vocal learners--songbirds, hummingbirds and parrots, suggesting the possibility that the androgen receptor might play a role in vocal development and that the molecular mechanism regulating the androgen receptor expression in the vocal areas might be important in the evolution of vocal learning.

Comparative analysis of gene expressions among avian brains: a molecular approach to the evolution of vocal learning.

Among avian species, three families of birds (songbirds, parrots, and hummingbirds) learn songs. In the brain of these vocal learners, there are neural networks called 'song systems' that specialize in song learning and production. To explore the evolution of the molecular basis of vocal learning, we conducted a comparative analysis of gene expression in vocal learners (Bengalese finches and budgerigars) and non-learners (quails and pigeons). The expression of one gene is similar in vocal learners, but that of other genes is highly diverse. In non-learners, by contrast, no nuclei-specific expression exists. These results suggest that songbirds and parrots acquired their song systems through both similar and different molecular mechanisms.

Repulsive guidance molecule plays multiple roles in neuronal differentiation and axon guidance.

Repulsive guidance molecule (RGM) is a membrane-bound protein originally isolated as a guidance molecule for retinal axons. Three RGM isoforms (RGMa-RGMc) exist in vertebrates. We showed previously that RGMa is a cell-survival factor in the neuroepithelium of chick embryos that suppresses the proapoptotic activity of its receptor neogenin. In the present study, we performed gain- and loss-of-function analysis of RGMa in chick embryos to further investigate RGMa function. We found that RGMa overexpression promotes neuronal differentiation, whereas RGMa small interference RNA represses it. Similar experiments conducted at later developmental stages using retroviral vectors reveal that perturbation of RGMa expression disturbs the retinotectal projection. Our work provides the first evidence for a role for RGMs in axon guidance in vivo. In addition, these results suggest that RGMa exerts multiple functions during neural development.

Cadherins: potential regulators in the faculty of language.

The cadherin superfamily is a large (now more than 100 proteins) protein family originally identified as cell adhesion molecules. Each cadherin shows distinct expression patterns in the nervous system, and their expressions are both spatially and temporally regulated and diverse among different species. Mounting evidence has suggested that cadherins play multiple roles in neural development and functions. Recently, using songbirds and mice, the potential role of cadherins in vocal behavior has been demonstrated. Here, we will briefly introduce general function of cadherins, and analyze the potential involvement of cadherins in vocal behaviors and their evolution.

Comparative analysis of protocadherin-11 X-linked expression among postnatal rodents, non-human primates, and songbirds suggests its possible involvement in brain evolution.

BACKGROUND: Protocadherin-11 is a cell adhesion molecule of the cadherin superfamily. Since, only in humans, its paralog is found on the Y chromosome, it is expected that protocadherin-11X/Y plays some role in human brain evolution or sex differences. Recently, a genetic mutation of protocadherin-11X/Y was reported to be associated with a language development disorder. Here, we compared the expression of protocadherin-11 X-linked in developing postnatal brains of mouse (rodent) and common marmoset (non-human primate) to explore its possible involvement in mammalian brain evolution. We also investigated its expression in the Bengalese finch (songbird) to explore a possible function in animal vocalization and human language faculties. METHODOLOGY/PRINCIPAL FINDINGS: Protocadherin-11 X-linked was strongly expressed in the cerebral cortex, hippocampus, amygdala and brainstem. Comparative analysis between mice and marmosets revealed that in certain areas of marmoset brain, the expression was clearly enriched. In Bengalese finches, protocadherin-11 X-linked was expressed not only in nuclei of regions of the vocal production pathway and the tracheosyringeal hypoglossal nucleus, but also in areas homologous to the mammalian amygdala and hippocampus. In both marmosets and Bengalese finches, its expression in pallial vocal control areas was developmentally regulated, and no clear expression was seen in the dorsal striatum, indicating a similarity between songbirds and non-human primates. CONCLUSIONS/SIGNIFICANCE: Our results suggest that the enriched expression of protocadherin-11 X-linked is involved in primate brain evolution and that some similarity exists between songbirds and primates regarding the neural basis for vocalization.

Differential cadherin expression in the developing postnatal telencephalon of a New World monkey.

Cadherins are cell adhesion molecules widely expressed in the nervous system, where they play various roles in neural patterning, nuclei formation, axon guidance, and synapse formation and function. Although many published articles have reported on cadherin expression in rodents and ferrets, there are limited data on their expression in primate brains. In this study, in situ hybridization analysis was performed for 10 cadherins [nine classic cadherins (Cdh4, -6, -7, -8, -9, -10, -11, -12, and -20) and T-cadherin (Cdh13)] in the developing postnatal telencephalon of the common marmoset (Callithrix jacchus). Each cadherin showed broad expression in the cerebral cortex, basal ganglia, amygdala, and hippocampus, as previously shown in the rodent brain. However, detailed expression patterns differed between rodents and marmosets. In contrast to rodents, cadherin expression was reduced overall and localized to restricted areas of the brain during the developmental process, suggesting that cadherins are more crucially involved in developmental or maturation processes rather than in neural functioning. These results also highlight the possibility that restricted/less redundant cadherin expression allows primate brains to generate functional diversity among neurons, allowing morphological and functional differences between rodents and primates.