Gene expression changes in aging retinal microglia: relationship to microglial support functions and regulation of activation

Microglia, the resident immune cells of the central nervous system (CNS), are thought to contribute to the pathogenesis of age-related neurodegenerative disorders. It has been hypothesized that microglia undergo age-related changes in gene expression patterns that give rise to pathogenic phenotypes. We compared the gene expression profiles in microglia isolated ex vivo from the retinas of mice ranging from early adulthood to late senescence. We discovered that microglial gene expression demonstrated progressive change with increasing age, and involved genes that regulate microglial supportive functions and immune activation. Molecular pathways involving immune function and regulation, angiogenesis, and neurotrophin signaling demonstrated age-related change. In particular, expression levels of complement genes, C3 and CFB, previously associated with age-related macular degeneration (AMD), increased with aging, suggesting that senescent microglia may contribute to complement dysregulation during disease pathogenesis. Taken together, senescent microglia demonstrate age-related gene expression changes capable of altering their constitutive support functions and regulation of their activation status in ways relating to neuroinflammation and neurodegeneration in the CNS.     

A potential association of RNF219-AS1 with ADHD: Evidence from categorical analysis of clinical phenotypes and from quantitative exploration of executive function and white matter microstructure endophenotypes

AimsAttention‐deficit/hyperactivity disorder (ADHD) is a neuropsychiatric disorder of substantial heritability, yet emerging evidence suggests that key risk variants might reside in the noncoding regions of the genome. Our study explored the association of lncRNAs (long noncoding RNAs) with ADHD as represented at three different phenotypic levels guided by the Research Domain Criteria (RDoC) framework: (i) ADHD caseness and symptom dimension, (ii) executive functions as functional endophenotype, and (iii) potential genetic influence on white matter architecture as brain structural endophenotype.MethodsGenotype data of 107 tag single nucleotide polymorphisms (SNP) from 10 candidate lncRNAs were analyzed in 1040 children with ADHD and 630 controls of Chinese Han descent. Executive functions including inhibition and set‐shifting were assessed by STROOP and trail making tests, respectively. Imaging genetic analyses were performed in a subgroup of 33 children with ADHD and 55 controls using fractional anisotropy (FA).ResultsOne SNP rs3908461 polymorphism in RNF219‐AS1 was found to be significantly associated with ADHD caseness: with C‐allele detected as the risk genotype in the allelic model (P = 8.607E‐05) and dominant genotypic model (P = 9.628E‐05). Nominal genotypic effects on inhibition (p = 0.020) and set‐shifting (p = 0.046) were detected. While no direct effect on ADHD core symptoms was detected, mediation analysis suggested that SNP rs3908461 potentially exerted an indirect effect through inhibition function [B = 0.21 (SE = 0.12), 95% CI = 0.02‐0.49]. Imaging genetic analyses detected significant associations between rs3908461 genotypes and FA values in corpus callosum, left superior longitudinal fasciculus, left posterior limb of internal capsule, left posterior thalamic radiate (include optic radiation), and the left anterior corona radiate (P _{FWE corrected} < 0.05).ConclusionOur present study examined the potential roles of lncRNA in genetic etiological of ADHD and provided preliminary evidence in support of the potential RNF219‐AS1 involvement in the pathophysiology of ADHD in line with the RDoC framework.     

High oxygen environment during pregnancy rescues sickle cell anemia mice from prenatal death

Several mouse models of sickle cell disease have been developed for the study of the pathophysiology of sickle cell disease and the investigation of drug and gene therapies. In previous years, we produced a sickle cell anemia mouse model in which the endogenous mouse α- and β-globin genes were knocked out and replaced by the human α- and β^s-globin transgenes. The β^s-globin gene was contained in a 240 kb YAC that preserved the entire native genomic context of the β-globin locus. These mice have hemolytic anemia, reticulocytosis and irreversible sickle cells in the peripheral blood, as well as other pathological features of sickle cell disease. However, in the embryo, the γ-globin, like the mouse embryonic globin, declined quickly, and was replaced by β^s-globin expression from 12 days of gestation. The low level of fetal hemoglobin expression in utero led to intrauterine sickling and fetal death so that very few live-born sickle cell anemia mice could be obtained. To rescue these mice from intrauterine death, we investigated the effect of placing the pregnant mothers in a high O₂ environment. From the tenth day of gestation onwards, we placed the mothers into a chamber containing 50% O₂ and kept them with the newborn pups in it for another 10 days after birth. The frequency of sickle cell anemia mice we obtained was increased from less than 2% to 35%. The survived sickle cell anemia mice develop congestion, atrophy, and infarcts in multiple organs similar to those found in patients with sickle cell disease. We conclude that a high oxygen environment can be used to obtain more sickle cell anemia mice in those models that have a high perinatal mortality. The higher yield of these mice has facilitated physiological and therapeutic studies of sickle cell anemia.     

Direct sequencing of SARS-coronavirus S and N genes from clinical specimens shows limited variation

Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) emerged, in November 2002, as a novel agent causing severe respiratory illness. To study sequence variation in the SARS-CoV genome, we determined the nucleic acid sequence of the S and N genes directly from clinical specimens from 10 patients--1 specimen with no matched SARS-CoV isolate, from 2 patients; multiple specimens from 3 patients; and matched clinical-specimen/cell-culture-isolate pairs from 6 patients. We identified 3 nucleotide substitutions that were most likely due to natural variation and 2 substitutions that arose after cell-culture passage of the virus. These data demonstrate the overall stability of the S and N genes of SARS-CoV over 3 months during which a minimum of 4 generations for transmission events occurred. These findings are a part of the expanding investigation of the evolution of how this virus adapts to a new host.     

Residues Distal to the Active Site Contribute to Enhanced Catalytic Activity of Variant and Hybrid β-Lactamases Derived from CTX-M-14 and CTX-M-15

A variety of CTX-M-type extended-spectrum β-lactamases (ESBLs), including hybrid ones, have been reported in China that are uncommon elsewhere. To better characterize the substrate profiles and enzymatic mechanisms of these enzymes, we performed comparative kinetic analyses of both parental and hybrid CTX-M enzymes, including CTX-M-15, -132, -123, -64, -14 and -55, that are known to confer variable levels of β-lactam resistance in the host strains. All tested enzymes were susceptible to serine β-lactamase inhibitors, with sulbactam exhibiting the weakest inhibitory effects. CTX-M-55, which differs from CTX-M-15 by one substitution, A⁷⁷V, displayed enhanced catalytic activity (k_cat/K_m) against expanded-spectrum cephalosporins (ESCs). CTX-M-55 exhibits higher structure stability, most likely by forming hydrophobic interactions between A⁷⁷V and various key residues in different helices, thereby stabilizing the core architecture of the helix cluster, and indirectly contributes to a more stable active site conformation, which in turn shows higher catalytic efficiency and is more tolerant to temperature change. Analyses of the hybrids and their parental prototypes showed that evolution from CTX-M-15 to CTX-M-132, CTX-M-123, and CTX-M-64, characterized by gradual enhancement of catalytic activity to ESCs, was attributed to introduction of different substitutions to amino acids distal to the active site of CTX-M-15. Similarly, the increased hydrolytic activities against cephalosporins and sensitivity to β-lactamase inhibitors, clavulanic acid and sulbactam, of CTX-M-64 were partly due to the amino acids that were different from CTX-M-14 and located at both the C and N termini of CTX-M-64. These data indicate that residues distal to the active site of CTX-Ms contributed to their enhanced catalytic activities to ESCs.