Generation and characterization of a novel shoulder contracture mouse model

Frozen shoulder is a relatively common disorder that leads to severe pain and stiffness in the shoulder joint. Although this disorder is self‐limiting in nature, the symptoms often persist for years, resulting in severe disability. Recent studies using human specimens and animal models have shown distinct changes in the gene expression patterns in frozen shoulder tissue, indicating that novel therapeutic intervention could be achieved by controlling the genes that are potentially involved in the development of frozen shoulder. To achieve this goal, it is imperative to develop a reliable animal joint contracture model in which gene expression can be manipulated by gene targeting and transgenic technologies. Here, we describe a novel shoulder contracture mouse model. We found that this model mimics the clinical presentation of human frozen shoulder and recapitulates the changes in the gene expression pattern and the histology of frozen shoulder and joint contracture in humans and other larger animal models. The model is highly reproducible, without any major complications. Therefore, the present model may serve as a useful tool for investigating frozen shoulder etiology and for identifying its potential target genes. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 33:1732–1738, 2015.     

Regulating emotion to improve physical health through the amygdala

The opinion of mind–body interaction has been increasingly acknowledged in recent years, as exemplified by accumulating evidence indicating that physical health (body) is associated with emotion and emotion regulation (mind). Yet, the neural basis linking emotion regulation with physical health remains largely uninvestigated. Here we used magnetic resonance imaging to investigate the neural basis of this pathway in a large population of healthy young adults. With a systematic study revealing the association of self-reported physical health and emotion traits of personality and general affective experiences, we further demonstrated that, for better physical health, individuals needed to regulate their emotion more effectively. Importantly, individuals who had larger gray matter (GM) volume in the amygdala reported not only a higher ability of emotion regulation but also better physical health. Further, GM volume in the amygdala mediated the correlation between emotion regulation ability and physical health. Our findings suggest that the amygdala plays a critical role in the neural circuit through which emotion regulation may influence physical health. Therefore, our study takes the first step toward exploring the neuroanatomical basis for body–mind interaction and may inform interventions aimed at promoting physical health by augmenting skills of emotion regulation.     

Neural indicators of emotion regulation via acceptance vs reappraisal in remitted major depressive disorder

Mood disorders are characterized by impaired emotion regulation abilities, reflected in alterations in frontolimbic brain functioning during regulation. However, little is known about differences in brain function when comparing regulatory strategies. Reappraisal and emotional acceptance are effective in downregulating negative affect, and are components of effective depression psychotherapies. Investigating neural mechanisms of reappraisal vs emotional acceptance in remitted major depressive disorder (rMDD) may yield novel mechanistic insights into depression risk and prevention. Thirty-seven individuals (18 rMDD, 19 controls) were assessed during a functional magnetic resonance imaging task requiring reappraisal, emotional acceptance or no explicit regulation while viewing sad images. Lower negative affect was reported following reappraisal than acceptance, and was lower following acceptance than no explicit regulation. In controls, the acceptance > reappraisal contrast revealed greater activation in left insular cortex and right prefrontal gyrus, and less activation in several other prefrontal regions. Compared with controls, the rMDD group had greater paracingulate and right midfrontal gyrus (BA 8) activation during reappraisal relative to acceptance. Compared with reappraisal, acceptance is associated with activation in regions linked to somatic and emotion awareness, although this activation is associated with less reduction in negative affect. Additionally, a history of MDD moderated these effects.     

Arteriolar responses to vasodilator stimuli and elevated P(O2) in renin congenic and Dahl salt-sensitive rats

OBJECTIVES: Angiotensin II suppression leads to impaired vascular relaxation in normotensive animals on a high-salt diet. The goal of this study was to determine whether normal vascular reactivity could be restored by transferring the chromosomal region carrying the Dahl salt-resistant (R) renin gene into the Dahl salt-sensitive (SS) genetic background in a strain of renin congenic rats (RGRR). METHODS: Male RGRR and SS rats were fed low-salt (0.4%) and high-salt (4%) diets for 4 weeks. The responses of cremaster muscle arterioles to acetylcholine (ACh), sodium nitroprusside (SNP), and elevated PO2 were assessed using video microscopy. RESULTS: ACh-induced dilation was significantly enhanced in RGRR on a high-salt diet compared to SS rats, while dilation to the NO donor SNP was similar in both strains. A high-salt diet significantly enhanced arteriolar constriction in response to elevated PO2, in both SS and RGRR rats. CONCLUSIONS: These data suggest that transfer of the chromosomal region containing the renin gene is crucial in the recovery of ACh-induced dilation of arterioles in RGRR rats vs. SS rats, and that factors in the SS genetic background contribute to an enhanced sensitivity to elevated PO2, independent of genes on chromosome 13.     

儿童Lowe综合征OCRL基因突变2例报告

目的探讨儿童Lowe综合征的临床特点和基因特征。方法分析2例Lowe综合征患儿的临床资料和OCRL基因检测结果,并复习相关文献。结果 2例患儿均为男性,均存在小分子蛋白尿、高钙尿症、佝偻病和肾结石。例2患儿还有轻度代谢性酸中毒、糖尿和隐睾症。例1患儿生后不久发现有视力异常和先天性白内障,并行手术治疗,同时存在精神运动发育落后,头颅磁共振成像(MRI)示胼胝体发育不良。例2患儿就诊时肾外症状不明显,但眼科检查发现有先天性白内障,头颅MRI示脑发育低下、脑白质髓鞘化延迟,且随访过程中逐渐出现智力发育落后。OCRL基因检测发现2个突变,例1为剪切位点突变NG 008638.1:g.46846-46848del TAA/ins C,例2为缺失移码突变NM000276.3:c.321del C,两种突变以前文献均未报道。结论 Lowe综合征的诊断主要通过临床表现和OCRL基因检测,对于有先天性白内障、肾小管病变的患儿需要与Lowe综合征鉴别。本研究发现2个OCRL基因的新突变。     

First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer

Improved understanding of the relationship among structure, dynamics, and function for the enzyme phenylalanine hydroxylase (PAH) can lead to needed new therapies for phenylketonuria, the most common inborn error of amino acid metabolism. PAH is a multidomain homo-multimeric protein whose conformation and multimerization properties respond to allosteric activation by the substrate phenylalanine (Phe); the allosteric regulation is necessary to maintain Phe below neurotoxic levels. A recently introduced model for allosteric regulation of PAH involves major domain motions and architecturally distinct PAH tetramers [Jaffe EK, Stith L, Lawrence SH, Andrake M, Dunbrack RL, Jr (2013) Arch Biochem Biophys 530(2):73–82]. Herein, we present, to our knowledge, the first X-ray crystal structure for a full-length mammalian (rat) PAH in an autoinhibited conformation. Chromatographic isolation of a monodisperse tetrameric PAH, in the absence of Phe, facilitated determination of the 2.9 Å crystal structure. The structure of full-length PAH supersedes a composite homology model that had been used extensively to rationalize phenylketonuria genotype–phenotype relationships. Small-angle X-ray scattering (SAXS) confirms that this tetramer, which dominates in the absence of Phe, is different from a Phe-stabilized allosterically activated PAH tetramer. The lack of structural detail for activated PAH remains a barrier to complete understanding of phenylketonuria genotype–phenotype relationships. Nevertheless, the use of SAXS and X-ray crystallography together to inspect PAH structure provides, to our knowledge, the first complete view of the enzyme in a tetrameric form that was not possible with prior partial crystal structures, and facilitates interpretation of a wealth of biochemical and structural data that was hitherto impossible to evaluate.Mammalian phenylalanine hydroxylase (PAH) (EC 1.14.16.1) is a multidomain homo-multimeric protein whose dysfunction causes the most common inborn error in amino acid metabolism, phenylketonuria (PKU), and milder forms of hyperphenylalaninemia (OMIM 261600) (1). PAH catalyzes the hydroxylation of phenylalanine (Phe) to tyrosine, using nonheme iron and the cosubstrates tetrahydrobiopterin and molecular oxygen (2, 3). A detailed kinetic mechanism has recently been derived from elegant single-turnover studies (4). PAH activity must be carefully regulated, because although Phe is an essential amino acid, high Phe levels are neurotoxic. Thus, Phe allosterically activates PAH by binding to a regulatory domain. Phosphorylation at Ser16 potentiates the effects of Phe, with phosphorylated PAH achieving full activation at lower Phe concentrations than the unphosphorylated protein (5, 6). Allosteric activation by Phe is accompanied by a major conformational change, as evidenced by changes in protein fluorescence and proteolytic susceptibility, and by stabilization of a tetrameric conformer (3).There are >500 disease-associated missense variants of human PAH; the amino acid substitutions are distributed throughout the 452-residue protein and among all its domains (Fig. 1A) (7–9). Of those disease-associated variants that have been studied in vitro (e.g., ref. 10), some confound the allosteric response, and some are interpreted as structurally unstable. We also suggest that the activities of some disease-associated variants may be dysregulated by an altered equilibrium among conformers having different intrinsic levels of activity, arguing by analogy to the enzyme porphobilinogen synthase (PBGS) and its porphyria-associated variants (11). Consistent with this notion, we have recently established that PAH can assemble into architecturally distinct tetrameric conformers (12), and propose that these conformers differ in activity due to differences in active-site access. This idea has important implications for drug discovery, as it implies that small molecules could potentially modulate the conformational equilibrium of PAH, as has already been demonstrated for PBGS (e.g., ref. 13). Deciphering the relationship among PAH structure, dynamics, and function is a necessary first step in testing this hypothesis.Open in a separate windowFig. 1.The structure of PAH. (A) The annotated domain structure of mammalian PAH. (B) The 2.9 Å PAH crystal structure in orthogonal views, colored as in part A, subunit A is shown in ribbons; subunit B is as a Cα trace; subunit C is in sticks; and subunit D is in transparent spheres. In cyan, the subunits are labeled near the catalytic domain (Top); in red, they are labeled near the regulatory domain (Bottom). The dotted black circle illustrates the autoregulatory domain partially occluding the enzyme active site (iron, in orange sphere). (C) Comparison of the subunit structures of full-length PAH and those of the composite homology model; the subunit overlay aligns residues 144–410. The four subunits of the full-length PAH structure (the diagonal pairs of subunits are illustrated using either black or white) are aligned with the two subunits of 2PAH (cyan) and the one subunit of 1PHZ (orange). The catalytic domain is in spheres, the regulatory domain is in ribbons, and the multimerization domain is as a Cα trace. The arrow denotes where the ACT domain and one helix of 2PAH conflict.Numerous crystal structures are known for one- and two-domain constructs of mammalian PAH (14).

Table S1.

Mammalian PAH structures available in the PDB (August 2015)

神经纤毛蛋白及其配体的免疫调节机制

神经纤毛蛋白(NRP)是信号素(Sema)家族和血管内皮生长因子(VEGF)家族的共同受体.通过Sema家族和VEGF家族,NRP分别在神经元和心血管发育中起着重要作用.NRP通过与其配体的其他受体结合,形成复合体,如Sema家族的神经丛蛋白和VEGF家族的VEGF受体(VEGFR)2,进而加强其信号传导,从而调节一些生理过程,其中最重要的为胞迁移.研究证实,NRP在一些疾病病理相关过程中亦起着重要作用.NRP可高表达于多种肿瘤细胞,这提示NRP是肿瘤治疗的一个潜在靶点.此外,在免疫系统中,NRP通过Sema3A参与免疫负调节,为治疗自身免疫疾病提供新思路.笔者拟就NRP及其配体的免疫调节机制进行综述如下.     

Alterations of gene profiles in Leydig-cell-regenerating adult rat testis after ethane dimethane sulfonate-treatment

Only occupying about 1%–5% of total testicular cells, the adult Leydig cell (ALC) is a unique endocrine cell that produces androgens. Rat Leydig cells regenerate after these cells in the testis are eliminated with ethane dimethane sulfonate (EDS). In this study, we have characterized Leydig cell regeneration and messenger ribonucleic acids (mRNA) profiles of EDS treated rat testes. Serum testosterone, testicular gene profiling and some steroidogenesis-related proteins were analyzed at 7, 21, 35 and 90 days after EDS treatment. Testicular testosterone levels declined to undetectable levels until 7 days after treatment and then started to recover. Seven days after treatment, 81 mRNAs were down-regulated greater than or equal to two-fold, with 48 becoming undetectable. These genes increased their expression 21 days and completely returned to normal levels 90 days after treatment. The undetectable genes include steroidogenic pathway proteins: steroidogenic acute regulatory protein, Scarb1, Cyp11a1, Cyp17a1, Hsd3b1, Cyp1b1 and Cyp2a1. Seven days after treatment, there were 89 mRNAs up-regulated two-fold or more including Pkib. These up-regulated mRNAs returned to normal 90 days after treatment. Cyp2a1 did not start to recover until 35 days after treatment, indicating that this gene is only expressed in ALCs not in the precursor cells. Quantitative polymerase chain reaction, western blotting and semi-quantitative immunohistochemical staining using tissue array confirmed the changes of several randomly picked genes and their proteins.