Primary osteoarthritis chondrocyte map of chromatin conformation reveals novel candidate effector genes.

OBJECTIVES: Osteoarthritis is a complex disease with a huge public health burden. Genome-wide association studies (GWAS) have identified hundreds of osteoarthritis-associated sequence variants, but the effector genes underpinning these signals remain largely elusive. Understanding chromosome organisation in three-dimensional (3D) space is essential for identifying long-range contacts between distant genomic features (e.g., between genes and regulatory elements), in a tissue-specific manner. Here, we generate the first whole genome chromosome conformation analysis (Hi-C) map of primary osteoarthritis chondrocytes and identify novel candidate effector genes for the disease. METHODS: Primary chondrocytes collected from 8 patients with knee osteoarthritis underwent Hi-C analysis to link chromosomal structure to genomic sequence. The identified loops were then combined with osteoarthritis GWAS results and epigenomic data from primary knee osteoarthritis chondrocytes to identify variants involved in gene regulation via enhancer-promoter interactions. RESULTS: We identified 345 genetic variants residing within chromatin loop anchors that are associated with 77 osteoarthritis GWAS signals. Ten of these variants reside directly in enhancer regions of 10 newly described active enhancer-promoter loops, identified with multiomics analysis of publicly available chromatin immunoprecipitation sequencing (ChIP-seq) and assay for transposase-accessible chromatin using sequencing (ATAC-seq) data from primary knee chondrocyte cells, pointing to two new candidate effector genes SPRY4 and PAPPA (pregnancy-associated plasma protein A) as well as further support for the gene SLC44A2 known to be involved in osteoarthritis. For example, PAPPA is directly associated with the turnover of insulin-like growth factor 1 (IGF-1) proteins, and IGF-1 is an important factor in the repair of damaged chondrocytes. CONCLUSIONS: We have constructed the first Hi-C map of primary human chondrocytes and have made it available as a resource for the scientific community. By integrating 3D genomics with large-scale genetic association and epigenetic data, we identify novel candidate effector genes for osteoarthritis, which enhance our understanding of disease and can serve as putative high-value novel drug targets.

On pharyngealized vowels in Northern Horpa: An acoustic and ultrasound study.

In the Northern Horpa (NH) language of Sichuan, vowels are divided between plain and pharyngealized sets, with the latter pronounced with auxiliary articulatory gestures involving more constriction in the vocal tract. The current study examines how the NH vocalic contrast is manifested in line with the process of pharyngealization both acoustically and articulatorily, based on freshly gathered data from two varieties of the language (i.e., Rtsangkhog and Yunasche). Along with formant analyses, ultrasound imaging was employed to capture the tongue postures and positions during vowel production. The results show that in contrast with plain vowels, pharyngealized vowels generally feature lower F2 values and higher F1 and F3 values. Mixed results for F2 and F3 suggest that the quality contrasts are vowel-dependent. Ultrasound images, on the other hand, reveal that the vocalic distinction is affected by different types of tongue movements, including retraction, backing, and double bunching, depending on the inherent tongue positions for each vowel. The two NH varieties investigated are found to display differential formant changes and different types of tongue displacements. The formant profiles along with ultrasound images support the view that the production of the NH phonologically marked vowels is characteristic of pharyngealization.

Geometric stability of topological lattice phases.

The fractional quantum Hall (FQH) effect illustrates the range of novel phenomena which can arise in a topologically ordered state in the presence of strong interactions. The possibility of realizing FQH-like phases in models with strong lattice effects has attracted intense interest as a more experimentally accessible venue for FQH phenomena which calls for more theoretical attention. Here we investigate the physical relevance of previously derived geometric conditions which quantify deviations from the Landau level physics of the FQHE. We conduct extensive numerical many-body simulations on several lattice models, obtaining new theoretical results in the process, and find remarkable correlation between these conditions and the many-body gap. These results indicate which physical factors are most relevant for the stability of FQH-like phases, a paradigm we refer to as the geometric stability hypothesis, and provide easily implementable guidelines for obtaining robust FQH-like phases in numerical or real-world experiments.

Model characterization of gapless edge modes of topological insulators using intermediate Brillouin-zone functions.

We characterize gapless edge modes in translation invariant topological insulators. We show that the edge mode spectrum is a continuous deformation of the spectrum of a certain gluing function defining the occupied state bundle over the Brillouin zone. Topologically nontrivial gluing functions, corresponding to nontrivial bundles, then yield edge modes exhibiting spectral flow. We illustrate our results for the case of chiral edge states in two-dimensional Chern insulators, as well as helical edges in quantum spin Hall states.

Theory of minimum spanning trees. I. Mean-field theory and strongly disordered spin-glass model.

The minimum spanning tree (MST) is a combinatorial optimization problem: given a connected graph with a real weight ("cost") on each edge, find the spanning tree that minimizes the sum of the total cost of the occupied edges. We consider the random MST, in which the edge costs are (quenched) independent random variables. There is a strongly disordered spin-glass model due to Newman and Stein [Phys. Rev. Lett. 72, 2286 (1994)], which maps precisely onto the random MST. We study scaling properties of random MSTs using a relation between Kruskal's greedy algorithm for finding the MST, and bond percolation. We solve the random MST problem on the Bethe lattice (BL) with appropriate wired boundary conditions and calculate the fractal dimension D=6 of the connected components. Viewed as a mean-field theory, the result implies that on a lattice in Euclidean space of dimension d , there are of order W(d-D) large connected components of the random MST inside a window of size W , and that d=d(c)=D=6 is a critical dimension. This differs from the value 8 suggested by Newman and Stein. We also critique the original argument for 8, and provide an improved scaling argument that again yields d(c)=6 . The result implies that the strongly disordered spin-glass model has many ground states for d>6 , and only of order one below six. The results for MSTs also apply on the Poisson-weighted infinite tree, which is a mean-field approach to the continuum model of MSTs in Euclidean space, and is a limit of the BL. In a companion paper we develop an epsilon=6-d expansion for the random MST on critical percolation clusters.

Theory of minimum spanning trees. II. Exact graphical methods and perturbation expansion at the percolation threshold.

Continuing the program begun by the authors in a previous paper, we develop an exact low-density expansion for the random minimum spanning tree (MST) on a finite graph and use it to develop a continuum perturbation expansion for the MST on critical percolation clusters in space dimension d . The perturbation expansion is proved to be renormalizable in d=6 dimensions. We consider the fractal dimension D(p) of paths on the latter MST; our previous results lead us to predict that D(p)=2 for d>d(c)=6 . Using a renormalization-group approach, we confirm the result for d>6 and calculate D(p) to first order in epsilon=6-d for d<6 using the connection with critical percolation, with the result D(p)=2-epsilon/7+O(epsilon(2)) .

Bax, Bcl-2, and cyclin expression and apoptosis in rat substantia nigra during development.

Naturally occurring cell death via apoptosis has been reported in the substantia nigra of rats during development, culminating during the perinatal period. Cellular pathways leading to apoptotic death of developing nigral dopamine neurons remain unknown, although the apoptotic mediator, caspase 3, has been shown to be activated during this process. Our previous results demonstrated the inability of antioxidants to rescue the nigral dopamine neurons that undergo apoptosis during development. In the present study, we investigated using immunohistochemistry the expression of cyclins D1, D3, and E in the substantia nigra during pre- and postnatal development, since their re-expression in postmitotic neurons has been proposed to contribute to developmental apoptosis. We also investigated by Western blot analysis of nigral tissue isolated during the first postnatal week the expression of the anti- and pro-apoptotic proteins, Bcl-2 and Bax, respectively, since altered Bcl-2 expression during developmental apoptosis has been described. During apoptotic death of nigral dopamine neurons in development, we detected a significant increase in the Bax:Bcl-2 ratio, which is consistent with enhanced apoptosis. There were no changes in the expression of the cyclins during the same apoptotic period. These novel findings suggest that nigral dopamine neurons undergo developmental apoptotic death through a Bax:Bcl-2-sensitive pathway that does not involve cyclin mediation.

Lipid peroxidation-mediated oxidative stress and dopamine neuronal apoptosis in the substantia nigra during development.

The cellular pathways underlying naturally occurring neuronal apoptosis in the rat substantia nigra (SN) during the perinatal period remain largely unknown. Determining the mediators of this process in development may shed light on causes of premature neuronal death in adult neurodegenerative disorders, including the loss of dopamine neurons in Parkinson's disease. In the present study, we investigated whether lipid peroxidation-mediated oxidative stress mediates developmental death of nigral neurons by (1) establishing the profile of lipid peroxidation and other oxidative stress markers throughout the postnatal period both in the SN and striatum, and (2) examining whether the inhibitor of lipid peroxidation, alpha-tocopherol, protects these neurons from death. In addition to monitoring, the level of lipid peroxidation throughout development, we also measured the activities of three antioxidant enzymes, namely superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx). We have shown that lipid peroxidation and SOD activity progressively increased from postnatal day (PND) 3 to PND 42 in both SN and striatum. During this period, GPx activity remained stable, while catalase activity transiently increased at PND 8 only in the SN. Furthermore, alpha-tocopherol treatment from embryonic day 18 to PND 2 did not reduce the number of apoptotic neurons at PND 3. These results do not support the hypothesis that lipid peroxidation-mediated oxidative stress is the major mediator of nigral dopamine neuronal apoptosis during the perinatal period.

Ascorbate prevents the interaction of superoxide and nitric oxide only at very high physiological concentrations.

The bioactivity of nitric oxide (.NO) depends, in part, on its interaction with superoxide. Usually, superoxide dismutase (SOD) preserves .NO bioactivity by limiting the availability of superoxide. Ascorbic acid also effectively scavenges superoxide, but the extent to which this interaction is necessary for intact .NO bioactivity is not known. Therefore, the present study examined the effect of ascorbic acid on .NO bioactivity with isolated rabbit arterial segments. A steady flux of superoxide (1.15 to 2.3 micromol . L-1 . min-1) produced either by pyrogallol autoxidation or a hypoxanthine/xanthine oxidase system inhibited endothelium-derived .NO-mediated arterial relaxation elicited by acetylcholine. This effect of superoxide was completely blocked by SOD (300 IU/mL) and the manganese SOD mimic EUK-8 (300 micromol/L) and partially inhibited by ascorbic acid (10 mmol/L). Lower concentrations of ascorbic acid were ineffective despite scavenging >90% of superoxide. We increased the endogenous flux of superoxide (3.2+/-0.3-fold) by inhibiting vascular copper-zinc SOD with diethyldithiocarbamate. This increased endogenous flux of superoxide produced an impairment of .NO-mediated arterial relaxation that was reversed by EUK-8 (300 micromol/L) but not ascorbic acid (10 mmol/L) despite equivalent scavenging of the endogenous superoxide flux. We used 3-nitrotyrosine formation (from peroxynitrite) as an indicator of .NO interaction with superoxide and found that SOD and EUK-8 compete more effectively with .NO for superoxide than does ascorbic acid. These data indicate that preservation of .NO bioactivity by superoxide scavengers depends not only on superoxide scavenging activity, but also on the rate of superoxide scavenging. Normal extracellular concentrations of ascorbic acid (30 to 150 micromol/L) are not likely to prevent the interaction of .NO with superoxide under physiological conditions.

Vasodilatory properties of recombinant maxadilan.

Maxadilan is a peptide from the salivary gland of the sand fly Lutzomyia longipalpis, a vector for leishmaniasis. Cutaneous injection of femtomolar quantities of maxadilan produces long-lasting erythema, making it the most potent vasodilator known. Isolated rabbit thoracic and abdominal aorta, carotid artery, and iliac artery demonstrated dose-dependent arterial relaxation in response to maxadilan with a mean effective concentration (EC50) of 2.7 +/- 1.5, 2.1 +/- 0.5, 2.6 +/- 0.4, and 1.9 +/- 0.5 nM, respectively. Maxadilan proved to be at least sevenfold more potent than nitroglycerin in each arterial bed (EC50 = 25 +/- 12, 32 +/- 9, 37 +/- 10, and 22 +/- 13 nM, respectively; P < 0.05 for each vs. maxadilan). Arterial relaxation to maxadilan was independent of endothelium and was equipotent in the thoracic and abdominal aorta, carotid artery, and iliac artery. Arterial relaxation to maxadilan was not inhibited by K(+)-channel antagonists, methylene blue, quinacrine, or ouabain. Maxadilan-mediated arterial relaxation was found to be adenosine 3',5'-cyclic monophosphate (cAMP) dependent, as it was potentiated by the phosphodiesterase inhibitors 3-isobutyl-1-methylxanthine and theophylline, and it was inhibited by the protein kinase A inhibitor H-89. Consistent with this observation, incubation of thoracic aorta with maxadilan (0.1 microM) produced a time-dependent increase in arterial cAMP content coincident with arterial relaxation. Using rabbit aortic smooth muscle cells, we also observed a time-dependent reduction in intracellular calcium in response to maxadilan. Thus these data indicate that maxadilan, a peptide from the sand fly salivary gland, is a potent vasodilator that reduces intracellular calcium through a cAMP-dependent mechanism.

Sentence processing by adolescents with and without mental retardation.

A more thorough test of the hypothesis that persons with mental retardation are less likely to construct semantically integrated representations of sentences that they hear than are subjects without mental retardation (Merrill & Bilsky, 1989; Merrill & Mar, 1987) was provided. A series of sentences were presented to adolescents with and without mental retardation. Their memory for the object nouns of the sentences was then tested when they were provided with either the subject noun, the verb, or the subject plus verb of the sentence as a retrieval cue. The two-word cue was relatively better if an integrated semantic representation was constructed. Manipulations included decreasing the processing time given to subjects (expected to inhibit the construction of integrated representations) and presenting a picture with the sentence (expected to facilitate the construction of integrated representations). The reduction in time decreased performance for the subjects without mental retardation to the level normally observed for those with mental retardation; presenting a picture increased performance of subjects with mental retardation to the level of comparison subjects. Results support the suggestion that previously observed differences in sentence processing between individuals with and without mental retardation may be due to differences in generating integrated representations of the sentences during processing.

Degree of associative relatedness and sentence processing by adolescents with and without mental retardation.

Previous research has suggested that individuals with mental retardation are less likely than individuals without mental retardation to access and incorporate information about the relations between words of sentences in the representations of those sentences in memory (e.g., Merrill & Bilsky, 1990; Merrill & Jackson, in press). A cued recall study and a semantic verification study were conducted to determine whether the magnitude of this group difference could be made smaller by increasing the degree to which the words in the sentences were semantically related. In both experiments, individuals with mental retardation exhibited an ability to utilize contextual information to a greater extent when the words were related. In the highly related conditions, the differences between groups was virtually eliminated.