Adipose-derived stromal cells protect intervertebral disc cells in compression: implications for stem cell regenerative disc therapy

INTRODUCTION: Abnormal biomechanics plays a role in intervertebral disc degeneration. Adipose-derived stromal cells (ADSCs) have been implicated in disc integrity; however, their role in the setting of mechanical stimuli upon the disc's nucleus pulposus (NP) remains unknown. As such, the present study aimed to evaluate the influence of ADSCs upon NP cells in compressive load culture. METHODS: Human NP cells were cultured in compressive load at 3.0MPa for 48 hours with or without ADSCs co-culture (the ratio was 50:50). We used flow cytometry, live/dead staining and scanning electron microscopy (SEM) to evaluate cell death, and determined the expression of specific apoptotic pathways by characterizing the expression of activated caspases-3, -8 and -9. We further used real-time (RT-) PCR and immunostaining to determine the expression of the extracellular matrix (ECM), mediators of matrix degradation (e.g. MMPs, TIMPs and ADAMTSs), pro-inflammatory factors and NP cell phenotype markers. RESULTS: ADSCs inhibited human NP cell apoptosis via suppression of activated caspase-9 and caspase-3. Furthermore, ADSCs protected NP cells from the degradative effects of compressive load by significantly up-regulating the expression of ECM genes (SOX9, COL2A1 and ACAN), tissue inhibitors of metalloproteinases (TIMPs) genes (TIMP-1 and TIMP-2) and cytokeratin 8 (CK8) protein expression. Alternatively, ADSCs showed protective effect by inhibiting compressive load mediated increase of matrix metalloproteinases (MMPs; MMP-3 and MMP-13), disintegrin and metalloproteinase with thrombospondin motifs (ADAMTSs; ADAMTS-1 and 5), and pro-inflammatory factors (IL-1beta, IL-6, TGF-beta1 and TNF-alpha). CONCLUSIONS: Our study is the first in vitro study assessing the impact of ADSCs on NP cells in an un-physiological mechanical stimulation culture environment. Our study noted that ADSCs protect compressive load induced NP cell death and degradation by inhibition of activated caspase-9 and -3 activity; regulating ECM and modulator genes, suppressing pro-inflammatory factors and preserving CK8. Consequently, the protective impact of ADSCs found in this study provides an essential understanding and expands our knowledge as to the utility of ADSCs therapy for intervertebral disc regeneration.published_or_final_versio

Immune cascades in human intervertebral disc: the pros and cons

The unique structural hallmark of the intervertebral disc has made its central composition, the nucleus pulposus (NP), excluded from the immunologic tolerance. Consequently, the intervertebral disc is identified as an immune-privileged organ. Traditionally, local detrimental immune activities caused by NP at the lesion sites of the disc are noted as a significant factor contributing to disc degeneration. However, given the beneficial activities of immune cells in other immune-privileged sites on basis of current evidence, the degenerate disc might need the assistance of a subpopulation of immune cells to restore its structure and lessen inflammation. In addition, the beneficial impact of immune cells can be seen in the absorption of the herniated NP, which is an important factor causes the mechanical compression of nerve roots. Consequently, a modulated immune network in degenerate disc is essential for the restoration of this immune-privileged organ. Until now, the understandings of immune response in disc degeneration still rest on the harmful aspect. Further studies are needed to explore its beneficial influence. Accordingly, there are no absolutely the pros and cons in terms of immune reactions caused by NP.published_or_final_versio

Magnetic field effects on the electroluminescence of organic light emitting devices: A tool to indicate the carrier mobility

The magnetoelectroluminescence (MEL) of organic light emitting devices with a N, N′ -bis(l-naphthyl)- N, N′ -diphenyl- 1, l′ -biphentl- 4, 4′ -diamine:tris-(8-hydroxyquinoline) aluminum (NPB: Alq 3) mixed emission layer (EML) has been investigated. We find that MEL is maximized when the volume ratio of NPB of the mixed EML reaches 30% and the EML thickness is 40 nm. The features of MEL under various magnetic field strengths are insensitive to the change in EML thickness and mixing ratio. Meanwhile, MEL has a close relationship with the carrier mobility. We have conducted a theoretical study to further verify the relationship. Our experimental and theoretical results confirm that MEL can function as a tool to indicate the mobility. © 2010 American Institute of Physics.published_or_final_versio

Down-regulated CK8 expression in human intervertebral disc degeneration

As an intermediate filament protein, cytokeratin 8 (CK8) exerts multiple cellular functions. Moreover, it has been identified as a marker of notochord cells, which play essential roles in human nucleus pulposus (NP). However, the distribution of CK8 positive cells in human NP and their relationship with intervertebral disc degeneration (IDD) have not been clarified until now. Here, we found the percentage of CK8 positive cells in IDD (25.7+/-4.14%) was significantly lower than that in normal and scoliosis NP (51.9+/-9.73% and 47.8+/-5.51%, respectively, p<0.05). Western blotting and qRT-PCR results confirmed the down-regulation of CK8 expression in IDD on both of protein and mRNA levels. Furthermore, approximately 37.4% of cell clusters were CK8 positive in IDD. Taken together, this is the first study to show a down-regulated CK8 expression and the percentage of CK8 positive cell clusters in IDD based upon multiple lines of evidence. Consequently, CK8 positive cells might be considered as a potential option in the development of cellular treatment strategies for NP repair.published_or_final_versio

From Nonspecific DNA–Protein Encounter Complexes to the Prediction of DNA–Protein Interactions

©2009 Gao, Skolnick. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.doi:10.1371/journal.pcbi.1000341DNA–protein interactions are involved in many essential biological activities. Because there is no simple mapping code between DNA base pairs and protein amino acids, the prediction of DNA–protein interactions is a challenging problem. Here, we present a novel computational approach for predicting DNA-binding protein residues and DNA–protein interaction modes without knowing its specific DNA target sequence. Given the structure of a DNA-binding protein, the method first generates an ensemble of complex structures obtained by rigid-body docking with a nonspecific canonical B-DNA. Representative models are subsequently selected through clustering and ranking by their DNA–protein interfacial energy. Analysis of these encounter complex models suggests that the recognition sites for specific DNA binding are usually favorable interaction sites for the nonspecific DNA probe and that nonspecific DNA–protein interaction modes exhibit some similarity to specific DNA–protein binding modes. Although the method requires as input the knowledge that the protein binds DNA, in benchmark tests, it achieves better performance in identifying DNA-binding sites than three previously established methods, which are based on sophisticated machine-learning techniques. We further apply our method to protein structures predicted through modeling and demonstrate that our method performs satisfactorily on protein models whose root-mean-square Ca deviation from native is up to 5 Å from their native structures. This study provides valuable structural insights into how a specific DNA-binding protein interacts with a nonspecific DNA sequence. The similarity between the specific DNA–protein interaction mode and nonspecific interaction modes may reflect an important sampling step in search of its specific DNA targets by a DNA-binding protein

Measurement of the Branching Fraction of J/psi --> pi+ pi- pi0

Using 58 million J/psi and 14 million psi' decays obtained by the BESII experiment, the branching fraction of J/psi --> pi+ pi- pi0 is determined. The result is (2.10+/-0.12)X10^{-2}, which is significantly higher than previous measurements.Comment: 9 pages, 8 figures, RevTex

First observation of psi(2S)-->K_S K_L

The decay psi(2S)-->K_S K_L is observed for the first time using psi(2S) data collected with the Beijing Spectrometer (BESII) at the Beijing Electron Positron Collider (BEPC); the branching ratio is determined to be B(psi(2S)-->K_S K_L) = (5.24\pm 0.47 \pm 0.48)\times 10^{-5}. Compared with J/psi-->K_S K_L, the psi(2S) branching ratio is enhanced relative to the prediction of the perturbative QCD ``12%'' rule. The result, together with the branching ratios of psi(2S) decays to other pseudoscalar meson pairs (\pi^+\pi^- and K^+K^-), is used to investigate the relative phase between the three-gluon and the one-photon annihilation amplitudes of psi(2S) decays.Comment: 5 pages, 4 figures, 2 tables, submitted to Phys. Rev. Let

CK8 phosphorylation induced by compressive loads underlies the downregulation of CK8 in human disc degeneration by activating protein kinase C

Cytokeratin 8 (CK8) is a member of the cytokeratins family with multiple functions on the basis of its unique structural hallmark. The aberrant expression of CK8 and its phosphorylation are pertinent with various diseases. We have previously shown that CK8 exists in normal human nucleus pulposus (NP) cells and decreases as the intervertebral disc degenerates. However, the underlying molecular regulatory machinery of CK8 in intervertebral disc degeneration (IDD) has not been clarified. Here, we collected NP samples from patients with idiopathic scoliosis as control and IDD as degenerate groups. We found that CK8 expression decreased in IDD with an increased phosphorylation in degenerate NP cells. Moreover, NP cells were cultured under different compressive load schemes for diverse time duration. We found that compressive loads resulted in phosphorylation and disassembly of CK8 in a time-dependent and degree-dependent manner in vitro. The activation of protein kinase C was a significant molecular factor contributing to this phenomenon. Taken together, this study is the first to address the molecular mechanisms of CK8 downregulation in NP cells. Importantly, our findings provide clues regarding a molecular link between compressive loads and CK8 alterations, which shed a novel light on the etiology of IDD.published_or_final_versio

Unsupervised Learning of Category-Specific Symmetric 3D Keypoints from Point Sets

Automatic discovery of category-specific 3D keypoints from a collection of objects of a category is a challenging problem. The difficulty is added when objects are represented by 3D point clouds, with variations in shape and semantic parts and unknown coordinate frames. We define keypoints to be category-specific, if they meaningfully represent objects’ shape and their correspondences can be simply established order-wise across all objects. This paper aims at learning such 3D keypoints, in an unsupervised manner, using a collection of misaligned 3D point clouds of objects from an unknown category. In order to do so, we model shapes defined by the keypoints, within a category, using the symmetric linear basis shapes without assuming the plane of symmetry to be known. The usage of symmetry prior leads us to learn stable keypoints suitable for higher misalignments. To the best of our knowledge, this is the first work on learning such keypoints directly from 3D point clouds for a general category. Using objects from four benchmark datasets, we demonstrate the quality of our learned keypoints by quantitative and qualitative evaluations. Our experiments also show that the keypoints discovered by our method are geometrically and semantically consistent

First Measurements of eta_c Decaying into K^+K^-2(pi^+pi^-) and 3(pi^+pi^-)

The decays of eta_c to K^+K^-2(pi^+pi^-) and 3(pi^+pi^-) are observed for the first time using a sample of 5.8X10^7 J/\psi events collected by the BESII detector. The product branching fractions are determined to be B(J/\psi-->gamma eta_c)*B(eta_c-->K^+K^-pi^+pi^-pi^+pi^-)=(1.21+-0.32+- 0.23)X10^{-4}$,B(J/\psi-->gamma eta_c)*B(eta_c-->K^{*0}\bar{K}^{*0}pi^+pi^-)= (1.29+-0.43+-0.32)X10^{-4}$, and (J/\psi-->gamma eta_c)* B(eta_c-->pi^+pi^-pi^+pi^-pi^+pi^-)= (2.59+-0.32+-0.48)X10^{-4}. The upper limit for eta_c-->phi pi^+pi^-pi^+pi^- is also obtained as B(J/\psi-->gamma eta_c)*B(eta_c--> phi pi^+pi^-pi^+pi^-)< 6.03 X10^{-5} at the 90% confidence level.Comment: 11 pages, 4 figure