Prediction of a Potential Mechanism of Intervertebral Disc Degeneration Based on a Novel Competitive Endogenous RNA Network

Low back pain which resulted from intervertebral disc degeneration (IDD) is a common health problem that afflicts people all over the world. Due to the lack of an overall understanding of the molecular interactions involved in IDD, we hope to better understand the pathogenetic mechanisms that drive the degenerative process. The purpose of this study is to obtain mRNAs, miRNAs, lncRNAs, and circRNAs associated with IDD gained from public databases and to establish an interaction network. According to the results of microarray analysis and bioinformatics analysis from the contrast of IDD and normal nucleus pulposus tissues, a total of 49 mRNAs, 10 miRNAs, 30 lncRNAs, and 4 circRNAs were obtained and a lncRNA/circRNA–miRNA–mRNA interaction network was constructed. NEAT1–miR-5100–COL10A1 and miR663AHG/HEIH/hsa-circ-0003600–miR-4741–HAS2/HYAL1/LYVE1 might be potential interaction axes of the molecular mechanism in IDD. The increased expression of NEAT1 might inhibit miR-5100 and subsequently upregulate the expression of COL10A1, which leads to IDD, while the increased expression of miR663AHG/HEIH/hsa-circ-0003600 might inhibit miR-4741 and indirectly upregulate HAS2/HYAL1/LYVE1, and leads to the protection from IDD. More interaction axes are to be exploited to provide theoretical bases for further study on IDD.

Integrated transcriptome and proteome analyses identifies novel genes and regulatory networks in intervertebral disc degeneration

Background: Intervertebral disc degeneration is a major cause of symptoms like low back pain and neck pain. Many groups have tried to reveal the regulatory network using either transcriptome or proteome profiling technologies, however, the relationship between these differentially expressed proteins and mRNAs are not elucidated. Since post-transcriptional regulation and other mechanisms may affect the translation of mRNA to protein, a combined transcriptome and proteome study may give more precise data on unveiling important regulatory network and key genes of Intervertebral disc degeneration. Results: In the present study, we used the label-free quantification proteomic approach and identify 656 proteins expressed in either degenerated or normal nucleus pulposus, of which 503 proteins are differentially expressed. Taking advantage of the existing nucleus pulposus transcriptome data, we combine and reanalyze the data and find 105 differentially expressed mRNA between degenerated and normal nucleus pulposus. By comparing these data, only 9 genes show significant changes in both protein and mRNA data, while 6 genes (TNFAIP6, CHI3L1, KRT19, DPT, COL6A2 and COL11A2) show concordant changes in both protein and mRNA level. Further functional analyses show different functions of the altered mRNAs and proteins in degeneration, indicating great difference between protein network and mRNA network. Using the gene co-expression network method, we uncover novel regulatory network and potential genes that may play vital roles in intervertebral disc degeneration by combining protein and mRNA data. Conclusions: This is the first study to identify novel regulatory network of intervertebral disc degeneration using combined analysis of both transcriptome and proteome, which may give new insight into the molecular mechanism of intervertebral disc degeneration.

miR-155 Inhibits Nucleus Pulposus Cells’ Degeneration through Targeting ERK 1/2

We first investigated the difference in microRNA expression between normal NP cells and degenerative NP cells using gene chip. We have found that the expression of ERK1/2 was decreased with overexpression of miR-155 in normal nucleus pulposus cell. Expression of ERK1/2 was increased with inhibition of miR-155. Overexpression or inhibition of miR-155 had no effects on the expression level of mRNA ERK1/2 in nucleus pulposus cell, which showed that miR-155 affected the expression of pERK1/2 after transcription of ERK1/2 mRNA indicating that ERK1/2 was a new target protein regulated by miR-155. In the degeneration of intervertebral disc, inhibited miR-155 decreased the expressions of extracellular main matrix collagen II and glycosaminoglycan and increased expression of ERK1/2. Taken together, our data suggested that miR-155 was the identified miRNA which regulated NP cells degenerated through directly targeting ERK1/2.

312 MORPHOLOGICAL AND MORPHOMETRICS CHARACTERISTICS OF NUCLEAR SPERM IN THE EPIDIDYMAL TRANSIT OF BULLS

During the transit through the epididymis, many morphological, physiological, and biochemical characteristics of spermatozoa are modified, as part of the maturation process. Nuclear maturation continues in the epididymis through an increase in formation of protamine disulfide. Thus, penetration through the oocyte membranes could be facilitated for elongated spermatozoa with dramatically condensed chromatin and nuclear integrity. Therefore, size, shape, and nuclear defects could be used to estimate the stage-related nuclear transformations from early spermiogenesis to the end of epididymal transit. Nellore bulls (n = 9), 30-36 months old, with high seminal quality (>80% motile and morphologically normal sperm) were submitted to orchiectomy. Impressions in slides of the caput, corpus, and caudal regions of the epididymis were prepared for evaluation of morphology (Feulgen stain, phase-contrast microscopy at 1000 ×) and morphometry of the nuclear sperm. The slides were captured in a Motic 2300 camera adapted to the microscope and digitally assessed. Nuclear morphology was considered normal (without visible alterations), head defect (variations in shape and form), and nuclear defects (abnormal chromatin condensation and presence of vacuoles). Base, width, length (μm), and area (μm2) were estimated in least 60 sperm nuclei. No difference (P > 0.05) among epididymal regions for normal nuclei (70.3 ± 3.1%), head defects (3.4 ± 0.5%), and nuclear defects (5.3 ± 1.3%) were seen. The base was higher (2.68 ± 0.5 μm, P < 0.01) in the caput than corpus (2.44 ± 0.4 μm) and caudal regions (2.41 ± 0.4μm). Normal nuclei were associated (P < 0.01) with width (r = 0.20), length (r = 0.27), and area (r = 0.44) in the caput and with width (r = 0.21), length (r = 0.40), and area (r = 0.33) in the corpus of epididymis. Epididymal transit affected (P < 0.001) the measures and nuclear status that accounted for regression analysis: normal nucleus (40.254 + 3.027, length; R2 = 0.20), head defects (0.922 + 1.097, width + 0.093 × area; R2 = 0.26), and nuclear defects (6.993-0.496, length + 0.454, base; R2 = 0.23). The results suggest that important variations occur in the nuclear status during the epididymal transit in the bovine spermatozoa. The higher measures in the sperm base of the caput suggest a narrowing probably indicating the continuity of nuclear remodeling from the final steps of spermiogenesis. The area accounts for 20% of the nuclear shape variations along the epididymal segment. These events characterize the adaptation of nuclear membranes and chromatin structure surrounded by epididymal environment and comprise a part of the maturation process. Additionally, morphometric variations are associated with defects in the nuclear structures and can be used to determine the conditions of spermatogenesis and sperm maturation based on the evaluation of ejaculated semen. We thank CNPq/PROPP and Fundect for financial support.

Prenatal disruption of binocular interactions creates novel lamination in the cat's lateral geniculate nucleus

The elimination of retinogeniculate afferents from one eye on embryonic day 44 (E44) has pronounced effects on the formation of the cellular laminae in the cat lateral geniculate nucleus (LGN). Only two laminae form: a dorsal, “magnocellular” layer, and a ventral, “parvocellular” layer. Soma size measurements and previously reported patterns of termination of retinogeniculate axons suggest that the dorsal lamina is a coalescence of the normal A-laminae and the dorsal, magnocellular division of layer C, while the ventral layer is a composite of the parvocellular sublamina of layer C and the remaining C-laminae. This is a novel pattern of lamination in the LGN that differs from that found in the normal nucleus, not only in that there are now only two cell layers rather than the normal five, but also in that the interlaminar zone occurs in an abnormal location. This result is markedly different from that observed in other species where interlaminar zones present after early monocular enucleation are a subset of the ones which would normally be present. We suggest that, in the absence of ongoing binocular interactions, interactions between functionally distinct retinal ganglion cell classes from the remaining eye may direct the formation of cell laminae in the LGN, even when such interactions are not normally operative.