Citrus is a globally consumed fruit with great popularity. Mandarin (Citrus reticulata cv. ‘Shatangju’) is a local variety, and its planting area and yield are the greatest regarding fruit tree planting in Guangdong Province, China. However, its resistance to Huanglongbing (HLB) is weak. After infection by HLB, the fruits cannot develop normally. In this study, four kinds of fruits were classified as HBG, XQG, ZQG, and DHG, according to the color of their peels. The metabolomes of the three abnormally colored groups (HBG, XQG, and ZQG) and the normally colored group (DHG) were compared using a UPLC-QQQ-MS-based metabolomics approach. In total, 913 metabolites were identified and classified into 23 different categories, including phenylpropanoids and flavonoids; among them, 215 (HBG, 177; XQG, 124; and ZQG, 62) metabolites showed differential accumulation in the three comparison groups (HBG/XQG/ZQG versus DHG). A total of 2 unique metabolites, O-caffeoyl maltotriose and myricetin were detected only in DHG samples. When comparing HBG with DHG, there were 109 decreased and 68 increased metabolites; comparing XQG with DHG, there were 88 decreased and 36 increased metabolites; comparing ZQG with DHG, 41 metabolites were decreased, and 21 metabolites were increased. Metabolic pathway enrichment analysis of these differential metabolites showed significant enrichment of the “phenylpropanoid biosynthesis” pathway in all comparison groups. The hierarchical cluster analysis of the differential metabolites of the four groups showed a clear grouping patterns. The relative contents of three phenylpropanoids, four flavonoids, two alkaloids, one anthocyanin, and two other metabolites were significantly different between each comparison group. This study might provide fundamental insight for the isolation and identification of functional compounds from the peels of citrus fruit infected with HLB and for in-depth research on the effect of HLB on the formation of fruits pigment and the development of HLB-resistant citrus varieties.

Intervertebral disk degeneration (IDD) is a major cause of low back pain and an important socioeconomic burden. Degradation of the extracellular matrix (ECM) of nucleus pulposus (NP) cells in the interverterbal disk is important for IDD. Stress of a suitable frequency and amplitude promotes the synthesis of the ECM of NP cells, however, the associated mechanisms remain to be fully elucidated The present study aimed to investigate the effect of integrin α1 on the migration and ECM synthesis of NP cells under soft periodic mechanical stress. Rat NP cells were isolated and plated onto slides, and were then treated with or without the use of a periodic mechanical stress system. The expression levels of integrin α1, α5 and αv, ECM collagen 2A1 (Col2A1) and aggrecan, and the phosphorylation of phospholipase C-γ1 (PLCγ1) were measured using reverse transcription-quantitative polymerase chain reaction and western blot analyses. Cell migration was assayed using a scratch experiment. The results showed that exposure to periodic mechanical stress significantly induced the mRNA expression levels of Col2A1 and aggrecan, cell migration, mRNA expression of integrin α1 and phosphorylation of PLC-γ1 of the NP, compared with the control (P<0.05). Inhibition of the PLCγ1 protein by U73122 significantly decreased the ECM expression under periodic mechanical stress (P<0.05). Small interfering RNA-mediated integrin α1 gene knockdown suppressed the mRNA expression levels of Col2A1 and aggrecan, and suppressed the migration and phosphorylation of PLCγ1 of the NP cells under periodic mechanical stress, compared with the control (P<0.05). In conclusion, periodic mechanical stress induced ECM expression and the migration of NP cells via upregulating the expression of integrin α1 and the phosphorylation of downstream PLCγ1. These findings provide novel information to aid the understanding of the pathogenesis and development of IDD.

Several studies have evaluated the association between vitamin D receptor, matrix metalloproteinase 3 (MMP-3) polymorphisms and the risk of intervertebral disc degeneration susceptibility. The findings were inconsistent. This meta-analysis aimed to systematically assess the association between vitamin D receptor, MMP-3 polymorphisms and the risk of intervertebral disc degeneration susceptibility. A search of various databases was done covering all papers published until December 31th, 2014. Eight, 4, 3 studies were finally included that addressed the risk of intervertebral disc degeneration susceptibility and vitamin D receptor FokI (rs2228570), ApaI (rs7975232), and MMP-3 (rs731236) polymorphisms, respectively. FokI (f vs. F: summary odds ratio [OR], 1.13; 95% confidence interval [CI], 0.76–1.69; ff vs. FF: OR, 1.02; 95% CI, 0.59–1.77; ff vs. Ff/FF: OR, 1.05; 95% CI, 0.70–1.58), ApaI (a vs. A: OR, 0.73; 95% CI, 0.45–1.19; aa vs. AA: OR, 0.53; 95% CI, 0.22–1.25 p=0.14; aa vs. AA/Aa: OR, 0.69; 95% CI, 0.53–0.89) in the vitamin D receptor gene and MMP3 polymorphisms (5A vs. 6A: OR, 1.92; 95% CI, 0.77–4.80; 5A5A vs. 6A6A: OR, 2.17; 95% CI, 0.75–6.24; 5A5A vs. 5A6A/6A6A: OR, 1.58; 95% CI, 0.72–3.44) were not obviously associated with risk of intervertebral disc degeneration susceptibility. FokI, ApaI polymorphisms in the vitamin D receptor gene and MMP-3 polymorphism are not obvious risk factors for intervertebral disc degeneration susceptibility.

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Abstract. The aim of the present study was to dynamically observe and discuss the significance of inflammatory cytokines in cervical degenerative disease induced by unbalanced dynamic and static forces in rats. A total of 60 Sprague Dawley rats were randomized into test (n=45) and control (n=15) groups, which were randomly subdivided into three groups corresponding to assessment at one, three and six months post-operation. The test group included 10, 15 and 20 rats at the corresponding post-operative stage and the control group had five rats at each time-point. By excising cervicodorsal muscles and ligaments, an unbalanced dynamic and static rat model was established in the test group. At one, three and six-months post-operation, venous serum of test and control group rats was collected and inflammatory cytokines in the serum of all rats were quantitatively determined by ELISA. The results revealed that compared with the control group, the interleukin (IL)-1β, IL-10 and tumor necrosis factor-α levels in the test group were significantly increased at one and three months (P<0.05, <0.01 or <0.001), and that IL-12 was significantly increased at three months (P<0.05). However, transforming growth factor-β1 increased at one month but was significantly decreased at three months (P<0.01). IL-6 did not change significantly throughout the observation period (P>0.05). In conclusion, cervical vertebral stability may be accompanied with changes of inflammatory cytokines.

Background/Aims: Periodic mechanical stress has been shown to promote extracellular matrix (ECM) synthesis and cell migration of nucleus pulposus (NP) cells, however, the mechanisms need to be fully elucidated. The present study aimed to investigate the signal transduction pathway in the regulation of NP cells under periodic mechanical stress. Methods: Primary rat NP cells were isolated and seeded on glass slides, and then treated in our self-developed periodic stress field culture system. To further explore the mechanisms, data were analyzed by scratch-healing assay, quantitative reverse transcription polymerase chain reaction (RT-qPCR) analysis, western blotting, and co-immunoprecipitation assay. Results: Under periodic mechanical stress, the mRNA expression of ECM collagen 2A1 (Col2A1) and aggrecan, and migration of NP cells were significantly increased (P < 0.05 for each), associating with increases in the phosphorylation of Src, GIT1, and ERK1/2 (P < 0.05 for each). Pretreatment with the Src inhibitor PP2 reduced periodic mechanical stress-induced ECM synthesis and cell migration of NP cells (P < 0.05 for each), while the phosphorylation of GIT1 and ERK1/2 were inhibited. ECM synthesis, cell migration, and phosphorylation of ERK1/2 were inhibited after pretreatment with the small interfering RNA for GIT1 in NP cells under periodic mechanical stress (P < 0.05 for each), whereas the phosphorylation of Src was not affected. Pretreatment with the ERK1/2 inhibitor PD98059 reduced periodic mechanical stress-induced ECM synthesis and cell migration of NP cells (P < 0.05 for each). Co-immunoprecipitation assay showed that there was a direct interaction between Src and GIT1 and between GIT1 and ERK1/2. Conclusion: In conclusion, periodic mechanical stress induced ECM expression and migration of NP cells via Src-GIT1-ERK1/2 signaling pathway, playing an important role in regulation of NP cells.

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Background Mandarin ‘Shatangju’ is susceptible to Huanglongbing (HLB) and the HLB-infected fruits are small, off-flavor, and stay-green at the maturity period. To understand the relationship between pericarp color and HLB pathogen and the effect mechanism of HLB on fruit pericarp coloration, quantitative analyses of HLB bacterial pathogens and carotenoids and also the integrative analysis of metabolome and transcriptome profiles were performed in the mandarin ‘Shatangju’ variety with four different color fruits, whole green fruits (WGF), top-yellow and base-green fruits (TYBGF), whole light-yellow fruits (WLYF), and whole dark-yellow fruits (WDYF) that were infected with HLB. Results the HLB bacterial population followed the order WGF > TYBGF > WLYF > WDYF. And there were significant differences between each group of samples. Regarding the accumulation of chlorophyll and carotenoid, the chlorophyll-a content in WGF was the highest and in WDYF was the lowest. The content of chlorophyll-b in WGF was significantly higher than that in other three pericarps. There were significant differences in the total content of carotenoid between each group. WGF and TYBGF pericarps were low in phytoene, γ-carotene, β-cryptoxanthin and apocarotenal, while other kinds of carotenoids were significantly higher than those in WDYF. And WLYF was only short of apocarotenal. We comprehensively compared the transcriptome and metabolite profiles of abnormal (WGF, TYBGF and WLYF) and normal (WDYF, control) pericarps. In total, 2,880, 2,782 and 1,053 differentially expressed genes (DEGs), including 121, 117 and 43 transcription factors were identified in the three comparisons, respectively. The qRT-PCR confirmed the expression levels of genes selected from transcriptome. Additionally, a total of 77 flavonoids and other phenylpropanoid-derived metabolites were identified in the three comparisons. Most (76.65 %) showed markedly lower abundances in the three comparisons. The phenylpropanoid biosynthesis pathway was the major enrichment pathway in the integrative analysis of metabolome and transcriptome profiles. Conclusions Synthesizing the above analytical results, this study indicated that different color pericarps were associated with the reduced levels of some carotenoids and phenylpropanoids derivatives products and the down-regulation of proteins in flavonoids, phenylpropanoids derivatives biosynthesis pathway and the photosynthesis-antenna proteins.