Advanced glycation end products promote intervertebral disc degeneration by transactivation of matrix metallopeptidase genes.

OBJECTIVE: Examine the mechanism by which advanced glycation end products (AGEs) induce intervertebral disc degeneration (IDD) in C57BL/6J mice. METHODS: Matrix metallopeptidase (MMP) gene mRNA levels were assessed using RT-qPCR. Immunoprecipitation and co-immunoprecipitation were performed to identify the transcriptional complex regulating MMP expression due to AGEs. The preventive effects of inhibitors targeting this complex were tested in mice on high AGE diets. RESULTS: IDD and AGE accumulation were evident in mice on high-AGE diets (HAGEs), persisting across dietary shifts but absent in mice exclusively on low-AGE diets. Molecularly, HAGEs activated p21-activated kinase 1 (PAK1), prompting peroxisome proliferator-activated receptor gamma coactivator-related protein 1 (PPRC1) phosphorylation. Ubiquitin-specific protease 12 (USP12) interacted with the phosphorylated PPRC1 (pPPRC1), safeguarding it from proteasomal degradation. This pPPRC1, in collaboration with two histone acetyltransferases p300/CREB-binding protein (CBP) and a transcription factor activator protein 1(AP1), enhanced the expression of 12 MMP genes (MMP1a/1b/3/7/9/10/12/13/16/19/23/28). In vitro AGE exposure on nucleus pulposus and annulus fibrosus cells replicated this gene activation pattern, driven by the PAK1/pPPRC1-p300/CBP-AP1 pathway. The application of PAK1, p300, and AP1 inhibitors reduced pPPRC1-p300/CBP-AP1 binding to MMP promoters, diminishing their expression. These inhibitors effectively thwarted IDD in HAGE mice. CONCLUSION: Our results revealed that HAGEs instigate IDD via the PAK1/pPPRC1-p300/CBP-AP1 signaling pathway. This insight can guide therapeutic strategies to slow IDD progression in prediabetic/diabetic patients.

Learning curve of junior surgeons in robot-assisted pedicle screw placement: a comparative cohort study.

OBJECTIVE: Robot-assisted technology has been gradually applied to pedicle screw placement in spinal surgery. This study was designed to detailedly evaluate the learning curve of junior surgeons in robot-assisted spine surgery. METHODS: From December 2020 to February 2022, 199 patients requiring surgical treatment with posterior pedicle screw fixation were prospectively recruited into the study. The patients were randomized to the robot-assisted group (the RA group) or the conventional freehand group (the CF group). Under the senior specialist's supervision, pedicle screws were placed by two junior fellows without prior experience. Cumulative summation (CUSUM) analysis was performed on the learning curve of pedicle screw placement for performing quantitative assessment based on the time of screw insertion. RESULTS: In total, 769 and 788 pedicle screws were placed in the RA and CF groups. Compared with the CF group, the learning duration in the RA group was shorter in the upper thoracic region (57 vs. 70 screws), but longer in the lower thoracic (62 vs. 58 screws) and the lumbosacral region (56 vs. 48 screws). The slope of learning curve was lower in the RA group than in the CF group. The screw accuracy in the RA group was superior to that in the CF group, especially in upper thoracic region (89.4% vs. 76.7%, P < 0.001). This disparity of accuracy became wider in deformity cases. In the upper thoracic region, the mean placement time was 5.34 ± 1.96 min in the RA group and 5.52 ± 2.43 min in the CF groups, while in the lower thoracic and lumbosacral regions, the CF group's mean placement times were statistically shorter. Three screw-related neural complications occurred in the CF group. CONCLUSION: Robot-assisted technique has its advantages in the upper thoracic region and deformity cases, which is easier and safer to insert pedicle screws. The robot-assisted technique allowed a short learning curve for junior surgeons and exhibited consistently excellent results even in the early application period.

HiJAKing Immunotherapy-Resistant Melanoma for a Cure.

Immune checkpoint blockers (ICBs) have brought great promise to patients with advanced melanoma, a tumor type that was claimed largely incurable not long ago. However, therapeutic resistance to ICBs has limited their utility in the clinic. Here, we provide a commentary on recent research endeavors concerning ICB resistance in melanoma patients.

C-type lectin domain-containing protein CLEC3A regulates proliferation, regeneration and maintenance of nucleus pulposus cells.

It is widely assumed that as connective tissue, the intervertebral disc (IVD) plays a crucial role in providing flexibility for the spinal column. The disc is comprised of three distinct tissues: the nucleus pulposus (NP), ligamentous annulus fibrous (AF) that surrounds the NP, and the hyaline cartilaginous endplates (CEP). Nucleus pulposus, composed of chondrocyte-like NP cells and its secreted gelatinous matrix, is critical for disc health and function. The NP matrix underwent dehydration accompanied by increasing fibrosis with age. The degeneration of matrix is almost impossible to repair, with the consequence of matrix stiffness and senescence of NP cells and intervertebral disc, suggesting the value of glycoproteins in extracellular matrix (ECM). Here, via database excavation and biological function screening, we investigated a C-type lectin protein, CLEC3A, which could support differentiation of chondrocytes as well as maintenance of NP cells and was essential to intervertebral disc homeostasis. Furthermore, mechanistic analysis revealed that CLEC3A could stimulate PI3K-AKT pathway to accelerate cell proliferation to further play part in NP cell regeneration.

Analysis of the epidemiological trends of enterovirus A in Asia and Europe.

BACKGROUND: Enteroviruses have been in massive, cyclical epidemics worldwide. An in-depth understanding of the international epidemiological characteristics of Enterovirus A (EVA) is critical to determining its clinical significance and total disease burden. Although much research has been conducted on EVA epidemiology, there is still a lack of a comprehensive overview of EVA epidemiological characteristics and trends. OBJECTIVE: EVA nucleic acid sequences from the NCBI virus database were used to summarize the epidemic time (based on the time of specimen collection), spatial and serotype distribution of EVA, and to analyze EVA isolated from cerebrospinal fluid specimens. METHODS: EVA sequences were searched in NCBI Virus by keyword ("Enterovirus A″ or "EVA") to screen sequences released before December 2021 and sort them to analyze EVA by year, geographic region and serotype prevalence. RESULTS: The results found 23,041 retrieved nucleic acid sequences with precise collection dates and geographical regions as of December 2021, with Asia accounting for 87%, Europe for 11% and Africa and the Americas for only 2%. Overall, EV-A71, CVA6 and CVA16 are a few of the main prevalent serotypes; and the prevalence characteristics of the different serotypes change over time from place to place. CONCLUSION: The prevalence of different serotypes of EVA varies considerably over time and space, and we focused on analysing the epidemiological characteristics of EVAs in Asia and Europe and EVAs that invade the nervous system. This study will likely provide important clues for prevention, control and future research in virological surveillance, disease management and vaccine development.

PSD95 as a New Potential Therapeutic Target of Osteoarthritis: A Study of the Identification of Hub Genes through Self-Contrast Model.

Osteoarthritis (OA) is a worldwide joint disease. However, the precise mechanism causing OA remains unclear. Our primary aim was to identify vital biomarkers associated with the mechano-inflammatory aspect of OA, providing potential diagnostic and therapeutic targets for OA. Thirty OA patients who underwent total knee arthroplasty were recruited, and cartilage samples were obtained from both the lateral tibial plateau (LTP) and medial tibial plateau (MTP). GO and KEGG enrichment analyses were performed, and the protein-protein interaction (PPI) assessment was conducted for hub genes. The effect of PSD95 inhibition on cartilage degeneration was also conducted and analyzed. A total of 1247 upregulated and 244 downregulated DEGs were identified. Significant differences were observed between MTP and LTP in mechanical stress-related genes and activated sensory neurons based on a self-contrast model of human knee OA. Cluster analysis identified DLG4 as the hub gene. Cyclic loading stress increased PSD95 (encoded by DLG4) expression in LTP cartilage, and PSD95 inhibitors could alleviate OA progression. This study suggests that inhibiting PSD95 could be a potential therapeutic strategy for preventing articular cartilage degradation.

Role of the receptor for activated C kinase 1 during viral infection.

Viruses can survive only in living cells, where they depend on the host's enzymatic system for survival and reproduction. Virus-host interactions are complex. On the one hand, hosts express host-restricted factors to protect the host cells from viral infections. On the other hand, viruses recruit certain host factors to facilitate their survival and transmission. The identification of host factors critical to viral infection is essential for comprehending the pathogenesis of contagion and developing novel antiviral therapies that specifically target the host. Receptor for activated C kinase 1 (RACK1), an evolutionarily conserved host factor that exists in various eukaryotic organisms, is a promising target for antiviral therapy. This review primarily summarizes the roles of RACK1 in regulating different viral life stages, particularly entry, replication, translation, and release.

Electrospun Fibers Improving Cellular Respiration via Mitochondrial Protection.

Cellular respiration is the prerequisite for cell survival and functions, and mitochondrial function and microcirculation oxygen supply are essential for cellular respiration. However, in diabetic fracture, cellular respiration of bone marrow stem cells (BMSCs) is disrupted because of the dysfunction of mitochondria and microcirculation disorders. Here, the electrospun fibers of GelMA loaded with Hif-1 pathway activator (DFO) are constructed to improve the cellular respiration of BMSCs via protecting mitochondrial function and reconstructing microcirculation. The sequential process of electrospinning and UV crosslinking endowed the electrospun fibers with breathability and the biomechanical properties like the periosteum. In vitro biomolecular experiments showed that by crosslinking grafted polyethylene glycol acrylate liposomes loaded with DFO, the functional electrospun fibers can release DFO locally to activate Hif-1 in BMSCs, which can regulate the balance of Bcl-2/Bax to protect mitochondria and upregulate the expression of VEGF to reconstruct microcirculation. Animal experiments confirmed that the functional electrospun fibers can promote the recovery of diabetic fracture in vivo. These suggested that the functional electrospun fibers can improve cellular respiration for cell survival and functions of BMSCs. This study provides a new treatment strategy for diabetic fracture and other tissue regeneration on basis of cellular respiration improvement.

A quadruple recombination event discovered in hepatitis E virus.

Hepatitis E virus (HEV) can infect humans, pigs, and many other animals, but recombination in HEV has rarely been reported. In the present study, phylogenetic and recombination analysis was performed on 557 complete HEV genome sequences from the GenBank database. A potentially significant quadruple recombination event was identified by recombination detection analysis. The recombinant progeny virus, HEV_32_Manchester_301214, was produced by inter-genotype recombination between the major parent HEPAC-44 and the minor parent HE-JA15-1335. HEV_32_Manchester_301214 and HEPAC-44 belong to genotype 3, while HE-JA15-1335 belongs to genotype 1, and these three strains were all isolated from humans. Three breakpoints of the four recombination events occurred in the ORF2 region, while another occurred in the ORF1 region. This quadruple recombination event was confirmed by phylogenetic analysis. The genotype, host, and recombination regions of the three strains were analyzed, and the analysis results provide valuable information for future research on HEV diversity.

Antinociceptive effect of intrathecal injection of miR-9-5p modified mouse bone marrow mesenchymal stem cells on a mouse model of bone cancer pain.

BACKGROUND: A growing body of studies have indicated that bone marrow mesenchymal stem cells (BMSCs) have powerful analgesic effects in animal models of bone cancer pain. Here, we explored the molecular mechanisms underlying how BMSCs alleviate pain sensation in a mouse model of bone cancer pain. METHODS: C3H/HeN adult male mice were used to generate a bone cancer pain model. BMSCs were isolated from mouse bone marrow, modified by transfection with microRNA-9-5p (miR-9-5p), and infused into the spinal cord. Spontaneous flinches, paw withdrawal latency, limb-use score, and weight-bearing score were used to assess pain-related behaviors. ELISA, RT-PCR, western blot, and luciferase assay were used to assess gene expressions. RESULTS: Our results show that miR-9-5p regulated the expression of both repressor element silencing transcription factor (REST) and µ-opioid receptors (MOR) by targeting REST in primary mouse BMSCs. Overexpression of miR-9-5p reversed the activation of inflammatory pathway in TNF-α- and IL-6-treated BMSCs. In addition, miR-9-5p modified BMSCs alleviated cancer pain in the sarcoma-inoculated mouse model. MiR-9-5p modified BMSCs suppressed cytokine expression in the spinal cord of sarcoma-inoculated mice by suppressing REST gene expression. CONCLUSIONS: Our results indicate that miR-9-5p modified BMSCs can relieve bone cancer pain via modulating neuroinflammation in the central nervous system, suggesting genetically modified BMSCs could be a promising cell therapy in pain management.

Poliovirus serotype 2 and coxsackievirus A promote the natural recombination of poliovirus.

Poliovirus (PV) is a member of the species Enterovirus C (EV-C), which may cause irreversible paralysis and death. So, for the purpose of analyzing the evolution of PV2 to help in eradicating PVs globally, a recombination analysis was performed to verify all viral genomes of EV-C, and we found 13 putative recombination events that produced PV1, 14 recombination events that can give rise to PV2, and 9 events that can lead to PV3. By analyzing our findings, we found that PV2 was involved in 25 of 36 PV recombination events, whereas coxsackievirus A (CVA) strains were involved in 12 of 36 PV recombination events, indicating that PV2 and CVAs play major roles in the natural recombination of PV. In addition, we found 11 of 36 breakpoint positions located in 2A region, which is the most active region of the recombination events.

Selective thoracolumbar/lumbar fusion for Syringomyelia-associated scoliosis: a case-control study with Lenke 5C adolescent idiopathic scoliosis.

BACKGROUND: Selective thoracolumbar/lumbar fusion technique was introduced to treat adolescent idiopathic scoliosis (AIS) patients with major thoracolumbar/lumbar curves. Theoretically, this surgical strategy could also be applied to syringomyelia patients. No previous study has specifically addressed the effectiveness of selective thoracolumbar/lumbar fusion for patients with syringomyelia-associated scoliosis. The aim of the study was to investigate the effectiveness of selective thoracolumbar/lumbar fusion for the surgical treatment of patients with syringomyelia-associated scoliosis. METHODS: From February 2010 to September 2016, 14 syringomyelia-associated patients with major thoracolumbar/lumbar curves were retrospectively reviewed. Besides, 30 Lenke 5C AIS patients were enrolled as a control group. Posterior selective thoracolumbar/lumbar fusion was performed for both groups. Patients' demographic, operative, radiological, and quality of life data were reviewed with follow-up. Intragroup comparisons were performed for each parameter. RESULTS: The two groups were matched by age, gender, curve characteristics, duration of follow-up, and all preoperative radiographic parameters except for thoracic kyphosis. After surgery, the average correction rate of the major thoracolumbar/lumbar curve was 82.2 ± 7.8% in the syringomyelia group, which was not significantly different from that of AIS group (82.5 ± 10.6%, P = 0.47). A similar improvement of unfused thoracic curve was observed between the two groups (50.1 ± 16.5% vs. 48.5 ± 26.9%, P = 0.29). During the follow-up, the correction effect of scoliosis was well maintained, without aggravation of the original neural symptoms or fresh permanent neurological deficits. Of note, the number of fusion levels was significantly larger in syringomyelia group than that in AIS group (7.6 ± 1.4 vs. 6.5 ± 1.2, P < 0.01). The average follow up was 47.6 months (36-81 months). CONCLUSION: Similar to AIS cases, syringomyelia-associated scoliosis can be effectively and safely corrected by selective thoracolumbar/lumbar fusion with satisfactory surgical outcomes. However, the syringomyelia group, on average, required an additional fused segment for treatment as compared to the AIS group (7.6 versus 6.5 in the AIS group).

Inflammation-dependent downregulation of miR-194-5p contributes to human intervertebral disc degeneration by targeting CUL4A and CUL4B.

Inflammation is one of the major causes of intervertebral disc degeneration (IDD). Emerging evidence has revealed that increase in the levels of pro-inflammatory cytokines, such as interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α), can activate a variety of signaling pathways, eventually resulting in IDD. Here, we show that the two cullin family genes, CUL4A and CUL4B, but not other cullins, are specifically overexpressed in IDD samples compared with healthy controls, and the CUL4A and CUL4B levels are positively correlated with the severity of IDD. In vitro analyses in human osteoblast cells (hFOB1.19), nucleus pulposus cells (hNPCs), and annulus fibrosus cells (hAFCs) indicated that treatment with IL-6 and TNF-α can increase CUL4A and CUL4B levels. By performing a microRNA-based microarray analysis, we found a set of microRNAs (miRNAs) that were differentially expressed in IDD samples compared with samples from healthy controls. Of these miRNAs, miR-194-5p, was significantly downregulated in IDD samples and could bind to the three prime untranslated regions (3'-UTRs) of both CUL4A and CUL4B, thereby downregulating their expression. The in vitro overexpression or downregulation of miR-194-5p, with a miR-194-5p-mimic or with anti-miR-194-5p, can cause the repression or induction of both CUL4A and CUL4B, respectively. Interestingly, treatment with IL-6 and TNF-α inhibitors in primary hNPCs and hAFCs that were isolated from patients with IDD led to the downregulation of CUL4A and CUL4B. Together, these findings provide insight into how the inflammation-dependent downregulation of miR-194-5p contributes to the pathogenesis of IDD, which may aid in the development of new therapeutic approaches for IDD by directly targeting miR-194-5p or CUL4A and CUL4B.

Echovirus plays a major role in natural recombination in the coxsackievirus B group.

Coxsakievirus (CV) B4, CVB5, and CVA9 belong to the species Enterovirus B. These viruses can cause viral encephalitis, aseptic meningitis, pancreatitis, flaccid paralysis, dilated myocarditis, and hand, foot, and mouth disease (HFMD). In order to analyze the evolution of CVB4, CVB5, and CVA9, we analyzed all of the available genome sequences of Enterovirus B (EVB) isolates and found that there were 12 putative recombination events that produced CVB4, 13 putative recombination events that produced CVB5, and 10 putative recombination events that produced CVA9. These recombination events involved 17 EVB serotypes as major or minor parents. The most active Echovirus (EchoV) appears to have been involved in 20 of the 35 recombination events, acting as one of the parental viruses of circulating CVB4, CVB5, and CVA9 strains. Our study indicates that EchoV plays a major role in recombination in the CVB group, and Echov_E30 is the most active in CVB4, whereas Echov_E3 and Echov_E25 are the most active in CVA9.

microRNA-29a-3p, Up-Regulated in Human Gastric Cells and Tissues with H.Pylori Infection, Promotes the Migration of GES-1 Cells via A20-Mediated EMT Pathway.

BACKGROUND/AIMS: Helicobacter pylori (H. pylori) infection is closely related to human gastric mucosa-associated diseases. Several recent studies on miRNAs have expanded our insights on H.pylori pathogenesis. This study aimed to investigate the biological roles and underlying molecular mechanisms of miR-29a-3p in human gastric cells and tissues with H.pylori infection. METHODS: miR-29a-3p expression was quantified by quantitative RT-PCR (qRT-PCR). A miR-29a-3p target gene was validated by bioinformatics analysis, western blotting and dual luciferase reporter gene assays. Western blotting and immunohistochemistry (IHC) assay were performed to detect the protein expression. Transwell assay was used to determine the cell migration ability. RESULTS: MiR-29a-3p was up-regulated in H.pylori-positive gastric mucosa tissues and H.pylori-infected gastric cells. The up-regulation of miR-29a-3p was dose-dependent in BGC-823 and GES-1 cells infected with H.pylori. Using gain- and loss-of-function experiments in vitro, we demonstrated that miR-29a-3p promoted the migration of gastric epithelial cells. We further characterized A20 as a direct target of miR-29a-3p. The expression of A20 was decreased in H.pylori-positive gastric mucosa tissues compared with H.pylori-negative gastric mucosa tissues. A20 downregulation was time- and dose-dependent in GES-1 and BGC-823 cells infected with H.pylori. In GES-1 and BGC-823 cells infected with H.pylori, the miR-29a-3p mimic significantly blocked A20 expression, which suggests that H.pylori decreased A20 expression through up-regulating miR-29a-3p in GES-1 and BGC-823 cells infected with H.pylori. The knockdown of A20 by siRNA enhanced the migration of human gastric epithelial cells and promoted the expression of Snail, Vimentin, and N-cadherin and inhibited the expression of E-cadherin. CONCLUSION: The miR-29a-3p may act as a tumor promotive miRNA by regulating cells migration through directly targeting of A20 gene in human gastric epithelial cells infected with H.pylori.

Zbtb20 regulates the terminal differentiation of hypertrophic chondrocytes via repression of Sox9.

The terminal differentiation of hypertrophic chondrocytes is a tightly regulated process that plays a pivotal role in endochondral ossification. As a negative regulator, Sox9 is essentially downregulated in terminally differentiated hypertrophic chondrocytes. However, the underlying mechanism of Sox9 silencing is undefined. Here we show that the zinc finger protein Zbtb20 regulates the terminal differentiation of hypertrophic chondrocytes by repressing Sox9. In the developing skeleton of the mouse, Zbtb20 protein is highly expressed by hypertrophic chondrocytes from late embryonic stages. To determine its physiological role in endochondral ossification, we have generated chondrocyte-specific Zbtb20 knockout mice and demonstrate that disruption of Zbtb20 in chondrocytes results in delayed endochondral ossification and postnatal growth retardation. Zbtb20 deficiency caused a delay in cartilage vascularization and an expansion of the hypertrophic zone owing to reduced expression of Vegfa in the hypertrophic zone. Interestingly, Sox9, a direct suppressor of Vegfa expression, was ectopically upregulated at both mRNA and protein levels in the late Zbtb20-deficient hypertrophic zone. Furthermore, knockdown of Sox9 greatly increased Vegfa expression in Zbtb20-deficient hypertrophic chondrocytes. Our findings point to Zbtb20 as a crucial regulator governing the terminal differentiation of hypertrophic chondrocytes at least partially through repression of Sox9.

Echovirus plays major roles in the natural recombination of Coxsackievirus B3.

Coxsakievirus B3 (CVB3) is a member of enterovirus B (EVB) group, which can cause serious heart diseases such as viral myocarditis. In order to analyze the evolution of CVB3, we performed a recombination analysis of all viral genomes of enterovirus B, and found that there were 19 putative recombination events that produced CVB3. A total of 11 serotypes were found to be involved in the generation of CVB3 progeny virus. These recombination events involved echovirus, EcoV (which includes EcoV6, EcoV9, EcoV14, EcoV15, EcoV17, EcoV21, EcoV24, and EcoV25), CVB4, CVB5, and EVB81, as major or minor parents. The most active, EcoV, which was involved in the 14 of 19 recombination events, acts as one of the parental viruses for CVB3 strains among molecular evolution and recombination events in circulating CVB3. Our study indicates that, EcoV plays major roles in CVB3 recombination, and is involved in the production of 11 new CVB3 recombinant strains.

Recombination analysis of coxsackievirus B5 genogroup C.

Coxsackievirus B5 (CVB5) is a member of the species Enterovirus B of the genus Enterovirus, family Picornaviridae. Based on its VP1 sequence, CVB5 is divided into four genogroups: A, B, C, and D. From 2002 to 2012, CVB5 serotype genogroup C caused an outbreak of aseptic meningitis in China. In order to study the evolution of CVB5 genogroup C, phylogenetic and recombination analysis was performed using the 399 available enterovirus B genome sequences in the GenBank database. The results indicated that 10 strains of CVB5 serotype genogroup C resulted from recombination between members of genogroup B and echovirus serotype E6, and another 5 strains resulted from recombination between members of genogroup C and serotype CVB4. These recombination events were confirmed by phylogenetic analysis.

Intergenotype Recombination among New Norovirus GII.4 Variants in the Complete Genome.

GII.4 noroviruses (NoVs) are a major cause of acute gastroenteritis in humans. A new variant of GII.4, the Sydney variant, has recently become more prevalent on a global scale. Intragenotype recombinations are widespread within the pandemic NoV GII.4 lineage, and are likely to be important forces driving the evolution and emergence of novel GII.4 viruses. In this study, we sought to examine the role that intergenotype recombination has played in the emergence of GII.4 Sydney 2012 variants. The results show that the GII.4 Sydney 2012 variants, Kawasaki194 and CA3477, were intergenotype recombination NoV strains with a GII.4 capsid and a GII.P16 polymerase gene. It has been reported for the first time that GII.4 new variant recombinants come from intergenotype recombination of GII.P16 and GII.4 strains in the complete genome.

Emodin inhibits coxsackievirus B3 replication via multiple signalling cascades leading to suppression of translation.

CVB3 (coxsackievirus 3) is a primary causal agent of viral myocarditis. Emodin is a natural compound isolated from certain plant roots. In the present study, we found that emodin inhibited CVB3 replication in vitro and in mice, and now we report an unrecognized mechanism by which emodin inhibits CVB3 replication through suppression of viral protein translation via multiple pathways. On one hand, emodin treatment inhibited Akt/mTOR (mammalian target of rapamycin) signalling and activated 4EBP1 (eukaryotic initiation factor 4R-binding protein 1), leading to suppression of translation initiation of ribosomal protein L32 encoded by a 5'-TOP (terminal oligopyrimidine) mRNA. On the other hand, emodin treatment differentially regulated multiple signal cascades, including Akt/mTORC1/p70(S6K) (p70 S6 kinase), ERK1/2 (extracellular-signal-regulated kinase 1/2)/p90(RSK) (p90 ribosomal S6 kinase) and Ca(2+)/calmodulin, leading to activation of eEF2K (eukaryotic elongation factor 2 kinase) and subsequent inactivation of eEF2 (eukaryotic elongation factor 2), resulting in inhibition of CVB3 VP1 (viral protein 1) synthesis. These data imply that eEF2K is a major factor mediating cross-talk of different arms of signalling cascades in this signal network. This notion was verified by either overexpressing eEF2K or treating the cells with siRNAs or eEF2K inhibitor A484954. We showed further that the emodin-induced decrease in p70(S6K) phosphorylation plays a dominant positive role in activation of eEF2K and in turn in conferring the antiviral effect of emodin. This finding was further solidified by expressing constitutively active and dominant-negative Akt. Collectively, our data reveal that emodin inhibits viral replication through impairing translational machinery and suppression of viral translation elongation.