PRMT5 promotes epithelial-mesenchymal transition via EGFR-ß-catenin axis in pancreatic cancer cells.

Protein arginine methyltransferase 5 (PRMT5) has been implicated in the development and progression of human cancers. However, few studies reveal its role in epithelial-mesenchymal transition (EMT) of pancreatic cancer cells. In this study, we find that PRMT5 is up-regulated in pancreatic cancer, and promotes proliferation, migration and invasion in pancreatic cancer cells, and promotes tumorigenesis. Silencing PRMT5 induces epithelial marker E-cadherin expression and down-regulates expression of mesenchymal markers including Vimentin, collagen I and ß-catenin in PaTu8988 and SW1990 cells, whereas ectopic PRMT5 re-expression partially reverses these changes, indicating that PRMT5 promotes EMT in pancreatic cancer. More importantly, we find that PRMT5 knockdown decreases the phosphorylation level of EGFR at Y1068 and Y1172 and its downstream p-AKT and p-GSK3ß, and then results in down-regulation of ß-catenin. Expectedly, ectopic PRMT5 re-expression also reverses the above changes. It is suggested that PRMT5 promotes EMT probably via EGFR/AKT/ß-catenin pathway. Taken together, our study demonstrates that PRMT5 plays oncogenic roles in the growth of pancreatic cancer cell and provides a potential candidate for pancreatic cancer treatment.

Linc-RoR promotes proliferation, migration, and invasion via the Hippo/YAP pathway in pancreatic cancer cells.

Large intergenic noncoding RNA regulator of reprogramming (Linc-RoR) was first identified as a regulator to increase the emergence of induced pluripotent stem cells through reprogramming differentiated cells and is abnormal expression in a variety of malignant tumors. However, the function of Linc-RoR in pancreatic cancer progression needs further clarification. The data from this study demonstrated that Linc-RoR knockdown suppressed cell proliferative capacity and colony formation, while Linc-RoR overexpression promoted these behaviors. In particular, Linc-RoR overexpression promoted the level of mesenchymal markers, inhibited the expression of epithelial markers, as well as enhanced pancreatic cancer cells migration and invasion, whereas Linc-RoR knockdown inhibited the expression of mesenchymal markers, promoted the expression of epithelial markers, as well as weakened pancreatic cancer cells migration and invasion. Further study revealed that Linc-RoR knockdown brought about a significant fall in YAP phosphorylation and a rise in total YAP, while Linc-RoR overexpression produced the opposite results. Specifically, Linc-RoR promoted YAP in the cytoplasm into the nucleus. Taken together, we conjectured that Linc-RoR promoted proliferation, migration, and invasion of pancreatic cancer cells by activating the Hippo/YAP pathway. YAP might be an underlying target of Linc-RoR and mediate epithelial-mesenchymal transition (EMT) in pancreatic cancer (PC); thus, Linc-RoR might be a very meaningful biomarker for PC.

Protein arginine methyltransferase 1 coordinates the epithelial-mesenchymal transition/proliferation dichotomy in gastric cancer cells.

Protein arginine methyltransferase 1 (PRMT1) is up-regulated and promotes migration, invasion and proliferation in wide range of cancers. However, we for the first time identify that PRMT1 promotes migration and invasion and inhibits proliferation in gastric cancer cells, a phenomenon called "migration-proliferation dichotomy". First, we find that PRMT1 overexpression promotes migration and invasion and inhibits proliferation, whereas PRMT1 knockdown reverses the above abilities. Next, PRMT1 reduces the expression of epithelial marker E-cadherin and increases the expression of mesenchymal markers including N-cadherin, Vimentin, snail and ß-catenin in gastric cancer cells. Furthermore, our studies show that PRMT1 silencing promotes the phosphorylation of LATS1, and then induces YAP phosphorylation, while overexpression of PRMT1 down-regulates the phosphorylation of LATS1 and YAP, indicating that PRMT1 inhibits EMT probably via Hippo signaling. Collectively, the present study reveals important roles of PRMT1 in progression of gastric cancer. Given the dual functions of PRMT1, it is as a potential drug target of gastric cancer with extreme caution.

LncRNA UCA1 promotes migration and invasion in pancreatic cancer cells via the Hippo pathway.

Although overexpression of the long non-coding RNA (lncRNA) UCA1 has been implicated in several human cancers, its biological function in pancreatic cancer remains to be clarified. In this study, we reported that UCA1 expression was significantly increased in pancreatic cancer tissues and correlated with clinicopathological features, tumor stage, and poorer patient outcome. We further showed that UCA1 promoted cell migration and invasion of pancreatic cancer cells. Importantly, we found that UCA1 overexpression inhibited YAP phosphorylation, and increased YAP expression. Mechanistically, UCA1 interacted with MOB1, Lats1, and YAP, forming shielding composites. Moreover, we demonstrated that UCA1 increased YAP nuclear localization and stabilization, and improved TEAD luciferase activity. In turn, YAP promotes UCA1 expression. Collectively, the present study provides insights into the mechanistic regulation of UCA1 promoting pancreatic cancer progression through the Hippo signaling pathway. UCA1 may serve as a candidate biomarker for poor prognosis and a target for new pancreatic cancer therapies.

TGF-ß1-miR-200a-PTEN induces epithelial-mesenchymal transition and fibrosis of pancreatic stellate cells.

Although the function of miR-200a has been discussed in many cancers and fibrotic diseases, its role in pancreatic fibrosis is still poorly understood. In this study, we for the first time confirm that miR-200a attenuates TGF-ß1-induced pancreatic stellate cells activation and extracellular matrix formation. First, we find that TGF-ß1 induces activation and extracellular matrix (ECM) formation in PSCs, and the effects are blocked by the inhibitor of PI3K (LY294002). Furthermore, we identify that miR-200a is down-regulated in TGF-ß1-activated PSCs, and up-regulation of miR-200a inhibits PSCs activation induced by TGF-ß1. Meanwhile, TGF-ß1 inhibits the expression of the epithelial marker E-cadherin, and increases the expression of mesenchymal markers vimentin, and the expression of ECM proteins a-SMA and collagen I, while miR-200a mimic reversed the above effects in PSCs, indicating that miR-200a inhibits TGF-ß1-induced activation and epithelial-mesenchymal transition (EMT). In addition, overexpression of miR-200a promotes the expression of PTEN and decreases the expression of matrix proteins and attenuates phosphorylation of Akt and mTOR. Taken together, our study uncovers a novel mechanism that miR-200a attenuates TGF-ß1-induced pancreatic stellate cells activation and ECM formation through inhibiting PTEN /Akt/mTOR pathway.

Scoparone Protects Against Pancreatic Fibrosis via TGF-ß/Smad Signaling in Rats.

BACKGROUND/AIMS: This study was to investigate the influence of scoparone on pancreatic fibrosis in vitro and in vivo. METHODS: Pancreatic stellate cells (PSCs) were isolated from pancreas tissue blocks, and cultured for 3-5 generations for the experiment. PSCs were treated with scoparone in different doses as experimental groups, salvianolic acid B as a positive control and PBS as a blank group. We measured the effects of scoparone on cellular proliferation, oxidative stress, epithelial-mesenchymal transition (EMT), and pancreatic fibrosis. Cellular oxidative stress was detected by using commercially available kits. The impact of scoparone on EMT and fibrosis was detected through immunofluorescence or western blotting. RESULTS: Compared with the control group, scoparone significantly inhibited stellate cell proliferation, and reduced MDA, the expression of mesenchymal makers, and increased the levels of SOD and the expression of E-cadherin (P < 0.05). Western blot analysis showed that scoparone downregulated the expression of TGF-ß and p-smad2/3, and upregultated the expression of smad7 (P < 0.05). CONCLUSION: Scoparone can reduce the levels of oxidative stress, repress pancreatic stellate cells activation, and alleviate fibrosis by regulating TGF-ß/Smad pathway.

[miR-15a and miR-16 modulate drug resistance by targeting bcl-2 in human colon cancer cells].

OBJECTIVE: To investigate the reversal effect of targeted modulation of bcl-2 expression by miR-15a and miR-16 on drug resistance of human colon cancer cells. METHODS: Mimics or inhibitors of miR-15a and miR-16 were transfected into HCT8 or HCT8/VCR cells with the help of Lipofectamine 2000. The expressions of miR-15a and miR-16 mRNA were detected by RT-qPCR. The levels of bcl-2 and P-gp proteins were measured by Western blot. The inhibitory effects of VCR on growth of HCT8 and HCT8/VCR cells were detected by CCK8. RESULTS: After transfection of the mimics, the expression of miR-15a in the blank control group, negative control group and miR-15a mimic group was 1.00, 0.87 ± 0.24, and 223.44 ± 59.07, respectively, and miR-15a was increased significantly (P < 0.001). The expression of miR-16 in the blank control group, negative control group and miR-16 mimic group was 1.00, 0.66 ± 0.19, and 107.32 ± 22.58, respectively, and miR-16 expression was increased significantly (P < 0.001). The Western blot assay showed that the relative expressions of bcl-2 protein in the blank control group, negative control group, miR-15a mimic group and miR-16 mimic group were 1.00, 0.97 ± 0.02, 0.51 ± 0.06, and 0.65 ± 0.03, respectively, and the expression of bcl-2 protein was decreased significantly (P < 0.05), however, the expressions of P-gp protein showed no significant difference. The CCK8 test showed that at 1, 5, 25 and 125 µg/ml concentration of VCR, the survival rates of HCT8/VCR cells were basically the same in the blank control group, negative control group, miR-15a mimic group and miR-16 mimic group, but the survival rate of HCT8/VCR cells was significantly decreased after transfection of mimics (P < 0.05). After transfection of the inhibitors, the expressions of both miR-15a and miR-16 were decreased significantly (P < 0.001). The Western blot showed that the expression of bcl-2 protein was increased (P < 0.05), while the expression of P-gp protein showed no significant difference. The CCK8 test showed that the survival rate of HCT8 cells which were transfected with inhibitors was significantly higher than that of the blank control group (P < 0.05). CONCLUSIONS: miR-15a and miR-16 may reverse the drug resistance in human colon cancer cells. A possible mechanism is regulating the expression of bcl-2.

The influence of management practices on plant diversity: a comparative study of three urban wetlands in an expanding city in eastern China.

Rapid urbanization has drawn some aquatic environments into the urban texture from the outskirts of cities, and the composition and distribution of plant species in urban wetlands along the urban gradient have changed. Understanding the drivers of these changes will help in the conservation and utilization of urban wetlands. This study investigated the differences in plant diversity and associated influencing factors in three wetlands, Xixi wetland, Tongjian Lake wetland, and Qingshan Lake wetland, which are located in a core area, fringe area, and suburban area of Hangzhou City, respectively. The results showed that a total of 104 families, 254 genera, and 336 species of plants were recorded in the Xixi wetland; 179 species, 150 genera, and 74 families were found in the Qingshan Lake wetland; and 112 species, 96 genera, and 57 families were collected in the Tongjian Lake wetland. The main plant species and flora distribution of the three urban wetlands showed similarities. Indigenous spontaneous vegetation was highest in the Xixi wetland, while cultivated plant species were most abundant in the Tongjian Lake wetland. The introduction of cultivated plants decreased the distance attenuation effect of plant communities, which led to a certain degree of plant diversity convergence among the three wetlands. Eight endangered plants were preserved in the Xixi wetland by planting them in suitable habitats. Ellenberg's indicator values showed that the proportion of heliophilous plants was higher in the Qingshan Lake wetland, while the proportion of thermophilous plants and nitrogen-loving plants in the Tongjian Lake wetland was higher than in the other two wetlands. The importance of artificial interference factors affecting the differences in plant diversity was significantly higher than that of natural environmental factors in urban wetlands. The preservation of spontaneous plants and the introduction of cultivated plants had an importance of 25.73% and 25.38%, respectively. These were the main factors influencing the plant diversity of urban wetlands. The management mode that did not interfere with spontaneous vegetation and confined maintenance to cultivated plants in the Xixi wetland was beneficial for improving wetland plant diversity. Scientific plant reintroduction can also improve wetland plant diversity.

Preparation of polypeptide-metal complexes-coated Hosenkoside A and its inhibitory effect in cervical cancer.

We reported the anti-cervical cancer effect of proprietary saponin content from seeds of Impatiens balsamina L., Hosenkoside A. Our study found that Hosenkoside A significantly promotes cell apoptosis and cell cycle arrest after administration, exhibiting anti-tumor effects. Then the transcriptome sequencing results after administration showed that Hosenkoside A had a significant inhibitory effect on Histone deacetylase 3 (HDAC3). After sufficient administration time, the inhibition of HDAC3 expression level leads to a significant decrease in lysine acetylation at histone 3 sites 4 and 9, blocking the activation of Signal transducer and activator of transcription 3 (STAT3) and achieving anti-tumor effects. In addition, we encapsulated Hosenkoside A into polypeptide metal complexes (PMC) to form slow-release spheres. This material breaks down in the tumor environment, not only does it solve the problem of low drug solubility, but it also achieves targeted sustained-release drug delivery. Under the same concentration of stimulation, the PMC complex group showed better anti-tumor effects in both in vitro and in vivo experiments.

Jatropholane-type diterpenes from Euphorbia sikkimensis.

Four new jatropholane-type diterpenes (1-4), named sikkimenoids A-D, were isolated from the aerial parts of Euphorbia sikkimensis. The structural elucidations of 1-4 were accomplished by extensive NMR analyses, and their absolute configurations were established by ECD calculations. Compound 2 exhibited weak antiangiogenic activity with an IC(50) value of 43.0 µM when evaluated using a zebrafish model.

[Morphological observation of human gastric cancer cell SGC-7901 clones and identification of gastric cancer stem cells].

OBJECTIVE: To dynamically investigate the morphology of human gastric cancer SGC-7901 cell clones, and then compare the tumorigenic ability of different clones in order to identify the tumor stem cell clones. METHODS: Clones derived from gastric cancer SGC-7901 cells were assessed by morphological observation, and the clone formation rate and proportion of each clone were calculated. The expression of CD44 and CDX2 in different clones was detected by immunofluorescence microscopy and Western blot. Furthermore, different clones were isolated and cultured, and their self-renewal property was assayed. Cells of different clones were subcutaneously inoculated into nude mice and the tumorigenic ability of each group was determined. RESULTS: Clones derived from gastric cancer SGC-7901 cells had three types, i.e. clones of tight, transitional and loose types. The total clone formation rate was (9.80 ± 1.07)%, and the proportion of tight, transitional and loose type clones was 10.2%, 56.0% and 33.8%, respectively. The results of immunofluorescence microscopic examination showed that the signal of CD44 was significantly stronger in the tight clones than in the transitional and loose clones, however, the signal of CDX2 was weakest in the tight colonies. The results of Western blot were consistent with that of immunofluorescence microscopic observation. SGC-7901 cells of tight clones possessed strong ability of self-renewal and in vivo tumorigenicity in the nude mice. CONCLUSION: SGC-7901 cell clones vary in morphology and differentiation, and the tight type clones may include rich gastric cancer stem cells.

Expression of Helicobacter pylori ggt gene in baculovirus expression system and activity analysis of its products.

The gamma-glutamyltranspeptidase (GGT) of Helicobacter pylori (HpGT) is a newly found virulence factor. In an approach to gain insight into the gene function, the four domains of the HpGT were cloned and expressed in baculovirus expression system. The results of a functional assay showed that the HpGT products acted as GGT, even when the N-terminal 380 amino acids were deleted. However, only the full length open reading frame (ORF) of the HpGT gene was apparently effective on cell growth. This result indicated that the products of the full length ORF might have an important role in gastric carcinogenesis. In this paper, we are the first to report that changes of mitochondrial membrane potential can be detected using 5, 5', 6, 6'-tetrachloro-1, 1', 3, 3'-tetraethylbenzimidazole carbocyanine iodide (JC-1) staining in insect cells.

lncRNA SNHG3 acts as oncogene in ovarian cancer through miR-139-5p and Notch1.

Ovarian cancer (OC) is a common malignant tumor of the female reproductive system. Long non-coding RNAs (lncRNAs) play an important role in OC occurrence and development. Thus, the function and potential mechanism of lncRNA small nucleolar RNA host gene 3 (SNHG3) was explored in the development of OC. The expression of SNHG3, microRNA (miR)-139-5p and Notch homolog 1, translocation-associated (Drosophila) (Notch1) in OC were detected by RT-qPCR or western blot assay. In addition, CCK-8 and wound-healing assays were used to detect OVCAR3 proliferation and migration ability. The targeting relationship of miR-139-5p with SNHG3 or Notch1 was verified through luciferase reporter assay. Rescue experiments were performed to confirm whether SNHG3 could mediate OVCAR3 proliferation and migration through miR-139-5p and Notch1. In OC tissues and cell lines, the expression of SNHG3 and Notch1 were significantly increased, and the expression of miR-139-5p was significantly decreased. SNHG3 inhibition suppressed the proliferation and migration of OVCAR3 cells. Luciferase reporter experiment confirmed that miR-139-5p could target SNHG3 and Notch1. Transfection of miR-139-5p inhibitor significantly reversed the inhibitory effect of SNHG3 knockdown on OVCAR3 proliferation and migration. Moreover, SNHG3 inhibition or miR-139-5p mimic abolished the promotion of Notch1 overexpression on OVCAR3 proliferation and migration. In conclusion, SNHG3 could accelerate the proliferation and migration of OC cells by regulating miR-139-5p and Notch1.

The KRAS/Lin28B axis maintains stemness of pancreatic cancer cells via the let-7i/TET3 pathway.

Increasing evidence demonstrates that Lin28B plays critical roles in numerous biological processes including cell proliferation and stemness maintenance. However, the molecular mechanisms underlying Lin28B nuclear translocation remain poorly understood. Here, we found for the first time that KRAS promoted Lin28B nuclear translocation through PKCß, which directly bound to and phosphorylated Lin28B at S243. Firstly, we observed that Lin28B was upregulated in pancreatic cancer, contributing to cellular migration and proliferation. Furthermore, nuclear Lin28B upregulated TET3 messenger RNA and protein levels by blocking the production of mature let-7i. Subsequently, increased TET3 expression could also promote the expression of Lin28B, thereby forming a Lin28B/let-7i/TET3 feedback loop. Our results suggest that the KRAS/Lin28B axis drives the let-7i/TET3 pathway to maintain the stemness of pancreatic cancer cells. These findings illuminate the distinct mechanism of Lin28B nuclear translocation and its important roles in KRAS-driven pancreatic cancer, and have important implications for development of novel therapeutic strategies for this cancer.

Analgesic effect of resveratrol on colitis-induced visceral pain via inhibition of TRAF6/NF-κB signaling pathway in the spinal cord.

Visceral pain is a complex and common symptom of inflammatory bowel disease (IBD) patients. Developing novel efficient therapeutics is still a common interest for clinicians. Increasing evidence have shown that tumor necrosis factor (TNF) receptor associated factor 6 (TRAF6) contributes to the pathological pain state in some pain models. Resveratrol (RSV) has showed promising potential for the treatment of neuropathic pain and inflammatory pain. However, whether RSV has analgesic effect on visceral pain and the underlying mechanisms remain unclear. In this study, we established the colitis model through intrarectal administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS), and found that TNBS induced colonic inflammation and visceral hypersensitivity. Meanwhile, astroglial marker glial fibrillary acidic protein (GFAP), TRAF6, phosphorylation of NF-κB (pNF-κB), tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) levels were increased in L6-S1 spinal cord after TNBS enema. Then, intrathecal injection of TRAF6 siRNA attenuated visceral pain, blocked the upregulation of pNF-κB, TNF-α and IL-1ß levels in the spinal cord in TNBS mice. Furthermore, spinal administration of NF-κB inhibitor, BAY11-7082 reversed the pain behavior and suppressed spinal TNF-α and IL-1ß expression in TNBS mice. Finally, repeated intrathecal injection of RSV reversed TNBS-induced visceral pain hypersensitivity in a dose-dependent manner. Meanwhile, TNBS-induced enhancement of spinal GFAP, TRAF6, pNF-κB, TNF-α and IL-1ß were reduced by the same treatment of RSV. In conclusion, our results suggest that RSV exerts the effects of antinociception on colitis-induced visceral hyperalgesia through inhibition of spinal TRAF6/NF-κB signaling pathway and the production of inflammatory mediators in the spinal cord, suggesting a new application of RSV for the treatment of visceral pain.

The UCA1/KRAS axis promotes human pancreatic ductal adenocarcinoma stem cell properties and tumor growth.

Emerging evidence indicates that the long noncoding RNA UCA1 is upregulated in multiple cancers, including pancreatic ductal adenocarcinoma (PDAC), and plays a critical role in various complex biological processes. However, the functional roles of UCA1 in PDAC remain to be clarified. In the current study, we showed that UCA1 significantly promoted cell proliferation and tumor growth both in vitro and in vivo, and enhanced stemness maintenance of PDAC cell lines. Moreover, we found that UCA1 overexpression increased the activity and expression of oncogenic KRAS. Mechanistically, upregulated UCA1 increased phospho-KRAS protein levels by interacting with hnRNPA2B1, and KRAS facilitated high cytoplasmic accumulation of hnRNPA2B1. Additionally, we identified that UCA1 functioned as a competing endogenous RNA (ceRNA) to increase the expression of KRAS via sponging miR-590-3p, and in turn, KRAS promoted UCA1 expression. Collectively, these findings suggest that the UCA1-KRAS axis plays a crucial role in PDAC progression and that UCA1 may serve as a target for new PDAC therapies.

Significance of Bcl-xL in human colon carcinoma.

AIM: To investigate the clinical significance of Bcl-xL gene in the pathogenesis of human colon carcinoma. METHODS: Fifty-six pair tissue samples from patients with colon cancer were collected, and protein level of the Bcl-xL gene was measured by immunohistochemistry method. The correlation of Bcl-xL expression with clinical index was evaluated. After human colon cancer cell line HT29 was transfected with Bcl-xL small interfering RNA (siRNA), the anchorage-independent growth of cancer cells was detected by colony formation in soft agar and invasion ability of cancer cells was determined by a transwell model. RESULTS: The Bcl-xL expression was higher in cancerous tissue samples than in normal tissue samples (38.78 +/-11.36 vs 0.89 +/- 0.35, P < 0.001), and was associated with the pathological grade, lymph node metastasis and Duke's stage of colorectal carcinoma. Transfection with Bcl-xL siRNA inhibited the colony formation and invasion ability of human colon cancer cell line HT29 in vitro. CONCLUSION: Bcl-xL gene plays an important role in carcinogenesis of human colorectal carcinoma and is associated with malignant biological behaviors of human colorectal carcinoma.

Inhibition of signal transducer and activator of transcription 3 expression by RNA interference suppresses invasion through inducing anoikis in human colon cancer cells.

AIM: To investigate the roles and mechanism of signal transducer and activator of transcription 3 (STAT3) in invasion of human colon cancer cells by RNA interference. METHODS: Small interfering RNA (siRNA) targeting Signal transducer and activator of transcription 3 (STAT3) was transfected into HT29 colon cancer cells. STAT3 protein level and DNA-binding activity of STAT3 was evaluated by western blotting and electrophoretic mobility shift assay (EMSA), respectively. We studied the anchorage-independent growth using colony formation in soft agar, and invasion using the boyden chamber model, anoikis using DNA fragmentation assay and terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling (TUNEL), respectively. Western blot assay was used to observe the protein expression of Bcl-xL and survivin in colon cancer HT29 cells. RESULTS: RNA interference (RNAi) mediated by siRNA leads to suppression of STAT3 expression in colon cancer cell lines. Suppression of STAT3 expression by siRNA could inhibit anchorage-independent growth, and invasion ability, and induces anoikis in the colon cancer cell line HT29. It has been shown that knockdown of STAT3 expression by siRNA results in a reduction in expression of Bcl-xL and survivin in HT29 cells. CONCLUSION: These results suggest that STAT3 siRNA can inhibit the invasion ability of colon cancer cells through inducing anoikis, which antiapoptotic genes survivin and Bcl-xL contribute to regulation of anoikis. These studies indicate STAT3 siRNA could be a useful therapeutic tool for the treatment of colon cancer.

MicroRNA-146a-5p attenuates visceral hypersensitivity through targeting chemokine CCL8 in the spinal cord in a mouse model of colitis.

Visceral pain, observed in inflammatory bowel disease (IBD) patients, is a challenging medical problem and remains poorly understood because the mechanisms underlying it are unclear. Emerging evidence indicates that microRNAs (miRNAs) play a crucial role in the pathogenesis of acute and chronic pain. In this study, we aimed to explore the potential role of miR-146a-5p (the mature form of miR-146a) in a mouse model of colitis induced by intracolonic injection of trinitrobenzene sulfonic acid (TNBS). We found that induction of colitis resulted in visceral hyperalgesia manifested by a decreased pain threshold to colorectal distension and upregulation of miR-146a-5p expression in the lumbosacral spinal cord. In situ hybridization and immunohistochemistry results showed that miR-146a-5p was colocalized with neuronal marker NeuN, but not with astrocytic marker GFAP or microglial marker IBA-1. Dual-luciferase reporter assay showed that miR-146a-5p directly targeted the 3'-untranslated region (UTR) of CCL8, which was previously identified as an important regulator of visceral pain. In cultured Neuro-2a cells, TNF-α-induced CCL8 upregulation was decreased by transfection of miR-146a-5p mimic dose-dependently. In vivo, exogenous supplementation of miR-146a-5p by intrathecal miR-146a-5p agomir significantly alleviated visceral pain and decreased CCL8 expression in colitis mice. Furthermore, inhibition of CCL8 expression by CCL8 siRNA relieved colitis-induced visceral nociception. Finally, in naïve mice intrathecal miR-146a-5p antagomir upregulated CCL8 expression and induced visceral pain hypersensitivity, which could be partially rescued by neutralization of CCL8. Taken together, the present findings indicate that miR-146a-5p may be an endogenous suppressor of visceral pain and exogenous supplementation of miR-146a-5p could exert an analgesic effect at least partly by targeting spinal CCL8 expression. Thus, miR-146a-5p may serve as a novel therapeutic target for visceral pain intervention in the context of colitis.