Effect of CX3CL1/CX3CR1 in cerebral ischemia and its mechanisms / 国际脑血管病杂志

CX3CL1, also known as Fractalkine, is the only member of chemokines CX3C subclass. It plays an important role in a variety of central nervous system diseases and ischemic cerebrovascular diseases by binding to its specific receptor CX3CR1. In recent years, a large number of studies have investigated the specific role and related molecular mechanism of CX3CL1/CX3CR1. This article reviews the effect and molecular mechanism of CX3CL1/CX3CR1 in ischemic cerebrovascular disease, aiming to expand the understanding of the mechanism of CX3CL1/CX3CR1, and provide new ideas and intervention targets for the prevention, diagnosis and treatment of ischemic cerebrovascular disease.

Research advances of chemokine receptor CX3CR1 and microglia in retinal degenerative diseases / 国际眼科杂志(Guoji Yanke Zazhi)

@#Retinal degenerative diseases such as retinitis pigmentosa and age-related macular degeneration are the main clinical blinding eye diseases with complex etiology and irreversible damage to vision. CX3CR1 is a specific receptor of the chemokine CX3CL1. Both of them participate in various physiological functions and pathological changes of the whole body through regulating the immune system of the body. In recent years, studies have pointed out that CX3CR1 regulates the activity and function of retinal microglia, which play an important role in the process of retinal degenerative diseases. In this paper, the structure and function of the chemokine receptor CX3CR1 and the role of microglia in retinal degenerative diseases were reviewed, so as to provide ideas and directions for future research and treatment of such diseases.

Dexmedetomidine alleviates cognitive dysfunction induced by tibial fracture in rats / 南方医科大学学报

OBJECTIVE@#To study the effect of dexmedetomidine (Dex) on cognitive dysfunction induced by tibial fracture in rats.@*METHODS@#Sixteen male SD rats were randomized into control group and tibial fracture group, and the behavior indicators were measured on days 1, 3, 5 and 7 after tibial fracture and the expressions of CX3L1 protein and mRNA in the hippocampus were detected. Another 24 male SD rats were randomly divided into control group, tibial fracture group, and tibia fracture with CX3CL1 antibody group, and the behavior indicators and hippocampal CX3L1 protein expression were evaluated after corresponding treatments. In another experiment, we randomized 24 male SD rats into control group, tibial fracture group and tibial fracture with Dex treatment, and tested their hippocampal CX3L1 protein and mRNA expressions as well as the behavior indicators after the treatments.@*RESULTS@#Compared with the control rats, the rats with tibial fracture spent significantly less time in the novel arm on days 1, 3, 5 and 7 after the fracture ( < 0.05) with obviously lowered expressions of CX3L1 protein and mRNA in the hippocampus ( < 0.05). In the rats with tibial fracture, treatment with CX3CL1 antibody further decreased the time spent in the novel arm ( < 0.05) and the expression level of CX3L1 protein in the hippocampus ( < 0.05); In contrast, treatment with Dex significantly increased the time spent time in the novel arm ( < 0.05) and enhanced the hippocampal expressions of CX3L1 protein and mRNA in rats with tibial fractures ( < 0.05).@*CONCLUSIONS@#Dex can alleviate cognitive dysfunction induced by tibial fracture in rats by increasing the expression of CX3CL1 in the hippocampus.

Signal transducer and activator of transcription 3 promotes vascular endothelial cell proliferation and migration by fractalkine / 生物工程学报

Signal transducer and activator of transcription 3 (STAT3) and Chemokine CX3C ligand 1 (Fractalkine/CX3CL1) play important roles in vascular inflammation and injury. To study if STAT3 promotes vascular endothelial cell proliferation and migration through fractalkine, we overexpressed or knocked down STAT3 in vascular endothelial cells, and used quantitative real-time PCR and Western blotting to determine the effect of STAT3 on fractalkine expression. The wild type and STAT3 binding site mutant fractalkine promoter luciferase reporter plasmids were constructed, and luciferase activity assays were used to explore the effect of STAT3 on the transcriptional activity of the fractalkine promoter. MTT assays were used to detect the effect of overexpression or knockdown of STAT3 or fractalkine on the proliferation rate of vascular endothelial cells. Scratch assays were used to detect the effect of overexpression or knockdown of STAT3 or fractalkine on vascular endothelial cell migration. There results showed that overexpression of STAT3 could promote fractalkine expression, and knockdown of STAT3 could down-regulate fractalkine expression. STAT3 could directly bind to the promoter of fractalkine to promote its transcriptional activity via binding the GAS site of the fractalkine promoter. Knockdown of STAT3 could inhibit the migration of vascular endothelial cell, and overexpression of fractalkine antagonized this inhibition. Our data concluded that STAT3 promotes the proliferation and migration of vascular endothelial cell by binding the GAS site of the fractalkine promoter to promote fractalkine transcriptional activity and expression.

Roles and mechanisms of chemokine CX3CL1 in cerebral ischemia / 国际脑血管病杂志

Chemokine CX3CL1 mainly participates in the physiological and pathological processes of nervous system by activating its receptor CX3CR1. Under physiological conditions, CX3CL1 can inhibit the activation of microglia; when cerebral ischemia and hypoxia, CX3CL1 can affect the expression of multiple downstream target genes involved in activation of adenosine receptor, inhibition of Ca2+ influx, and promotion of blood vessel growth. It is of great significance to improve the energy metabolism disorder and to establish microcirculation around infarcts after cerebral ischemia.

Roles and mechanisms of chemokine CX3CL1 in cerebral ischemia / 国际脑血管病杂志

Chemokine CX3CL1 mainly participates in the physiological and pathological processes of nervous system by activating its receptor CX3CR1.Under physiological conditions,CX3CL1 can inhibit the activation of microglia;when cerebral ischemia and hypoxia,CX3CL1 can affect the expression of multiple downstream target genes involved in activation of adenosine receptor,inhibition of Ca 2+ influx,and promotion of blood vessel growth.It is of great significance to improve the energy metabolism disorder and to establish microcirculation around infarcts after cerebral ischemia.

NF-kB mediated CX3CL1 activation in the dorsal root ganglion contributes to the maintenance of neuropathic pain induced in adult male Sprague Dawley rats

Abstract Purpose: To evaluate the role of CX3CL1 and NF-κB in the lumbar disc herniation induced neuropathic pain. Methods: After LDH induced by implantation of autologous nucleus pulposus (NP) on the left L5 nerve root was established, mechanical thresholds and thermal hyperalgesia were tested at relevant time points during an observation period of 28 days. Expression of CX3CL1 and NF-κBin the dorsal root ganglion (DRG) were performed by using Western blotting and RT-PCR. Results: Implantation of autologous nucleus pulposus (NP) induced neuropathic pain, associated with increased mRNA and protein expression of CX3CL1 in the DRG. Moreover, intrathecal injection of neutralizing antibody against CX3CL1 could attenuates LDH-induced persistent pain hypersensitivity. Interestingly, NF-κB activation in the DRGs were found in LDH-induced neuropathic pain. Furthermore, NF-κB downregulation by p65 inhibitor PDTC markedly alleviated LDH-induced mechanical allodynia and thermal hyperalgesia in rat. Importantly, CX3CL1 neutralizing antibody (10 μg/10 μl, i.t.) reduces p-p65 protein level in DRG Conclusions: CX3XL1 could regulate LDH-induced neuropathic pain through NF-κB pathway. Targeting CX3CL1 and NF-κB may represent a potential treatment for neuropathic pain caused by LDH.

Tissue-specific Role of CX₃CL1 Expressing Immune Cells and Their Relationships with Human Disease

Chemokine (C-X3-C motif) ligand 1 (CX₃CL1, also known as fractalkine) and its receptor chemokine (C-X3-C motif) receptor 1 (CX₃CR1) are widely expressed in immune cells and non-immune cells throughout organisms. However, their expression is mostly cell type-specific in each tissue. CX₃CR1 expression can be found in monocytes, macrophages, dendritic cells, T cells, and natural killer (NK) cells. Interaction between CX3CL1 and CX₃CL1 can mediate chemotaxis of immune cells according to concentration gradient of ligands. CX₃CL1 expressing immune cells have a main role in either pro-inflammatory or anti-inflammatory response depending on environmental condition. In a given tissue such as bone marrow, brain, lung, liver, gut, and cancer, CX₃CL1 expressing cells can maintain tissue homeostasis. Under pathologic conditions, however, CX₃CL1 expressing cells can play a critical role in disease pathogenesis. Here, we discuss recent progresses of CX3CL1/CX₃CL1 in major tissues and their relationships with human diseases.

Pooled analysis of prognostic significance of CX3CL1 mRNA expression level in patients with non-small cell lung cancer / 医学研究生学报

Objective The prognostic expression level and prognostic significance of CX3CL1 in patients with non-small cell lung cancer(NSCLC) need further investigation. The purpose of this paper was to investigate the effects of various CX3CL1 mRNA expression levels on patients with NSCLC.Methods By retrieving lung-cancer related gene expression profile data in NCBI GEO database and TCGA of UCSC Cancer Browser, 8 datasets were included based on inclusion and exclusion criteria. All the datasets were collated and standardized through R statistical software. Univariate and multivariate Cox models were conducted for prognosis analysis of CX3CL1 in each dataset. HR values of all the datasets were pooled by meta algorithm.Results High-expression of CX3CL1 mRNA in tumor tissues of lung adenocarcinoma was a positive prognostic factor for overall survival(pooled HR=0.53; 95% CI=0.43-0.65 in univariate analysis; pooled HR=0.52; 95% CI=0.42-0.64 in multivariate analysis). However, in lung squamous cell carcinoma, there was no significant association between CX3CL1 expression and overall survival (pooled HR=1.09; 95% CI=0.82-1.45 in univariate analysis; pooled HR=1.18; 95% CI=0.88-1.58 in multivariate analysis).Conclusion The mRNA level of CX3CL1 in lung adenocarcinoma was positively correlated with better prognosis, but there was no correlation between CX3CL1 mRNA level and prognosis in patients with lung squamous cell carcinoma. CX3CL1 may be used as a potential prognostic marker for patients with lung adenocarcinoma.

Renovascular hypertension increases serum TNF and CX3CL1 in experimental Trypanosoma cruzi infection

Trypanosoma cruzi triggers a progressive inflammatory response affecting cardiovascular functions in humans and experimental models. Angiotensin II, a key effector of the renin-angiotensin system, plays roles in mediating hypertension, heart failure, and inflammatory responses. T. cruzi and AngII can induce inflammatory responses by releasing inflammatory mediators. The aim of this study was to evaluate systemic AngII, tumor necrosis factor (TNF), and CX3CL1 mediators in a two-kidney one-clip (2K1C) renovascular hypertension model using Wistar rats infected with T. cruzi. Our data showed an increase in serum AngII in uninfected and T. cruzi-infected rats 1 week after 2K1C surgery compared to non-2K1C (Sham) animals. The baseline systolic blood pressure was higher in both uninfected and infected 2K1C rats. Despite no difference in circulating parasites in the acute phase of infection, elevated serum TNF and CX3CL1 were observed at 8 weeks post-infection in 2K1C rats in association with higher cardiac inflammatory infiltration. In summary, AngII-induced hypertension associated with T. cruzi infection may act synergistically to increase TNF and CX3CL1 in the 2K1C rat model, thereby intensifying cardiac inflammatory infiltration and worsening the underlying inflammation triggered by this protozoan.

Correlation analysis of chemokine CX3CL1 level and the prognosis of ST elevation myocardial infarction / 解放军医学杂志

Objective To investigate the correlation ofchemokine CX3CL1 and the major adverse cardiovascular events (MACCE) within 1 year in patients with ST elevation myocardial infarction (STEMI).Methods Five hundred patients with STEMI were continuously selected from the Department of Emergency,General Hospital of Shenyang Command,and 5ml venous blood was extracted at the time of admission.Plasma CX3CL1 levels were detected by ELISA.The median of CX3CL1 plasma concentration of all selected patients (2108.3pg/ml) was used to assign the patients to low expression group (＜2108.3pg/ml) and high expression group (≥ 2108.3pg/ml).The baseline information and the medical history of patients were recorded and followed up by telephone inquiry for 1 year.The endpoint events were defined as MACCE (including cardiac death,heart failure,nonfatal stroke and nonfatal myocardial infarction).Based on the MACCE in 1 year following up,patients were assigned into MACCE group and non-MACCE group.Multivariable Cox regression analysis was performed to analyze the correlation between CX3CL1 level and MACCE in STEMI patients.Results The plasma CX3CL1 level significantly increased in MACCE group compared with that in Non-MACCE group,the difference was statistically significant (P＜0.01).Kaplan-Meier and Cox regression analysis showed that the plasma concentration of CX3CL1 was independently associated with the occurrence of MACCE within 1 year (HR=1.124,95％CI:1.032-1.217,P=0.003).Conclusion Chemokine CX3CL1 concentration in plasma is positively correlated with the prognosis of patients with STEMI,and can be used in the prognosis assessment of STEMI patients.

Research progress of CX3CL1-CX3CR1 and the pathogenesis of sepsis / 天津医药

Sepsis has poor prognosis, and its pathogenesis is not clear. Chemokine CX3CL1 (Fractalkine, FNK) has many functions such as chemotaxis, adhesion and mediate immune injury. CX3CR1 is the only receptor of CX3CL1 and participates in the development of sepsis. Here we review the structure, biological function and possible mechanism of CX3CL1 and CX3CR1 in the pathogenesis of sepsis.

The influence of IL-18 on pancreatic stellate cells and CX3CL1 expression / 中华胰腺病杂志

Objective To evaluate the effect of IL-18 on the expression of E-cadherin,α-SMA and CX3 CL1 in pancreatic stellate cells (PSCs).Methods The human PSC line HPaSteC was routinely cultured and passaged.Five,25,50 and 100 μg/L IL-18 was used to treat PSCs for 72 h and the untreated PSCs were used as control.Treated and untreated cells were both collected,and RT-PCR and Western blot were used to detect mRNA and protein expression of E-cadherin,α-SMA and CX3CL1 respectively.Results The mRNA expressions of E-cadherin in control group and 5,25,50 and 100 μg/L IL-18 treated group were 1.03 ± 0.17,0.77 ±0.15,0.89 ± 0.12,0.54 ± 0.11 and 0.46 ± 0.06.The mRNA expression of α-SMA were 1.03 ± 0.19,0.85 ± 0.14,1.33 ± 0.22,1.60 ± 0.14 and 1.94 ± 0.09;The mRNA expression of CX3CL1 were 1.01 ±0.08,0.88 ±0.25,0.86 ±0.17,1.58 ±0.26 and 1.83 ±0.13.The mRNA expression of E-cadherin in IL-18 treated group were down-regulated,while the mRNA expression of α-SMA and CX3CL1 were up-regulated,and the differences between control and IL-18 100 μg/L treated group were statistically significant (P ＜ 0.05 or ＜ 0.01).The protein expression of E-cadherin in control group and 5,25,50 and 100 μg/L IL-18 treated group were 1.00 ±0.14,1.14 ±0.04,1.14 ±0.07,0.85 ±0.08 and 0.80 ±0.06.The protein expression of α-SMA were 1.00 ± 0.02,0.77 ± 0.07,1.29 ± 0.02,1.59 ± 0.07 and 1.70 ± 0.02;The protein expression of CX3CL1 were 1.00 ± 0.05,1.03 ± 0.05,1.37 ± 0.06,1.46 ± 0.18 and 1.45 ± 0.12.The protein expression of E-cadherin was down-regulated but no significant differences were observed among different groups.The protein expression of α-SMA was up-regulated and the differences between control group and 25,50 and 100 μg/L IL-18 treated groups were statistically significant (all P ＜0.01).The protein expression of CX3CL1 was up-regulated and the differences between control group and 100 ng/ml IL-18 treated group were statistically significant (P ＜0.05).Conclusions IL-18 can activate PSCs and up-regulate the expression of chemokine CX3CL1.

A calpain inhibitor protects against fractalkine production in lipopolysaccharide-treated endothelial cells

BACKGROUND: Fractalkine (CX3CL1) is a chemokine with a unique CX3C motif and is produced by endothelial cells stimulated with lipopolysaccharide (LPS), tumor necrosis factor (TNF)-α, interleukin (IL)-1, and interferon-γ. There have been several reports that the caspase/calpain system is activated in endotoxemia, which leads to cellular apoptosis and acute inflammatory processes. We aimed to determine the role of the caspase/calpain system in cell viability and regulation of fractalkine production in LPS-treated endothelial cells. METHODS: Human umbilical vein endothelial cells (HUVECs) were stimulated with 0.01–100 μg/mL of LPS to determine cell viability. The changes of CX3CL1 expression were compared in control, LPS (1 μg/mL)-, IL-1α (1 μg/mL)-, and IL-1β (1 μg/mL)-treated HUVECs. Cell viability and CX3CL1 production were compared with 50 μM of inhibitors of caspase-1, caspase-3, caspase-9, and calpain in LPS-treated HUVECs. RESULTS: Cell viability was significantly decreased from 1 to 100 μg/mL of LPS. Cell viability was significantly restored with inhibitors of caspase-1, caspase-3, caspase-9, and calpain in LPS-treated HUVECs. The expression of CX3CL1 was highest in IL-1β-treated HUVECs. CX3CL1 production was highly inhibited with a calpain inhibitor and significantly decreased with the individual inhibitors of caspase-1, caspase-3, and caspase-9. CONCLUSION: The caspase/calpain system is an important modulator of cell viability and CX3CL1 production in LPS-treated endothelial cells.

Protective effect of naringin on vascular endothelial injury induced by high glucose associated with reduced expression of CX3 CL1 and antioxidant / 中国药理学通报

Aim To study the protective effect of the naringin on chemotactic factor CX3 CL1 of human umbilical vein endothelial cells ( HUVEC) induced by high glucose. Methods The effect of different concen-trations of naringin on HUVEC cell viability was deter-mined by MTS. HUVECs were divided into 4 groups:① control group, ② high glucose group, ③ naringin group and④high glucose treated with naringin group. After treatment for 5 days, the concentration of nitric oxide ( NO ) in the culture medium was measured by nitrate reductase; intracellular reactive oxygen species ( ROS) was analyzed with fluorescence probe; the ex-pressions of CX3 CL1 mRNA and protein were deter-mined by the reverse transcription PCR ( RT-PCR ) and Western blot ( WB ) . Results NO content in the culture medium of high glucose group was markedly de-creased, which could be increased by naringin. Com-pared with the control group, intracellular ROS in the high glucose group was drastically elevated, but narin-gin decreased the elevated ROS induced by high glu-cose. The results of RT-PCR and WB showed that nar-ingin could downregulate the increased expressions of CX3CL1 mRNA and protein induced by high glucose. Conclusion Naringin has protective effect on the in-jury of the HUVEC induced by high glucose, which is associated with reducing the expression of CX3 CL1 and the antioxidative and anti-inflammatory action.

In vitro effects of bone marrow-derived mesenchymal stem cells on the biological behavior of lipopolysaccharide-activated retinal microglia / 中华实验眼科杂志

Background Retinal microglia (RMG) plays an important role in the pathogenesis of retinal degenerative diseases,while chemokine CX3CL1 participates in the regulation of steady-state of microglia.It has been determined that bone marrow-derived mesenchymal stem cells (BMSCs) have a remarkable role to modulate the immune response and protect the central nervous system through the release of soluble factors in a paracrine fashion and further affect the functional behavior of cells.However,whether BMSCs are able to interact with RMG and activate related signaling pathway for the maintaining of homeostasis in the retina is still unclear.Objective The aim of this study was to investigate the interaction between BMSCs and lipopolysaccharide (LPS)-activated RMG in vitro,and dissect the effects of CX3CL1/CX3CR1 signaling pathway on the biological behavior of BMSCs and RMG.Methods RMG was isolated from SD rats,cultured with mixed culture of retinal glial cells and purified by shaking.The cells were identified by detecting the expression of CD111b,Iba1 and glutamamine synthetase (GS) with indirect immunofluorescence assay.LPS (1 mg/ml,2 μl) was added in the medium for 24 hours to stimulate RMG,and then the cells were divided into LPS control group,BMSCs group (cocultured with BMSCs for 24 hours) and CB-BMSCs group (cocultured with CX3CL1-blocking-BMSCs for 24 hours).The cells without LPS stimulation served as the blank control group.The functions of RMG,including the release content of tumor necrosis factor-α (TNF-α) and interleukin-1 β (IL-1β),the proliferation,phagocytosis,and migration of RMG were examined.Results RMG was successfully isolated and harvested from SD rats by using mixed culture of retinal glial cells and purified by shaking.CD11b and Iba1 showed the positive expression with the green fluorescence in the cells and GS was absent.The contents of TNF-αt in the cell supernatant were (2.55 ±0.97) ng/ml,(24.91 ±3.07) ng/ml,(20.38 ±2.97) ng/ml and (24.90 ± 1.88) ng/ml in the blank control group,LPS control group,BMSCs group and CB-BMSCs group,respectively,showing a significant difference among the groups (F=119.90,P＜0.05).The contents of IL-1 β in the cell supernatant were (1.12±0.36) ng/ml,(10.40±2.76) ng/ml,(7.00± 1.75) ng/ml and (9.55 ± 1.11) ng/ml in the blank control group,LPS control group,BMSCs group and CB-BMSCs group,respectively,showing a significant difference among the groups(F =34.96,P＜0.05).The secretory volume of TNF-α and IL-1 β were evidently lower in the BMSCs group than those in the LPS control group (both at P＜0.05),and no significant differences were found in the secretory volume of TNF-α and IL-1β between CB-BMSCs group and LPS control group (both at P＞0.05).The proliferative rate of RMG was lower in the BMSCs group than that in the LPS control group (P＜0.05),while there was no statistical difference between BMSCs group and CB-BMSCs group (P＞0.05).The mean fluorescence intensity (MFI) and the number of migrated RMG were considerably different among the four groups (F=70.55,15.49,both at P＜0.05),and those in the BMSCs group were significantly increased in comparison with the LPS control group (both at P＜0.05),while there was no significant difference between CB-BMSCs group and LPS control group (both at P＞0.05).Conclusions BMSCs could suppress the proliferation of LPS-activated RMG.Moreover,BMSCs might inhibit proinflammatory cytokines releasing,enhance phagocytosis and migration capabilities of RMG via CX3CL1/CX3CR1 signaling pathway.

Effect of fractalkine, IP-10 and different signal pathway inhibitors on NK cells in the tumor microenvironment / 解放军医学杂志

Objective To investigate the inducing effects of chemokines [fractalkine (FKN), IP-10] and different signal pathway inhibitors on NK cells in the tumor microenvironment (TME). Methods Immunohistochemistry was performed using antibodies for CD56 and DAP10 respectively on human breast carcinoma. Murine macrophages (RAW 264.7) and breast cancer cells (4T1) were co-cultivated at a 1:4 ratio to imitate the TME with NK cells (KY-1) set as the object. RT-PCR was used to determine the mRNA expressions of CD16, NKG2D and NK1.1, and the content of CD107a in the supernatants was determined by ELISA. 10ng/ ml FKN and 10ng/ml IP-10 were added into the TME, NK1.1+CD16+KY-1 cells were counted with flow cytometry, migration and adhesion assays were used to assess the related function of KY-1 cells. 4T1 cells were incubated in 10nmol/L of rapamycin, 30μmol/ L of LY294002, 500ng/μl of andrographolide and 2mmol/L of wortmannin, the 4T1 tumor supernatants (TSNs) were harvested separately and used to incubate RAW 264.7 for 48h, then the expressions of Rae1α and H60a mRNA in 4T1, RAW 264.7 and their mixture were determined by RT-PCR. Results The related indicators of KY-1 cells such as NK1.1+ number, chemotaxis rate, and adhesion function decreased obviously in TME, and the above indices increased after the addition of FKN and IP-10, and some signal pathway inhibitors indirectly promoted NK cells' function in TME, and among them rapamycin was the most efficient one (P<0.05). Conclusion FKN and IP-10 may up-regulate the number and function of NK cells in TME, and rapamycin can promote NK cells' killing function by inducing high expression of NKG2DLs (Rae1, H60a) on tumor cells.

Effects of expression silencing of chemokine receptor CX3CR1 on human hepatocellular carcinoma Huh7 cells and its machanism / 肿瘤

Objective: To investigate the effect of chemokine CX3C receptor 1 (CX3CR1) on human hepatocellular carcinoma Huh7 cells and its probable machanism. Methods: Hepatocellular carcinoma Huh7 cells were transfected with the recombinant adenovirus Ad-siCX3CR1 targeting and silencing CX 3CR 1 gene. Then the expression of CX3CR1 was detected by reverse transcription-PCR (RT-PCR) and Western blotting. The proliferation, apoptosis, migration and invasion of Huh7 cells were tested by MTT, flow cytometry, wound-healing and Transwell assay, respectively. The expression and phosphorylation of Akt which was related to phosphoinositide 3-kinase (PI3K)-Akt signal pathway were detected by Western blotting. Meanwhile, Huh7 cells transfected with AdsiCX3CR1 were treated with different concentrations of PI3K inhibitor LY294002, then the expression of Akt and cell invasion ability induced by CX3CR1 were detected by Western blotting and Transwell assay, respectively. Results: After Ad-siCX3CR1 was stably transfected into Huh7 cells, the expression of CX3CR1 was significantly inhibited (P < 0.001), and the cell proliferation and migration and invasion abilities were significantly increased (P < 0.001), but the apoptosis rate was significantly decreased (P < 0.01). When the expression of CX3CR1 was silenced, the level of phosphorylated Akt (p-Akt) in Huh7 cells was increased (P < 0.001), and LY294002 could reverse CX3CR1-induced phosphorylation of Akt and invasion of Huh7 cells effectively (P < 0.001). Conclusion: CX 3CR 1 gene silencing can promote proliferation, migration and invasion of hepatocellular carcinoma Huh7 cells, and also can suppress their apoptosis. The activation of PI3K-Akt signaling pathway may be involved in this process.

Effects of interfering chemotactic factor Fractalkine RNA expression on biological functions of human pancreatic cancer cell lines / 中华消化杂志

Objective To explore the effects of Fractalkine (FKN) on the biological functions of human pancreatic cancer cell lines SW-1990 and PNAC-1.Methods Adenovirus mediated FKN-small interfering RNA (siRNA) was transfected into human pancreatic cancer cell lines SW 1990 and PNAC-1.The differences in proliferation and invasion ability between before and after FKN-siRNA transfection were determined by clone formation assay,MTT assay and cells invasion assay.After FKN-siRNA transfection,the expression of FKN,tumor necrosis factor (TNF)-α and interleukin (IL)-6 at protein and mRNA level in human pancreatic cancer cell were detected by Western blot and reverse transcription-polymerase chain reaction (RT-PCR).The data were analyzed by one way analysis of variance.Results After human pancreatic cancer cell lines SW-1990 and PNAC-1 transfected with FKN-siRNA,the clone numbers (5.27 ％ ± 0.35 ％ and 4.60 ％ ± 0.30％ ) increased compared with those of control group ( 1.97％ ±0.25％ and 1.77％ ± 0.25％ ) and negative FKN-siRNA group (2.10％±0.30％ and 1.97％±0.25％),and the difference was statistically significant (F=113.51,103.86; both P＜0.05).The clone size was also enlarged.After human pancreatic cancer cell lines SW-1990 and PNAC-1 transfected with FKN-siRNA for 48 hours and 72 hours,the MTT test results showed the absorbance value (48 h:1.28±0.07 and 1.19±0.14; 72 h:1.49±0.11 and 1.52±0.16) was higher than that of control group (48 h:0.80±0.03 and 0.74±0.11;72 h:0.89±0.03 and 0.93±0.04) and negative FKN-siRNA group (48 h:0.85±0.02 and 0.76±0.05; 72 h:0.89±0.02 and 1.07±0.09),and the difference was statistically significant (F=83.80,71.99,17.19,23.51; all P＜0.05).The invasion ability assay showed that the invasion ability of FKN-siRNA transfected cells was stronger than that of control group and negative FKN-siRNA group,and the difference was statistically significant (F=37.37,9.08; both P＜0.05).After FKN-siRNA transfection,the expression of FKN at protein and mRNA level in SW-1990 and PNAC-1 cell line decreased (protein:F=118.93 and 88.62,mRNA:F=47.91 and 72.59),at the same time the expression of TNF-α and IL-6 at protein and mRNA level increased (protein:FTNF-α =112.90 and 77.88,FIL-6 =165.27 and 286.49,mRNA:FTNF-α ==47.93 and 45.19,FIL-6 =36.41 and 23.67),and the differences were statistically significant (all P values＜0.05).Conclusion With siRNA technology to silent FKN function,the proliferation and invasion ability of pancreatic cancer cell lines increased,which indicated FKN might inhibit certain biological functions of pancreatic cancer cells.

The efficacy and impact of recombinant human cytotoxic T lymphocyte-associated antigen-4 fusion protein on human tumor necrosis factor-α and CX3CL1 in active rheumatoid arthritis patients / 中华风湿病学杂志

Objective To evaluate the efficacy,safety and impact of recombinant human cytotoxic T lymphocyte-associated antigen (CTLA)-4 fusion proteins (rhCTLA-4Ig) on serum human tumor necrosis factor (TNF)-α and CX3CL1 in active rheumatoid arthritis (RA) patients.Methods Forty-four RA patients were treated with rhCTLA-4Ig and placebo.Clinical response was assessed by American College of Rheumatology (ACR) criteria and disease activity score in 28 joints (DAS28).The levels of serum TNF-α and CX3CL1 were determined in 44 RA patients and 20 healthy controls by enzyme-linked immunosorbent assay (ELISA).Comparisons between groups were performed by t-test or x2 test.Results At week 12,ACR20,ACR50and ACR70 responses in RA patients with rhCTLA-4Ig were achieved by 95％(20/21 ),76％( 16/21 )and 19％(4/21) respectively,but no patient with placebo achieved ACR20,ACRS0 and ACR70 responses.There were significantly statistical differences in ACR20 and ACR50 responses (x2=39.17,26.69,P＜0.01 ).At week 12,the mean DAS28 in the rhCTLA4Ig group was 3.1±1.3 versus 6.2±1.1 at baseline (P＜0.01).Similarly,health assessment questionnaire (HAQ) improved significantly,declining from 1.4±0.5 at baseline to 0.4±0.5 at week 12 (P＜0.01).However,the mean DAS28 in the placebo group was 5.8±1.2 versus 6.0±0.7 at baseline (P＞0.05),HAQ declined from 1.6±0.4 to 1.6±0.6 (P＞0.05).In addition,there were higher levels of TNF-α and CX3CL1 in the active RA patients than those of the healthy controls (P＜0.01).After 12 weeks therapy,Serum TNF-α and CX3CL1 levels in the rhCTLA-4Ig group decreased significantly (P＜0.01).There weren't decline in the placebo group (P＞0.05).Conclusion This study has shown that rhCTLA-4Ig is very effective in reducing disease activity,improving function during the 12 weeks treatment.rhCTLA-4Ig therapy for 12 weeks can lead to significant decrease of serum TNF-α and CX3CL1.