Regulation of Baicalin on Growth of Extranodal NK/T Cell Lymphoma Cells through FOXO3/CCL22 Signaling Pathway / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of baicalin on the growth of extranodal NK/T cell lymphoma (ENKTCL) cells and its related mechanism.@*METHODS@#Normal NK cells and human ENKTCL cells lines SNK-6 and YTS were cultured, then SNK-6 and YTS cells were treated with 5, 10, 20 μmol/L baicalin and set control. Cell proliferation and apoptosis was detected by Edu method and FCM method, respectively, and expressions of BCL-2, Bax, FOXO3 and CCL22 proteins were detected by Western blot. Interference plasmids were designed and synthesized. FOXO3 siRNA interference plasmids and CCL22 pcDNA overexpression plasmids were transfected with PEI transfection reagent. Furthermore, animal models were established for validation.@*RESULTS@#In control group and 5, 10, 20 μmol/L baicalin group, the proliferation rate of SNK-6 cells was (56.17±2.96)%, (51.92±4.63)%, (36.42±1.58)%, and (14.60±2.81)%, respectively, while that of YTS cells was (58.85±2.98)%, (51.38±1.32)%, (34.75±1.09)%, and (15.45±1.10)%, respectively. In control group and 5, 10, 20 μmol/L baicalin group, the apoptosis rate of SNK-6 cells was (5.93±0.74)%, (11.78±0.34)%, (28.46±0.44)%, and (32.40±0.37)%, respectively, while that of YTS cells was (7.93±0.69)%, (16.29±1.35)%, (33.91±1.56)%, and (36.27±1.06)%, respectively. Compared with control group, the expression of BCL-2 protein both in SNK-6 and YTS cells decreased significantly (P<0.001), and the expression of Bax protein increased in SNK-6 cells only when the concentration of baicalin was 20 μmol/L (P<0.001), while that in YTS cells increased in all three concentrations(5, 10, 20 μmol/L) of baicalin (P<0.001). The expression of FOXO3 protein decreased while CCL22 protein increased in ENKTCL cell lines compared with human NK cells (P<0.001), but the expression of FOXO3 protein increased (P<0.01) and CCL22 protein decreased after baicalin treatment (P<0.001). Animal experiments showed that baicalin treatment could inhibit tumor growth. The expression of CCL22 protein in ENKTCL tissue of nude mice treated with baicalin decreased compared with control group (P<0.01), while the FOXO3 protein increased (P<0.05). In addition, FOXO3 silencing resulted in the decrease of FOXO3 protein expression and increase of CCL22 protein expression (P<0.01, P<0.001).@*CONCLUSION@#Baicalin can inhibit proliferation and promote apoptosis of ENKTCL cell lines SNK-6 and YTS, up-regulate the expression of Bax protein, down-regulate the expression of BCL-2 protein, and down-regulate the expression of CCL22 protein mediated by FOXO3. Animal experiment shown that the baicalin can inhibit tumor growth. Baicalin can inhibit the growth and induce apoptosis of ENKTCL cells through FOXO3/CCL22 signaling pathway.

Prospective study on the relationship between CCL22, a cord blood chemokine, and risk of atopic diseases / 临床儿科杂志

Objective To investigate the risk of atopic disease in infants with a atopic mothers. Methods The level of CCL22 and total IgE in the cord blood were measured using ELISA for 33 newborns with atopic mothers and for 44 newborns with non-atopic mothers. Correlation between the two factors was examined. Periodic follow-ups were conducted on the newborns to observe the risk of atopic diseases. Results The atopic group showed a higher level of CCL22 than that in non-atopic group, and the difference was statistically significant (Z=5.20, P=0.000). When 0.9 kU/L was taken as the threshold of an elevated IgE level in cord blood, the positive rates of the atopic group (11/33) was much higher than that of the non-atopic group (4/44) (χ2=7.07, P=0.008). Furthermore, the level of CCL22 and the level of IgE were significantly positively correlated (r=0.808, P=0.000; r=0.348, P=0.021) in the atopic group and the non-atopic group, respectively. During the 12 months of follow-up, the number of atopic diseases occurred in the infants in the atopic group (24/33) was much higher than that in the non-atopic group (10/44) (χ2=19.12, P<0.001).Significant correlation exists between levels CCL22 and total IgE in cord blood and infant atopic diseases (Z=5.36, P=0.000; Z=4.44, P=0.000). Conclusions At birth, the infants with an atopic mother are already in a sensitization state and have a tendency to develop potential atopic diseases. There is a correlation between the history of atopic diseases in the mothers and the elevated level of CCL22 in the cord blood of the newborns, and the probability of developing atopic diseases for the newborns is significantly higher when the level of CCL22 is elevated. The combined detection of CCL22 and IgE levels impact significantly on the prediction of the risk of atopic diseases clinically.

Effect of chemokine CCL20 and CCL22 combined with skin antigen-induced Treg on survival time of grafted skin / 中国免疫学杂志

Objective:To study the effect of chemokines CCL20 and CCL22 combined with skin-induced Treg on survival time of grafted skin.Methods: Skin grafting mice were divided into four groups, three mice per group, namely Treg group, Treg+CCL20 group,Treg+CCL22 group and control group.C57BL/6 mice were used as donor and BALB/c as acceptor, and the Treg cells were isolated from the mice induced by skin allograft.After skin grafted,CCL20 and CCL22 were subcutaneous injection every day,which lasted for 10 day.Survival time of skin in each group were observed and recorded.The Treg colonzation experiments were performed as follows.We firstly isolated Treg with Magnetic cell sorting system( MACS) and then labled them with 99 Tcm.After that we intravenously injected them into the mice.3 hours later,the mice were sacrifced and the radioactivity of organs were detected by GC-2016γradioim-munoassay counter.Results:①After Treg treated the survival time of skin grafted in antigen-induced Treg group was signifiantly longer than control group,when treg were cooperated with CCL20 and CCL22,the skin grafted showed more longer survival time than Treg and control groups( P<0.001 ).②After injection of induced Treg, Treg in autologous and allogeneic skin grafts goups were mainly distributed in autologous and allogeneic skin,accounting for 60% and 98% respectively.When cooperated with CCL20 or CCL22,the Treg were mainly distributed in liver.Conclusion:Chemokines CCL20 and CCL22 synergistically improved the effects of skin antigen induced Treg on survival time of skin graft,which probably related with the Treg colonization into the liver.

Dieckol, a Component of Ecklonia cava, Suppresses the Production of MDC/CCL22 via Down-Regulating STAT1 Pathway in Interferon-gamma Stimulated HaCaT Human Keratinocytes

Macrophage-derived chemokine, C-C motif chemokine 22 (MDC/CCL22), is one of the inflammatory chemokines that controls the movement of monocytes, monocyte-derived dendritic cells, and natural killer cells. Serum and skin MDC/CCL22 levels are elevated in atopic dermatitis, which suggests that the chemokines produced from keratinocytes are responsible for attracting inflammatory lymphocytes to the skin. A major signaling pathway in the interferon-gamma (IFN-gamma)-stimulated inflammation response involves the signal transducers and activators of transcription 1 (STAT1). In the present study, we investigated the anti-inflammatory effect of dieckol and its possible action mechanisms in the category of skin inflammation including atopic dermatitis. Dieckol inhibited MDC/CCL22 production induced by IFN-gamma (10 ng/mL) in a dose dependent manner. Dieckol (5 and 10 muM) suppressed the phosphorylation and the nuclear translocation of STAT1. These results suggest that dieckol exhibits anti-inflammatory effect via the down-regulation of STAT1 activation.

Mechanism of Macrophage-Derived Chemokine/CCL22 Production by HaCaT Keratinocytes

BACKGROUND: CC chemokine ligand 17 (CCL17) and CCL22 are the functional ligands for CCR4. We previously reported that inhibitors of nuclear factor-kappa B and p38 mitogen-activated protein kinase (p38 MAPK), but not of extracellular signal-related kinase (ERK), inhibited tumor necrosis factor (TNF)-alpha- and interferon (IFN)-gamma-induced production of CCL17 by the human keratinocyte cell line, HaCaT. Further, an inhibitor of epidermal growth factor receptor (EGFR) enhanced the CCL17 production by these keratinocytes. OBJECTIVE: To identify the mechanism underlying CCL22 production by HaCaT cells. METHODS: We investigated the signal transduction pathways by which TNF-alpha and IFN-gamma stimulate HaCaT cells to produce CCL22 by adding various inhibitors. RESULTS: TNF-alpha- and IFN-gamma-induced CCL22 production was inhibited by PD98059, PD153035, Bay 11-7085, SB202190, c-Jun N-terminal kinase (JNK) inhibitor II, and Janus kinase (JAK) inhibitor 1. CONCLUSION: Our results indicate that CCL22 production in HaCaT cells is dependent on ERK, EGFR, p38 MAPK, JNK, and JAK and is mediated by different signal pathways from those regulating CCL17 production. Altogether, our previous and present results suggest that EGFR activation represses CCL17 but enhances CCL22 production by these cells.

Anti-Inflammatory Effect of Quercetagetin, an Active Component of Immature Citrus unshiu, in HaCaT Human Keratinocytes

Citrus fruit contain various flavonoids that have multiple biological activities. However, the content of these flavonoids are changed during maturation and immature Citrus is known to contain larger amounts than mature. Chemokines are significant mediators for cell migration, while thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) are well known as the typical inflammatory chemokines in atopic dermatitis (AD), a pruritic and chronic inflammatory skin disease. We reported recently that the EtOH extract of immature Citrus unshiu inhibits TARC and MDC production. Therefore, we investigated the activity of flavonoids contained in immature Citrus on TARC and MDC levels. As a result, among the various flavonoids, quercetagetin has stronger inhibitory effects on the protein and mRNA expression of TARC and MDC than other flavonoids. Quercetagetin particularly has better activity on TARC and MDC level than quercetin. In HPLC analysis, the standard peak of quercetagetin matches the peaks of extract of immature C. unshiu. This suggests that quercetagetin is an anti-inflammatory component in immature C. unshiu.

Therapeutic effects of recombinant Salmonella typhimurium harboring CCL22 miRNA on atopic dermatitis-like skin in mice

Th-2-biased immune responses are known to play a key role in the pathogenesis of atopic dermatitis. In particular, the macrophage-derived chemokine CCL22 is directly implicated in Th-2-associated skin inflammatory reactions, and its levels are significantly elevated in serum and are correlated with disease severity in atopic dermatitis. In this study, we tested the development of genetic therapeutic options to treat atopic dermatitis using bacteria expressing miRNA. We constructed a recombinant strain of Salmonella typhimurium expressing CCL22 miRNA (ST-miRCCL22) for the in vivo knockdown of CCL22. The CCL22 gene was downregulated with CCL22 miRNA in activated lymphocytes. In mice with a cutaneous disease similar to atopic dermatitis, interleukin-4 was inhibited and interferon-gamma was induced after treatments with ST-miRCCL22. Furthermore, CCL22 levels were suppressed in the atopic mice treated with ST-miRCCL22. These results suggest that ST-miRCCL22 may be an effective genetic agent for treating atopic dermatitis.

Expression of Th1/Th2 chemokines and their receptors in the lesions of bullous pemphigoid / 中国免疫学杂志

Objective:To investigate the expression of Th1 chemokine CXCL9,CXCL10,CXCL11,Th2 chemokine CCL22 and their receptors in the lesions of bullous pemphigoid (BP).Methods:Immunohistochemical assay was performed to detect the expression of CXCL9,CXCL10,CXCL11,CCL22 and their receptors CXCR3 and CCR4 in BP lesions and normal control skin.Results:CXCL9,CXCL10,CXCL11,CCL22,CXCR3 and CCR4 were overexpressed in BP lesions than those in normal control skin (P＜0.01).The positive rates of CXCL9,CXCL10,CXCL11 and CXCR3 in BP lesions were 50%(15/30),46.7%(14/30),46.7%(14/30) and 53.3%(16/30),respectively.The positive rates of CCL22 and CCR4 were 66.7% (20/30) and 56.7% (17/30).Conclusion:The overexpression of Th1 chemokine CXCL9,CXCL10,CXCL11,Th2 chemokine CCL22 and their receptors may play important roles in the pathogenesis of BP.

Roles of the chemokine CCL22 and its receptor CCR4 in the pathogenesis of systemic lupus erythematosus / 中华皮肤科杂志

Objective To investigate the expression of cellular chemokine CCL22 and its receptor CCR4, as well as its clinical significance in systemic lupus erythematosus (SLE), along with its roles in the pathogenesis of this disease. Methods Forty-eight patients with SLE and 26 normal human controls were recruited into this study. The patient cohort included 2 males and 46 females with an average age of 33.98± 12.73 years and disease course of 1 month to 20 years. Blood samples were collected from the subjects. ELISA and flow cytometry were used to examine the plasma concentration of CCL22 together with the CCR4 expression on peripheral blood cells. SLEDAI was applied to evaluate the severity of SLE patients. Results The plasma concentration of CCL22 was 227.03±122.84 ng/L in SLE group, 369.53±79.10 ng/L in the control group, 168.09±61.83 ng/L in patients with lupus nephritis and 292.77±163.45 ng/L in patients without lupus nephritis; there was a significant difference between the SLE patients and normal con-trols (P < 0.05) as well as between patients with lupus nephritis and those without (P < 0.05). Increased percentage of CCR4-expressing cells were observed in the peripheral blood of patients with SLE compared with the controls (20.24%±13.86% vs 10.44%±3.07%, P < 0.01), and the percentage of CD3+CCR4+ cells was significantly higher than that of CD3-CCR4+ cells. Moreover, a decrease was noted in the plasma con-centration of CCL22 in severe patients (P < 0.05). In SLE patients, the percentage of CCR4 increased with the rise in SLEDAI score, whereas the plasma concentration of CCL22 negatively correlated with SLEDAI score (r = -0.308, P < 0.05). Conclusion CCL22/CCR4 may play a certain role in the development, pro-gression and organ involvement in SLE.