Résumé
Chemokine signal pathways are important for the regulation of tumour metastasis. Chemokine receptors CXCR4 (C-X-C chemokine receptor type 4) and XCR1 (chemokine XC receptor 1) are involved in the metastasis of breast cancer, while the interaction between them remains unclear. Here we first identified the interaction between CXCR4 and XCR1 based on membrane protein yeast two-hybrid assays. Bioluminescence resonance energy transfer (BRET) showed that XCR1 could competitively bind to CXCR4 to form a heterodimer (P < 0.01). Results of wound healing assays via transient transfection of XCR1 and CXCR4 into HEK293 cells showed that 41.55% of the migration area rate in the co-transformation group was lower than 58.75% in the CXCR4-alone group after adding 30 nmol/L S D F-β. The co-expression of XCR1 inhibited the cellular motility, possibly mediated by the SDF-1β (stromal cell-derived factor 1)/CXCR4 signal pathway (P < 0.05). Furthermore, CXCR4 on the cell surface after co-expression of XCR1 in CXCR4-EGFP transgenic HEK293 cells was detected by flow cytometry. And the result suggested that XCR1 could accelerate the internalization of CXCR4 into the heterodimer induced by 30 nmol/L SDF-1β (P<0.05), which increased the internalization rate from 14.38% to 64.10%. Finally, the phosphorylation of Akt and ERK, which were involved in cell proliferation and migration, respectively, were examined. After 10 minutes of SDF-1β stimulation, ERK phosphorylation in the CXCR4-alone group showed a 3.59-fold increase, whereas the increase of ERK phosphorylation in the co-transfected group was only 2.08-fold. Interestingly, heterodimer formation reduced the phosphorylation level of ERK and shortened the activation time, whereas the phosphorylation level of Akt remained unchanged. Collectively, our findings revealed the hetero-dimerization of CXCR4 and XCR1 and its effects on CXCR4-mediated cellular motility, receptor internalization, and ERK pathway phosphorylation. Therefore, XCR1-targeting drugs could be candidates for cross-desensitization of CXCR4 and might represent a possible option for inhibiting breast cancer metastasis.
Résumé
Blocking immune checkpoint programmed cell death receptor 1 (PD-1) or programmed death receptor-ligand 1 (PD-L1) can enhance anti-tumor activity of effector T cells. However, the lack of response in many patients to PD-1/PD-L1 therapy remains a question. Improving the immunosuppressive tumor microenvironment (TME) to enhance the efficacy of immune checkpoint inhibitors has become a promising cancer treatment strategy. We constructed a liposome system (PD-L1/siCXCL12-Lp) of CXCL12 siRNA and anti-PD-L1 peptide with matrix metalloproteinases (MMPs) responsiveness, which combined the TME regulation of siCXCL12 and the immune regulation of anti-PD-L1 peptide. All animal experiments were approved by the Biomedical Ethics Committee of Peking University. The authors found that PD-L1/siCXCL12-Lp directly down-regulated the expression of CXCL12 in vitro (33.8%) and in vivo (15.5%). It also effectively increased the ratio of CD8+/Treg by 20.0%, which helped the anti-PD-L1 peptide to better exert its immune effect. The combination therapy significantly inhibited tumor growth (52.08%) with great safety, which explored a new idea for cancer immunotherapy.
Résumé
Objective:To investigate the role and mechanism of chemokine receptor type 4 (CXCR4) in renal injury and fibrosis caused by calcium oxalate crystals in mice.Methods:In June 2021, Fifteen male C57/BL6 mice were divided into control group (5 mice), model group (5 mice), and AMD3100 intervention group (5 mice) by random number table method. In model group and AMD3100 intervention group, glyoxylate (100 mg/kg) was injected intraperitoneal for 7 consecutive days for modeling. Meanwhile, the AMD3100 intervention group was also given intraperitoneal injection of AMD3100 (1 mg/kg) for 7 days. The control group was continuously injected with equal volume saline intraperitoneally. After 7 days, peripheral blood was collected from each group to determine the levels of serum urea nitrogen (BUN) and creatinine (Scr) to assess the renal function; HE, Von-Kossa, Picrosirius Red staining was also taken from the left kidney to observe the pathological changes of renal tissue. CXCR4, transforming growth factor β1 (TGF-β1) were detected by immunohistochemistry and western blot. The expression levels of PI3K/AKT pathway-related proteins were detected by western blot.Results:The results of biochemical indexes showed that the serum Scr [(108.03±13.56) μmol/L vs. (39.50±4.48)μmol/L, P<0.01)] and BUN[(5.66±0.48)mmol/L vs. (0.77±0.10)mmol/L, P<0.01) levels were significantly increased in model group compared with the control group. The AMD3100 intervention group was significantly lower than the model group in terms of Scr [(65.77±3.27)μmol/L vs. (108.03±13.56)μmol/L, P<0.05) and BUN [(2.97±0.44)mmol/L vs. (5.66±0.48)mmol/L, P<0.05) levels. The results of kidney pathology in mice showed that renal tubules were significantly dilated with inflammatory cell infiltration in the model group compared with the control group, and a large number of calcium oxalate crystals and collagen fibers were deposited. The extent of kidney damage, calcium oxalate crystals and collagen fibers deposition were significantly reduced in the AMD3100 intervention group compared with the model group. The results of western blotting showed that the relative expression of CXCR4(0.639±0.019 vs. 0.158±0.012, P<0.01) and TGF-β1(0.698+ 0.018 vs. 0.314+ 0.015, P<0.05) was significantly increased in the model group compared with the control group. The relative expression of CXCR4(0.322±0.231) in the AMD3100 intervention group compared with the model group (0.322±0.231 vs. 0.639±0.019, P<0.05) and TGF-β1(0.445+ 0.017 vs. 0.698+ 0.018, P<0.05) were significantly decreased. The results of immunohistochemical staining showed the trend of CXCR4 and TGF-β1 expression in each group consistent with the results of protein blotting assay. Western blotting results showed that the expression of p-PI3K (0.613±0.016 vs. 0.213±0.011, P<0.01) and p-AKT(0.149±0.013 vs. 0.047±0.014, P<0.01) was significantly increased in the model group compared with the control group. The expression of p-PI3K in the AMD3100 intervention group compared with the model group (0.292±0.020 vs. 0.613±0.016, P<0.05) and p-AKT (0.098±0.021 vs. 0.149±0.013, P<0.05)was significantly decreased. Conclusion:CXCR4 inhibits calcium oxalate crystal-induced kidney injury and interstitial fibrosis in mice by targeting the PI3K/AKT pathway.
Résumé
Objective:To observe the pattern change of chemokine (C-X-C motif) ligand 12 (CXCL12)/C-X-C chemokine receptor type 4 (CXCR4) in systemic lupus erythematosus (SLE) by comparing the condition of SLE patients between pre-treatment and post-treatment.Methods:Peripheral blood mononuclear cells were obtained from 15 healthy controls and 8 first visit SLE patients before and after treatment. The expression of CXCR4, CD3, and CD19 on lymphocytes were detected by flow cytometry. CXCL12 concentration in serum was measured by enzyme-linked immunosorbent assay. For statistical analysis, the independent sample t test was used for independent samples of normally distributed data, the paired sample t test was used for paired samples of normally distributed data, the Mann-Whitney U test was used for independent samples of nonnormal distribution data, and the Wilcoxon test was used for comparison of related samples of non-normal distribution data. In the correlation analysis, Pearson correlation coefficient was used for data conforming to the normal distribution, and Spearman correlation coefficient was used for data not conforming to the normal distribution. Results:① The percentage of CXCR4 + lymphocyte subsets was significantly increased after treatment (90±7)% when compared with that before treatment (75±17)% ( t=-2.440, P<0.05). ② The mean fluorescence intensity of CXCR4 + lymphocyte subsets was significantly decreased after treatment [119.00(92.93, 142.50)] compared with that before treatment [177.00(130.75, 280.25)] ( Z=-2.100, P<0.05). ③ The plasma concentration of CXCL12 was significantly decreased after treatment (540±243) pg/ml when compared with that before the treatment (1 146±570) pg/ml ( t=3.219, P<0.05). ④ Moreover, there were significant correlations between the trends of the above three indicators after treatment. Conclusion:The proportion, the mean fluorescence intensity of CXCR4 + lymphocyte in peripheal blood subsets and the concentration of CXCL12 in plasma of first visit SLE patients are significantly changed after treatment. That suggests CXCL12/CXCR4 is closely associated with SLE.
Résumé
Cell membranes have recently emerged as a new source of materials for molecular delivery systems. Cell membranes have been extruded or sonicated to make nanoscale vesicles. Unlike synthetic lipid or polymeric nanoparticles, cell membrane-derived vesicles have a unique multicomponent feature, comprising lipids, proteins, and carbohydrates. Because cell membrane-derived vesicles contain the intrinsic functionalities and signaling networks of their parent cells, they can overcome various obstacles encountered
Résumé
Objective::To observe the clinical efficacy of Duhuo Xuduan Tang for oral administration and iontophoresis in the treatment of knee osteoarthritis (KOA) with liver and kidney deficiency and its effect on stromal cell-derived factor-1 (SDF-1)/C-X-C chemokine receptor type 4 (CXCR4) signaling pathway. Method::Totally 150 KOA patients with deficiency of liver and kidney diagnosed in the Teaching Hospital of Tianjin University of Traditional Chinese Medicine(TCM) were randomly divided into control group, oral TCM group and iontophoresis group, with 50 cases in each group. The control group was given glucosamine sulfate capsule, 0.5 g/time, twice a day, while the oral TCM group was given Duhuo Xuduan Tang, 150 mL/time, twice a day. In the iontophoresis group, Duhuo Xuduan Tang was administered at Kuangu acupoint, Xiguan acupoint, Xiyan acupoint and Dubi acupoint for iontophoresis for 30 minutes, once a day. All of the three groups were treated for 4 weeks. The swelling degree and the pain degree of knee joint before and after treatment were observed, and the clinical efficacy was recorded. The protein contents of SDF-1, CXCR4, matrix metalloproteinase-3 (MMP-3) and matrix metalloproteinase-13 (MMP-13) in knee joint fluid before and after treatment were detected by enzyme-linked immunosorbent assay (ELISA). Result::The efficacy of oral TCM group was better than that of iontophoresis group and control group, and the recurrence rate was the lowest (P<0.05). Compared with before treatment, the tenderness increased, whereas visual analogue scale(VAS) score, knee swelling score, The Western Ontario and McMaste Universities (WOMAC) score and SDF-1, CXCR4, MMP-3 and MMP-13 protein content in knee joint fluid decreased in oral TCM group after treatment, which were better than those in iontophoresis group and control group (P<0.05). Conclusion::Duhuo Xuduan Tang for oral administration and iontophoresis has an obvious effect on KOA with liver and kidney deficiency, with the best effect through oral administration. Its mechanism may be related to the inhibition of SDF-1/CXCR4 inflammatory signaling pathway and cartilage decomposition.