[Effect of TRPM8 on pain symptom and neuronal proliferation in mice with interstitial cystitis/bladder pain syndrome].

Objective: To explore the effects between transient receptor potential melastatin 8 (TRPM8) on pain symptom and neuronal proliferation in mice with interstitial cystitis/bladder pain syndrome (IC/BPS). Methods: Female wild-type C57BL/6 mice (8-10 weeks old) were divided into control group, IC/BPS model group, and IC/BPS model+menthol group (6 mice each) by random number table method; TRPM8 knockout mice were randomly divided into TRPM8 knockout group and TRPM8 knockout model group (6 mice each). The IC/BPS model group, the IC/BPS model+menthol group, and the TRPM8 knockout model group were injected subcutaneously with residues 65-84 of murine uroplakin 3A (UPK3A65-84). The IC/BPS model+menthol group continued to be injected with menthol. After successful modeling, the differences in pain thresholds between the groups were assessed by mechanosensitivity. The bladder wall was injected with a cell membrane red fluorescent probe (Dil), and the dorsal root ganglion (DRG) tissues were collected 10 days later. The differences in the protein and mRNA levels of TRPM8 and GAP43 in the groups were detected by Western blotting and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. Immunofluorescence was used to detect the distribution of TRPM8 expression with GAP43 or Dil in DRG tissues. The relationship between TRPM8 and pain symptom and its role in neuronal proliferation in IC/BPS mice were analyzed. Results: The models were all constructed successfully. Compared with the control group, the pain thresholds of mice in the IC/BPS model group and IC/BPS model+menthol group were reduced [(8.50±1.22), (5.50±1.05) vs (11.67±2.16), respectively, all P<0.001]. Compared with the control group, the expression of TRPM8 mRNA was elevated in the IC/BPS model group and IC/BPS model+menthol group, while TRPM8 was not expressed in the TRPM8 knockout group [(3.16±0.05), (6.46±0.21), and 0 vs (1.00±0.06), respectively, all P<0.001]. The expression of TRPM8 protein and mRNA in each group had the same trend (P<0.001). Compared with the control group, the expression of GAP43 mRNA in the DRG of the IC/BPS model group and the IC/BPS model+menthol group was increased, whereas the expression of GAP43 mRNA in the TRPM8 knockout model group was decreased (all P<0.001). The trend of GAP43 protein expression was the same as that of mRNA expression (P<0.001). Immunofluorescence results showed an increase in the number of GAP43-positive neurons in the DRG of the IC/BPS model group and the IC/BPS model+menthol group, and a decrease in the TRPM8 knockout group compared with the control group (all P<0.001). Compared with the control group, the number of Dil-positive neurons in the DRG of the IC/BPS model group and the IC/BPS model+menthol group was increased, while the TRPM8 knockout group was decreased (all P<0.001). Conclusion: TRPM8 can exacerbate pain symptom in IC/BPS mice, and the mechanism may be related to the induction of sensory nerve proliferation at the DRG level.

[The roles of IL-17 and CXCR4 in chronic allograft nephropathy in rat models].

Objective: To investigate the role and mechanism of IL-17 and CXCR4 in chronic graft nephropathy (CAN) in rat models. Methods: CAN rat models were established using Fisher 344 to Lewis rats. In the control group(n=10), Lewis rats were performed isotransplantation. CAN rat models were established in experimental group(n=10). All the rats were havested 6 weeks after transplantation. Kidneys were examined by pathology to evaluate the injury of the renal allograft. SDF-1/CXCR4, IL-17 and α-SMA expression level in serum and renal graft were detected by immunohistochemical staining and Enzyme-linked immunosorbent assays(ELISA). The percent and the absolute amount of CD4(+) cells、CXCR4(+) cells and IL-17(+) cells were detected by flow cytometry. Results: The graft of the experiment group showed obvious pathological features of CAN. The protein expression levels of SDF-1/CXCR4 and IL-17 in the experiment group are significantly higher than in the control group. We could also observe the number of Th17 cells has a significant increase in the experiment group[blood (0.19±0.05)% vs (0.12±0.03)%; kidney (0.52±0.21)% vs (0.17±0.12)%]. The IL-17 level also showed the same differences between the experiment[blood (243.12±17.63) pg/ml, kidney (521.54±21.76) pg/ml]and the control group[blood (35.78±7.3) pg/ml, kidney (77.34±11.1) pg/ml]. Conclusions: The Th17 cells increase in the CAN rats model, maybe the SDF-1/CXCR4 has a chemotaxis to collect Th17 cells to the injured kidney. And its expression of IL-17 may promote the renal cells to transform into fibroblasts.