958 20 2 CJITWN November 20 Vol. 2 No. * MCP - - MCP - AIN SD 4 + A - + B ADR 6. 5 mg /kg 2 24 h > 00 mg A B + - + 2 8 24 h 4 8 MCP - 2 24 h P < 0. 05 6. 5 mg /kg MCNS 2 RT - PCR MCP - MCP - MCP - A B MCP - MCP - mrna MCP - 2 3 MCP - - Experimental Study on Effects of Addition and Subtraction of Fangjihuangqi Decoction on the Expressions of MCP - mrna and Protein in Renal Cortext of Adriamycin - induced Nephropathic Rats CHEN Hongyu JIE Leqin WANG Yongjun et al Department of Nephrolog Hangzhou Hospital of Traditional Chinese Medicine Hangzhou 30007 ABSTRACT Objective To investigate mechanism of action of MCP - in minimal lesion nephrotic syndrome and effects of addition an subtraction of Fangjihuangqi decoction on the expressions of MCP - mrna and protein in renal cortext of adriamycin - induced nephropathic rats. Methods The rats were randomly divided into four groups normal control group nephropathy control group Fangjihuangqi decoction adding Leigongteng group Fangjihuangqi decoction adding Leigongteng and substracting Gancao group. ADR 6. 5 mg /kg was intravenously administered to the nephropathy group while saline was intravenously administered to the normal control group. Two weeks after the administration the nephropathic model of 24 - hour protein urine quantity > 00 mg was formed. Treatment group were given intragastric administration with Fangjihuangqi decoction adding Leigongteng and Fangjihuangqi decoction adding Leigongteng and substracting Gancao respectively while normal control group and nephropathy control group were given intragastric administration with equivalent physiological saline. The quantitation of 24 - hour proteinuria was tested on first second and eighth week respectively. Blood and kidneys were harvested at the end of experimention. Histology of kidney was tested by HE staining. The expression levels of MCP - were tested by immunohistology and RT - PCR. Results Two weeks after adriamycin - induced administration the quantitation of 24 - hour proteinuria in the nephropathy control group increased significantly in comparison with those in the normal control group P < 0. 05. This illuminated that MCNS model can be successfully prepared by Adriamycin single intravenous injection of 6. 5 mg /kg. 2 The expression levels of MCP - were tested by immunohistology and RT - PCR. The expression levels of MCP - in the nephropathy control group increased in comparison with those in the normal control group. In comparison with the nephropathy control group the expressions of MCP - decreased significantly in two traditional Chinese medicine - treated groups. There was no statistics signification in the expression of MCP - in Fangjihuangqi decoction adding Leigongteng group and in the Fangjihuangqi decoction adding Leigongteng and substracting Gancao. Conclusion There was adnormity in expressions of MCP - mrna and protein in Adriamycin - induced nephropathic rats. The increase of MCP - expression is likely to be one of import facts in the production of proteinuria in Adriamycin - induced nephropathic rats. 2 The healing effects were * No. 2009ZDJB02 No. 2007B0062 30007
20 2 CJITWN November. 2 No. 959 not markedly changed in two treatment groups in this experiment. 3. It is possible that addition an subtraction of Fangjihuangqi decoction have effects on chronic nephropathy by decreasing expressions of MCP - mrna reducing protein and lessening pathological lesion. KEY WORDS Addition and subtraction of fangjihuangqi Adriamycin - induced nephropathy Monocyte chemotactic factor - experimental studyurine protein the injury of renal pathology and the expression of MCP 3 3. 24 h 7 4 56 24 h 3. 2 28 56 6 BUN Scr Alb ALT 3. 3 RT - PCR 0% HE - MCP - 3. 4 MCP - MCP - 3 μm 60 PBS 5 min 3. SD 48. 2 20 ± 20 g DAB 080206 5 MCP - Image - Pro - Plus 2 2. SD 6. 5 mg /kg 7 3% H 2 O 2 0 min 37 20 min 50 μl PDGF - BB 20 4 PBS 5 min 3 37 30 min PBS 5 min 3 3. 5 RT - PCR MCP - - 80 2 mm 2 mm 2 mm Trizol Promega RNA 4 24 h 24 h RNA M - MLV Pro- > 00 mg mega cdna PCR BIO - 2. 2 2 RAD MCP - 36 A 5' - CAGGTCTCTGTCACGCT- B 2 A TCT - 3' 5' - CTAGTATTCATGGAAGGGAATAG - 3' B 527 bp 60 32 PCR. 7% BIO - RAD 2 ml /d Quantity - one 8 A = K W 2 /3 2 4 SPSS 3. 0 0. g 0. 2 g x 珋 ± s t 0. g
960 20 2 2. 24 h 7 4 24 h 3 ~ 4 d > 00 mg P < 0. 05 28 56 24 h 7 4 28 56 x 珋 ± s mg /24 h 7 d 2 4 d 2 28 d 2 56 d 6 3. 82 ± 5. 36 28. 48 ± 5. 36 37. 80 ± 6. 34 43. 64 ± 6. 34 86. 6 ± 5. 80 592. 27 ± 73. 52 789. 94 ± 8. 26 96. 4 ± 29. 0 A 8. 72 ± 8. 46 548. 80 ± 6. 5 665. 85 ± 73. 45 72. 03 ± 57. 74 B 68. ± 8. 58 550. 9 ± 54. 66 633. 09 ± 63. 7 80. 84 ± 98. 67 P < 0. 05 2. 2 4 8 BUN Alb 4 Scr BUN Alb Alt 4 P < 0. 05 ALT Scr 8 4 8 P < 0. 05 3 2 4 8 Scr BUN Alb Alt x 珋 ± s n Scr μmol /L BUN mmol /L Alb g /L AIT Iu /L 4 29. 5 ±. 32 5. 22 ± 0. 88 35. 48 ± 2. 07 58. 83 ± 4. 49 6 8 58. 4 ± 8. 5 5. 72 ± 0. 59 34. 0 ±. 3 53. 57 ± 4. 99 4 36. 33 ± 5. 24 9. 24 ±. 79 27 ± 3. 2 56. 20 ± 5. 06 6 8 02. 33 ± 8. 29 0. 3 ±. 2 26. 45 ± 2. 57 ± 2. 60 A 6 B 6 4 33. 83 ± 4. 8 9. 35 ± 2. 07 25. 43 ± 3. 44 0. 53 ± 0. 67 8 89. 83 ± 5. 65 0. 53 ± 0. 67 26. 08 ±. 63 60. 83 ± 9. 83 4 36. 67 ± 3. 84 9. 33 ±. 42 26. 85 ± 3. 34 52. 50 ± 4. 70 8 90 ± 0. 07 0. 5 ± 0. 69 27. 39 ± 2. 20 58. 25 ± 6. 56 P < 0. 05 3 HE 4 4 MCP - MCP - A B P < 0. 05 A B 8 PI / Image - Pro - Plus 3 4 5 RT - PCR MCP - mrna 2 P < 0. 05 3 4 8 MCP - x 珋 ± s n 4 8 6 4. 89 ± 2. 94 * 5. 08 ± 2. 74 * 6 3 568. 95 ± 93. 65 3 549. 9 ± 334. 4 A 6 33. 9 ± 8. 44 * 00. 64 ± 60. 32 * B 6 577. 24 ± 57. 55 * 247. 7 ± 60. 98 * P < 0. 05 * P < 0. 2 CJITWN November 20 Vol. 2 No. A B P < 0. 05 quanity one GAP- DH 4 5 6
20 2 CJITWN November. 2 No. 96 4 图 5 图 6 GAPDH MCP- 407%bp 527 bp RT-PCR 法检测实验 4 周末时 MCP-% mrna 在各组肾组织中的表达 GAPDH MCP- 407%bp 527 bp RT-PCR 法检测实验 8 周末时 MCP-% mrna 在各组肾组织中的表达 RT - PCR 4 8 MCP - mrna x 珋 ± s n 4 8 6. 00 ±. 74 * 3. 05 ± 0. 27 * 6 30. 33 ±. 52 34. 5 ±. 20 A 6 2. 87 ±. 35 * 28. 57 ± 0. * B 6 2. 96 ± 0. 6 * 28. 46 ± 0. 87 * P < 0. 05 * P < 0. MC- NS Wistar SD ADR 5 ~ 7. 5 mg /kg 4 ~ 5 d 2 6 ~ 9 3 h ~ 56 d 2 9 3 5 000 mg /d 5 d > 0. 75 mg /kg 2 6 SD 220 ± 20 g ADR 6. 5 mg /kg 3 7 B2 TXB2 ADR 7 d 4 4 d 24 h 8 > 00 mg P < 0. 05 UUO - MCP - 76 CCR2 CCR9 3 MCP - - IL - MCP - 3 MCP - 5 MCP - T 4 2 MCP - 4 3 MCP - / IL - - α TNF - α - β TGF - β MCP - MCP - α α - SMA FN 7 α MCP - G G α β γ C C 4 5 9 4 5 4 5
962 20 0 MCP - MCP - P < 0. 05 MCP - + + P < 0. 05 MCP - MCP - Scr BUN Alb AIT ~ 4 - A B MCP -... P > 0. 05 992 3 5 48-50. 2... 2. MCP - 200. 75. 3. Morri T Fujitu H Narita T et al. Association of monocyte chemoattractant protein - with renal tubular damage in diabetic ne- T phropathy. J Diabetics Complication 2003 7-5. 4. Viedt C Dechend R Fei J et al. MCP - induces inflammatory activation of human tubular epithelial cells involvement of the transcription factors nuclear factor - kappab and activating protein -. J Am Soc Nephrol 2002 3 6 534-547. 5... 2007 9 4 63-64. 6.. AIT. 2006 8 2 4-5. 7... 2003 25 5 296. MCP - 8.. 28 56. 2008 26 5 000-002. 9. 24 h.. 20 26 33-35. 0... 2009 6 46-49. 20-08 - 7 20-09 - 8 RT - PCR 檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱檱 2 CJITWN November 20 Vol. 2 No. sx7965258@ 26. com 2 4. 5 A4 3 20 4