Effect of ligustrazine on chronic allograft nephropathy in rats

OBJECTIVES: Our previous study demonstrated that Ligustrazine reduced renal dysfunction associated with ischemia-reperfusion injury in mice. In this study, we investigated whether Ligustrazine herbal injection had any preventive and therapeutic effectiveness against chronic allograft nephropathy in rats. METHODS: Male Lewis rats that received left renal grafts from male Fisher 344 rats were randomly divided into five groups: group A only received cyclosporine (CsA; 10 mg.kg(-1).d(-1)) every day. Groups B, C, and D received low-dose Ligustrazine (50 mg.kg(-1).d(-1))+CsA (10 mg.kg(-1).d(-1)); high-dose Ligustrazine (100 mg.kg(-1).d(-1))+CsA (10 mg.kg(-1).d(-1)); and CsA (10 mg.kg(-1))+mycophanolate mofetil (10 mg.kg(-1).d(-1)), respectively. Group E was an isografted group (Fisher 344 to Fisher 344). Grafts were preserved in 4 degrees C University of wisconsin solution for 1 hour prolonged cold ischemia. All recipient animals were unilaterally right nephrectomized. Serum creatinine (Scr), blood urea nitrogen (BUN), urinary protein, kidney malondialdehyde (MDA) level, and superoxide dismutase (SOD) were determined, as well as examining kidney histology with immunohistochemistry for Bcl-2, and ICAM-1. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were tested to show whether Ligustrazine has side effects. RESULTS: Compared with group E, ischemia and rejection produced a ninefold increase in GSR and Banff score, as well as significant increases of Scr, BUN, and urinary protein levels in group A. High doses of Ligustrazine significantly slowed the increase (P<.01). Ligustrazine also decreased MDA levels and ameliorated the down-regulation of SOD activity. Bcl-2 was up-regulated posttransplantation, especially in the Ligustrazine-treated group (P<.01). The up-regulation of ICAM-1 was greatly diminished by Ligustrazine (P<.01). Furthermore, there was little difference in ALT/AST between the Ligustrazine-treated and the isograft group. CONCLUSION: These findings suggested that Ligustrazine could postpone chronic renal allograft dysfunction associated with cold ischemia injury and chronic allograft rejection but had no evident hepatic side effects.     

大鼠肾移植后抗波形蛋白抗体升高与慢性移植肾肾病的相关性

目的 研究大鼠肾移植术后抗波形蛋白抗体的表达水平与慢性移植肾肾病(CAN)的相关性.以及不同免疫抑制剂对其的影响.方法 选取近交系F344大鼠作为同系肾移植的供、受者(共9对),选取F344和Lewis大鼠分别作为同种肾移植的供、受者(共27对).同系移植组受者术后不给予免疫抑制剂;同种移植组受者术后10 d内给予环孢素A(CsA),然后将同种移植组受者随机平均分为生理盐水(NS)组、CsA组和霉酚酸酯(MMF)组(每组9只),分别给予NS、CsA和MMF灌胃.术后第4、8和12周时分别处死每组受者3只,观察CAN的进展、波形蛋白及其基因的表达以及抗波形蛋白抗体的水平,取正常大鼠(包括F344和Lewis大鼠)作为对照.结果 观察期内同系移植组未发生CAN;而同种移植组发生了CAN,且不断加重,其中CsA组和NS组的CAN病理改变非常明显,而MMF组明显较轻.术前所有受者血清中均未检测到抗波形蛋白IgM和IgG抗体,术后也未检测到抗波形蛋白IgM抗体;术后同系移植组仅检测到微量的抗波形蛋白IgG抗体,同种移植组检测到大量的抗波形蛋白IgG抗体.随着CAN的进展,同种移植中,CsA组和NS组血清抗波形蛋白IgG抗体的水平逐渐增高,而MMF组抗体水平的增高显著低于NS组(P<0.05),但仍显著高于同系移植组(P<0.05).结论 大鼠同种肾移植术后,受者体内可产生抗波形蛋白IgG抗体,且产生的时间早于CAN,抗波形蛋白IgG抗体水平也会随着CAN的进展而增高.MMF可抑制抗波形蛋白IgG抗体的产生,CsA无此作用.     

Campath-1H (alemtuzumab) as an induction agent for the prevention of graft rejection and preservation of renal function in kidney transplant patients: Philippine 3-year follow-up

OBJECTIVE: The purpose of this study was to evaluate the safety and efficacy of induction with Campath-1H with low-dose cyclosporine (CsA) monotherapy using outcome measures of acute rejection episodes (ARE), chronic allograft nephropathy (CAN), graft and patient survivals, as well as malignancies and infections. MATERIALS AND METHODS: Fourteen kidney transplant recipients were randomized to receive either Campath 1H induction with CsA monotherapy (9 patients) or immunosuppression with CsA, azathioprine, and steroids (5 patients). Campath (20 mg IV) was administered within 6 hours after the anastomosis and repeated 24 hours later. Cyclosporine was started 72 hours after the first Campath dose (10 mg/kg on the first day, then 4 mg/kg/d), seeking to achieve target trough CsA levels of 90 to 110 ng/mL. This is a 3-year follow-up of the 9 patients who received Campath-1H induction. RESULTS: Six of 9 (67%) patients developed ARE (borderline ARE to Banff IB) in the Campath group compared with 1 of 5 (20%) in the other group (ARE Banff IIA). They all received methylprednisolone for 3 days with good responses. One of the 6 patients in the Campath group with ARE also displayed CAN and was converted to sirolimus; 2 others had mycophenolate mofetil and steroids added to their immunosuppression after the ARE. Creatinine levels ranged from 1 to 1.7 mg/dL at 24 to 36 months posttransplantation in both groups. Among the Campath group, 1 patient died 6 months posttransplantation with sepsis secondary to infectious diarrhea. Upper respiratory tract infections comprised the majority of infections at 24 to 36 months. No malignancies were observed. CONCLUSIONS: Three years posttransplantation, patients given Campath induction with CsA monotherapy showed a greater incidence of ARE, although renal function remained comparable to CsA-azathioprine-prednisone therapy. AREs were easily reversed with steriods. Infections were minor.     

红细胞生成素对慢性肾衰竭大鼠残肾组织归巢因子表达的影响

目的 观察红细胞生成素(EPO)对慢性肾衰竭(CRF)大鼠肾组织归巢因子表达的影响.方法 采用分阶段5/6肾切除制备大鼠CRF模型.实验动物随机分为3组:假手术组、CRF模型组和EPO治疗组.从第3周开始,治疗组大鼠每次皮下注射重组人EPO 50 IU/kg,每周3次,共6周.8周后检测各组大鼠血肌酐(Scr)、血尿素氮(BUN)、尿蛋白、血红蛋白(Hb)；采用实时荧光定量PCR、Western印迹和免疫组化方法检测残肾组织EPO及其受体(EPOR)、归巢因子及其受体(SDF-1、CXCR4、Ang-1、Tie2、SCF、c-Kit)的表达.结果 与模型组比较,EPO治疗可上调残肾组织归巢因子及其受体(SDF-1、CXCR4、Ang-1、Tie2、SCF、c-Kit)mRNA和蛋白的表达(均P＜0.05)；同时,EPO治疗还可上调残肾组织EPO及EPOR的mRNA和蛋白的表达(均P＜ 0.05).此外,EPO治疗还能下调大鼠Scr、BUN和尿蛋白水平(均P＜0.05),上调Hb水平(P＜0.05).结论 EPO能改善慢性肾衰竭大鼠的肾功能,这种作用可能与其激活残肾组织归巢因子而参与损伤肾脏的修复有关.     

Combination therapy of mycophenolate mofetil and rapamycin in prevention of chronic renal allograft rejection in the rat

BACKGROUND: Chronic rejection is the leading cause of long-term allograft loss. Until now, no therapy has been recognized as being efficient in its prevention. In addition to their immunosuppressive activity, mycophenolate mofetil (MMF) and rapamycin (RAPA) show diverse properties against vascular smooth muscle cell activity, cell-adhesion molecule expression, and ischemia-reperfusion injury. The combination effect of MMF and RAPA was tested to prevent chronic renal allograft rejection in the rat in this study. METHODS: Nephrectomized Lewis recipients underwent orthotopic transplantation with Fisher (F344) kidneys (allograft groups) or Lewis kidneys (isograft control). The initial episode of acute rejection was controlled with a short course of cyclosporine A (CsA) (1.5 mg/kg/day for 10 days). From weeks 4 to 20, animals were thereafter treated every other day either with vehicle, MMF (20 mg/kg), RAPA (0.8 mg/kg), or MMF (20 mg/kg) plus RAPA (0.8 mg/kg) in combination. Animals were sequentially killed at serial intervals over a follow-up of 50 weeks, and histologic study was performed on harvested kidneys according to the Banff working classification for allograft pathology. RESULTS: Animals treated with MMF or RAPA alone showed a Banff sum score similar to the allograft control group (6.31+/-1.01 and 7.27+/-1.14 vs. 7.21+/-1.14, respectively; P>0.05). When the recipient rats were treated with MMF and RAPA in combination, it resulted in a clinically and statistically significant reduction of Banff sum score (4.21+/-0.79, P<0.01), with specific inhibition of vascular fibrous intimal thickening, allograft glomerulopathy, and interstitial fibrosis. CONCLUSION: Over a 50-week study, concomitant therapy of MMF and RAPA prevents chronic renal allograft rejection, probably through reduction of ischemic and cytotoxic degenerative changes. These results warrant further investigation in the combination of MMF and RAPA as anti-chronic rejection therapy in clinical transplantation.     

SDF-1 plays a key role in the repairing and remodeling process on rat allo-orthotopic abdominal aorta grafts

BACKGROUND: Our previous study demonstrated that prolonged cold preservation promoted neointima formation and remodeling but delayed the subsequent arteriosclerosis of rat abdominal aorta grafts. The mechanisms of this phenomenon remain obscure. In this study, we investigated whether stromal cell derived factor-1 (SDF-1) could play a role in recruiting stem cells to repair and remodel the damaged intima of abdominal aorta grafts. METHODS: Male Spague-Dawley rats received abdominal aorta grafts from male Wistar rats. Hematoxylin and eosin staining was performed to assess the structure of graft aortas by measuring the neointimal thickness. Immunohistochemical staining detected SDF-1 expression. RT-PCR demonstrated the expression of CXCR4, the only known natural receptor of SDF-1 expression on stem cells. RESULTS: The neointimal thickness of the SDF-1 antibody-treated group was inconspicuous; a significant relationship existed between the expression of SDF-1 and the neointimal thickness of the grafts. Furthermore, no CXCR4 was detected in normal abdominal aortas, but it was observed in the grafted abdominal aorta. CONCLUSION: Prolonged cold ischemia may delay the graft's arteriosclerosis by selectively chemoattracting stem cells to the damaged intima through SDF-1, the presence of which may predict graft arteriosclerosis and the subsequent development of chronic graft dysfunction (CGD). The SDF-1 antibody slowed the endothelial chimerism by blocking this chemoattration. In addition to mycophenolate mofetil and FK 506, SDF-1 antibody might be a new potential effective strategy to decrease the frequency of CGD.     

Detection of Citrate Synthase Autoantibodies in Rats with Chronic Allograft Nephropathy

Citrate synthase (CS) is the one of the key enzymes in the citric acid cycle and an important mitochondrial autoantigen. The autoimmune responses against CS have not been studied in chronic allograft nephropathy (CAN). This study investigated the role of specific CS autoantibodies in rats bearing renal allografts affected with CAN.

Methods

Fisher344 rat renal grafts were orthotopically transplanted into Lewis rats following the procedure of Kamada with our modification. Lewis-to-Lewis and Fisher344-to-Fisher344 kidney transplantations were also performed as autologous control groups (each n = 9). All the allograft recipients given cyclosporine (10 mg/kg⁻¹d⁻¹ × 10 d) were divided into four groups (each n = 9): (1) vehicle: normal saline orally; (2) cyclosporine: 6 mg/kg⁻¹d⁻¹; (3)FK506: 0.15 mg/kg⁻¹d⁻¹; (4) mycophenolate mofetil (MMF): 20 mg/ kg⁻¹d⁻¹. At 4, 8, and 12 weeks posttransplantation, the animals were sacrificed to harvest sera and renal allografts. The serum creatinine (SCr) was measured and pathological changes assessed according to Banff 97 criteria. IgM and IgG isotypes of CS antibodies were detected in all recipient sera by enzyme linked immunosorbent assays.

Results

Both IgM and IgG isotype CS autoantibodies were observed in the sera of all the recipients before and after transplantation, but the levels of IgM CS autoantibody were obviously higher than IgG isotype in all the blood samples. It was stable not only in autologous but also in allograft groups. In both autologous groups, the SCr and IgM and IgG isotype CS autoantibodies showed no obvious change before and after transplantation, and no typical CAN occurred. The values of IgG isotype of CS autoantibody (ΔOD) at 4, 8 and 12 weeks were stable. At 4 weeks, the values of SCr, Banff score, and IgG isotype CS autoantibody (ΔOD) were not significantly different (P > .05) among the allograft groups. At 8 and 12 weeks, with progression of CAN in vehicle, cyclosporine and FK506 groups' values of SCr, Banff score, and IgG (ΔOD) also increased dramatically (P = .005) in all three groups when compared with the baseline and 4 week values, but the differences among the three groups were not significant (P > .05). At 8 and 12 weeks, the MMF group suffered mild-to-moderate CAN, but the values of SCr and Banff score were significantly lower than those in the other three groups. MMF significantly inhibited the formation of IgG (ΔOD) when compared with the other three groups (P = .02).

Conclusion

This study suggested that the IgG isotype of CS autoantibody contributes to CAN after kidney transplantation. The IgM isotype is physiological. MMF significantly inhibited the formation of IgG isotype CS autoantibody, which may be related to its effects to alleviate CAN.     

The effects of different immunosuppressants on chronic allograft nephropathy by affecting the transforming growth factor-beta and Smads signal pathways

OBJECTIVE: This study investigated the effects of various immunosuppressants on chronic allograft nephropathy (CAN) by affecting transforming growth factor-beta (TGF-beta) and Smads signal pathway. METHODS: Vascular smooth muscle cells (VMSC) from rat aorta were incubated for 6 or 12 hours with various immunosuppressants. Cyclosporine (CsA) (3 microg/mL), FK506 (1 microg/mL), mycophenolate mofetil (MMF) (0.3 microg/mL), rapamycine (Rapa) (10 microg/mL), CsA (1 microg/mL/MMF 0.3 microg/mL). We used the Sprague-Dawley Wistar rat accelerated kidney sclerosis model. Before transplantation, the kidney was preserved 1 hour in 0 degrees C to 4 degrees C heparin sodium chloride solution to reinforce the cold ischemia injury. The rats were divided into eight groups (each group n = 8): group A, pseudo-OP; group B, isotransplantation; group C, CsA 6 mg/kg . d; group D, FK506 0.15 mg/kg . d; group E, MMF 20 mg/kg . d; group F, Rapa 0.8 mg/kg. d; group G, CsA 3 mg/kg . d + MMF 20 mg/kg . d. The serum creatinine levels and pathological changes, according to the Banff scheme, were observed at 2, 4, 6, 8 and 12 weeks posttransplantation. Immunohistochemistry and quantitative fluorescence polymerase chain reactions were used to end localize and quantitate the expression of TGF-beta1 and Smad 2, 3, 7 in VMSC and in the transplanted kidney. RESULTS: CsA and FK506 stimulated gene expression and protein production of TGF-beta1, smad2, and smad3, but inhibited expression of smad7 both in VSMC and in the transplanted kidney. In contrast, MMF and Rapa down-regulated gene expression and protein production of TGF-beta1, smad2, 3 while up-regulating expression of smad7. There was no significant difference between the CsA group and the FK506 group, as well as the MMF group and the Rapa group. The group treated with CsA + MMF was similar to the MMF and the Rapa groups. CONCLUSION: Our study suggested that various immunosuppressants affected differentially TGF-beta1 and Smads signal pathways in rat VSMC and kidney grafts. CsA and FK506 can cause CAN, owing to up-regulated expression of smad2 and smad3, and down-regulation of smad7 expression. MMF and Rapa can prevent the CAN progression, because of down-regulation of the expression of smad2 and smad3, with increased smad7 production.     

SDF-1 expression is elevated in chronic human renal allograft rejection

The exact mechanism of acute and chronic allograft rejection still remains unclear. The chemokine SDF-1 as mediator of allograft rejection has been under intensive investigation in liver, cardiac and bone marrow transplantation, whereas in renal transplantation, there are no reports about SDF-1 to date. This study was performed to evaluate if SDF-1 might also play an important role in human renal graft biopsies. One hundred and ninety formalin-fixed, paraffin-embedded renal allograft biopsies were included in the analysis from patients with normal renal graft morphology (according to Banff 97 classification grade 1, n = 84), with acute interstitial rejection (Banff grade 4 type I, n = 10), with acute vascular rejection (Banff grade 4 type II, n = 21), with chronic allograft nephropathy (CAN, Banff grade 5, n = 23), and without rejection but with various other lesions (Banff grade 6, n = 42). SDF-1 was localized by immunohistochemistry. In biopsies with CAN, SDF-1 expression was significantly elevated in interstitial infiltrates and infiltrating neointimal cells of arteries compared with biopsies with normal renal graft morphology. This is the first study describing a role of SDF-1 in human renal allograft rejection. We were able to demonstrate in a large number of biopsies an upregulation of SDF-1 in patients with CAN. Whether SDF-1 has pro-inflammatory or protective properties in this setting has to be evaluated in further trials.     

C4d deposition is correlated with the level of antivimentin antibody in rat kidneys undergoing chronic allograft nephropathy

Aims

Antivimentin antibody is often produced as an autoantibody after transplantation. C4d deposition, a marker of humoral immunity during transplantation, is believed to reflect alloantibodies. This study investigated the relationship between C4d deposition and humoral immunity to vimentin among rat kidneys undergoing chronic allograft nephropathy (CAN).

Methods

Fisher 344 rat renal grafts were orthotopically transplanted into Lewis rats following the procedure of Kamada with our modification. All recipients were administered cyclosporine (CsA) (10 mg/kg⁻¹ · d⁻¹ × 10 d) before being divided into 3 groups of oral treatments: (1) vehicle, (2) CsA (6 mg/kg⁻¹ · d⁻¹), and (3) mycophenolate mofetil (MMF; 20 mg/kg⁻¹ · d⁻¹). At 4, 8 and 12 weeks after transplantation, the rats were killed, the renal allografts harvested, and the sera collected. Serum creatinine (SCr) was measured and pathologic changes assessed according to the Banff 97 criteria. The antivimentin antibody was quantified by enzyme-linked immunosorbent assay. The deposition of C4d detected by immunofluorescence was analyzed by integrated optical density (IOD).

Results

Antivimentin antibody was observed in sera of all transplanted rats. The level of antivimentin antibody (IgGΔOD) increased gradually during the development of CAN from 4 weeks. Simultaneously, C4d deposition in peritubular capillaries also progressively strengthened. There was a strong positive correlation between the content of antivimentin antibody and C4d deposition (r = 0.892; P = .000). MMF simultaneously decreased antivimentin antibody formation and C4d deposition. In contrast, CsA had no significant effect.

Conclusions

We demonstrated the production of antivimentin antibodies and the deposition of C4d during the development of CAN. There was a positive correlation between them. Whether humoral immunity to vimentin contributes to C4d deposition is not clear and further studies are needed to elucidate this issue.     

Comparison of cyclosporine versus mycophenolate mofetil on expression of Fractalkine and CX3CR1 in chronic allograft nephropathy

INTRODUCTION: We sought to investigate whether there was a difference between cyclosporine (CsA) and mycophenolate mofetil (MMF) to affect the expression of Fractalkine/CX3CR1 in chronic allograft nephropathy (CAN). METHODS: The Sprague-Dawley Wistar rat accelerated kidney sclerosis model was performed as modified from the procedure of Kamada. Recipients were divided into three oral treatment groups (each group n = 8): group A was CsA 10 mg/kg . d for 10 days followed by vehicle; group B was CsA 10 mg/kg . d for 10 days followed by CsA 6 mg/kg.d; group C was CsA 10 mg/kg . d for 10 days followed by MMF 20 mg/kg . d. Pathological changes graded according to Banff 97 Standards were observed at 4, 8, and 12 weeks posttransplantation. The immunohistochemistry and quantitative real-time fluorescence polymerase chain reaction (PCR) were used to assess the distribution and expression of Fractalkine/CX3CR1 in the grafted kidney. RESULTS: Fractalkine/CX3CR1 were mostly expressed in the tubulointerstitium and tubular epithelial cell basolateral membrane. A proportion of the vessel showed positive staining for Fractalkine/CX3CR1, occasionally in glomerular parietal wall cells. The expression of Fractalkine/CX3CR1 in grafted kidneys at all the time points was significantly less in the MMF than in the CsA group or the control group (P < .05). Real-time fluorescence quantitative PCR revealed similar outcomes as immunohistochemistry. The expression of Fractalkine coincided with CX3CR1. CONCLUSION: Fractalkine/CX3CR1 may play an important role in the development of interstitial fibrosis in CAN. Different immunosuppressants have various effects on expression of the Fractalkine/CX3CR1.     

SDF-1/CXCR4在肝移植急性排斥反应中的表达

目的 通过检测基质细胞衍生因子-1(stromal-dirived factors-1,SDF-1)的受体CXCR4在移植肝脏急性排斥反应中的表达,探讨其在肝移植急性排斥反应中的作用机制.方法 根据Banff病理分级将原位肝移植术后肝穿刺活检标本分成4组,应用RT-PCR方法 半定量分析各组标本中CXCR4的mRNA水平.结果 对照组和未排斥组肝穿标本大多呈现CXCR4的轻、中度表达;排斥组肝穿标本CXCR4高度表达的例数较多,而且随着排斥程度的加重,肝穿标本CXCR4表达的相对R值呈现递增趋势.结论 CXCR4在人类肝移植急性排斥反应中起着重要作用,通过各种生物学手段抑制CXCR4在移植肝脏中的表达有可能成为防治急性排斥反应有效途径.     

Anti-apoptotic and anti-oxidant effects of grape seed proanthocyanidin extract in preventing cyclosporine A-induced nephropathy

Aim: Although the pathogenesis of cyclosporine (CsA) nephropathy is not completely understood, it is attributed to oxidative damage and apoptosis. Grape seed proanthocyanidin extract (GSPE) is a molecule with anti‐oxidant and anti‐apoptotic properties. Our aim was to demonstrate the effects of GSPE in preventing CsA nephropathy. Methods: Twenty‐four Sprague–Dawley rats were divided into four groups. The control, GSPE, CsA and CsA+GSPE groups were given 1 mL olive oil, 100 mg/kg GSPE, 25 mg/kg CsA and 100 mg/kg GSPE+25 mg/kg CsA, respectively. On day 21, blood samples were taken for blood urea nitrogen (BUN), creatinine and CsA levels, and renal tissue was used for total oxidant system (TOS), total anti‐oxidant system (TAS), oxidative stress index (OSI) and malondialdehyde (MDA) measurements. In addition to renal histopathology, apoptosis staining was performed on renal tissue. Results: The BUN, creatinine, TOS, OSI, MDA, histopathological score, and apoptotic index exhibited increases in the CsA group. In the CsA+GSPE group, however, BUN, creatinine, OSI, MDA, renal histopathological score and apoptotic index (AI) decreased and TAS levels increased. In addition, there was no difference between the CsA and CsA+GSPE groups with regard to CsA levels. Conclusion: We demonstrated that GSPE prevents CsA nephropathy and that this effect is achieved by anti‐apoptotic and anti‐oxidant activity. We also achieved a significant recovery in kidney functions without affecting CsA plasma levels.     

肾缺血-再灌注损伤大鼠SDF-1、ICAM-1表达与肾小管坏死评分的相关性研究

目的探讨肾缺血-再灌注损伤(IRI)大鼠基质细胞衍生因子(SDF)-1、细胞间黏附分子-1(ICAM-1)与肾小管坏死评分的相关性。方法将60只SD大鼠随机分成手术组、假手术组两组,每组各30只。根据手术后检测时间不同,每组再分为6个不同时段的亚组(1、6、12、24、48、72 h组),每个亚组有5只大鼠。手术组建立大鼠肾IRI模型,假手术组大鼠仅予游离双侧肾动脉后缝合切口。检测各个时间点肾功能、肾小管坏死评分及肾脏组织SDF-1、ICAM-1表达变化。对手术组大鼠肾组织SDF-1、ICAM-1表达与肾功能和肾小管坏死评分进行Pearson直线相关分析。结果手术组术后血尿素氮(BUN)、血清肌酐(Scr)较术前及相应时间段假手术亚组明显升高(均为P0.05),且于术后12 h显著升高,高峰期在术后48 h。手术组肾小管坏死评分随时间的延长逐渐增高(均为P0.05);肾小管坏死评分最高在手术48 h组(P0.05)。与手术1 h组比较,手术6 h组大鼠肾组织SDF-1、ICAM-1表达开始明显增多(均为P0.05);手术后48 h达高峰,于手术后72 h开始下降。手术组的肾组织SDF-1、ICAM-1表达与术后各时间段BUN、Scr、肾小管坏死评分呈正相关(r=0.614、0.662、0.751;0.640、0.703、0.785;均为P0.05)。结论当大鼠肾组织发生IRI时,SDF-1、ICAM-1表达上调,BUN、Scr升高,肾小管坏死评分升高,而且SDF-1、ICAM-1的表达与BUN、Scr、肾小管坏死评分呈正相关,提示SDF-1、ICAM-1表达增高程度可以作为反映肾IRI后严重程度的指标。     

Chronic allograft nephropathy in athymic nude rats after adoptive transfer of primed T lymphocytes

The impact of presensitized T lymphocytes on the development of chronic allograft nephropathy (CAN) was investigated in nude athymic LEW.RNU recipients of F344 renal allografts. The recipients (n = 8) were reconstituted with 5 x 10(7) T lymphocytes primed against donor skin grafts to induce graft rejection. LEW.RNU (n = 8) and euthymic LEW recipients (n = 6) which underwent no further intervention after transplantation served as control groups. Adoptive transfer of primed T cells induced CAN in LEW.RNU rats. Their kidney function decreased progressively. After 90 days a moderate glomerulopathy, tubular atrophy and interstitial fibrosis were observed, vascular changes were only mild or absent. Cellular infiltrates were predominated by CD4+ T cells and ED1+ macrophages. Deposition of tenascin and laminin was enhanced. Grafts of euthymic recipients displayed only mild signs of CAN according to the Banff criteria. These data implicate an important role for the cellular immune response in the development of CAN.     

Combined FTY720/cyclosporine A treatment promotes graft survival and lowers the peripheral lymphocyte count in DA to lewis heart and skin transplantation models

OBJECTIVE: The immunomodulator, FTY720, lowers the peripheral lymphocyte count (PLC) by inducing migration of circulating lymphocytes to secondary lymphoid organs. We investigated the efficacy of mono- vs. combined-FTY720/CsA therapy on graft survival (GS) and on lowering the PLC in a solid organ and a skin graft model, using strains with strong MHC disparity. Methods: Heterotopic cardiac or tail skin grafting was performed using the DA (RT1a) to Lewis (RT1(1)) rat strain combination. FTY720 was administered as a single daily dose by gavage alone or in combination with subcutaneously delivered CsA. PLC, body weight and drug concentrations were determined on day 7, 28, or the day of rejection. MAIN FINDINGS: In placebo-treated animals the heart and skin allografts rejected after 6 and 8 days. FTY720 delayed rejection of both the solid organ and skin grafts. The maximal effect was achieved at 1 mg x kg(-l) x day(-1) FTY720, resulting in a median survival time (MST) of 14 days for both allotransplants comparable to the effect achieved by 1 mg x kg x day(-1) CsA in both models. In the cardiac graft experiment with CsA co-administration, doses of 0.3 and 1 mg/kg were used. Under these conditions very small doses of FTY720 were effective in maintaining grafts throughout the treatment period. Adding higher FTY720 doses to the 1 mg x kg(-1) x day(-1) CsA was needed to effectively extend the skin GS, e.g. 0.3 mg x kg(-l) x day(-1) FTY720 prolonged GS from 13 to 47.5 days MST, i.e. well beyond the 28 day-treatment period. CsA did not influence the PLC at clinically relevant doses. FTY720 lowered the PLC significantly and dose-dependently, at doses lower than those needed for the prolongation of both cardiac and skin GS with FTY720 monotherapy. In rats with skin grafts the PLC was markedly lowered up to 1 mg x kg(-1) x day(-1) FTY720, whereas, in the heart model, it was lowered up to 0.1 mg x kg(-1) x day(-1). Independently of the graft type, within the combination regimens 0.3 mg x kg(-1) x day(-1) FTY720 achieved a maximal PLC depletion. CONCLUSIONS: Combining FTY720 and CsA was very well tolerated with respect to weight gain and lack of any clinically detectable infections. In the strain combination used FTY720 monotherapy was less effective than previously reported in maintaining grafts. The two-drug regimens extended strikingly the GS for both models. However, the prolongation of the heart GS was smoothly dose-related with FTY720 doses ranging from 0.01 to 1 mg x kg(-1) x day(-1) , whereas, the skin graft prolongation was modest at doses up to 0.1 mg x kg(-1) x day(-1) and remarkably enhanced at 0.3 and 1 mg x kg(-1) x day(-1) FTY720.     

Upregulation of connective tissue growth factor in a rat model of chronic allograft nephropathy

AIM: To study the expression of connective tissue growth factor (CTGF) in transplanted rat kidney and its relationship with chronic allograft nephropathy (CAN). METHODS: Kidney transplantation was performed from Lewis to Fisher 344 allogeneic rat strain, and kidney grafts were harvested at the eighth, 12th and 16th week. The morphological changes were studied, and collagen deposition was determined by Masson trichrome stain. Serum creatinine was examined. The fibrotic process and the CAN grades were evaluated according to Banff 97 schema. The expressions of transforming growth factor beta, CTGF and alpha-smooth muscle actin were detected to assess the development of grafted kidney fibrosis and to discuss their relationships. Spearman correlation was used for correlation study between CTGF expression and development of CAN. RESULTS: Serum creatinine was promoted in a time-dependent manner. Morphological changes suggested that the grafted kidneys were under abnormalities. At the end stage, focal segmental glomerulosclerosis was seen; tubular epithelial cells lost their phenotype and interstitial fibrosis was notable. Masson trichrome stain showed significant collagen accumulation in a time-dependent manner. Immunohistochemistry and western blotting results showed that the transforming growth factor beta, CTGF and alpha-smooth muscle actin expression were markedly promoted compared with the control group. CTGF was mainly expressed in the plasm of proximal tubular epithelial cells based on the severity of CAN. CONCLUSION: Connective tissue growth factor might play an important role in the pathological changes of CAN after kidney transplantation. The expression of CTGF in epithelial cells could act as a molecular marker of interstitial fibrosis and CAN.