Differential expression of interleukin-15, a pro-inflammatory cytokine and T-cell growth factor, and its receptor in human prostate.

BACKGROUND: Pro-inflammatory interleukin (IL)-15 plays a major role in host defense and chronic inflammation by stimulating T-lymphocyte recruitment and growth. Expression of IL-15 and IL-15 receptor (IL-15R) in human prostate was examined. METHODS: Normal and benign hyperplastic (BPH) prostate specimens (n = 23) were analyzed for IL-15 and IL-15Ralpha-chain expression by immunohistochemistry and Real-Time-PCR/RT-PCR. Regulation of prostatic stromal cell (PSC) IL-15 mRNA and effect of IL-15 on prostatic cell growth were analysed in vitro. RESULTS: In normal prostate, anti-IL-15 and anti-IL-15Ralpha-chain reactivity were restricted to smooth muscle and stromal cells. However, in BPH, in addition epithelial cells frequently exhibited discrete anti-IL-15R and often intense, membranous anti-IL-15 reactivity. IL-15/IL-15R mRNA were detected in all prostatic cells types. In BPH tissues, IL-15 mRNA content was variable (15-fold). IL-15 mRNA synthesis of PSC was significantly up-regulated by IFN-gamma. Furthermore IL-15 strongly stimulated the growth of BPH-T-lymphocytes and weakly that of carcinoma cell lines, but not of stromal cells. CONCLUSIONS: Overexpression of IL-15 and IL-15Ralpha-chain in BPH and massive proliferation of BPH-T-lymphocytes induced by IL-15 suggest a role for IL-15 in prostatic inflammation. Since IFN-gamma, a T-lymphocyte product, stimulates prostatic IL-15 production; chronic inflammation might be triggered by this paracrine loop.     

亚致死性过氧化氢损伤致肾小管上皮细胞整合素受体分布的改变

目的探讨急性肾功能衰竭（ＡＲＦ）时活性肾小管上皮细胞脱落现象及管型阻塞的机制。方法用猪肾小管上皮细胞株（ＬＬＣＰＫ１）观察亚致死性Ｈ２Ｏ２损伤致细胞与基质粘附性及整合素受体α３分布的改变。结果细胞与基质间粘附力显著降低（Ｐ＜００５），细胞表面整合素受体α３由原先基侧部分布变为半随机分布（Ｐ＜００５）。结论细胞受亚致死性Ｈ２Ｏ２损伤后，可使整合素受体α３分布改变，细胞基质粘附力降低，致使活性细胞脱落和管腔阻塞。     

Induction of interleukin-1 and interleukin-1 receptor antagonist during contaminated in-vitro dialysis with whole blood

BACKGROUND: Previous studies on the permeability of cellulosic and syntheticdialysers for bacterial-derived cytokine-inducing substancesgave conflicting results. We tried to study this issue as closeto the in-vivo situation as possible. METHODS: An in-vitro dialysis circuit with whole human blood presentin the blood compartment of cuprophane (Cup), polysulphone (PS),and polyamide (PA) dialysers was employed; sterile filtratesderived from Pseudomonas aeruginosa cultures were added to thedialysate. We studied the induction of interleukin-1ß(IL-1ß) by plasma samples taken from the blood compartmentas well as the induction of IL-1ß and interleukin-1receptor antagonist (IL-1Ra) in mononuclear cells separatedfrom whole blood after circulation by radioimmunoassay and polymerasechain reaction. RESULTS: Plasma samples from the blood side of all dialysers inducedIL-1ß from non-circulated mononuclear cells afteraddition of pseudomonas filtrates to the dialysate; the maximalamount of IL-1ß induced by samples from the bloodcompartment was 4.8±1.2 ng/ml for Cup, 1.9±0.5ng/ml for PS, and 2.0±0.6 ng/ml for PA. Mononuclear cellsseparated after contaminated dialysis with all types of dialysersexpressed increased mRNA levels for IL-1ß and IL-1Ra.Production of IL-1Ra by cells separated after contaminated dialysiswas determined after Cup and PS dialysis; there was increasedproduction of IL-1Ra by these cells (Cup, 10.3±4.2; PS,7.3±2.5 ng/ml) compared to cells separated after steriledialysis (Cup, 5.6±2.1, P<0.05; PS, 4.5±1.1ng/ml, n.s.) or from non-circulated blood (Cup experiments,4.7±1.5, P<0.05; PS experiments, 4.1±1.2 ng/ml,n.s.). CONCLUSIONS: These data suggest penetration of cytokine-inducing substancesthrough both cellulosic and synthetic dialysers. Differencesbetween dialysers may exist regarding extent and time courseof penetration. The detection of cytokine mRNA as well as themeasurement of IL-1Ra synthesis is more sensitive substancesthrough dialyser membranes than the measurement of IL-1ßprotein synthesis.     

Administration of interleukin-1 receptor antagonist ameliorates renal ischemia-reperfusion injury.

Interleukin (IL)-1 is a major contributor to inflammation and apoptosis during ischemia/reperfusion (I/R) injury. Its deleterious effects are primarily mediated by the activation of nuclear factor-kappaB (NF-kappaB). Receptor-binding and signaling of IL-1 can be blocked by the IL-1 receptor antagonist (IL-1ra). The aim of our study was to characterize effects and mechanisms of IL-1ra administration on inflammation, apoptosis, and infiltration in renal I/R injury. Renal ischemia was induced in Lewis rats by clamping of the left renal artery for 45 min. Kidneys were removed for histological and molecular analysis 24 h or 5 days after reperfusion. IL-1ra ameliorated I/R induced renal injury and inflammation. Furthermore, the number of apoptotic tubular cells was lower in IL-1ra-treated animals 24 h after ischemia, which was paralleled by a Bax/Bcl-2 mRNA ratio towards anti-apoptotic effects. IL-1ra reduced the expression of monocyte chemoattractant protein-1 (MCP-1) mRNA at 24 h and 5 days and that of intracellular adhesion molecule-1 (ICAM-1) expression at 24 h in the ischemic reperfused kidneys. Our results indicate that IL-1ra treatment ameliorates renal I/R injury and this protective effect might be mediated by reduced induction of NF-kappaB mediated MCP-1, ICAM-1, and a decreased ratio between Bax and Bcl-2 mRNA expression.     

Levels of serum-soluble receptor for interleukin-2 in patients with colorectal cancer

P < 0.05). Markedly elevated levels of serum-soluble IL-2R were recognized in patients with stage IV cancer, those with Dukes' stage D cancer, and those with liver metastasis. Moreover, the prognosis of patients with low levels of IL-2R (<531 U/ml) was significantly better than that of those with high levels (P < 0.05). These findings demonstrate that an elevated concentration of soluble IL-2R might be a useful indicator of liver metastasis and poor prognosis in patients with colorectal carcinoma. (Received for publication on June 17, 1997; accepted on Jan. 6, 1998)     

病理性因素对体外肾小管上皮细胞表型转化的影响

目的 通过低血清，高糖，高白蛋白的刺激，观察肾小管上皮细胞的表型变化。方法 以体外培养的人近端肾小管上皮细胞系HKC为对象，分别用低血清（2%小牛血清），高糖（25mmol/L)，高白蛋白（1.5g/L)刺激，用透射电镜检测细胞形态变化；用免疫组织化学（组化）检测角蛋白（CK），波形蛋白（vimentin),α-平滑肌肌动蛋白（SMA），Ⅰ型胶原和转化生长因子（TGF）-β1的蛋白表达。Western blot进一步检测I型胶原蛋白表达的动态变化；细胞原位杂交检测I型胶原基因表达。结果 在低血清，高糖，高白蛋白直接作用下，肾小管上皮细胞形态变化明显，包括呈梭形改变，透射电镜下细胞表面微绒毛及细胞内线粒体明显减少，粗面内质网明显增加，免疫组化染色显示CK减弱，vimentin,α-SMA，Ⅰ型胶原和TGF-β1增强，Western blot显示I型胶原表达增加，且与损伤程度正相关。原位杂交显示I型胶原基因表达增加。结论 在低血清，高糖，高白蛋白直接作用下，体外培养的正常人近端肾小管上皮细胞系HKC发生了上皮向间叶的表型转化。     

FSGS患者尿蛋白对肾小管上皮细胞活力及转分化的影响

目的：探讨局灶性-节段性肾小球硬化症（focal segmental glomerulosclerosis,FSGS）患者尿蛋白对肾小管上皮细胞（TECs）活力及转分化的影响。方法：用硫酸铵沉淀法提取FSGS患者尿液中的总蛋白成分,应用不同浓度尿蛋白处理体外培养人近端肾小管上皮细胞,然后应用全自动生化分析仪检测不同浓度尿蛋白刺激后HK-2细胞LDH值并计算其释放率、免疫细胞化学（ICC）法检测不同浓度尿蛋白刺激后HK-2细胞E-钙黏蛋白（e—cadherin）的表达水平和Western blotting法检测HK-2细胞α-平滑肌肌动蛋白（Q—SMA）蛋白表达水平。结果：所提取的尿蛋白主要成分为白蛋白、转铁蛋白和IgG等,分别占59．3％、15．3％和13．8％;肾小管上皮细胞LDH释放率和α—SMA蛋白的表达水平随尿蛋白浓度的升高而升高;E—cadherin的表达水平随着尿蛋白浓度的升高而下降。结论：FSGS尿蛋白可通过诱导肾小管上皮细胞损伤和转分化而促进肾小管-间质纤维化。     

移植肾排异反应中肾小管上皮细胞的表型转化

目的 观察人移植肾肾穿刺标本不同排异反应病变中肾小管上皮细胞表型转化状态,探讨排异反应与肾小管上皮细胞表型转化的相关性。 方法 免疫组织化学SP法检测55例移植肾穿刺不同病变组中肾小管上皮细胞α平滑肌肌动蛋白（α-SMA）的表达。 结果 各组萎缩病变的肾小管上皮细胞均有α-SMA阳性表达,表现为近基底膜处胞质阳性染色,提示出现了表型转化。在无萎缩病变的肾小管中,仍有部分肾小管细胞呈α-SMA阳性染色。7例基本正常病例组均无肾小管上皮细胞的表型转化。28例急性T细胞介导排异 IA级病例组中,1例肾小管上皮细胞α-SMA阳性表达率为25%~50%,3例为10%~25%。14例排异反应IB 级中,1例α-SMA阳性表达率达50%以上,2例25%~50%,2例10%~25%。 结论 随着急性T细胞介导性排异反应加重,肾小管上皮细胞发生表型转化的现象明显增强。     

血清IL-2/SIL-2R系统平衡紊乱在梗阻性黄疸围手术期中作用的临床研究

目的 探讨梗阻性黄疸（梗黄）病人围手术期血清白细胞介素-2（IL-2）活性和可溶性白细胞介素-2受体（SIL-2R）表达状况及IL-2／SIL-2R系统平衡紊乱的临床意义。方法39例梗黄病人（良性21例，恶性18例），分别采用放免计数器和双抗体夹心ELISA法测定血清IL-2和SIL2R水平。结果（1）良恶性两组术前血清IL-2较对照组明显降低，SIL-2R明显高于对照组，术后逐渐恢复，但良恶性两组变化幅度差异明显；（2）血清总胆红素与血清IL-2及SIL-2R水平之间密切相关；（3）胆道外引流术式较内引流术式相对不利于梗黄病人术后血清IL-2和SIL-2R水平的恢复。结论梗黄病人体内存在血清IL-2／SIL-2R系统平衡紊乱，监测血清IL-2和SIL-ZR有助于估价病情、判断预后及鉴别梗阻的性质，胆道外引流相对不利于血清IL-2和SIL-2R的恢复。     

番茄红素抑制原代前列腺上皮细胞增殖和雄激素受体基因片段活性

目的 观察类胡萝卜素族中番茄红素对原代前列腺上皮细胞增殖和雄激素受体基因片段活性的影响.方法 分离培养获得原代前列腺上皮细胞后,加入1、5、10μmol/L番茄红素溶液,免疫荧光法测定其对原代上皮细胞增殖的影响.将含有构建的雄激素受体基因片段和荧光素酶报告基因的质粒DNA导入培养中的原代细胞,转染后加入1、5、10μmol/L番茄红素溶液,通过荧光光度计的结果评价番茄红素对雄激素受体基因活性的影响.结果与细胞培养基对照组比较,加入1、5、10μmol/L番茄红素溶液后细胞增殖分别减少了3.04%、6.88%和18.42%(P<0.01).与作为番茄红素溶剂的1%四氢呋喃溶剂(THF)对照组比较,加入5、10μmol/L番茄红素溶液后细胞增殖分别减少了3.86%和15.40%(P<0.05).将上述不同浓度番茄红素溶液加入转染后的含有构建的雄激素受体基因片段和荧光素酶报告基因的质粒DNA的原代细胞后,加入1、5、10μmol/L番茄红素溶液后荧光光度计数值分别降低了52.38%、68.10%和95.24%.结论 番茄红素对原代前列腺上皮细胞的增殖有显著抑制作用,它对雄激素受体基因片段的活性也有着抑制作用,并且与剂量呈正相关.     

Cellular interleukin-1 receptor antagonist production in patients receiving on-line haemodiafiltration therapy.

BACKGROUND: Repetitive exposure to cytokine-inducing substances (pyrogens) results in chronic inflammation, which may significantly contribute to some of the long-term complications in dialysis patients. On-line dialysis modalities, such as on-line haemodiafiltration (HDF), raise particular concerns because of the administration of infusate prepared from potentially contaminated dialysis fluid. Hence, great retention capability for pyrogens is of critical importance for the safe performance of on-line systems. METHODS: The microbiological safety of a novel on-line system, ONLINEplus(TM), was assessed in clinical practice in five centres for 3 months. Infusate and dialysis fluid were regularly monitored for microbial counts, endotoxins, and cytokine-inducing activity. Levels of interleukin-1 receptor antagonist (IL-1Ra) were determined in supernatants of whole blood incubated either under pyrogen-free conditions (spontaneous cytokine production) or following low-dose endotoxin exposure (LPS-stimulated cytokine production). RESULTS: We failed to detect microorganisms or endotoxin contamination of infusate during the entire study period. Moreover, neither infusate nor dialysis fluid demonstrated cytokine-inducing activity. Intradialytic IL-1Ra induction was not detected, as there was no difference between pre- and post-session values for both spontaneous and LPS-stimulated IL-1Ra production (115+/-26 vs 119+/-27 and 2445+/-353 vs 2724+/-362 pg/10(6) white blood cells (WBC), respectively). Neither the number of immunocompetent cells nor their capacity to produce IL-1Ra declined during this period, indicating that cells were not significantly stimulated during treatment. Spontaneous and LPS-induced exvivo IL-1Ra generation remained unchanged after 3 months of on-line HDF therapy as compared with the start of the study (71+/-30 pre- vs 48+/-14 post-study, and 2559+/-811 vs 2384+/-744 pg/10(6) WBC, respectively). CONCLUSIONS: The present on-line system performed safely from a microbiological view-point as both the dialysis fluid and infusate were consistently free of microorganisms, endotoxins, and cytokine-inducing substances. As a result, on-line HDF therapy had no effect upon the chronic inflammatory responses in end-stage renal disease patients.     

YWHAZ对肾小管上皮细胞增殖的影响

目的探索调节蛋白酪氨酸/色氨酸羟化酶激活蛋白ζ(YWHAZ)对正常氧条件以及缺氧复氧处理的肾小管上皮细胞增殖的影响。 方法利用人近端肾小管上皮细胞系(HK－2)通过RNAi技术以及质粒转染的方式建立YWHAZ低表达和过表达的细胞模型,观察干预YWHAZ表达后细胞的一般情况,检测细胞增殖活性;而后给予细胞缺氧复氧处理,检测细胞增殖情况的改变。 结果成功建立YWHAZ低表达和过表达的细胞模型,低表达YWHAZ后细胞的增殖能力较对照组下降,在缺氧复氧条件下增殖能力下降更为显著,过表达YWHAZ则缓解了缺氧复氧造成的细胞增殖抑制。 结论YWHAZ可能为维持近端肾小管上皮细胞增殖活性的重要因子。     

Angiotensin II receptor blocker inhibits tumour necrosis factor-alpha-induced cell damage in human renal proximal tubular epithelial cells

Aim: We investigated the effect of angiotensin II (AII) type 1 (AT1) and angiotensin II type 2 (AT2) receptor blockers on tumour necrosis factor alpha (TNF‐α)‐induced cell damage in human renal proximal tubular epithelial cells (RPTEC). Methods: The lactate dehydrogenase (LDH) and N‐acetyl‐beta‐glucosaminidase (NAG) release into the medium after TNF‐α treatment in RPTEC were determined using modified commercial procedures. In addition, the levels of caspase 3/7 activity in RPTEC were measured after TNF‐α treatment with ΑΤ1 or AT2 receptor blockers. Finally we investigated the change of p22phox protein levels after TNF‐α with ΑΤ1 or AT2 receptor blockers in RPTEC. Results: Tumour necrosis factor alpha (10^?8 mol/L) significantly increased LDH and NAG release into the medium from RPTEC. ΑΤ1 receptor blockers, olmesartan and valsartan (10^?9?10^?6 mol/L) showed a significant reduction on TNF‐α‐induced LDH and NAG release in RPTEC. AT2 receptor blocker, PD123319 (10^?7?10^?5 mol/L) also decreased TNF‐α‐induced LDH and NAG release in RPTEC. Blockade of both ΑΤ1 and AT2 receptor indicated additional reduction on TNF‐α‐induced LDH and NAG release. TNF‐α (10^?8 mol/L) treatment showed small but significant increases of caspase 3/7 activity in RPTEC, and AT1 and AT2 receptor blockers (10^?8 mol/L) comparably decreased TNF‐α‐induced caspase 3/7 activity. Significant increases of p22phox protein levels were observed in TNF‐α‐treated group in RPTEC. However, only ΑΤ1 (10^?8 mol/L) but not AT2 (10^?5 mol/L) receptor blocker significantly decreased TNF‐α‐induced p22phox protein levels. Conclusion: The present study demonstrates that TNF‐α induces renal tubular cell damage in RPTEC and AT1/AT2 receptor blockers showed cytoprotective effects probably via at least partly different mechanism.     

Ganoderma extract prevents albumin-induced oxidative damage and chemokines synthesis in cultured human proximal tubular epithelial cells.

BACKGROUND: Ganoderma lucidum (Ganoderma or lingzhi) is widely used as an alternative medicine remedy to promote health and longevity. Recent studies have indicated that components extracted from Ganoderma have a wide range of pharmacological actions including suppressing inflammation and scavenging free radicals. We recently reported that tubular secretion of interleukin-8 (IL-8) induced by albumin is important in the pathogenesis of tubulointerstitial injury in the proteinuric state. In this study, we explored the protective effect of Ganoderma extract (LZ) on albumin-induced kidney epithelial injury. METHODS: Growth arrested human proximal tubular epithelial cells (PTECs) were incubated with 0.625 to 10 mg/ml human serum albumin (HSA) for up to 72 h. HSA induced DNA damage and apoptosis in PTEC in a dose- and time-dependent manner. Co-incubation of PTEC with 4-64 microg/ml LZ significantly reduced the oxidative damage and cytotoxic effect of HSA in a dose-dependent manner (P<0.001). Increased release of IL-8 and soluble intercellular adhesion molecules-1 (sICAM-1) in PTEC induced by HSA was ameliorated by co-incubation with Ganoderma (16 microg/ml). To explore the components of LZ that exhibited most protective effect in HSA-induced PTEC damages, LZ was further separated into two sub-fractions, LZF1 (MW <30 kDa) and LZF2 (MW <3 kDa), by molecular sieving using millipore membrane. PTEC were incubated with 5 mg/ml HSA in the presence of different doses of LZF1, LZF2 or unfractionated LZ. RESULTS: There was no difference in the degree of protection from HSA-induced cytotoxicity or oxidative DNA damage between different fractions of LZ. However, low molecular weight LZ (<3 kDa) was most effective in reducing sICAM-1 released from HSA-activated PTEC whereas the high molecular weight LZ (unfractionated LZ) was more effective in diminishing IL-8 production. CONCLUSIONS: Our results suggest that Ganoderma significantly reduces oxidative damages and apoptosis in PTEC induced by HSA. The differential reduction of IL-8 or sICAM-1 released from HSA-activated PTEC by different components of the LZ implicates that components of Ganoderma with different molecular weights could play different roles and operate different mechanisms in preventing HSA-induced PTEC damage.     

Functional endothelin 1 receptors on human glomerular podocytes and mesangial cells.

To determine if endothelin 1 (Et1) receptors are present in human glomeruli, and which glomerular cells possess these receptors, 125I Et1 binding to isolated glomeruli and cultured glomerular mesangial and epithelial cells was studied. The latter were identified as podocytes. We demonstrated that Et1 binds specifically and reversibly to isolated human glomeruli and to cultured glomerular mesangial and epithelial cells. Scatchard analysis of competitive inhibition of 125I Et1 binding gave the following results (m +/- SEM, n = 3): isolated glomeruli, Kd = 4.2 +/- 2.1 x 10(-10) M, Bmax = 8.1 +/- 1.2 x 10(10) sites/mg protein; mesangial cells, Kd = 5.2 +/- 1.5 x 10(-10) M, Bmax = 1.87 +/- 0.49 x 10(4) sites/cell; epithelial cells, Kd = 7.2 +/- 1.5 x 10(-10) M, Bmax = 2.46 +/- 0.15 x 10(4) sites/cell. These receptors seem to be functional, since in both mesangial and epithelial cells Et1 induces a rapid and transient increase in intracellular [Ca2+]i. All these results indicate that Et1 may regulate glomerular filtration rate through an autocrine-paracrine pathway on mesangial cells and on podocytes.     

肾小管上皮细胞转分化在慢性移植肾肾病中的作用

目的探讨肾小管上皮细胞-肌成纤维细胞转分化在慢性移植肾肾病(CAN)的发生、发展中的作用。方法36例移植肾穿刺标本分为正常组(N组),CAN组(C组),依Banff 97标准将C组按CANⅠ、Ⅱ、Ⅲ级分为C_1、C_2、C_3组,每组9例。免疫组化染色半定量分析比较α-平滑肌肌动蛋白(α-SMA)、波形蛋白(VIM)、角质蛋白(CK_(AEI/AE3))和转化生长因子β1(TGF-β1)在各组中的表达,线性相关分析TGF—β1和α-SMA、VIM、CK_(AE1/AE3)表达的相关性。结果C_1、C_2、C_3组α-SMA表达积分分别为0．95±0．07、1．78±0．12、2．42±0．31,VIM表达积分分别为0．74±0．05、1．31±0．18、2．34±0．25,组间呈递增趋势,均明显高于N组α-SMA表达积分0．07±0．02、VIM表达积分0．09±0．02(P＜0．05)；移植肾组织中TGF-β1表达与α-SMA、VIM呈正相关(r分别为0．73、0．68,P＜0．05),而与CKAE1/AE3表达呈负相关(r=-0．71,P＜0．05)。结论肾小管上皮细胞-肌成纤维细胞转分化参与了CAN的发生、发展,而局部肾组织中TGF-β1的表达上调可能是介导肾小管上皮细胞转分化的重要原因。     

缬沙坦抑制人类肾小管上皮细胞转分化的初步研究

目的：探讨血管紧张素ⅡⅠ型受体拮抗剂缬沙坦（ｖａｌｓａｒｔａｎ,Ｖａｌ）在人类肾小管上皮细胞系（ＨＫＣ）转分化中的作用。方法将培养的ＨＫＣ细胞分为（１）无血清培养培养对照组;（２）阳性对照组（ＭＣＰ－１＋ＡＡＩ组）：培养液中加入马兜铃酸－Ｉ（ＡＡＩ）和单核细胞趋化蛋白－１）ＭＣＰ－１）：（３）Ｖａｌ组：培养液中加入Ｖａｌ;（４）ＭＣＰ－１＋ＡＡＩ＋Ｖａｌ组。应用间接酶标免疫组织化学方法（ＩＥＩ）检