Abnormal High Expression of B-Cell Activating Factor Belonging to the TNF Superfamily (BAFF) Associated With Long-Term Outcome in Kidney Transplant Recipients

B-cell activating factor belonging to the tumor necrosis factor superfamily (BAFF) is a critical regulator of B-cell maturation and survival. We investigated the expression of BAFF in peripheral blood sample from kidney transplant recipients. Results of flow cytometry showed that at 5 years or more posttransplantation, cell-surface BAFF was significantly expressed on peripheral CD3+ T lymphocytes kidney transplant recipients in compared with other groups (P < .05). BAFF expression was noted on CD4+ and CD8+ T cells. The BAFF messenger RNA level in peripheral mononuclear cells was consistent with the protein level. However, the serum soluble BAFF level varied among individuals in each group. Stratified by renal function, the cell-surface BAFF level was significantly higher in recipients with abnormal renal function compared with those with normal renal function (P < .05). Enzyme-linked immunosorbent assay results showed that expression level of cell-surface BAFF significantly correlated with panel reactive antibody. These results indicate that BAFF may be involved in the development of graft loss.     

B Cell Activating Factor (BAFF) in Long-term Kidney Transplant Recipients Is Not a Prognostic Marker for Allograft Dysfunction or Survival

Objective

B cell activating factor (BAFF) has been shown to play a role in B cell survival, maturation, and activation, and has been linked with renal transplant outcome. BAFF signaling has been associated with plasmablast survival, anti-HLA immunization, and loss of graft function. We aimed to analyze the interplay between BAFF, memory B cells, and plasmablasts in relation to allograft function in long-term kidney transplant (KTx) recipients and their anti-HLA sensitization.

慢性排斥移植肾TNF_α和IL-2R原位表达

本文应用免疫组化技术对18例慢性排斥移植肾活检组织中肿瘤坏死因子α(TNFα)及白细胞介素Ⅱ受体(IL-2R)表达进行检测。结果表明TNFα和IL-2R在移植肾中表达较正常肾脏明显增强,且主要分布于移植肾间质浸润炎性细胞(淋巴细胞和巨噬细胞),提示慢性排斥移植肾中浸润淋巴细胞和巨噬细胞处于活化状态,其可能通过产生和释放各种细胞因子发挥其各种生物学作用,从而介导细胞免疫损伤,因而认为细胞免疫可能在慢性排斥发病中起作用,细胞因子及其受体则是这一过程中的重要介质。     

肿瘤坏死因子相关细胞凋亡诱导配体受体-4在人胰腺癌组织中的表达

目的：通过检测肿瘤坏死因子相关细胞凋亡诱导配体受体-4(TRAIL-R4)在胰腺癌组织中的表达，探讨胰腺癌细胞抵抗细胞凋亡的机理。方法：应用mRNA印迹法(Northern blotting)、蛋白质印迹法(Western blotting)和免疫组织化学技术，定性、定位分析TRAIL-R4在正常胰腺组织和胰腺癌组织中的表达。结果：TRAIL-R4 mRNA和蛋白在正常胰腺组织中呈弱表达或不表达，而在胰腺癌组织中呈高表达；免疫组织化学检测显示，在胰腺癌细胞中TRAIL-R4蛋白呈强染色。结论：TRAIL-R4表达水平在正常胰腺组织与胰腺癌组织中存在显著差异，提示胰腺癌细胞中可能存在对TRAIL介导的细胞凋亡抵抗新机理。     

Synergistic Antitumor Effect of an Angiogenesis Inhibitor (TNP-470) and Tumor Necrosis Factor in Mice

Purpose We investigated the potentiation of combination therapy using tumor necrosis factor (TNF) with TNP-470, a potent angiogenesis inhibitor. Methods We evaluated the antitumor effect in vivo against subcutaneous (s.c.) MC38 mouse colon adenocarcinoma tumors in C57BL/6 mice. The mice were treated with a single bolus injection via the tail vein of 3 or 8 μg rhTNF in 0.5% bovine serum albumin/normal saline (BSA/NS), or with 0.5% BSA/NS alone as a control, with or without TNP-470 pretreatment, given as 30 or 60 mg/kg × 2 days, s.c. DNA synthesis in human umbilical endothelial cells (HUVEC) was assessed by [³H]thymidine uptake after incubation with TNF, with or without TNP-470. Results The antitumor effect of TNP-470 pretreatment combined with 3 μg recombinant human (rh) TNF injection resulted in an 80% reduction of tumor volume compared with the control. This was significantly better than that induced by 3 μg rhTNF alone (P < 0.005). DNA synthesis in HUVEC was inhibited by TNF with TNP-470 in a dose-dependent manner, but there was no enhanced effect against MC38 in vitro. Conclusions These results suggest that the combination of the angiogenesis inhibitor TNP-470 and TNF might have a synergistic antitumor effect on solid tumors in vivo.     

Humoral Heart Rejection (Severe Allograft Dysfunction with no Signs of Cellular Rejection or Ischemia): Incidence, Management, and the Value of C4d for Diagnosis

Severe allograft dysfunction after heart transplant (HT), without ischemia or evidence of cellular rejection upon endomyocardial biopsy (EMB), is a rare but potentially fatal condition that suggests humoral rejection (HR). Its incidence, and the methods of choice for its diagnosis and management, remain uncertain. We retrospectively studied 445 HT patients (April 1991-December 2003) to determine incidence of HR diagnosed by clinical and conventional histopathological criteria. We used immunofluorescence (IF) techniques to test archived frozen EMB issue for IgM, IgG, C1q, C3, fibrin and C4d. Twelve patients (2.7%) fulfilled the criteria for HR after a mean time post-HT of 21.3 +/- 24.7 months (range: 2-72 months). Patients were treated with high doses of steroids and plasmapheresis, with successful recovery in 11 cases. IF studies using classical markers were mainly negative for the six patients with enough EMB tissue for testing. All six patients showed positivity for C4d during the HR episode but not before or after. Humoral rejection was observed in less than 3% of HT patients. Plasmapheresis treatment was highly effective. Classical IF tests were not useful for diagnosis, but C4d appears to be useful both for confirmation of diagnosis and for monitoring response to treatment.     

心肌细胞NAD（P）H荧光成分分析法监测心脏移植排斥反应

目的为早期诊断和有效预防心脏移植排斥反应,研究在细胞水平用心肌NAD（P）H荧光成分分析监测排斥反应的可行性。方法用NAD（P）H荧光标记监测线粒体功能,从14例儿童心脏移植患者心内膜活检组织中分离活体心肌细胞和大鼠（n=5,13～14周龄）心肌细胞,光谱分辨的时间相关单光子计数（TCSPC）记录375nm紫外激光激发的心肌自发荧光（AF）光谱和荧光寿命。将鼠心肌细胞分为4组,无缺氧对照组,无缺氧鱼藤酮组,缺血对照组和缺血鱼藤酮组;对比无排斥反应（R0）人心肌细胞,无缺氧鼠细胞和缺血鼠细胞间,R0人心肌细胞和轻度排斥反应（R1）间的AF动力学变化。结果成功分离并测定了人心肌细胞AF光谱,在420～560nm光谱区域,至少存在0.5～0.7ns,1.9～2.4ns和9.0～15.0ns3个AF寿命池。与无缺氧对照组比较,缺血对照组鼠心肌细胞荧光强度明显增强（P〈0.05）;无缺氧鱼藤酮组和缺血鱼藤酮组心肌AF强度均显著增强,两组比较差异无统计学意义（P〉0.05）。R0组人心肌细胞AF强度显著弱于无缺氧对照组鼠心肌细胞（P〈0.05）;人心肌AF强度与排斥反应相关,R1的荧光强度显著增强（P〈0.05）,但光谱形态无改变,与缺血对照组鼠心肌细胞光谱改变一致。结论光谱分辨的荧光光谱结合荧光寿命分析是监测心肌线粒体氧化代谢的敏感方法,有良好的重复性。人心肌AF表现出较高的代谢活性,排斥反应时活性降低。该技术结合多光子共聚焦显微镜,可直接在体评估线粒体功能,是监测早期排斥反应的新方法。     

Immunohistochemical Expression of Fatty Acid Synthase (FASN) is Correlated to Tumor Aggressiveness and Cellular Differentiation in Salivary Gland Carcinomas

Fatty acid synthase (FASN) expression is closely related to cancer progression, in particular, tumor aggressiveness and poor prognosis. This study aimed to analyse the expression of FASN in carcinomas of the salivary glands and correlate it with Ki-67 expression. We analysed by immunohistochemistry the expression of FASN and Ki‐67 on tissue sections from 7 cases of adenocarcinoma, not otherwise specified (AdNOS), 6 cases of polymorphous adenocarcinoma (PAC), 16 cases of acinic cell carcinoma (AcCC), 19 cases of adenoid cystic carcinoma (AdCC), 15 cases of epithelial-myoepithelial carcinoma (EMC); 10 cases of secretory carcinoma (SC), 13 cases of mucoepidermoid carcinoma (MEC), 10 cases of salivary duct carcinoma (SDC) and 7 cases of myoepithelial carcinoma (MC). These carcinomas were classified into aggressive and indolent regarding their biological behaviour. Additionally, MEC and AdCC were also classified according to the histological grade. High expression of FASN was found in SDC (100%), SC (100%), AcCC (68.7%) and AdNOS (57.2%). No association was found between FASN and Ki-67 expression. Aggressive carcinomas showed a higher rate of Ki-67 proliferation (p < 0.001) and greater expression of FASN when compared to indolent carcinomas (p < 0.05). With regards to carcinomas categorized as indolent, FASN expression was much higher in the lesions that presented cell differentiation (SC and AcCC). Also, FASN expression was significantly higher in high-grade AdCC and MEC when compared to low-grade tumors (p < 0.05). We concluded that FASN expression was correlated to tumor aggressiveness and cellular differentiation in salivary gland carcinomas.     

Comparison of C4d Detection on Erythrocytes and PTC‐C4d to Histological Signs of Antibody‐Mediated Rejection in Kidney Transplantation

C4d on erythrocytes (EC4d), C4d peritubular capillary deposition (PTC‐C4d) staining and histology were compared in a cross‐sectional cohort of 146 renal allograft biopsies (132 patients). EC4d levels paralleled PTC‐C4d staining, but were more predictive of peritubular capillaritis (PTC). Donor‐specific antibodies (DSA), PTC‐C4d, EC4d and PTC were analyzed in an independent longitudinal follow‐up cohort (96 biopsies, 76 patients). Seventy‐six samples were PTC and EC4d concordant, 11 positive and 65 negative, 7 PTC‐EC4d+ and 13 PTC+EC4d?. EC4d levels were related to DSA occurrence. With ABMR defined by PTC and DSA, all apparently discordant patients, EC4d negative, were correctly reassigned comparing EC4d level curves with rejection kinetics, with positive EC4d samples predating biopsy or late biopsies compared with ABMR flare‐ups. All EC4d‐positive patients without PTC or DSA had permanent high EC4d levels unrelated to rejection. EC4d was more abundant in PTC‐positive (mean = 108.5%± 3.4; n = 50) than PTC‐negative samples (mean = 88.1%± 1.3; n= 96; p < 0.0001). Sensitivity, specificity, positive predictive value and negative predictive value of PTC‐C4d and EC4d for PTC were, respectively, 75%, 79%; 64%, 76% (p < 0.05); 28%, 46% (p < 0.05) and 93%, 94%. Values were similar for DSA. A noninvasive blood test, EC4d, and particularly longitudinally monitoring EC4d levels, may increase surrogate ABMR testing options.     

Increased Expression of Valosin-Containing Protein (p97) Is Correlated With Disease Recurrence in Follicular Thyroid Cancer

Background Valosin-containing protein (VCP) is involved in the ubiquitin/proteasome-degradation pathway, which works in proliferation and antiapoptosis in human cancer cells. Our previous study showed that VCP expression levels correlated with the recurrence and prognosis of several human cancers, such as hepatocellular carcinoma, gastric carcinoma, and colorectal carcinoma. In this study, the correlation of VCP expression with the prognosis of differentiated thyroid carcinoma was examined. Methods VCP expression in 332 patients who underwent operation for differentiated thyroid carcinoma—257 with papillary thyroid carcinoma and 75 with follicular thyroid carcinoma (FTC)—was analyzed by immunohistochemistry. The staining intensity in tumor cells was categorized as weaker than (low expression), equal to (intermediate expression), or stronger than (high expression) that in endothelial cells in noncancerous tissue. Results One hundred ten (33.5%) cases showed low VCP expression, 117 (28.0%) showed intermediate expression, and 101 (30.8%) showed high expression. VCP expression significantly correlated with histological subtype (P < .05) and lymph node metastasis (P < .01). However, it correlated with neither any clinicopathologic factor nor prognosis in papillary thyroid carcinoma. VCP expression correlated with extrathyroidal extension (P < .05), pT classification (P < .05), and lymph node metastasis (P < .01) in FTC. Patients with low VCP expressing FTC showed better disease-free and overall survival rates than those with intermediate or high expression (P < .01 and P < .05, respectively). Multivariate analysis revealed VCP expression and extracapsular extension to be independent prognostic factors for disease-free survival in cases of FTC. Conclusions The prognostic utility of VCP expression in FTC was demonstrated.     

介入治疗激素性股骨头坏死血管内皮细胞生长因子表达的实验研究

目的：观察股动脉药物灌注治疗实验性激素股骨头坏死过程中血管内皮细胞生长因子的表达，探讨介入治疗对骨坏死再血管化和再骨化的机理，方法：新西兰兔48只，利用Shwartman反应和大剂量甲基强的松龙造成兔股骨头坏死模型后，随机分为动脉组、静脉组和对照组。在治疗后的1、2、3和4周，观察股骨头VEGF阳性成骨细胞率，阳性软骨细胞率和阳性血管率，结果：动脉比静脉治疗能更快，更有率提高VEGF阳性成骨细胞率，阳性软骨细胞率和阳性血管率，结论：股动脉介入股骨坏死比静脉治疗更快速有效，其机理在于疏通已经阻塞的血管，促进VEGF等细胞因子的分泌从而加速股骨头的再血管化和再骨化。     

参附注射液对单侧输尿管梗阻大鼠肾脏肝细胞生长因子表达的影响

目的：探讨单侧输尿管梗阻后大鼠肾间质纤维化发生过程中肝细胞生长因子（HGF）的表达及中药参附注射液对其的影响。方法：采用单侧输尿管结扎（UUO）制造梗阻性肾病模型，将56只大鼠随机分为对照组（假手术组）、手术组（UUO组）和治疗组（UUO＋参附），术后7d、14d观察肾组织病理改变，应用免疫组织化学方法检测肾组织中HGF的表达。结果：与对照组相比，手术组肾间质出现了明显的纤维化，HGF的表达在术后第7天明显增加，第14天较第7天减弱，与手术组相比，治疗组肾间质纤维化明显减轻，而且HGF的表达在术后第7天明显上调，第14天较第7天上调更明显，有统计学差异（P〈0．05）。结论：参附注射液可以上调肾组织HGF的表达，减轻肾小管一间质纤维化，发挥肾保护作用。     

Immediate Antibody-Mediated (Hyperacute) Rejection in Small-Bowel Transplantation and Relationship to Cross-Match Status and Donor-Specific C4d-Binding Antibodies: Case Report

Background

The role of preformed donor-specific antibodies (DSAs) as a barrier to isolated intestinal transplantation (ITx) remains ambiguous; thus, a positive cross-match has not been a contraindication to ITx.

Objective

To report the case of a patient with Crohn's disease who underwent ITx and developed immediate antibody-mediated rejection on reperfusion of the allograft.

Methods

Percent reactive antibody testing was performed using pretransplantation serum samples and at transplantation using bead-based assays (Luminex, Luminex Corp, Austin, Tex) and flow cytometry solid-phase assays (FlowPRA single-antigen beads (One Lambda, Inc, Canoga Park, Calif). Serologic tests, flow cytometry cross-matching, and flow cytometry assays of C4d-binding serum antibodies were also performed. Histologic and immunofluorescent analysis of biopsy specimens was performed.

Results

HLA typing revealed no sharing of class I or II antigens between donor and recipient. Pretransplantation donor-specific antibodies (DSA) were present at transplantation. Cross-matching (performed during surgery) was positive for class I and II by serologic testing and flow cytometry. After reperfusion, the graft immediately developed severe ischemic injury and arteritis on mucosal biopsy specimens, with immunoglobulin deposition. The DSA C4d binding antibodies were also present. After intense immunosuppression and plasmapheresis, the graft and the biopsy histologic findings showed marked improvement (day 2). By day 7 posttransplantation, patient and graft status were stable. The patient has remained clinically stable for more than a year after transplantation.

Conclusions

Pretransplant DSA in ITx can be a risk factor for immediate (hyperacute) but potentially reversible antibody-mediated rejection. Thus, pretransplantation DSA and cross-match results are critical components to be considered in patients awaiting or undergoing ITx.     

The role of superextended lymphadenectomy (D4) in gastric cancer

AIM: The outcome of surgery in gastric cancer differs in Japan and Western countries and the extension of lymphadenectomy may play a crucial role in survival. In Japan the choice of performing extended (D2) and superextended (D4) lymphadenectomies is based on retrospective studies, and a prospective randomized study comparing D2 and D4 is still in course. In Western countries the randomized trials comparing D1 and D2 could not provide definite indications, D2 is not yet performed as a routine procedure and D4 is accepted only by few surgeons. We report our experience and discuss indications and results. METHODS: Since January 2000 through December 2002 we performed 27 superextended lymphadenectomies for the radical treatment of advanced gastric cancer. Early gastric cancers and patients over 80 years of age received conventional D2 gastrectomies. Selection of patients for D4 was made after laparotomy, when intraoperative peritoneal lavage cytology could rule out the presence of malignant cells, while D2 was done in case of peritoneal micrometastases. RESULTS: Every patients had 39.5 nodes removed on average (range 17-94), and micrometastases in tier 16 were found in 7 cases (26%). Early post-operative surgical morbidity was 18% (5 patients) and mortality was 3.7% (1 patient). As much as 30% of patients complained of diarrhea as a late complication. The follow up could demonstrate a 3 year overall actuarial survival of 76%. Actuarial survival was 100% for N- and 70% for N+. A remarkable data was that 4 out of 5 patients who died from recurrence in the follow-up, were N4+. Actuarial survival at 3 years for N4+ patients was 34%, and the difference in survival between N4+ and other N+ was statistically significant (p<0.05). CONCLUSIONS: Superextended lymphadenectomy in gastric cancer is feasible with postoperative morbidity and mortality rates not exceeding the rates of other lymphadenectomies. Actuarial survival at 3 years with D4 was better than in previous personal experience with D2, although the patients who underwent D4 were selected by intraperitoneal lavage cytology, while D2 patients had not been selected. The prognosis for N4- patients was better than for N4+ with micrometastases in tier 16. The presence of N4 micrometastases worsens the prognosis, but it is still uncertain whether D4 does improve survival: it is undoubtedly a new means of more accurate staging in gastric cancer surgery. The newer TNM classification regards the number of nodes removed as an indicator of radicality. Every surgeon should consider that superextended lymphadenectomies could comply with R0 radicality, and perform it within the ranges of low morbidity and mortality, until randomized trials with definitive results are available.