Hepatitis B Immunoglobulins Inhibit Dendritic Cells and T Cells and Protect Against Acute Rejection After Liver Transplantation

The efficacy of anti-viral intravenous immunogobulins (anti-HBs Ig and anti-CMV Ig) in preventing acute rejection after liver transplantation was assessed in a retrospective analysis, and correlated to their effects on immune cells in vitro. HBs Ag-positive liver graft recipients (n = 40) treated prophylactically with anti-HBs Ig had a significantly lower incidence of acute rejection compared with recipients without viral hepatitis (n = 147) (12% vs. 34%; p = 0.012), while the incidence of rejection in HCV-positive recipients (n = 29) was similar to that in the control group. Treatment with anti-CMV Ig (n = 18) did not protect against rejection. In vitro, anti-HBs Ig suppressed functional maturation of and cytokine production by human blood-derived dendritic cells (DC) at concentrations similar to the serum concentrations reached during anti-HBs Ig treatment of liver graft recipients. In addition, anti-HBs Ig inhibited allo-antigen- and lectin-stimulated proliferation of peripheral T cells. Anti-CMV Ig suppressed functional DC-maturation and alloantigen-stimulated T-cell proliferation, but not lectin-driven T-cell proliferation. In conclusion, anti-HBs Ig protects against acute rejection after liver transplantation, probably by functional inhibition of the two principal immune cells involved in allograft rejection, DC and T cells.     

Bisphosphonates Induce Apoptosis of Stromal Tumor Cells in Giant Cell Tumor of Bone

Giant cell tumour of bone (GCT) is an aggressive primary neoplasm that results in the production of osteolytic lesions. Stromal cells, which form the main neoplastic component of this tumor, regulate the formation of osleoclast-like giant cells that are ultimately responsible for bone destruction. Bisphosphonates prevent bone resorption by inhibiting osteoclast activity and promoting osteoclast apoptosis, and they have been known to induce apoptosis of primary neoplastic cells such as those in breast and prostate cancers. We hypothesized that in bisphosphonates may induce apoptosis not only in osteoclast-like giant cells but also in neoplastic stromal cells of GCT both in vitro and in vivo. Twelve patients with GCT were treated with weekly injections of pamidronate for a period of 6 weeks prior to surgery. GCT specimens were collected at the time of biopsy and during definitive surgery. TUNEL assay was used to evaluate apoptotic DNA fragmentation in cells. In addition, twelve GCT primary cultures from these patients were treated with zoledronate, pamidronate, or alendronate for 48 hours at different doses (3, 30, or 150 M) and subjected to apoptosis assay by flow cytometry following fluorescent Annexin-V labeling. The results showed that pamidronate significantly induced apoptosis in both osteoclast-like giant cells and stromal tumor cells, in vivo. All three bisphosphonates caused substantial apoptosis of stromal tumor cells in cultures. Zoledronate was the most potent reagent, resulting in an average cell death of 27.41% at 150 M, followed by pamidronate (22.23%) and alendronate (15.3%). Our observations suggest that these drugs may be considered as potential adjuvants in the treatment of GCT.Both authors (Y.Y. Cheng and L. Huang) contributed equally to this work.     

肿瘤浸润性树突状细胞与粘蛋白MUC1在前列腺癌与前列腺增生中的表达研究

目的 :探讨上皮特异性肿瘤蛋白 (MUC1)与肿瘤浸润性树突状细胞 (TIDC)在前列腺癌及前列腺增生组织中的表达。　方法 :采用免疫组织化学SP法检测MUC1和TIDC在 30例前列腺癌、2 0例前列腺增生组织的表达。结果 :MUC1在前列腺癌、前列腺增生组织中均表达 ,MUC1的染色分型同肿瘤的病理分级密切相关 (P <0 .0 0 1)。TIDC的数量在高分化与低分化前列腺癌间差异有显著性 (P <0 .0 0 1)。　结论 :MUC1的表达模式和TIDC数量的监测可以作为前列腺癌恶性程度和预后的判断指标。TIDC的减少可能是肿瘤免疫逃逸和耐受的重要环节 ,肿瘤细胞表面高密度的MUC1可能参与前列腺癌侵袭转移和激素耐受机制。     

应用热诱导凋亡法制备靶向胶质瘤细胞的树突状细胞疫苗

目的 应用热诱导凋亡U251细胞致敏树突状细胞,观察其诱导特异性细胞毒性T细胞的能力,探讨其对肿瘤细胞的杀伤机制.方法 应用改良Inaba法,从小鼠骨髓细胞中诱导出树突状细胞,应用电镜和流式细胞技术行表型鉴定,然后和经热诱导凋亡的胶质瘤细胞共培养获得树突状细胞疫苗,细胞计数试剂盒( CCK-8)检测疫苗在体外促T细胞增殖和杀伤肿瘤细胞作用,将疫苗经尾静脉注入荷瘤裸鼠体内,检测体内肿瘤生长抑制作用.结果 44℃孵育3h能有效诱导胶质瘤细胞凋亡,凋亡率达(40.10±1.08)％.负载凋亡细胞抗原的树突状细胞在体外能有效促进T细胞增殖,增加干扰素(IFN)-γ的分泌,并随效靶比增加,对胶质瘤细胞的杀伤率增大.体内注射疫苗4周后,治疗组肿瘤体积(301.704±21.659) mm3,空白对照组为(487.116 ±65.975) mm3,差异有统计学意义(P＜0.01).结论 热诱导凋亡胶质瘤细胞致敏的树突状细胞能在体外有效促进T细胞增殖,诱导特异性细胞毒性T细胞杀伤胶质瘤细胞,有效抑制体内肿瘤生长.     

Prednisolone Suppresses the Function and Promotes Apoptosis of Plasmacytoid Dendritic Cells

Organ transplant recipients are highly susceptible to viral infections early after transplantation. Plasmacytoid dendritic cells (PDC) play a major role in antiviral immunity. Therefore, we determined the numbers of circulating PDC after liver transplantation (LTX) and established the effects of immunosuppressive drugs on PDC survival and function. PDC were determined longitudinally in 13 LTX recipients treated with prednisone and cyclosporin or tacrolimus. Purified PDC were cultured with or without clinically relevant concentrations of cyclosporin, tacrolimus or prednisolone. Apoptosis induction was monitored by determination of active caspase-3, nuclear condensation and annexin-V/7AAD staining. After LTX, a 4-fold reduction in the number of circulating PDC was observed (p < 0.01), which recovered partially after discontinuation of prednisone treatment. In vitro, prednisolone induced apoptosis in PDC, while cyclosporin and tacrolimus did not. Higher doses of prednisolone were needed to induce apoptosis in Toll-like receptor (TLR)-stimulated PDC. However, non-apoptosis inducing concentrations of prednisolone suppressed interferon-alpha production, upregulation of co-stimulatory molecules and allo-stimulatory capacity of TLR-stimulated PDC. In conclusion, prednisolone induces apoptosis in PDC, which explains the decline in circulating PDC numbers after transplantation. Moreover, prednisolone suppresses the functions of TLR-stimulated PDC. Therefore, corticosteroid-free immunosuppressive therapy may reduce the number and severity of viral infections after transplantation.     

体外培养的未成熟树突状细胞对延长小鼠同种异体移植心存活时间的影响

目的观察术前输注用低剂量粒细胞巨噬细胞集落刺激因子(GM-CSF)培养的小鼠骨髓源性未成熟树突状细胞(DC)对小鼠同种异体移植心存活时间的影响。方法分别用常规剂量和低剂量GM-CSF培养小鼠骨髓源性成熟DC和未成熟DC,采用流式细胞术检测细胞表型,比较其在体外刺激同种异体T细胞增殖的能力,并将培养的未成熟DC约1.0×106个于术前7d输注受者体内,观察同种异体移植心存活的时间。结果与常规剂量GM-CSF下培养出的成熟DC不同,低剂量GM-CSF培养出的DC为较单纯的未成熟DC,表型为CD11c+,CD80-,CD86-,MHCⅡlow,在体外不能有效刺激同种异体T细胞活化增殖,将其于术前输注受者体内,移植心的存活时间由6.3±1.2d延长至14.3±1.9d(P<0.01)。结论用低剂量GM-CSF可培养出表型和功能均未成熟的小鼠骨髓源性DC,将其在术前7d输注受者体内,可明显延长移植心的存活时间。     

抗人IgG抗体与丝裂霉素联合应用治疗膀胱癌的实验研究

目的 探讨抗人IgG抗体和丝裂霉素(MMC)联合应用对人膀胱癌细胞株的抑制作用及机制.方法 抗人IgG抗体和MMC单独或联合作用于人膀胱癌T24细胞,噻唑盐(MTT)法检测细胞生长抑制率,流式细胞仪检测T24细胞的凋亡率,蛋白质印迹法检测半胱氨酸天冬氨酸蛋白酶3(Caspase-3)和PARP的表达.采用T24细胞裸鼠移植瘤模型进行抗人IgG抗体和MMC的体内抗瘤实验.结果 单独应用抗人IgG抗体和MMC对T24细胞生长抑制率分别为(25.02±6.71)%和(32.31±6.46)%,而二者联用的抑制率达(73.66±5.81)%.PBS、25mg/L羊IgG、25mg/L羊抗人IgG抗体、2mg/L MMC、2mg/L MMCIk 25mg/L羊IgG和2mg/L MMC加25mg/L羊抗人IgG抗体处理T24细胞72 h后细胞凋亡率分别为1.7%、2.3%、20.7%、22.4%、28.3%和53.8%.羊抗人IgG抗体、MMC、MMC加羊IgG和MMC加羊抗人IgG抗体处理T24细胞72 h后出现17 000的Caspase-3和85 000的PARP剪切片断.体内实验显示羊IgG组、羊抗人IgG抗体组、MMC组、MMC加羊抗人IgG抗体组抑瘤率分别为2.31%、12.73%,36.81%和50.51%.HE切片观察见生理盐水组和羊IgG组移植瘤细胞基本无凋亡和坏死,羊抗人IgG抗体组肿瘤细胞凋亡和坏死增多,MMC组和MMC加羊抗人IgG抗体组则见大量肿瘤细胞凋亡和坏死.结论 抗人IgG抗体和MMC联合应用对膀胱癌具有体内外双重抗瘤作用,其抗瘤机制可能与诱导肿瘤细胞凋亡有关.     

胰腺癌患者外周血中树突状细胞的分离、纯化与扩增

目的 探讨从胰腺癌患者外周血中分离、纯化与扩增树突状细胞（DC）的有效方法。方法 分别从健康人（10例，对照组）和胰腺癌患者（12例，实验组）外周血中分离出单核细胞，而后将实验组的单核细胞与GM－CSF及IL－4共同培养，用免疫荧光法和流式细胞仪检测培养前、后的DC数量及DC表面HLA－DR及B7－2的表达水平，并与对照组比较。结果 与对照组相比，实验组DC表面HLA－DR及B7－2表达水平较低（P＜0.01）；经GM－CSF及IL－4联合培养7天后，其单核细胞中的DC数较培养前明显增多（P＜0.01），且DC表面HLA－DR及B7－2表达水平较培养前明显增高（P＜0.01），与对照组相比则无明显差异（P＞0.05）。结论 GM－CSF与IL－4的联合应用能有效地从胰腺癌患者外周血中制备出大量具有功能活性的DC。     

磁流体热疗对荷Lewis肺癌小鼠肿瘤细胞凋亡和周期的影响

目的探讨磁流体热疗对荷Lewis肺癌小鼠肿瘤细胞的凋亡和周期的影响。方法接种Lewis肺癌细胞悬液于C57BL／6小鼠的皮下，等肿瘤长至直径约为（0．8±0．1）cm时，随机分为4组：对照组、磁场组、磁流体组、实验组。加温治疗后48h，眼球取血，检测血中白细胞的变化，切取肿瘤标本，流式细胞仪检测细胞凋亡率和周期的变化。结果热疗后4组血中自细胞没有明显的变化（F=0．62，P=0．613）；肿瘤细胞凋亡率实验组为（63．55±8．39）％，对照组、磁场组、磁流体组分别为（28．43±6．29）％，（32．75±5．07）％，（32．42±6．15）％，实验组肿瘤细胞凋亡率明显高于其他3组（q=11．925，P〈0．05；g=10．458。P〈0．05；g=10．570，P〈0．05）；实验组细胞周期出现明显G1／G0期阻滞为（68．13±5．73）％显著高于对照组（47．95±9．98）％（q=5．501，P〈0．05），磁场组（49．23±6．62）％（q=5．152，P〈0．05），磁流体组（52．28±9．64）％（q=4．320，P〈0．05）。结论磁流体热疗可明显提高Lewis肺癌细胞的凋亡率，抑制Lewis肺癌细胞G1期向S期的进程。     

Intrahepatic Cholangiocarcinoma with Lymph Node Metastasis Successfully Treated by Immunotherapy with CD3-Activated T Cells and Dendritic Cells After Surgery: Report of a Case

Intrahepatic cholangiocarcinoma (ICC) with lymph node (LN) metastasis is generally associated with a poor prognosis. However, we treated ICC with LN metastasis successfully by surgery and postoperative immunotherapy in a 59-year-old woman. The immunotherapy consisted of CD3-activated T cells and tumor lysate- or peptide-pulsed dendritic cells. Pathological examination confirmed a diagnosis of moderately differentiated adenocarcinoma with LN metastasis and portal vein invasion. The patient has been alive without recurrence for 3 years 6 months since her operation.     

Plasmacytoid Dendritic Cells: No Longer an Enigma and Now Key to Transplant Tolerance?

Plasmacytoid (p) dendritic cells (DC) are a specialized subset of DC whose primary role was initially defined by the production of type I interferons in response to viral infection. They are now known to also possess a repertoire of functions capable of determining T cell fate and activation. Under homeostatic conditions, non‐lymphoid tissue‐resident pDC play a critical role in the regulation of mucosal immunity, as well as the development of central and peripheral tolerance. Although these cells display a number of characteristics that differ from conventional DC, particularly altered costimulatory molecule expression and poor allostimulatory capacity when interacting with T cells, this phenotype favors the generation of alloantigen‐specific regulatory CD4⁺ or CD8⁺ T cells critical to the development of graft tolerance. In this minireview, we discuss pDC ontogeny, functional biology and the emerging data that demonstrate the importance of pDC in the induction of tolerance, as well as recent studies that define mechanisms underlying pDC‐mediated tolerance to both solid organ and haematopoietic stem cell transplants. We also highlight their use in clinical settings and the potential of pDC both as targets and cellular therapeutic agents to improve the outcome of organ transplantation.     

Rapamycin‐Conditioned Donor Dendritic Cells Differentiate CD4+CD25+Foxp3+ T Cells In Vitro with TGF‐β1 for Islet Transplantation

Dendritic cells (DCs) conditioned with the mammalian target of rapamycin (mTOR) inhibitor rapamycin have been previously shown to expand naturally existing regulatory T cells (nTregs). This work addresses whether rapamycin‐conditioned donor DCs could effectively induce CD4⁺CD25⁺Foxp3⁺ Tregs (iTregs) in cell cultures with alloantigen specificities, and whether such in vitro‐differentiated CD4⁺CD25⁺Foxp3⁺ iTregs could effectively control acute rejection in allogeneic islet transplantation. We found that donor BALB/c bone marrow‐derived DCs (BMDCs) pharmacologically modified by the mTOR inhibitor rapamycin had significantly enhanced ability to induce CD4⁺CD25⁺Foxp3⁺ iTregs of recipient origin (C57BL/6 (B6)) in vitro under Treg driving conditions compared to unmodified BMDCs. These in vitro‐induced CD4⁺CD25⁺Foxp3⁺ iTregs exerted donor‐specific suppression in vitro, and prolonged allogeneic islet graft survival in vivo in RAG^?/‐ hosts upon coadoptive transfer with T‐effector cells. The CD4⁺CD25⁺Foxp3⁺ iTregs expanded and preferentially maintained Foxp3 expression in the graft draining lymph nodes. Finally, the CD4⁺CD25⁺Foxp3⁺ iTregs were further able to induce endogenous naïve T cells to convert to CD4⁺CD25⁺Foxp3⁺ T cells. We conclude that rapamycin‐conditioned donor BMDCs can be exploited for efficient in vitro differentiation of donor antigen‐specific CD4⁺CD25⁺Foxp3⁺ iTregs. Such in vitro‐generated donor‐specific CD4⁺CD25⁺Foxp3⁺ iTregs are able to effectively control allogeneic islet graft rejection.     

导入以腺病毒为载体多药耐药基因的脐血有核细胞对化疗药物的耐受性

通过腺病毒载体介导使多药耐药基因转染入脐血有核细胞,提高其对化疗药物的耐受性.结果表明,与未转染的脐血有核细胞相比,转染MDR1基因的脐血有核细胞对化疗药物有明显的抵抗性(P<0.01).通过腺病毒介导的MDR1基因转染脐血有核细胞能有效的抵抗化疗药物对其损伤,从而有望对临床肿瘤的大剂量化疗提供一种有效防护细胞损伤的方法,提高化疗效果.     

Missing Self-Induced Activation of NK Cells Combines with Non-Complement-Fixing Donor-Specific Antibodies to Accelerate Kidney Transplant Loss in Chronic Antibody-Mediated Rejection

Open in a separate window     

Serum cystatin C as an endogenous marker of renal function in patients with mild to moderate impairment of kidney function.

BACKGROUND: Estimation of the glomerular filtration rate (GFR) is essential for the evaluation of patients with chronic kidney disease (CKD). Recently, serum cystatin C was proposed as a new endogenous marker of GFR and in our study its diagnostic accuracy was compared with that of other markers of GFR. METHODS: In this study, 164 patients with CKD stages 2-3 (GFR 30-89 ml/min/1.73 m2), who had performed 51Cr-labelled ethylenediaminetetra-acetic acid clearance, were enrolled. In each patient, serum creatinine and serum cystatin C were determined. Creatinine clearance was calculated using the Cockcroft-Gault (C&G) and the modification of diet in renal disease (MDRD) formulas. RESULTS: The mean 51CrEDTA clearance was 57 ml/min/1.73 m2, the mean serum creatinine 149 micromol/l and the mean serum cystatin C 1.74 mg/l. We found significant correlation between 51CrEDTA clearance and serum creatinine (R = -0.666), serum cystatin C (R = -0.792), reciprocal of serum creatinine (R = 0.628), reciprocal of serum cystatin C (R = 0.753) and calculated creatinine clearance from the formulas C&G (R = 0.515) and MDRD formulas (R = 0.716). The receiver operating characteristic (ROC) curve analysis (cut-off for GFR 60 ml/min/1.73 m2) showed that serum cystatin C had a significantly higher diagnostic accuracy than serum creatinine (P = 0.04) and calculated creatinine clearance from the C&G formula (P < 0.0001), though only in female patients. No difference in diagnostic accuracy was found between serum cystatin C and creatinine clearance calculated from the MDRD formula. CONCLUSIONS: Our results indicate that serum cystatin C is a reliable marker of GFR in patients with mildly to moderately impaired kidney function and has a higher diagnostic accuracy than serum creatinine and calculated creatinine clearance from the C&G formula in female patients.     

How to Comply with the Food and Drug Administration's "Investigational Device Exemption (IDE)" Regulations Including an Application Form (Revised January 1982) Prepared by the ASAIO Standards Committee,

This is a revision of the IDE Application Form, "How to Comply with the Food and Drug Administration's New 'Investigational Device Exemption (IDE)' Regulations, Including an Application Form" originally published in Artificial Organs , Volume 4, Number 4, November 1980.
This publication updates the IDE application form to facilitate compliance with the amended investigational device exemption regulations now in effect.