The effect of antisera to thymosin alpha 1 on the course of autoimmune ovarian dysgenesis in neonatally thymectomized mice

Thymectomy at 3 days of age (Tx-3) in B6A female mice results in an autoimmune oophoritis that has only been successfully overcome by the transplant of an intact thymus or an injection of T cells. In Tx-3 mice levels of thymosin alpha 1 (TSN alpha 1), a potent thymic hormone involved in the development of helper T cells, was previously shown to be high after 7 days. By 60 days levels of TSN alpha 1 returned to levels found in intact mice. By this age ovarian dysgenesis was also complete and accompanied by high circulating levels of auto-oocyte antibody (AOA), estradiol-17 beta (E2) and testosterone (T). In the present study injections of antisera to TSN alpha 1 were given to Tx-3 mice in an attempt to decrease circulating TSN alpha 1 levels. We reasoned that this treatment should inhibit lymphocyte differentiation, and possibly in turn aid in overcoming the ovarian dysgenesis. After treatment of the Tx-3 mice, dysgenic ovaries persisted and high levels of AOA remained similar to the untreated Tx-3 mice. Levels of E2 and T, however, were returned to those found in intact mice. These results suggest that there is a sensitive balance between the thymus and the ovary that may not be related to changes in only a single thymic hormone.     

Aromatase activity in cultured ovaries from neonatally thymectomized mice with ovarian dysgenesis

The autoimmune oophoritis resulting from thymectomy at 3 days of age (Tx-3) in B6A female mice is characterized by dysgenic ovaries and circulating auto-antibodies against the oocyte (AOA). Dysgenesis of the ovaries starts around 24 days of age with a decline in numbers of the oocytes and follicles and is accompanied by lymphocytic infiltration. By 60 days of age the ovarian dysgenesis (OD) is complete with a preponderance of interstitial cells associated with elevated levels of testosterone (T). From 60-120 days of age the ovaries become progressively smaller in size and T levels rise. Since ovarian interstitial cells can produce T, assessment of aromatase activity was determined using cultured ovaries from 20-, 30-, 60-, 90- and 120-day-old mice. Similar or enhanced ability in aromatizing T to estradiol-17 beta (E2) was demonstrated by the ovaries from all Tx-3 mice compared with those from intact mice. At 30 and 60 days of age Tx-3 mice had increased circulating levels of E2 then the levels of E2 returned to those of intact mice at 90 and 120 days of age. The results indicate that the ovaries in Tx-3 mice may have an ability to aromatize T to E2 in culture, but apparently are not doing so at 90 and 120 days of age in situ. Further, ovaries of Tx-3 animals are able to aromatize T to E2 in the absence of organized follicular cells. These abnormal responses of ovarian hormones clearly demonstrate that the presence of AOA have a damaging effect on the endocrine activity as well as the morphology of the ovary.     

Murine pregnancy-associated modulations in lymphocyte reactivity to mitogens: identification of the cell populations affected

Lymphocytes from the thymus, spleen and inguinal lymph nodes of syngeneically pregnant and non-pregnant mice were compared in their responsiveness to polyclonal stimulation by mitogen. Pregnancy-associated changes in mitogen reactivity were detected, on a cell-per-cell basis, in thymocytes (increased) and spleen cells (decreased) but not in lymph node cells. The hyperreactivity of thymocytes during pregnancy correlated with physiological involution of the thymus occurring through the selective loss of relatively immature, non-mitogen-reactive, Lyt 1+2+ cells. The remaining cells were found largely to be mature Lyt 1+2- T cells with the capacity to respond to mitogenic stimulation. It is most likely the relative increase in the proportion of these Lyt 1+2- cells that causes the hyper-responsiveness of thymocytes to mitogens observed during pregnancy. On the other hand, while spleen cells from pregnant animals gave lower responses to mitogens than those from control virgin females, isolated splenic T cells from the two groups proved equally reactive to T cell mitogens. This supports the contention that at least some aspects of immunity during pregnancy are down-regulated by inhibitory cells within the non-T cell compartment. The results demonstrate the importance of identifying the reactive cell population in studies on changes in lymphocyte responsiveness in pregnancy.     

HPV16E7肽疫苗对人免疫重建荷人宫颈癌细胞株SiHa细胞SCID鼠免疫功能的影响

目的:观察HPV16E7多肽疫苗对人免疫重建荷人宫颈癌细胞株SiHa细胞严重联合免疫缺陷(SCID)鼠移植瘤的影响。方法:对SCID鼠腹腔注射人外周血淋巴细胞(PBL)后24小时,皮下接种HPV16阳性的人宫颈癌细胞株SiHa细胞,建立人免疫重建荷SiHa细胞的SCID鼠模型,当移植瘤最小体积达100mm3时,三次背部皮下接种疫苗,每次接种间隔二周,观察荷瘤鼠一般生物学特性,检测血浆中人IgG含量、外周血中人CD3+、CD4+、CD8+T细胞、脾重、肿瘤浸润淋巴细胞(TIL)以及脾细胞毒杀伤功能。实验设立空白组,仅为腹腔内注射PBL,疫苗成份仅为不完全佐剂(IFA);随机多肽组,为腹腔内注射PBL,皮下接种SiHa细胞,疫苗成份仅为随机多肽+T辅助多肽+IFA;T辅助多肽组,为腹腔内注射PBL,皮下接种SiHa细胞,疫苗成份仅为T辅助多肽+IFA;多肽治疗组,为腹腔内注射PBL,皮下接种SiHa细胞,疫苗成份为HPV16E711-20(YMLDLQ-PETT),HPV16E786-93(TLGIVCPI)+T辅助多肽+IFA。结果:各组SCID鼠血浆中人IgG水平,8周内随重建时间延长而升高(P<0.05),多肽治疗组达2957.85μg/m l,但各组间无显著性差异;外周血中均可检测到人CD3+、CD4+、CD8+T细胞,但各组间无显著性差异(P>0.05);与空白组比较,其他三组SCID鼠脾重均显著增加(P<0.05)。组织学观察到移植瘤局部TIL浸润及明显的出血和坏死,但经免疫组化未检测到人CD8+T细胞。与空白组比较,多肽治疗组、随机多肽组、辅助多肽组的脾细胞毒杀伤功能均显著降低(P<0.05)。结论:人免疫功能重建的荷SiHa细胞的SCID鼠体内重建的人免疫功能随荷瘤时间的延长受到抑制,多肽疫苗在一定程度上能减轻免疫抑制状态。     

Trafficking of syngeneic murine lymphokine activated killer T cells following intraperitoneal administration in normal and tumor bearing mice.

Nongenetically restricted T cells may be important host effector cells in women with ovarian cancer receiving intraperitoneal (ip) IL-2 therapy. We developed an in vitro technique to produce murine lymphokine-activated killer T cells. Murine splenocytes were cultured in the presence of 1000 U/ml IL-2 for 10 to 15 days. Phenotypical analysis showed 95% of total cells to express the pan T phenotype Thy 1.2 and no NK cell phenotypes by Day 7 in culture. These cells were labeled with 51Cr and their trafficking pattern after ip administration into normal and M5067 tumor bearing mice was examined. Various organs and tissues were collected at different timepoints and monitored for radioactivity. Within 4 hr., about 60% of the counts were associated with the bowel, peritoneum, and omentum of both normal and tumor bearing mice. About 15% of counts were associated with the blood, lung, kidney, spleen, and liver of both normal and tumor bearing mice.     

Characterization of hormone-dependent suppressor cells in the uterus of mated and pseudopregnant mice

The survival of the implanted "fetal allograft" has been attributed to the action of local decidua-associated suppressor cells. These suppressor cells are Fc-receptor positive small lymphocytic cells lacking T-cell markers which arise following implantation, are localized at the implantation site, and block the action of IL-2 that stimulates NK and T effector cells. Kinetic studies have demonstrated the occurrence of an earlier peak of suppressive activity occurring 2-3 days after mating prior to implantation. The cells associated with pre-implantation suppression differs significantly from post-implantation suppressor cells. Velocity sedimentation studies show that early suppression is associated with large cells sedimenting at a modal velocity of 6-7 mm/hr. Suppressive activity from cells of similar size is also present in the uterine lining of hormonally-treated, pseudopregnant mice. In addition, suppressor cells can be demonstrated in the non-pregnant uterus at the time of estrus. These observations suggest suppressor cell activity may be hormonally regulated. The suppressor cell(s) in pseudopregnant mice bears Lyt 2.1 and Thy 1.2 surface antigens and suppressor(s) present in the pregnant animals bears Lyt 2.1 and Lyt 1.1. Although the suppressor cell was large, it did not appear to be a macrophage because it was resistant to antibodies to Mac-1 and FcR cell surface markers but susceptible to anti-T cell reagents. Furthermore, suppression was not mediated by a soluble factor that has been associated with the small lymphocytic suppressor. Thus, the suppressor activity present in the pre-implantation uterine lining appears to differ significantly from the suppressor cell activity found after implantation. The possible role of a hormone-activated suppressor T cell in the success of the pregnancy is discussed.     

Significance of early pregnancy factor (EPF) on reproductive immunology

We studied the significance of Early Pregnancy Factor (EPF), which appeared very soon after fertilization, on reproductive immunology. The results were as follows: EPF inhibited dose-dependently mixed lymphocyte reaction (MLR). EPF suppressed recognition phase of MLR. And EPF induced nonadherent, Thy 1.2+, Lyt 1-, Lyt 2+ regulator cells as suppressor T cells. Immunosuppressive effect of EPF was not restricted by major histocompatibility complex (MHC). Administration of anti EPF antibody in period before implantation of mice decreased the number of implantations and neonates. It suggested that EPF was important in this stage of reproduction.     

Change in leukocyte count in peripheral blood of nude mouse bearing transplanted gynecological malignant tumors

The cells of 25 cell lines derived from malignant tumors in the gynecological field were transplanted subcutaneously into nude mice. The growth of the grafted tumors and changes in the number of leukocytes and leukocyte hemograms of the peripheral blood were examined, and the following conclusions were obtained. (1) Leukocytosis (more than 20,000/mm3) was observed in only 2 cell lines (ovarian poorly differentiated clear cell carcinoma (HUOCA-II) and ovarian anaplastic carcinoma (HTBOA] among 24 cell lines transplanted (SKN cell line could not be transplanted). (2) A positive correlation was found between the growth of the grafted tumor and leukocytosis. The leukocyte hemogram revealed that 98% of the cells were granulocytes (neutrophils). (3) After resection of the tumor, the number of leukocytes and the leukocyte hemograms returned to normal. (4) No metastasis to the bone marrow or inflammation was found in the nude mice bearing tumors and showing leukocytosis. Further when bone marrow cells of mouse and human were cultured in soft agar containing conditioned medium of HUOCA-II and HTBOA cells colonies of leukocytes were observed in which 98% were granulocytes (neutrophils). It is concluded from the results above that the marked granulocytosis found in the tumor-bearing mice was caused in part by granulocyte-colony stimulating factor (G-CSF).     

Premature menopause: monoclonal antibody defined T lymphocyte abnormalities and antiovarian antibodies

The presence of other organ-specific autoimmune disorders in some patients with premature menopause has supported the concept of an autoimmune etiology. The authors analyzed the peripheral blood of 23 women with the diagnosis of premature menopause to detect the presence of monoclonal antibody-defined T-lymphocyte abnormalities and/or antiovarian antibodies. All subjects were less than 40 years of age with the duration of menopause ranging from less than 1 year to 11 years at the time of study. Thirty-five percent of the subjects had an elevated percentage of Ia+ (Dr-activated) T cells using monoclonal antibody L243. The percent T4 (helper) T8 (suppressor/cytotoxic) T cells and T4/T8 ratio were normal in the study group. Four subjects (approximately 17%) had elevated percentages of the age-related 3G5+ T cell subset. Two of the subjects with increased 3G5+ T cells also exhibited increased Ia+ T cells. Antiovarian steroid cell antibodies and antiadrenal cortical antibodies were present in approximately 9% of subjects. Anti-islet cell antibodies were not present. Thyroid antimicrosomal antibodies were present in 17% of subjects. Study subjects exhibited immunologic abnormalities that the authors hypothesize may play a role in the development of premature menopause in a larger percentage of patients than was previously suspected.     

The effect of intraperitoneal administration of OK-432 with recombinant interleukin-2 to patients with peritonitis carcinomatosa of recurrent gynecological cancer and changes in ascitic lymphocyte subsets

The effect of intraperitoneal administration of OK-432 followed by intraperitoneal instillation of recombinant interleukin-2(rIL-2) was examined in tumor bearing animals and in four recurrent gynecological cancer patients with peritonitis carcinomatosa which had been resistant to chemotherapeutic drugs. Seven days after intraperitoneal inoculation of tumor cells (10(6) cells/body of MH134 hepatoma into C3H/He mice and 10(5) cells/body of Meth-A fibrosarcoma into BALB/C mice, respectively), OK-432 was administered intraperitoneally. Two days later, a 14 day course of daily intraperitoneal instillation of rIL-2 followed. The survival time for animals treated with OK-432 combined with rIL-2 was significantly prolonged. Three of the 8 C3H/He mice and one of the 8 BALB/C mice in this group survived more than 150 days without forming ascites. However, the group treated by rIL-2 alone did not survive for more than 40 days. The group treated by OK-432 alone as well as the untreated group did not survive for more than 20 days. Ascitic fluid disappeared clinically in two of four cases and decreased in the rest. Ascitic cancer cells disappeared in one case and decreased in three cases. The serum CA125 level declined significantly in all cases. The surface markers of ascitic lymphocytes were analyzed by flow cytometry on day 8. The CD4+ subset accounted for 70-90% whereas the CD8+ subset accounted for only 7-17%. In three cases in which two color analysis was performed, the CD4+, CD29+ helper inducer T cell was dominant. We could conclude that LAK cells were not the main effector cells.     

Conversion of circulating estrone sulfate to 17beta-estradiol by ovarian tumor tissue: a possible mechanism behind elevated circulating concentrations of 17beta-estradiol in postmenopausal women with ovarian tumors.

AIM: Elevated serum levels of 17beta-estradiol (E2) are frequently found in postmenopausal women with ovarian tumors not classified as estrogen-producing. Conversion of circulating estrone sulfate (E1S) to E2 is one alternative way of E2 formation in target tissues in postmenopausal women. Our aim was to find out if conversion of circulating E1S to E2 by the tumor tissue could be a reason for elevated serum E2 levels in postmenopausal women with 'non-estrogen-producing' ovarian tumors. METHOD: Serum E2 was measured in 12 postmenopausal women with 'non-estrogen-producing' ovarian tumors (nine benign, three malignant). Total hydrolysis of and [3H]E2 formation from [3H]E1S by the tumor tissue homogenates was studied in vitro. RESULTS: Serum E2 showed significant positive correlations with total hydrolysis of and [3H]E2 formation from [3H]E1S in the total material as well as in the benign tumor subgroup. [3H]E2 formation was the most important independent variable. CONCLUSION: Conversion of circulating E1S to E2 by the tumor tissue could be one important reason for elevated S-E2 levels in postmenopausal women with 'non-estrogen-producing' ovarian tumors.     

CD137L基因转染对小鼠体内卵巢癌细胞生长的影响及其免疫调节研究

目的:初步探讨转染CD137L基因的小鼠卵巢癌细胞株OVHM细胞体内的致瘤性及其对免疫功能的影响。方法:以脂质体为载体将重组质粒pcDNA3.1和pcD-NA3.1/CD137LL分别导入OVHM细胞中,经HygroB筛选,用RT-PCR法检测目的基因的表达,筛选建立高表达的细胞株。计数法绘制体外培养细胞生长曲线。将转染前后的肿瘤细胞,分别接种于B6C3F1小鼠的腹部皮下观察其致瘤性。LDH法测定其CTL细胞的杀伤活性。结果:与野生型OVHM细胞相比,所建立的OVHM/CD137L细胞株可高表达CD137L。转染前后肿瘤细胞的体外生长速度无明显变化。OVHM/CD137L细胞在小鼠体内的致瘤性较OVHM/pcDNA3.1细胞和野生型OVHM细胞明显降低。接种OVHM/CD137L细胞小鼠的CTL细胞杀伤活性明显增强。结论:转染CD137L基因后OVHM细胞在小鼠体内的致瘤性显著降低,且诱导产生了明显的抗肿瘤免疫。CD137L有望成为卵巢癌基因治疗的候选基因。     

The assessment of dendritic cells cultured from peritoneal fluid macrophages: first report and new perspectives in the treatment of endometriosis

Dendritic cells represent discrete leukocyte subpopulation of specialist or "professional" antigen-presenting cells (APC). They play a crucial role in the activation of naive T cells "in vivo" They have monocyte/macrophages origin. There are no data in literature on the presence of dendritic cells derived from peritoneal fluid monocytes/macrophages. In our study we tried to culture PF macrophages from patients who undergone surgery so that to obtain dendritic cells. PF was aspirated during laparoscopy from patients with endometriosis, unexplained infertility or benign noninflammatory ovarian tumor. Peritoneal macrophages were isolated using adherence method then were cultured and stimulated with GM-CSF and IL-4. Phenotype of cultured cell was estimated using flow cytometry after incubation with monoclonal antibodies CD45/14, CD 40/HLA-DR, CD28/3, CD3/40L, CD25/5 and CD69/HLA-DR. Morphology of cultured cells was confirmed microscopically after May-Grunvald-Giemsa staining. PF leukocytes concentration varied from 1.2 x 10(6) cells/mm3 to 22.6 x 10(6) cells/mm3. Cultured monocytes/macrophages from PF had morphology typical for dendritic cells. We also found that only dendritic cells from patients with endometriosis had higher expression HLA-DR antigen (93.6% of cells) and low expression of CD40 (2.7% of cells) on their surface in comparison to reference group. It is worthy to notify that dendritic cells from patients with endometriosis expressed also CD25 antigen characteristic for T leukocytes. To our data it is the first report in literature on dendritic cells obtained from PF macrophages.     

卵巢肿瘤原位移植动物模型的建立

目的:建立能够模拟临床肿瘤发展过程的卵巢癌动物模型。方法:将处于对数生长期的人卵巢癌细胞株SKOV3(2×106个/只)、A2780多水平细胞数(2×106个/只、5×106个/只、1×107个/只、1.5×107个/只和2×107个/只)裸鼠皮下注射,A2780(1.5×107个/只)腹腔注射。取SKOV3第3代连续传代皮下瘤组织进行裸鼠右侧卵巢包膜下移植。结果:卵巢癌细胞株SKOV3皮下注射、组织块原位移植成瘤率均100%。原位移植卵巢癌裸鼠6～8周时剖腹探查见局部形成较大包块,与周围组织发生粘连,腹腔暗红色血性积液伴脏器广泛性转移,淋巴结转移。卵巢癌A2780细胞株多水平细胞数皮下注射均未成瘤;腹腔注射8周时剖腹探查见腹腔内生成2～3 mL无色澄清腹水,双侧卵巢增大,脾脏明显增大,未能发现任何腹腔瘤块。结论:成功建立了临床转移模式的SKOV3卵巢癌肿瘤细胞原位移植动物模型;卵巢癌A2780细胞接种未能成瘤,可能是因其对宿主体内非特异性免疫反应较敏感。     

C57BL/6小鼠上皮性卵巢癌模型的建立及其生物学特性

目的在免疫功能正常的C57BL/6小鼠体内建立上皮性卵巢癌腹腔转移瘤模型及皮下瘤模型,为卵巢癌的诊断、治疗及预防的相关研究提供基础。方法体外培养近交系C57BL/6小鼠卵巢上皮低分化腺癌细胞株ID-8,将对数生长期的ID-8细胞按1×10^7、5×10^6、1×10^6和1×10^5个细胞/只的剂量,分别接种于6～8周雌性SPF级C57BL/6小鼠腹腔及左侧肩部皮下,共8组,每组6只。观察腹腔瘤及皮下瘤的成瘤时间、成瘤率、腹水、腹腔肿瘤转移情况及小鼠生存期;处死小鼠时留取主要脏器、腹腔肿瘤及皮下肿瘤标本,行病理学检查。另外6只小鼠接种5×10^6个ID-8细胞,分别在4、8、16周后处死进行系统解剖,做常规病理学检查。结果将不同数量的ID-8细胞接种C57BL/6小鼠腹腔及皮下后,成瘤率均为100%,腹腔瘤模型组：腹腔注射1×10^5,1×10^6,5×10^6,1×10^7个ID-8细胞,平均生存时间分别为（141±6.7）d、（122.8±4.5）d、（83.4±7.2）d和（74.4±4.5）d,随着肿瘤细胞接种负荷增加,动物生存期明显缩短（P〈0.05）。皮下瘤组：1×10^7细胞组和5×10^6细胞组,1周左右成瘤;1×10^6细胞组,3周左右成瘤;1×10^5细胞组,6周左右成瘤。随着肿瘤接种负荷的增加,肿瘤直径和体积明显增大（P〈0.05）。结论 C57BL/6小鼠腹腔瘤模型类似人类Ⅲ、Ⅳ期卵巢上皮癌患者的临床特点。皮下瘤模型更易于观察免疫治疗或药物治疗的疗效。在免疫功能正常的C57BL/6小鼠建立的ID-8细胞卵巢癌肿瘤模型,是适合于卵巢癌分子和免疫治疗研究的模型。     

ZP3 peptides administered orally suppress murine experimental autoimmune ovarian disease

Experimental autoimmune ovarian disease (AOD) is a T cell-mediated chronic inflammatory disease that may lead to premature ovarian failure. Autoimmune disease can be suppressed by oral administration of autoantigens leading to tolerance. One of the major mechanisms of oral tolerance is induction of regulatory CD4+ T cells that can mediate active suppression by producing immunomodulatory cytokines. However, the role of oral tolerance as a treatment for experimental AOD has received little attention. Therefore, the purpose of this study was to examine the conditions necessary to produce oral tolerance in experimental AOD in B6AF1 female mice. In this study, mice received different doses of peptides of the mouse zona pellucida 3 (pZP3) via gastric intubation for 7 times. After 4 times of oral administration, AOD was induced by immunization with pZP3. The optimal tolerating regimen for oral administration of pZP3 in mice was 10 microg, which decreased morbidity of oophoritis compared to the control group. In this moderate-dose therapeutic group (MD), alterations in the estrous cycle were normalized and CD4+ T cells that were CD25+ increased while those that were CD25- decreased. The severity of autoimmune oophoritis and the titer of ZP autoantibodies were also significantly reduced. These findings suggest that oral administration of pZP3 may be successfully used as an oral tolerance strategy for suppression of AOD.     

多西紫杉醇对人卵巢癌SKOV3裸鼠移植瘤的抑制作用及其与顺铂、曲妥珠单抗联合作用的探讨

目的:观察多西紫杉醇(Docetaxel,又名多西他赛,商品名泰素帝)对人卵巢癌SKOV3裸鼠移植瘤的抑制作用,并观察其与顺铂和(或)曲妥珠单抗(Trastuzumab,商品名赫赛汀)联用的抑制作用差异。方法:将SKOV3细胞悬液接种于裸鼠右侧腋窝皮下,成功建立人卵巢癌SKOV3裸鼠移植瘤模型后,随机将荷瘤裸鼠分为8组,即顺铂组、赫赛汀组、泰素帝组、顺铂+赫赛汀组、顺铂+泰素帝组、赫赛汀+泰素帝组、顺铂+赫赛汀+泰素帝组和对照组,每组4只。顺铂按3 mg/kg、赫赛汀按30 mg/kg、泰素帝按5 mg/kg每周1次经尾静脉给药,连续6周,每周测量1次小鼠肿瘤的长、短径及小鼠体质量,用药结束后1周处死小鼠,计算抑瘤率,肿瘤组织HE染色观察细胞凋亡,TUNEL技术检测凋亡指数,免疫组织化学方法检测肿瘤组织中Ki-67的表达情况。结果:(1)泰素帝能明显抑制人卵巢癌SKOV3细胞裸小鼠移植瘤的生长,且分别与顺铂、赫赛汀联合后对肿瘤生长的抑制作用更明显,顺铂+赫赛汀+泰素帝组对肿瘤生长的抑制作用最明显;(2)泰素帝单药组肿瘤细胞凋亡指数较对照组显著升高,且分别与顺铂、赫赛汀联合后肿瘤细胞凋亡指数更高,顺铂+赫赛汀+泰素帝组肿瘤细胞凋亡指数最高;(3)泰素帝单药组肿瘤组织中Ki-67的表达率较对照组显著下降,且分别与顺铂、赫赛汀联合后肿瘤组织中Ki-67的表达率下降更明显,而顺铂+赫赛汀+泰素帝三联组肿瘤组织中Ki-67的表达率下降最明显。结论:(1)泰素帝能够有效抑制人卵巢癌SKOV3裸小鼠移植瘤的生长,其可能的机制是通过下调Ki-67等的表达而抑制肿瘤的增殖活性,并诱导肿瘤细胞凋亡;(2)泰素帝分别与顺铂、赫赛汀间具有协同作用,而泰素帝、顺铂、赫赛汀三药联合应用能更有效地抑制肿瘤生长。     

Implications of circulating autoantibodies and peripheral blood lymphocyte subsets for the genesis of premature ovarian failure

Several kinds of circulating autoantibodies and peripheral blood lymphocyte subsets were studied in 20 patients with secondary amenorrhea manifesting hormonal and clinical features of premature ovarian failure (POF). More than one kind of autoantibody was detected in 14 patients (70%). Seven patients (35%) had anti-thyroglobulin antibody, 6 (30%) had anti-parietal cell antibody, 8 (40%) had anti-nuclear antibody, and one patient with chronic thyroiditis had anti-TSH receptor antibody. Anti-adrenal cortex antibody and RA test were negative in all patients. Two patients had clinically evident autoimmune disease; one had myasthenia gravis and the other had chronic thyroiditis. Examination of peripheral blood lymphocyte subsets of 19 patients by flow cytometry revealed an increase in the percentage of OKT3+ cells and OKT4+ cells and a decrease in OKT8+ cells in POF patients compared with age-matched controls, but these differences were not significant. An increase in the OKT4/OKT8 ratio was, however, significant. It has been suggested that an autoimmune mechanism may participate in the genesis of POF, at least in patients with autoimmune diseases; however, the findings in this study support the hypothesis that some pure POF may also be caused by an autoimmune process resulting from a subclinical imbalance in the immunoregulatory system before manifestation of the autoimmune disease.     

Effects of lymphokine-activated killer cells and interleukin-2 on the ascites formation and the survival time of nude mice bearing human ovarian cancer cells

Effect of intraperitoneal instillations of interleukin-2 (IL-2) and/or lymphokine-activated killer (LAK) cells on the ascites formation and the survival time was examined by using a nude mice model with malignant ascites by intraperitoneal inoculation of human ovarian cancer cells (HRA) derived from ascites of a patient with serous cystadenocarcinoma of the ovary. On 28 days after tumor inoculation, all nude mice in both untreated and spleen cells only treated groups formed ascites. Two of 10 nude mice treated with IL-2 only after tumor inoculation survived without forming ascites during the experimental period. On the other hand, all nude mice treated with LAK cells only formed ascites by 14 days after tumor inoculation. When LAK cells and IL-2 were combined 5 of 10 mice survived without forming ascites during the experimental period. The survival time of the IL-2 only treated group was significantly prolonged, compared to that of medium only, spleen cells only and LAK cells only treated groups. When administration of LAK cells was followed by IL-2, the survival time was further prolonged.     

In vitro induction of tumor-specific human lymphocyte antigen class I-restricted CD8 cytotoxic T lymphocytes by ovarian tumor antigen-pulsed autologous dendritic cells from patients with advanced ovarian cancer

OBJECTIVE: The purpose of this study was to evaluate the potential of dendritic cells pulsed with whole-tumor extracts derived from autologous ovarian cancer cells in eliciting a tumor-specific cytotoxic T-cell response in vitro from patients with advanced ovarian cancer. STUDY DESIGN: CD8(+) T lymphocytes stimulated in vitro with autologous ovarian tumor lysate-pulsed dendritic cells were tested for their ability to induce a human leukocyte antigen class I-restricted cytotoxic T-lymphocyte response able to specifically kill autologous tumor cells in standard 6-hour chromium 51 cytotoxicity assays. In addition, to correlate cytotoxic activity by cytotoxic T-lymphocytes with a particular lymphoid subset, 2-color flow cytometric analysis of intracellular cytokine expression (interferon gamma and interleukin 4) at the single-cell level was performed. RESULTS: Cytotoxic T lymphocytes specific for autologous ovarian tumor cells were elicited from 3 patients with advanced ovarian cancer. Although cytotoxic T-lymphocyte populations expressed strong cytolytic activity against autologous tumor cells, they did not lyse concanavalin A-stimulated autologous lymphocytes or autologous Epstein-Barr virus-transformed lymphoblastoid cell lines and showed negligible cytotoxicity against the natural killer cell-sensitive cell line K-562. Cytotoxic effect against the autologous tumor cells was inhibited by an anti-human leukocyte antigen class I monoclonal antibody (W6/32). It is interesting that CD8(+) cytotoxic T lymphocytes expressed variable levels of CD56, a marker that may be associated with high cytotoxic activity. Finally, most of the tumor-specific CD8(+) T cells exhibited a T(H)1 cytokine bias, and a high percentage of interferon gamma expressors among cytotoxic T lymphocytes was correlated with higher cytotoxic activity. CONCLUSION: These data show that tumor lysate-pulsed dendritic cells can consistently induce in vitro specific CD8(+) cytotoxic T lymphocytes able to kill autologous tumor cells from patients with advanced stage ovarian cancer. This novel approach may have important implications for the treatment of residual or resistant disease with active or adoptive immunotherapy after standard surgical and cytotoxic treatment.