Peripheral tolerance in transgenic mice expressing class I MHC L(d) only on cardiac cells

BACKGROUND: The fate of autoreactive T cells exposed to extrathymic self-antigen is examined in a double transgenic (DTG) mouse [(L(d+) cardiacx2C)F1], where cardiac myocytes alone express L(d) and T cells express an antigen receptor (2C TCR) against L(d). METHODS: Na?ve cardiac L(d+) single transgenic (STG) mice (before breeding with 2C) and DTG mice were examined for evidence of autoimmunity. The L(d+) STG hearts were then transplanted to syngeneic L(d-) wild type C57BL/6 to evaluate the heart's immunogenicity. L(d+) skin grafts were transplanted to non-transgenic B6, transgenic 2C, STG, and DTG mice. Phenotype analysis of peripheral 1B2+(identifies 2C T cells), CD4+, and CD8+ T cells was performed by FACS. In vitro MLC and CTL, with and without the addition of IL-2 and suppressor cell assays, were evaluated. RESULTS: Neither STG nor DTG hearts developed any evidence of autoimmunity by histology. In contrast, B6 mice rejected the L(d+) STG heart in 17+/-9.7 days (P<0.01), while a syngeneic B6 heart transplant was accepted indefinitely. Survival of L(d+) skin grafts was prolonged in both STG and DTG mice. FACS quantitation revealed that while there was no deletion of peripheral 2C cells in the DTG, these 2C T cells did have a significantly reduced proliferative and cytotoxic response to H-2L(d). Restoration of the proliferative and cytotoxic response of the DTG cells by the addition of IL-2 was consistent with a state of anergy. CONCLUSIONS: These findings suggest that the expression of extrathymic class I MHC expression alone did not trigger autoimmune reactions but that the T cells can be rendered anergic to the specific 'self' antigen.     

Split tolerance in chimeric GVH mice

Abstract The i.v. inoculation of parental spleen cells into unirradiated adult F1 hybrid mice results in a graft-versus-host reaction (GVHR). In the strain combination B10D2±(B10.BRx B10.D2) F1, this reaction is associated with thymic injury and transient but profound cellular immune deficiency. We further analysed the immune status of these mice 60 days after GVHR induction. Phenotypic studies of spleen cells showed that these mice were re-populated with parental lymphocytes resulting in a high degree of chimerism (85%). At this time, the mice looked healthy and recovered a normal cytotoxic T cell response (CTL) against allogeneic cells. GVH chimeric splenocytes were unresponsive against F 1 hybrid cells in mixed lymphocyte culture (MLC), but exhibited anti-F1 CTL reactivity. We also analysed the anti-F 1 reactivity of these mice in vivo. GVH chimeric splenocytes were unable to induce GVHR after injection into a new F1 hybrid and F1 GVH mice specifically rejected F1 bone marrow (BM) cells after lethal irradiation. Grafting a neonatal parental thymus prevented the rejection of F1 BM cells and restored CTL alloreactivity. It is concluded that the chimeric state induced by GVHR is associated with a split tolerance and that a radiosensitive thymic-dependent mechanism is involved in maintaining self-tolerance in these mice.     

Drug-induced tolerance to allografts in mice. VI. Tolerance induction in H-2-haplotype-identical strain combinations in mice

When C3H/HeN (C3H) mice were primed with viable AKR/J (AKR) spleen cells and treated with cyclophosphamide (CP) two days later, a profound tolerance to AKR skin grafts was induced. This tolerance was induced also in other combinations disparate only at minor histocompatibility (H) antigens (AKR-C3H and BALB/c[BALB]-DBA/2[DBA]). In C3H mice made tolerant to AKR, delayed foot-pad reaction (DFR), cytotoxic lymphocytes (CTL), and cytotoxic antibodies (CTAb) against AKR spleen cells were abrogated completely. Tolerance to AKR mice was also observed in C3H mice primed with viable AKR and C57BL/6 (B6) spleen cells and treated with CP, but tolerance to B6 was not induced because a cell population responsible for DFR and CTL against B6 H-2 antigens remained after tolerance induction. These results suggest that there is a lymphocyte population responsible for DFR and CTL against antigens allogeneic at both major and minor H that is less proliferative than the population responsible for DFR and CTL against minor H antigens.     

自发性炎性疾病双转基因小鼠的研究

目的 通过建立强直性脊柱炎HLA－B2704和hβ2m双转基因动物模型,证实HLA－B2704和hβ2m基因在双转基因小鼠自发性炎性疾病发病过程中的作用,为研究B27相关性疾病的病因学、预防和治疗提供有力的工具。方法 将HLA－B2704和hβ2m转基因阳性小鼠交配后出生的子代,应用PCR、斑点杂交、Southern杂交、RT－PCR、流式细胞术和免疫组织化学等方法进行筛选鉴定和表达检测。同时隔日观察小鼠的发病情况,发病鼠行HE染色观察病理变化。结果 有8只高拷贝双转基因小鼠2周左右出现皮肤病变,关节炎和趾甲变化等自发性炎性疾病表现,正常小鼠、B27单转基因小鼠和HLA－B27／hβ2m双转基因小鼠流式细胞计的检测结果分别为0．63％、7．87％、35．87％,双转基因小鼠的细胞膜表面可见HLA－B2704高表达,而在B27单转基因小鼠表达不明显。结论 HLA－B2704重链可诱发转基因小鼠发生自发性炎性疾病,hβ2m可与B27形成稳定的复合体,从而稳定和增强HLA－B2704在细胞膜表面的表达。     

Early castration reduces prostatic carcinogenesis in transgenic mice

Objectives. To test the hypothesis that transgenic mouse models of prostate cancer could be useful for testing chemoprevention strategies by evaluating the effects of early castration on prostate carcinogenesis in TRAMP mice. Human prostate cancer, unlike other cancers, requires androgens for oncogenesis yet acquires partial androgen independence in the castrated milieu. This paradigm is the basis for an ongoing clinical trial using selective androgen deprivation for prostate cancer chemoprevention. However, preclinical correlates for hormonal prevention or other chemoprevention strategies of prostate cancer have not previously been demonstrated in autochthonous models of prostate carcinogenesis.Methods. Magnetic resonance imaging was used to longitudinally measure prostate growth in castrated and noncastrated TRAMP mice, and mice were prospectively examined for the onset of advanced, palpable prostate cancer. Modulation of androgen-responsive oncogene expression, as well as oncogene expression in refractory cancers, was evaluated by Western blot.Results. Early castration significantly reduced prostate tumor growth as measured by magnetic resonance imaging and improved cancer-free survival. Prevention of prostate cancer development in these mice was associated with durable suppression of androgen-responsive oncogene expression (T-antigen expression not detectable by Western blot); prostate cancers refractory to the hormonal prevention strategy demonstrated androgen-independent oncogene expression.Conclusions. These findings suggest that carcinogenesis related to androgen-responsive oncogene expression can be prevented in some cases by hormonal manipulation and that transgenic TRAMP mice are useful for the preclinical evaluation of hormonal and possibly other strategies of prostate cancer chemoprevention.     

Use of transgenic and knockout strategies in mice

Over 2 decades ago mouse models were generated with an exogenous gene integrated into its genome to create the first transgenic mice. Since that time, new methods have been developed and old methods improved, allowing investigators more flexibility to answer important questions about physiology and gene function. Transgenic and knockout technology have been particularly useful in the kidney as various transgenic mouse models have been successfully generated to gain a better understanding of renal physiology at the gene level. Now with the sequencing of mammalian genomes at the forefront of science, the need for transgenic technology to understand gene function in the context of the whole animal will become increasingly more important. Therefore, this article focuses on some of the strategies that can be used when generating transgenic mouse models.     

Complete diabetes protection despite delayed thymic tolerance in NOD8.3 TCR transgenic mice due to antigen-induced extrathymic deletion of T cells

Prevention of autoimmunity requires the elimination of self-reactive T cells during their development in the thymus and maturation in the periphery. Transgenic NOD mice that overexpress islet-specific glucose 6 phosphatase catalytic subunit-related protein (IGRP) in antigen-presenting cells (NOD-IGRP mice) have no IGRP-specific T cells. To study the relative contribution of central and peripheral tolerance mechanisms to deletion of antigen-specific T cells, we crossed NOD-IGRP mice to highly diabetogenic IGRP206-214 T-cell receptor transgenic mice (NOD8.3 mice) and studied the frequency and function of IGRP-specific T cells in the thymus and periphery. Peripheral tolerance was extremely efficient and completely protected NOD-IGRP/NOD8.3 mice from diabetes. Peripheral tolerance was characterized by activation of T cells in peripheral lymphoid tissue where IGRP was expressed followed by activation-induced cell death. Thymectomy showed that thymic output of IGRP-specific transgenic T cells compensated for peripheral deletion to maintain peripheral T-cell numbers. Central tolerance was undetectable until 10 weeks and complete by 15 weeks. These in vivo data indicate that peripheral tolerance alone can protect NOD8.3 mice from autoimmune diabetes and that profound changes in T-cell repertoire can follow subtle changes in thymic antigen presentation.     

Type I diabetes manifested solely by 2-h oral glucose tolerance test criteria

The clinical presentation of type 1 diabetes usually involves symptoms such as polyuria and polydipsia. However, investigators in the Diabetes Prevention Trial of Type 1 Diabetes (DPT-1) have detected a group of subjects with type 1 diabetes who have a different phenotype. These subjects are asymptomatic, have normal (<6.1 mmol/l) (group A) or impaired (6.1- <7.0 mmol/l) (group B) fasting glucose, but have 2-h glucose values >11.1 mmol/l on their oral glucose tolerance tests (OGTT). Of the 585 OGTTs performed on islet cell antibody (ICA)-positive relatives with insulin autoantibodies (IAA) or low first-phase insulin response (FPIR), normal glucose tolerance (NGT) was found in 427 subjects; impaired glucose tolerance (IGT) was found in 87 subjects, and diabetes was found by 2-h OGTT criteria alone in 61 subjects. Despite marked differences in 2-h glucose values (NGT 5.8 +/- 1.1 mmol/l, IGT 8.9 +/- 0.9 mmol/l, and group A 13.5 +/- 2.5 mmol/l), there were no significant differences in fasting glucose values among NGT (4.8 +/- 0.5 mmol/l), IGT (5.03 +/- 0.5 mmol/l), and group A (4.99 +/- 0.7 mmol/l) categories. Mean FPIR was higher in subjects with NGT compared with subjects with IGT and subjects diagnosed by 2-h OGTT criteria alone. However, the correlation between FPIR and 2-h glucose value was low (r2 = 0.114). Multivariate analysis demonstrated that additional independent variables provide smaller contributions to the 2-h glucose value. In conclusion, there are asymptomatic type 1 diabetic subjects whose diabetes was diagnosed by the 2-h criteria on OGTT alone. Despite the importance of beta-cell dysfunction in the pathogenesis of type I diabetes, factors other than impaired FPIR must also contribute to postprandial glucose tolerance in these subjects.     

大豆异黄酮在不同的雌激素环境下对MMTV-erbB-2转基因小鼠的作用

目的 观察大豆异黄酮在不同的雌激素环境下,对MMTV-erbB-2转基因小鼠的乳腺肿瘤发生及发展的影响.方法 选取鼠龄5周的健康MMTV-erbB-2转基因雌性小鼠150只,分为对照组、低雌激素组1、低雌激素组2、高雌激素组1和高雌激素组2五组,观察各组小鼠乳腺癌的发病率和潜伏期,并采用免疫组织化学染色SP法检测各组小鼠乳腺组织中雌激素受体(ER)、孕激素受体(PR)、增殖细胞核抗原(PCNA)的表达.结果 对照组、低雌激素组1、低雌激素组2、高雌激素组1和高雌激素组2的乳腺癌发病率分别为73.3%、96.7%、30.3%、40.0%和83.3%,对照组与高雌激素组2、低雌激素组1与高雌激素组2发病率比较差异无统计学意义(P＞0.05),其他各组间比较差异有统计学意义(P＜0.05).各组小鼠乳腺癌平均潜伏期比较差异无统计学意义(P＞0.05).低雌激素组1的TEB数量与其他各组比较差异有统计学意义(P＜0.05).erbB-2表达在各组之间表达差异无统计学意义(P＞0.05).低雌激素组1、低雌激素组2、高雌激素组1和高雌激素组2与对照组乳腺ER和PR的表达比较差异无统计学意义(P＞0.05).低雌激素组1实验小鼠发生肿瘤的乳腺组织内PCNA的表达明显高于其他各组(P＜0.05).对照组与高雌激素组2,低雌激素组1与高雌激素组2比较差异有统计学意义(P＜0.05),其他各组间比较差异有统计学意义(P＜0.05).结论 大豆异黄酮在不同的雌激素环境下,对MMTV-erbB-2转基因小鼠乳腺肿瘤的发生及发展起不同的作用.低雌激素环境下大豆异黄酮可促进乳腺肿瘤的发生及发展,高雌激素环境下大豆异黄酮可抑制乳腺肿瘤的发生及发展.

Abstract：

Objective To investigate the effect of the soybean isoflavones at different estrogen environments on the pathogenesis of breast cancer in MMTV-erbB-2 transgenic mice. Methods 150 fiveweek-old MMTV-erbB-2 transgenic female mice were phosen and divided into five groups; control group,low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2. The incidence and latent period of breast cancer were observed, and the expression of estrogen receptor (ER) ,progesterone receptor (PR), and proliferating cell nuclear antigen (PCNA) proteins was detected by using immunohistochemistry SP method. Results The incidence of breast cancer in control group, low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2 was 73. 3% , 96. 7% ,30. 3% , 40. 0% and 83. 3% , respectively, with the difference being not significant between control group and high estrogen group 2, and between low estrogen group 1 and high estrogen group 2 (P ＞ 0. 05 ) , but with the difference being significant among the other groups (P ＜ 0. 05 ). There was significant difference in the average latent period of breast cancer among all groups (P ＞ 0. 05 ). There was significant difference in the number of TEB between low-estrogen group 1 and other groups (P ＜ 0.05). There was significant difference in the erbB-2 expression among the groups ( P ＞ 0. 05 ). There was no significant difference in the expression of breast ER and PR between control group and other groups ( P ＞ 0. 05 ). The PCNA expression in breast tumor tissue in low-estrogen group 1 was significantly higher than other groups (P ＜0. 05) , and there was significant difference in the PCNA expression between control group and high estrogen group 2, between low estrogen group 1 and high estrogen group 2 (P ＜ 0. 05). Conclusion The soybean isoflavones at different estrogen environments play different roles in the occurrence and development of MMTV-erbb-2 transgenic mouse mammary tumor. In the context of low estrogen, soybean isoflavones could even promote breast cancer formation and development. In the context of high estrogen, soybean isoflavones could inhibit breast cancer and development.     

大豆异黄酮在不同的雌激素环境下对MMTV-erbB-2转基因小鼠的作用

Objective To investigate the effect of the soybean isoflavones at different estrogen environments on the pathogenesis of breast cancer in MMTV-erbB-2 transgenic mice. Methods 150 fiveweek-old MMTV-erbB-2 transgenic female mice were phosen and divided into five groups; control group,low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2. The incidence and latent period of breast cancer were observed, and the expression of estrogen receptor (ER) ,progesterone receptor (PR), and proliferating cell nuclear antigen (PCNA) proteins was detected by using immunohistochemistry SP method. Results The incidence of breast cancer in control group, low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2 was 73. 3% , 96. 7% ,30. 3% , 40. 0% and 83. 3% , respectively, with the difference being not significant between control group and high estrogen group 2, and between low estrogen group 1 and high estrogen group 2 (P ＞ 0. 05 ) , but with the difference being significant among the other groups (P ＜ 0. 05 ). There was significant difference in the average latent period of breast cancer among all groups (P ＞ 0. 05 ). There was significant difference in the number of TEB between low-estrogen group 1 and other groups (P ＜ 0.05). There was significant difference in the erbB-2 expression among the groups ( P ＞ 0. 05 ). There was no significant difference in the expression of breast ER and PR between control group and other groups ( P ＞ 0. 05 ). The PCNA expression in breast tumor tissue in low-estrogen group 1 was significantly higher than other groups (P ＜0. 05) , and there was significant difference in the PCNA expression between control group and high estrogen group 2, between low estrogen group 1 and high estrogen group 2 (P ＜ 0. 05). Conclusion The soybean isoflavones at different estrogen environments play different roles in the occurrence and development of MMTV-erbb-2 transgenic mouse mammary tumor. In the context of low estrogen, soybean isoflavones could even promote breast cancer formation and development. In the context of high estrogen, soybean isoflavones could inhibit breast cancer and development.     

大豆异黄酮在不同的雌激素环境下对MMTV-erbB-2转基因小鼠的作用

Objective To investigate the effect of the soybean isoflavones at different estrogen environments on the pathogenesis of breast cancer in MMTV-erbB-2 transgenic mice. Methods 150 fiveweek-old MMTV-erbB-2 transgenic female mice were phosen and divided into five groups; control group,low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2. The incidence and latent period of breast cancer were observed, and the expression of estrogen receptor (ER) ,progesterone receptor (PR), and proliferating cell nuclear antigen (PCNA) proteins was detected by using immunohistochemistry SP method. Results The incidence of breast cancer in control group, low estrogen group 1, low estrogen group 2, high estrogen group 1 and high estrogen group 2 was 73. 3% , 96. 7% ,30. 3% , 40. 0% and 83. 3% , respectively, with the difference being not significant between control group and high estrogen group 2, and between low estrogen group 1 and high estrogen group 2 (P ＞ 0. 05 ) , but with the difference being significant among the other groups (P ＜ 0. 05 ). There was significant difference in the average latent period of breast cancer among all groups (P ＞ 0. 05 ). There was significant difference in the number of TEB between low-estrogen group 1 and other groups (P ＜ 0.05). There was significant difference in the erbB-2 expression among the groups ( P ＞ 0. 05 ). There was no significant difference in the expression of breast ER and PR between control group and other groups ( P ＞ 0. 05 ). The PCNA expression in breast tumor tissue in low-estrogen group 1 was significantly higher than other groups (P ＜0. 05) , and there was significant difference in the PCNA expression between control group and high estrogen group 2, between low estrogen group 1 and high estrogen group 2 (P ＜ 0. 05). Conclusion The soybean isoflavones at different estrogen environments play different roles in the occurrence and development of MMTV-erbb-2 transgenic mouse mammary tumor. In the context of low estrogen, soybean isoflavones could even promote breast cancer formation and development. In the context of high estrogen, soybean isoflavones could inhibit breast cancer and development.