靶向IGF1R的siRNA抑制人肝癌细胞SMMC7721裸鼠移植瘤的实验研究

背景与目的:利用小干扰RNA(small interfering RNA,siRNA)抑制哺乳动物基因表达已成为研究基因功能的一种有效方法.本研究探讨人胰岛素样生长因子1类受体(insulin-like growth factor 1 receptor,IGF1R)的短发夹环RNA对人肝癌细胞株SMMC7721裸鼠移植瘤的抑制作用.方法:设计并合成特异性靶向IGFIR的siRNA片段,构建SMMC7721-IGFlR-siRNA表达质粒,将其转入SMMC7721细胞.通过G418筛选出稳定株,移植裸鼠成瘤.同时设立对照组.根据体积的均数值绘制肿瘤生长曲线,以HE染色法观察质粒处理后瘤组织的病理改变,Western blot检测肿瘤组织中IGF1R的表达变化,免疫组化SP法检测检测瘤组织内微血管密度,原位末端标记法(TUNEL法)检测肿瘤细胞的凋亡情况.结果:SMMC7721-IGF1R-siRNA组肿瘤体积与SMMC7721-IGF1R-mutation组、SMMC7721组相比差异有统计学意义(P＜0.05).病理学检查及TUNEL检测发现,SMMC7721-IGF1R-siRNA组肿瘤生长受到抑制,细胞凋亡指数[(50.2±6.4)%]明显高于SMMC7721.IGF1R-mutation 组[(5.4±1.0)%]和SMMC7721组[(6.0±2.1)%](P＜0.05).SMMC7721-IGF1R-siRNA组与SMMC7721-IGF1R-mutation 组、SMMC7721相比,瘤内IGFIR蛋白表达明显下调.SMMC7721-IGF1R-siRNA组微血管密度(11.3±4.4)与SMMC7721-IGF1R-mutation组(36.7±7.6)、SMMC7721组(28.4±6.5)相比明显下降(P＜0.05).结论:构建的SMMC7721-IGF1R-siRNA具有RNA干扰作用,能抑制SMMC7721细胞裸鼠移植瘤的生长.     

Aurora kinase a inhibitor MLN8237 suppresses pancreatic cancer growth

Pancreatic ductal adenocarcinoma (PDAC) is notorious for high mortality due to limited options of appropriate chemotherapy drugs. Here we report that Aurora kinase-A expression is elevated in both human and mouse PDAC samples. MLN8237, an inhibitor of Aurora kinase-A, efficiently reduced the proliferation and motility of PDAC cells in vitro as well as tumor growth in orthotropic xenograft model and genetic pancreatic cancer animal models (p53/LSL/Pdx-Cre mice) in vivo. MLN8237 exhibited tumor inhibitory effect through inhibiting proliferation and migration, and inducing apoptosis and senescence. These results provide the molecular basis for a novel chemotherapy strategy for PDAC patients.     

Is induced membrane technique effective in reconstruction of mandibular segmental bone defects? An experimental study

This study aimed to compare the effectiveness of different graft materials using induced membrane technique for reconstruction of mandibular segmental bone defects.New Zealand rabbits were used as the experimental animal. As first-stage surgical procedure, segmental bone defects were created at the lower border of the mandibula in all groups. Polymethylmethacrylate (PMMA) cement was inserted into the defects. After 6 weeks, PMMA cement was removed in all groups. In the Control group, defect areas were left empty. Defects were filled with autogenous graft in the Autograft group, xenograft in the Xenograft group, and a mixture of autogenous graft and xenograft in the Autograft + Xenograft group. Histopathological, stereological, and immunohistochemical analyses were performed.A total of 40 New Zealand rabbits were used. Rabbits were randomly divided into four subgroups as Control, Autograft, Xenograft and Autograft + Xenograft groups (n = 10). When the groups were compared in terms of newly formed bone tissue volumes, significant difference was found between the Control group and Autograft group, Xenograft group and Autograft + Xenograft group (p < 0.001, p < 0.001, p = 0.003). The results of immunohistochemical examination were consistent with this finding.Stereological and immunohistochemical results can be used as a justification to adopt the induced membrane technique on an experimental basis in humans when it comes to the reconstruction of small segmental mandibular defects.     

平阳霉素和吡柔比星联合用药抗人食管癌的实验研究

重复实验两次研究平阳霉素（ＢＬＭＡ５）和吡柔比星（ＴＨＰ）联合用药对裸鼠移植的人体食管癌的疗效，结果：在裸鼠可耐受的剂量下，ＢＬＭＡ５、ＴＨＰ、ＢＬＭＡ５＋ＴＨＰ等各组抑瘤率分别为５６．９０％～６１．５４％（Ｐ＜０．０１）、３４．６２％～３９．６６％（Ｐ＜０．０５）、７１．１５％～７５．８６％（Ｐ＜０．０１），证明ＢＬＭＡ５和ＴＨＰ联合用药可明显提高疗效。研究结果提示ＢＬＭＡ５和ＴＨＰ合用有可能用于食管癌的临床化疗。     

Methylprednisolone prevents rejection of intrastriatal grafts of xenogeneic embryonic neural tissue in adult rats

We studied the effects of high-dose methylprednisolone on the survival of intrastriatal neural xenografts and the host responses against them. Dissociated mesencephalic tissue from inbred mouse (CBA-strain) embryos was transplanted to the intact striatum of adult Sprague-Dawley rats. The rats received either daily injections of methylprednisolone (30 mg/kg), or cyclosporin A (10 mg/kg), or no immunosuppressive treatment. Two or six weeks after transplantation, there was good survival of xenografts in both the methylprednisolone- and cyclosporin A-treated rats. In contrast, the xenografts in untreated control rats were all rejected by six weeks. There was no marked difference in the degree of expression of MHC class I and II antigens and the accumulation of activated astrocytes and microglial cells/macrophages between the three groups. However, both methylprednisolone and cyclosporin A reduced infiltration of T lymphocytes to the transplantation sites. The expression of pro-inflammatory cytokines (interferon-λ, tumour necrosis factor-a, interleukin-6) in and around the grafts was lower in the methylprednisolone- and cyclosporin A-treated groups than in untreated control rats. Although high-dose methylprednisolone caused significant body weight loss, we conclude that this treatment can prevent rejection of intrastriatal grafts of xenogeneic embryonic neural tissue in the adult.     

Sequential,morphological, and antidonor antibody analysis in a hamster-to-rat heart transplantation model

The pathogenesis and the mechanism of accelerated graft rejection in concordant xenotransplantation are unclear. The histopathological features and kinetics neither fulfill the criteria of classic hyperacute rejection nor resemble an accelerated type of first-set allograft reaction. The aim of this study was to investigate the mechanism of concordant xenograft rejection in relation to the early morphological changes in hamster hearts transplanted to unmodified rat recipients by sequential, immunohistological analysis of grafts, regional lymph nodes, and spleens and to correlate these results to the production of antidonor antibodies, as determined by a flow cytometric assay. Histopathological features were characterized by a gradually increasing myocytolysis with fragmentation and loss of myofilaments. The first slight signs were observed a few hours after transplantation. Later, vascular changes developed, evolving into a leukocytoclastic type of vasculitis, eventually with thrombosis. No significant interstitial lymphocyte infiltration was present, but neutrophilic granulocytes and macrophages appeared. In addition, a distinct increase in B cells in spleens and lymph nodes was noted. Low levels of preformed antidonor antibodies did not increase during the first 48 h; however, significant amounts of species-, but not donor-, specific antibodies were demonstrated at the time of rejection. These data, together with the morphological observations, indicate a primarily humoral xenograft rejection in this model. Minor damage to graft myocytes a few hours after transplantation, progressing to vascular changes within 24–48 h, further suggests that preformed antidonor antibodies directed against endothelial or myocyte determinants may play an initiating role in the pathogenesis of unmodified, concordant xenograft rejection.     

Membrane Immunoisolation of a Diffusion Chamber for a Bioartificial Pancreas

Immunoisolation is a potentially important approach to transplanting islets without any immunosuppressive therapy. The concept of immunoisolation is outlined in systems in which the transplanted tissue is separated from the immune system of the host by an artificial barrier. We previously described a diffusion chamber as a bioartificial endocrine pancreas (Bio-AEP), which was constructed by placing pancreatic endocrine cells, trapped in a mixed matrix, in the center of a ring holder sandwiched between nucleopore membranes, which were shielded by silicone. This experiment was designed to evaluate a suitable pore size for the nucleopore membrane to ensure immunoisolation during xenoimplantation of the Bio-AEP in vitro and in vivo. A nucleopore membrane of pore size 0.1 μm or 0.2 μ m was employed as the semipermeable membrane which provided a mechanical barrier between the endocrine pancreas graft and the host immune system. The protective effect of the Bio-AEP from humoral immunity was determined in vitro, using sensitized sheep erythrocytes (EAs). A complement protein did not destroy the cell membranes of the EAs in the diffusion chamber containing the mixed matrix with the nucleopore membrane of 0.1 μm pore size. In an in vivo experiment, 6 streptozotocin (STZ) induced diabetic rats were implanted with Bio-AEPs constructed with nucleopore membranes of pore size 0. 1 μm and containing MIN6 cells in the mixed matrix. In the STZ diabetic rats with Bio-AEPs, a return to normoglycemia was observed up to 50 weeks after implantation without the use of any immunosuppressant. Also, the body weights of the rats gradually increased. During the observation, when the Bio-AEPs were removed from the STZ diabetic rats, the blood glucose immediately returned to preimplantation levels, and the body weights of the rats also decreased. The membranes of the Bio-AEPs removed from the STZ diabetic rats showed a very thin layer of fibroblastic cells on the outer surfaces. The results indicated that the Bio-AEP, in which pancreatic endocrine cells were trapped in a mixed matrix and with a 0.1 μm pore size membrane, should be useful for xenoimplantation into diabetic animals and may open a new field in the therapy of human diabetics.