Frequency and Significance of Antibodies to Soluble Liver Antigen/Liver Pancreas in Variant Autoimmune Hepatitis

Background: Antibodies to soluble liver antigen/liver pancreas are highly specific markers of autoimmune hepatitis. Aims: Determine the frequency and clinical significance of these antibodies in the variant syndromes. Methods: Antibodies to soluble liver antigen/liver pancreas were determined in 28 patients with variant forms, including 10 with cryptogenic chronic hepatitis and 18 with cholestatic variants. One hundred and seventy-two patients with classical autoimmune hepatitis were similarly tested. Results: Seven of the 28 patients with variant forms had the antibodies, and this frequency was not statistically different than that in classical disease (25 vs. 12%, p = 0.08). Antibodies were most common in patients with cryptogenic chronic hepatitis (40%). Seropositive patients were indistinguishable from seronegative patients with variant forms, and they responded as well to corticosteroid therapy as patients with autoimmune hepatitis. Relapse after corticosteroid withdrawal invariably occurred in the seropositive patients whether with variant or classical disease, and HLA DR3 was more common in the seropositive patients with variant forms than in normal subjects (60 vs. 15%, p = 0.03). Conclusions: Antibodies to soluble liver antigen/liver pancreas occur commonly in the variant forms of autoimmune hepatitis and identify patients that closely resemble classical disease. Seropositivity is associated with relapse after corticosteroid withdrawal and HLA DR3. The antibodies may be surrogate markers of a genetic propensity to relapse that is independent of clinical phenotype.     

p53 Protein expression in laryngeal squamous cell carcinomas bearing wild-type and mutated p53 gene

We performed an immunohistochemical analysis to investigate the expression of p53 protein in a panel of 18 laryngeal squamous cell carcinomas, 15 primary tumours and three in relapse, previously analysed by us for the presence of p53 gene mutations. Dysplastic and/or normal surrounding mucosa was evaluated in 15 different tumours. The results of our study are the following: (1) expression of p53 protein was observed in one out of five tumours positive for p53 gene mutations (20%) and in 10 out of 13 (80%) negative cases; (2), p53 protein over-expression was frequently observed in normal and/or dysplastic mucosa surrounding either wild-type (7/11) or mutated p53 tumours (2/4); (3), p53 immunoreactive cells showed a pattern of distribution in normal and mildly/moderately dysplastic mucosa (basal layers), different from that in severely dysplastic mucosa (whole thickness). These data further support the hypothesis that p53 protein over-expression may be a marker of the earliest phases of multistep tumorigenesis in laryngeal squamous cell carcinoma.     

The Prevalence of Anti-acetylcholinesterase Antibodies in Autoimmune Disease

A robust and precise enzyme linked immunosorbent assay (ELISA) with proven sensitivity and specificity has been employed to detect human antibodies (allogenic/autogenic) to human acetylcholinesterase (AChE). The sensitivity of the method has been established using mouse monoclonal antibodies (0.8?ng/ml) and uniquely, human sera positive for anti-Yt^a allogenic antibodies, to one phenotypic form (most common) of human AChE. The latter was also used as the positive human control to ensure functionality of the assay. The ELISA method was used to establish a normal distribution curve for absorbance values employing sera from healthy blood donors Subsequently, the ELISA was employed to investigate the prevalence of anti-AChE antibodies in patients with confirmed autoimmune disease and patients with non-autoimmune thyroid disease (diseased control). The results indicate that there is not a high prevalence of anti-AChE antibodies in patients with confirmed autoimmune disease. The lack of anti-AChE autoantibodies in patients' with clinically apparent Graves' ophthalmopathy, mitigates against there being a causal role of such antibodies in Graves' associated eye disease.     

P53蛋白在肺癌中表达的研究

应用免疫组化技术,检测了82例原发性肺癌组织标本P53蛋白的异常表达,阳性表达率为53.65％(44/82),其与组织学分型、分级无关。P53蛋白表达与肺腺癌有无局部淋巴结癌转移及患者预后有明显关系(P<0.05),而在肺鳞癌以上各组间差异均无显著性(P>0.05)。提示P53基因突变在肺腺癌、肺鳞癌的演变过程中可能发挥的不同作用。     

自身免疫性肝炎和乙型肝炎患者自身抗体检测及对比分析

目的:检测分析自身免疫性肝炎(AIH)与乙型肝炎(HB)患者血清自身抗体特点及诊断应用价值。方法:采用间接免疫荧光法检测AIH组(n=43)和HB组(n=100)血清抗核抗体(ANA)、抗平滑肌抗体(SMA)、抗线粒体抗体(AMA)及抗中性粒细胞胞浆抗体(pANCA、cANCA),比较两组自身抗体检出率、ANA滴度、荧光模型及肝功能和免疫球蛋白。结果:AIH组自身抗体以ANA、SMA为主,阳性率分别为93.02和67.44%,两种抗体同时阳性的检出率为55.80%,AMA及pANCA、cANCA的检出率分别为13.95%和6.97%、2.32%;而HB组只检测出16例(16.0%)ANA,无一例SMA阳性。AIH组ANA以高滴度(≥1:320)抗体为主,荧光核型主要以核仁、核膜型为主,丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)及IgG含量明显高于HB组。HB组ANA以低滴度(≤1:100)抗体为主,荧光核型以颗粒型所占比例较高。结论:检测AIH和HB患者自身抗体相关指征对提高诊断准确性,制定治疗方案有重要作用。     

Correlation between DNA alterations and p53 and p16 protein expression in cancer cell lines

To investigate the interaction between DNA abnormalities, p53 and p16 gene mutations, and methylation and protein expression, 20 cancer cell lines were examined by Western blotting. A clear relation was found to exist between p53 accumulation and mutation status. Of 20 cell lines examined, 14 demonstrated p53 homozygous mutations in exons 3-10, including 12 missense mutations, one nonsense mutation, and one frameshift mutation. Overexpression of p53 was always linked to missense mutations in exons 6-8. Intermediate expression of p53 was noted in cells with missense mutations or polymorphism to proline at codon 72 in exons 4-5, whereas there was slight or no visible expression in wild type cells and in cells with nonsense and frameshift mutations. DNA aberration in the p16 promoter gene correlated significantly with protein expression of the p16 suppressor gene. Overexpression was noted in six cell lines, intermediate expression in two, and slight or no visible expression in 12. Methylation-caused disappearance of p16 protein was noted in 40% (8/20) of the cell lines. Of six cell lines overexpressing p16 protein, two could be amplified with primers for both unmethylated and methylated forms in a methylation-specific RCP analysis. One cell line with no visible expression could also be amplified with both primers. Overexpression or disappearance of p16 protein may readily occur when one of two alleles has been methylated.     

HBxAg增加p53蛋白在肝癌细胞内积聚

目的 进一步研究ＨＢＶｘ基因产物ＨＢｘＡｇ与ｐ５３蛋白的相互关系,探讨其在原发性肝癌发生中的作用机制。方法 以肝癌细胞株Ｈｅｐ３Ｂ为靶细胞,应用薄层层析法做报道基因氯霉素乙酰转移酶（ＣＡＴ）试验与ｐ５３和ＨＢｘ基因共转染,并构建了地塞米松诱导表达的ＨＢｘ基因质粒,用免疫荧光法观察ＨＢｘＡｇ表达对细胞内ｐ５３蛋白的影响。结果 应用ＨＢｘ基因与ｐＧ１３ＣＡＴ共转染,随ＨＢｘ量的增加而ＣＡＴ信号增强,免     

Identification of a Unique Form of p53 in Human Cord Blood Associated with the Development of Maternal Autoantibodies

PROBLEM: The possible link between p53-reactive antibodies in multiparous women and exposure to a unique p53 protein during pregnancy was examined. METHOD OF STUDY: p53-reactive antibodies were evaluated in sera from nulligravid and multiparous women and patients with ovarian cancer by Western immunoblot. Furthermore, the presence of p53 protein was assayed in cord blood by enzyme-linked immunosorbent assay. Cord blood-derived p53 was compared structurally by protein fingerprinting and functionally by gel mobility shift assay to other isolates of p53. RESULTS: Antibodies reactive with wild-type p53 were observed in 92% of multiparous women and 42% were reactive with one tumor-derived p53 protein. p53 protein was detected in 27 of 154 samples of cord blood. Structural analysis indicated that the fetal p53 resembled the UL-1 p53. Functionally, the fetal and UL-1 proteins failed to bind DNA. CONCLUSIONS: Fetal p53 protein seems to be distinct from wild-type p53, characterized by enhanced stability, structural differences and inability to bind DNA, analogous to alternatively spliced variants. Exposure to fetal p53 protein may form the basis for immunologic protection against cancer induced by multiparity.     

Immunohistochemical detection of the p53 oncoprotein in tumours of melanocytic origin

We employed the polyclonal anti-p53 antibody NCL-CM1 to cultured cells and pathological tissues in order to investigate the expression of p53 oncoprotein in human malignant melanomas. The results in the cultured cells showed that the antigenic determinant was sensitive to formalin fixation, resulting in a lower reactivity than with fixation by alcohol. In pathological tissues, the expression of p53 oncoprotein increased with progression of the tumour. Among 79 melanomas 37 (47%) showed distinct nuclear labelling and the highest proportion of reactive cells was observed in metastatic melanomas (mean 4.8%). An immunocytochemical study also revealed the presence of mutant-type oncoprotein in human melanoma cell lines, which was recognized by monoclonal antibody P240, and we confirmed that the molecular weight of the antigens recognized by both antibodies was 53 kDa by Western blot analysis. Therefore, although the presence of point mutations in human melanomas is yet to be confirmed our data suggest that the antigen detected by NCL-CM1 is a mutanttype or a complex of mutant and wild-type p53 oncoproteins. This antibody may be useful in retrospective studies of tumours of melanocytic origin.     

脓毒症大鼠心肌细胞凋亡和p53蛋白表达相关性的研究

目的研究脓毒症大鼠心肌细胞凋亡的变化及其与p53蛋白表达的关系。方法以盲肠结扎穿刺法复制脓毒症大鼠模型,以电镜和凋亡原位末端标记法（TUNEL法）检测其心肌细胞凋亡变化,用免疫组化方法检测p53蛋白的表达。结果一定时间内脓毒症大鼠心肌细胞凋亡率均明显高于正常对照组和假手术对照组（P均〈0.05）,p53蛋白表达阳性数均较正常对照或假手术组明显升高（P均〈0.05）,其变化与TUNEL法检测凋亡的结果一致（P〈0.05）。结论细胞凋亡可能是脓毒症时心肌损害的机制之一,其调控基因p53的改变或许可以作为脓毒症病情改变的标志,可利用它对脓毒症进行干预,以改善脓毒症的预后。     

乙型肝炎病毒X蛋白转录抑制抑癌基因p53表达的研究

目的 探讨乙型肝炎病毒（HBV）X蛋白对抑癌基因p53基因表达的调节作用。方法 以寡核苷酸芯片技术筛选的HBx反式调节基因结果为基础，利用生物信息学技术确定抑癌基因p53的启动子区域（p53p），构建真核报告基因表达载体pCAT3-p53p；以该质粒单独及与pcDNA3．1（-）-X共转染HepG2细胞，用酶联免疫吸附法（ELISA）检测氯霉素乙酰转移酶（CAT）的表达活性。进一步采用半定量RT-PCR技术检测HBV X蛋白对p53基因表达的调节。结果 成功构建报告载体pCAT3-p53p，克隆的p53启动于有顺式激活下游基因的活性；pCAT3-p53p与pcDNA3＋1（-）-X共转染时CAT的表达活性明显下降，说明HBVX蛋白抑制p53基因启动子的转录。RT-PCR结果表明，转染了pcDNA3．1（-）-X的HepG2细胞中p53基因mRNA表达减少。结论 HBV X蛋白在转录水平上反式抑制抑癌基因p53的表达，本实验不仅验证了基因芯片的筛选结果．而且为HBx在致肝细胞癌发生中的作用提供分子基础。     

Long-term Treatment Outcomes for Autoimmune Hepatitis in Korea

Immunosuppressive therapy can improve clinical, biochemical and histological features and considerably prolong survival in patients with autoimmune hepatitis. Although ethnicity may affect disease severity and presentation, the long-term outcome of immunosuppression in Korean populations is unknown. This study was aimed to assess the efficacy of immunosuppressive therapy and determine the prognosis of autoimmune hepatitis in Korean populations. We reviewed the medical records of 86 patients diagnosed as having autoimmune hepatitis at the Samsung Medical Center between 1994 and 2008. Seventy-two (83.7%) patients reached remission after a median treatment duration of 3.5 months (range 1 to 44 months). Attempts to withdraw medications were made in 24 cases after the median treatment duration of 36 months (median 6 to 125 months). Thirteen of 24 (54.1%) patients relapsed after treatment withdrawal. Of the 86 patients, 6 (7.2%) experienced disease progression and the overall 5-and 10-yr progression-free survival rates were 91.2% and 85.5%, respectively. In conclusion, immunosuppressive therapy for autoimmune hepatitis results in a favorable rate of remission and excellent progression-free survival, but the relapse rate after treatment withdrawal is high. This suggests that long-term immunosuppressive therapy may be particularly important for treatment of Korean patients.     

非小细胞肺癌患者p53蛋白和MMP2表达水平与同步放化疗的相关性

目的 研究非小细胞肺癌(NSCLC)患者p53蛋白、MMP2的表达与同步放化疗疗效的相关性.方法 选取2013年2年至2014年1月在我院接受同步放化疗的NSCLC患者89例.采用免疫组化方法检测患者组织中P53蛋白的表达情况,并根据P53蛋白表达与否分为阳性组(41例)与阴性组(48例).分别比较两组患者的疗效、生存率及治疗前后血清MMP2的表达,并进行相关性分析.结果 阳性组患者RR为53.66%(22/41),显著低于阴性组的75.00%(36/48),P<0.05差异有统计学意义.治疗后阴性组MMP2水平为60.9±23.4μg/mL,显著低于阳性组的75.5±31.7μg/mL,P<0.05差异有统计学意义.阳性组平均生存期为17.3±4.6月,显著低于阴性组的31.4±5.3月,差异有统计学意义(t=12.865,P<0.001).经Spearman相关性分析可得,NSCLC患者组织p53蛋白表达、血清MMP2的表达与同步放化疗疗效及生存期均呈显著负相关.结论 NSCLC患者p53蛋白、血清MMP2的表达与同步放化疗疗效均呈显著负相关,且与生存期也呈显著负相关,对治疗NSCLC具有参考价值.     

Epstein-Barr virus infection correlates with the expression of COX-2, p16INK4A and p53 in classic Hodgkin lymphoma

Classic Hodgkin lymphoma (cHL), a germinal-center related B cell neoplasm in almost all cases, is characterized by scarcity of the neoplastic Hodgkin/Reed-Sternberg (H/RS) cells. Epstein-Barr virus (EBV) has been shown to affect cell cycle and regulation of apoptosis. In total, 95 cases of cHL were studied. Five-micrometer sections were prepared and stained with hematoxylin and eosin and immunohistochemical streptavidin-biotin methods for EBV-LMP-1, COX-2, p53, p16, ki-67 and cleaved caspase-3. In-situ hybridization for EBV encoded RNA was used to confirm the detection of EBV in H/RS. There were 49 nodular sclerosis, 32 mixed cellularity, 8 lymphocyte-rich, and 6 lymphocyte-depleted subtypes in this series of cases. EBV, COX-2, p16^INK4A and p53 were detected in 55% (52/95), 64% (61/95), 62% (59/95), and 65% (62/95) of the cases respectively. EBV was detected in 62% (38/61), 70% (41/59), and 69% (43/62) of COX2, p16 and p53 positive cases respectively. On the other hand, EBV-non-infected cases of cHL are associated with 59% (20/34), 69% (25/36), and 73% (24/33) of COX2, p16 and p53 negative cases respectively. In conclusion, EBV infection is associated with the expression of COX-2, p16^INK4A and p53. EBV might be the dominant factor in determining the expression of these three proteins.     

Autoantibodies to p53 in sera of patients with autoimmune thyroid disease

Mutations in the tumor suppressor gene, p53, lead to intracellular accumulation of abnormal p53 protein and are associated with p53 autoantibodies. p53 also accumulates in autoimmune diseases and Hashimoto's thyroiditis, but it is unknown if p53 autoantibodies occur in the latter. We measured p53 autoantibodies in the sera of 93 patients with thyroid disease and 19 patients without thyroid disease. Anti-p53 antibodies were detected in the sera from 4.2% (2/48) of patients with autoimmune thyroid disease, including one patient with Hashimoto's thyroiditis (3.7%, 1/27) and one with Graves' disease (4.8%, 1/21). A third patient with pseudohypoparathyroidism, but without thyroid disease, was also positive (1/19; 5.2%). None of 19 patients with differentiated thyroid cancer had anti-p53 antibodies. We conclude that anti-p53 antibodies can be detected in the sera from approximately 4% of patients with autoimmune thyroid disease. This finding suggests that increased DNA damage and apoptosis may be associated with autoimmune thyroid disease.     

Absence of p53 gene mutation and infrequent overexpression of p53 protein in hepatoblastoma

Ten cases of hepatoblastoma were studies for overexpression of p53 protein by immunohistochemistry and for possible p53 gene mutation by single strand conformation polymorphism (SSCP) analysis and direct DNA sequencing of the polymerase chain reaction products. Only one case of the macrotrabecular type at stage IV showed overexpression of p53 protein. No DNA mobility shift was found in any of the cases studies by SSCP analysis. DNA sequencing performed on the case showing overexpression of p53 protein revealed no mutation within exons 5 to 8. The associated adrenal cortical carcinoma of the same case also showed overexpression of p53 protein, but no mutation of the p53 gene. These results indicate that mutation of the p53 gene is infrequent in hepatoblastoma. This observation supports the view that mutation of the p53 gene is not as important in the oncogenesis of childhood neoplasms as in adult cancers.     

肺鳞状细胞癌中p53蛋白表达及p53基因突变的检测

目的 通过检测肺鳞状细胞癌及癌旁组织中Ｐ５３蛋白积聚及相同癌组织中Ｐ５３基因的突变，探讨Ｐ５３蛋白积聚及Ｐ５３基因突变在肺鳞癌发病中的意义。方法 采用免疫组化和银染－ＰＣＲ－ＳＳＣＰ方法检测１２０例肺鳞癌及癌旁肺组织中Ｐ５３蛋白的状况及相同鳞癌组织中，Ｐ５３基因５、６、７、８外，显子突变的情况。结果 Ｐ５３蛋白了性率为５２．５％，Ｐ５３基因突变率为５６．７％，突变便数在第５、６、７、８外显子的分布     

Autoimmune and inflammatory responses in Kashin-Beck disease compared with rheumatoid arthritis and osteoarthritis

To examine plasma levels of arthritis-related autoantibodies and inflammatory factors in Kashin-Beck disease (KBD) patients compared with rheumatoid arthritis (RA) patients, osteoarthritis (OA) patients, and healthy controls, the plasma levels of autoantibodies to types II, IX, and XI collagen and cyclic citrullinated peptide (CCP) and immunoglobulin (Ig)-G and IgM rheumatoid factors (IgG-RF and IgM-RF) from 45 KBD patients, 39 RA patients, 46 OA patients, and 30 healthy controls were determined by enzyme-linked immunosorbent assay. The plasma concentrations of nitric oxide (NO) and tumor necrosis factor-α (TNF-α) were measured using the Griess method and bioassay, respectively. Statistical analysis was performed using one-way analysis of variance followed by the least significant difference t test for differences among groups. Results indicated that the plasma levels of collagen IX antibodies, IgG-RF, and NO significantly increased in KBD patients compared with patients with RA and OA and the control group. The levels of collagen XI antibodies, CCP antibodies, and IgM-RF but not collagen II antibodies and TNF-α were significantly increased in the plasma of the KBD group compared with that of the control group. We conclude that autoimmunity and inflammation may be involved in the pathogenesis of KBD, in particular in the advanced stage.     

P53在细胞缺氧调控中的作用

缺氧损伤是常见而又重要的病理过程,P53蛋白在缺氧调控中发挥的作用也逐渐为人们所认识。P53可在细胞核内富集并协助细胞快速应答缺氧信号,还能根据缺氧程度等因素间接决定缺氧损伤细胞的命运,促使细胞周期停滞、衰老或凋亡等。因此,P53是细胞缺氧损伤与保护过程中的重要影响因素。