The Study Of The Expression Of BAFF In Chronic HBV Infected Persons And Pathological Features Of Liver Tissue

PartⅠThe study of immune status in HBV-infected paitientsBackground:Hepatitis B(HBV) is the most common infectious diseases of the world. According to WHO estimates,there are 20 million that infected with hepatitis B virus(hepatitisB virus,HBV),more than 350 million people belong to chronically infected,75%of people belonging to chronically infected happened in Asia.Our country are hepatitis B-prone areas.National sample of more than 3-year-old HBsAg-positive population standardized rate is 9.09%and HBV infection rate is 50.04%in 2002.Our country is estimated that chronic HBV infection is about 1.2 million.Each year the population die of liver failure,liver cirrhosis and hepatocellular carcinoma related HBV-induced was estimated to be about 300,000 cases,accounting for the death of the first infectious disease.HBV infection in the body are to be cleared or the formation of persistent infection depends largely on host immune response.In recent years cytokines may play a very important role in deciding the outcome of HBV infection.According to the type of cytokines and biological function,CD4 + T cells(Th) at least can be divided into Th1,Th2 and TH0 three subtypes.Th1 cells produce IL-2, IFN-γ,TNF-βand other cytokines and mainly involve in cell-mediated immune response;while Th2 cells secrete IL-4,IL-10,IL-13 and other cytokines,and mainly promote the humoral immune reaction;Th0 cell is precursors of Th1/Th2 cells, while secretes low levels of Th1/Th2 type cytokines In recent years,the pathogenesis of hepatitis B studied from Th1/Th2 cells was much concerned.Some authors consider,self-limiting infection occurred when HBV antigen-specific Th1 cells dominated,Th2 cells is concerned with the chronic infection of HBV. In recent years the auto-immune phenomenon of hepatitis caused much concern of researchers.Substantial research shows that the body's own immune can be caused by almost all of anopheles liver virus,including HAV,HBV,HCV,HEV and EBV.The body's own immune is mainly expressed a various of autoantibodies can be detected from serum.It is reported that the total positive rate of domestic HBV-infected paitients autoantibodies is 18.80%～61.3%.Because the different cases and the different test items of autoantibodies,the reported positive rate is also different,but have confirmed the CHB patients have a variety of autoantibodies and the autoimmune disorders.All that caused liver cell damage and caused gradually severe hepatitis.Some studies suggest that the imbalance of Th1/Th2 can caused autoimmune disorders,and pathogenic THL can lead to tissue-specific autoimmune diseases,while Th2 mediate allergic reactions and major asthma.The interferon(interferon) is the first antiviral drug that be approved to treat chronic hepatitis.It has been listed more than 20 years.It is almost safe.However,with the clinical application of interferon,some adverse reactions are concered in the course of treatment.With the application of interferon to enhance immune function,own antibody titers of the patient that the basic level is high may have gradually increased. It can caused poor treatment and even give rise to autoimmune diseases. B lymphocyte stimulator(BLys) are first discovered and cloned by Shu etcin 1999.It is the first 13 members of tumor necrosis factor(TNF) family.BAFF play an important role in activation of B cells and plasma cell,antibody-independent type conversion of CD154(CD 40L),the formation of GC,as well as non-dependent T cells and T cell-dependent antibody response.It can enhance the function of B cells, CD4 + T lymphocytes and NK cell and enhance the immune response. BAFF-activated T lymphocytes,so that CD4 + and CD8 + T cells showed activation status,in particular,can stimulate T cell secretion of Th1-type cytokines IFN-γ,and IL-2.Therefore,BAFF may regulate Th1/Th2 cell imbalance has an important role.The absence of BAFF can lead to lower immune function,over-expression of BAFF can lead to much self-reactive B cells appeared.Experiments show that: BLyS over-expression concerned with autoimmune diseases such as systemic lupus erythematosus(SLE),rheumatoid arthritis(RA) and autoimmune liver diseases and concerned with viral infection diseases such as HCV,HIV.However, But BAFF in the HBV infection expression situation still not clear.In conclusion,the disease outcome is closely related to the progress of disease, immune response intensity,and immune disorders.In recent years cytokines is considered play an important role in deciding the outcome of HBV infection. Th1/Th2 type cytokine imbalance is considered one of the reasons of chronic HBV infection.Tumor necrosis factor BAFF involved in immune regulation of B,T cells and play an important role in the body's humoral and cellular immunity.The abnormal expression of BAFF cause immune imbalance and lead to the occurrence of many diseases.It is hot spots of various diseasese since it was found.Thus,the target of this study is Th1/Th2 type cytokines and tumor necrosis factor BAFF for as to the study of immune response after HBV infection.Research objective 1 Detect expression of IL--10(IL-10),interferon-γ(IFN-γ) and B lymphocyte stimulator(BAFF) under different conditions that is infected by HBV in vivo. Explore the immune situation of HBV infected patients;2 Detect expression of IL-10(IL-10),interferon-γ(IFN-γ) and B lymphocyte stimulator(BAFF) of CHB.Discuss the impact of immune status by a- interferon.3 Detect the expression of IL-10,IFN-γand BAFF in CHB patients with IFN-antibody,Discuss the impact of immune status of interferon.Research MethodsFasting collected serum of 20 cases of hepatitis B virus carriers(AsC),10 Example of interferon-positive and 10 cases of interferon-antibody negative,20 cases of chronic hepatitis B(CHB) patients with interferon before treatment and 3 months treatment and 6 months treatment and 20 healthy people,using enzyme-linked immunosorbent assay to detect the level of IFN-γ,IL-10 and BAFFThe Results1 The expression of cytokines of HBV-infected paitients:The IFN-γexpression level of CHB patients and AsC patients is lower,The IFN-γexpression level of CHB patients have the most significant decline,and there is significant difference compared with the healthy people(P = 0.04);The IL-10 expression level is higher, The IL-10 expression level of CHB patients have the most significant increase,and there is significant difference compared with the healthy people(P = 0.02);The IFN-γ/IL-10 ratio is lower,The IFN-γ/IL-10 ratio of CHB patients have the most significant decline,and there is significant lower compared with AsC patients and the healthy people(P = 0.02,0.00);The BAFF expression level is lower,The BAFF expression level have the most significant decline,and there is significant difference compared with the healthy people(P = 0.01).HBV-infected paitients exist the disorder expression of Th1/Th2 type cytokine,The disorder expression of Th1/Th2 type cytokine of CHB patients is more Serious than AsC patients,The BAFF expression level of AsC patients is lower than CHB,That may be related to immune tolerance of AsC patients.2 The effect of interferon therapy on the expression of cytokines:The expression levels of IFN-γin CHB patients after interferon treatment is higher,The expression levels of IFN-γafter treatment of June were significantly higher than pre-treatment and treatment of 3 months(P = 0.00,0.04),and there is no significant difference with the health of people(P = 0.20);IL-10 expression level has a downward trend, but there is no significant statistical difference before and after the treatment(P＞0.05);With the treatment time extended,IFN-γ/IL-10 ratio after treatment is gradually increased than before treatment(P = 0.04,0.00),but still lower than the healthy people(P = 0.00);BAFF expression levels of before and after treatment has no significant change(F = 0.71,P= 0.55).All these Shows that interferon therapy may enhance the Th1-type cytokines IFN-γexpression,and decreased Th2-type cytokines IL-10 expression,and improve the imbalance in the ratio of Th1/Th2 cells, and tumor necrosis factor BAFF had no significant impact.3 The effects of NAb to cytokines:IFN-γexpression level of NAb-positive patients is lower than the negative patients and healthy people,but had no significant statistical difference among the three groups(F = 2.93,P = 0.07);IL-10 expression level of NAb positive is higher than NAb-negative patients,and significantly higher than those in healthy people(P = 0.00),had significant statistical difference among the three groups(F=7.51,P = 0.00);IFN-γ/IL- 10 ratio of NAb-positive patients was significantly lower than Nab negative patients and healthy people(P＜0.05); BAFF expression level of NAb-positive patients is higher than NAb-negative patients and the healthy people,and had no significant statistical difference among the three groups among the three groups(F = 0.69,P = 0.51).NAb have a visible weakening of the promote of interferon to Th1-type cytokines and the inhibition to Th2-type cells,the BAFF expression level of NAb-positive patients is increased,but still lower than the level of healthy people.Conclusion of the study1 HBV-infected paitients exist the imbalance expression of Th1/Th2 type cytokine,which is related to the decreased expression of Th1-type cytokines and the enhanced expression of Th2-type cytokine;interferon therapy can enhance the expression of Th1-type cytokineγ-IFN,decreased the expression of Th2-type factor IL-10 and interferon therapy can improve the situation of the immune disorder,and interferon-neutralizing antibody reduced the immune regulation of interferon on Th1/Th2 cell.2 The BAFF expression of HBV-infected paitients is lower,the BAFF expression of AsC was significantly lower than the level of the health of people.Because BAFF can enhance immune,promote the expression of Th1-type cytokine,adjust the imbalance expression of Th1/Th2 type cytokine,the lower expression of BAFF may be associated with HBV-related chronic infection;3 Research has shown that IL-10 andα-IFN can stimulate BAFF expression,the expression levels of IL-10 in CHB patients is higher and given high-doseα-IFN treatment,but BAFF expression levels remained low.All that shows HBV-infected paitients may exist generated obstacles of BAFF;There is no correlation between BAFF and the produce of autoantibodies in HBV-infected paitients. PartⅡThe meaning of hepatic rosette forming in CHB patientsBackground:alpha-fetoprotein(AFP) is a sugary protein,molecular weight is about 70kD, mainly is synthetized by ribosome of rough endoplasmic reticulum in the liver cells, in the fetal period it is produced by the yolk sac and liver,1 week after birth;AFP rapidly decreased to normal(<10ng/ml),normal liver tissue of adult did not express AFP.When the liver cells or embryonic gonad tissue malignant lesions,the relevant genes were activated,the cell of lost the ability of synthesis of AFP re-start synthesis of AFP,leading to AFP concentration in blood was significantly increased, but in clinic,we often seen AFP elevated in patients with hepatitis.At present,most scholars view elevated AFP in the viral hepatitis patients due to liver cells reactivation after necrosis,resulting in naive cells,while naive cells have the ability to produce AFP,so that AFP elevated.Serum AFP levels have consistently been higher than normal,and with the increase in inflammatory activity was significantly increased,emerged a tendency cirrhosis,and AFP levels were significantly elevated in patients suggests there is a tendency to malignant transformation with liver cell itself,with the the extension of course eventually develop into liver cancer. Flower colorectal pf liver cells(hepatic rosette):refers to substantial necrosis of liver cells,the regeneration of liver cells formed gland-like structure,often occured in in the vicinity of severe necrotic area,it is enveloped by squeezed liver sinusoidal and connective Organize,liver cells around the capillary bile duct to form a hepatic rosette that are considered primitive liver cells,it can produce AFP.It has not yet been confirmed that liver cells with hepatic rosette can produce AFP.Research objective1 To observe the expression of alpha-fetoprotein(AFP) in different clinical types of viral hepatitis;2 To explore clinical significance of the hepatic rosette in liver tissue(the relevance of serum AFP);Research methods:1 To detect AFP levels of serum by 378 cases viral hepatitis which includes 140 cases of chronic hepatitis B(CHB),140 hepatitis B cirrhosis(CIR),115 Example cirrhosis and 17 cases liver cancer after hepatitis(HCC)and analyze AFP level.2 Observa expression of AFP by specific immunohistochemistrythat can specific marke AFP of liver tissue with the help of 65 cases confirmed hepatic rosette formation in liver cells and 42 cases confirmed no hepatic rosette formation in liver cells;The results1.253 cases of 378 cases viral hepatitis patients have abnormal AFP,positive rate is 66.9%,AFP expression level increased slightly in the main;AFP positive rate increased gradually with the develop of the disease,hepatitis group and post-hepatitis liver cancer group were significantly difference(P=0.02),AFP positive rate is significant difference between CHB group and the CIR,CIR decompensated group,liver cancer after hepatitis group(P＜0.05).With the disease gradually increased,the level of serum AFP increased slightly,the difference of the serum AFP level of inter-group has statistical significance(F=8.00,P = 0.00),there is significantly difference between the various clinical types of viral hepatitis and HCC group(P=0.00),but there is no significant difference in all clinical types of viral hepatitis(P＞0.05).2.AFP positive rate of flower node group was significantly higher than non-flower node group,there is statistical significance between the two groups (χ~2=7.33,P = 0.01),and positive rate of AFP have increased trend with an increase in flower node;AFP levels of flower node group is higher than those without flower node group,but there is not statistical significance between the two groups(P = 0.15),AFP level is increased with increasing of flower node,and AFP level of a large number of flower node group is higher than a small number of flower node group(P=0.01,0.04).The formation of hepatic rosette is related to the positive rate of AFP and the increased of AFP levels.3.With an increase of hepatic rosette,cirrhosis was significantly higher(P=0.00), moderate chronic hepatitis significantly reduced the(P=0.04),we can see hepatic rosette volume has a certain relevancewith the develop of disease;at the same circumstances(moderate chronic hepatitis,severe chronic hepatitis group,active cirrhosis group) hepatic rosette group compared with no hepatic rosette group,there were no significant difference about AFP,ALT,AST,total bilirubin(TBIL), albumin(ALB)(P＞0.05).However,in the same circumstances,the level of AFP of hepatic rosette group is higher than that of non-hepatic rosette group,with the developing of the disease,AFP level have elevated in two groups;and ALT levels have no significant changes and even there is a downward trend.All these shows that the expression level of AFP has a positive correlation trend with the forming hepatic rosette.4.Immunohistochemical results showes:AFP expression in liver tissue are not clear when the AFP level＜1000ng/ml of hepatic rosette group or no hepatic rosette group;two cases tissue samples of liver cancer after hepatitis show:serum AFP level of one cases is 501.86ng/ml,there was no obvious expression of AFP in liver tissue, serum AFP level of other cases is 69137.17ng/ml,there is a large number of AFP expression in liver tissue.We can see the expression of AFP in liver tissue is about with serum AFP level,there is not direct relation with the volume of hepatic rosette.Conclusion of the study 1.AFP expression levels are elevated in all clinical types of Hepatitis B,there is positive correlation between AFP positive rate and the developing of the disease.There is higher positive rate and higher expression level in the condition of more serious disease.2.There were positive correlation among hepatic rosette with serum AFP levels and the progress of disease,with the amount of hepatic rosette increased,serum AFP levels exaltation,seriously ill patients was increased in the proportion;We think chronic hepatitis especially patients with cirrhosis have gene expression changes. AFP increased expression levels is related to both liver cell necrosis and regeneration and the change of liver cell gene expression.3.Liver tissue examination showed that low serum AFP level(＜1000ng/ml),the formation of flower-end AFP expression in liver cells was not obvious,the expression of liver tissue and serum AFP level related to AFP. PartⅢThe study of liver damage in idiopathic inflammatory myopathies patientsBackground:Idiopathic inflammatory myopathy(IIM) are a group of unknown etiology of non-suppurative inflammation of striated muscle,including polymyositis(PM), dermatomyositis(DM),both about 70%of inflammatory myopathies,PM / DM in the etiology and pathogenesis is not very clear,the current PM/DM are possible a group of muscle tissue damage diseases of a specific genetic susceptibility to a certain individual which is immune response caused by environmental factors.IIM can affect the skeletal muscle,smooth muscle,heart,brain and cardiovAsCular system,etc.,but the impact on the liver is not yet clear.CD8 + T cell-mediated immune play an important role in the PM pathogenic mechanisms,Atsushi Takahashi,etc.found that PM-related liver injury is related to CD8 + T cell-mediated cellular immunity,and recent studies found that CD138 + plasma cell-mediated humoral immunity in the pathogenesis of PM play an important role, but it is unclear that the CD138 + plasma cells is relevant to PM-related liver injury. Therefore,this study used the method of immunohistochemical marker CD138 + plasma cells in liver tissue of IIM patients to observe with the CD138 + plasma cell infiltration in liver tissue of patients.Research Objective1.To investigate the source of serum ALT in Idiopathic Inflammatory Myopathy(IIM) patients with hyperaminotransferase.2.To observe the IIM patients with elevated transaminase Whether or not the existence of liver tissue injury and to explore the possible mechanisms of liver tissue injury in the IIM patients with elevated transaminase; 3.To observe the infiltrating CD138 positive plasma cells in liver tissue of IIM and autoimmune hepatitis patients.Research Methods:1.Collected 28 cases ofⅡM patients and 11 cases of AIH patients with liver function(ALT,AST),myocardial enzymes(CK,CK-MB,HBDH,LDH), autoantibodies(ANA,) and other clinical data,analysis the reasons of serum ALT increased;2.6 cases of IIM patients and 11 cases of AIH patients with liver biopsy: conventional HE staining and the CD138+ plasma cells was staining with the method of immunohistochemical in liver tissue.To observe the liver damage and expression of CD 138+ plasma cell.The result:1 Serological target:The mean of serum ALT level is 198.8±209.0U/L inⅡM patients and ALT elevated gently between 40～120U/L in 50%(14/28).Compare with AIH patients,there was no significant difference(p＞0.05),Inversely,compare with the healthy controls,notable diference was be founding(p＜0.01),The abnormal rate of Lactate Dehydrogenase(LDH) and Hydroxybutyrate Dehydrogenase(HBDH) are 100%and 85.7%,higher than 78.6%of the Creatinkinase(CK),ANA-positive rates of IIM and in patients with AIH were 50%and 54.5%,The total antibody positive rates were 61.1%and 63.6%,There was no significant difference between the IIM and AIH(P＞0.05);2 Histopathologic examination of liver tissue:liver damage was mild and a few to middle quantity inflammatory cells infiltrated in portal tract In the PM patients, immunohistochemical staining did not find clear-specific CD138;positive plasma cells were found In AIH patients,there is often accompanied by piecemeal necrosis, bridging necrosis,liver damage was appreciably heavily,massive inflammatory cells infiltrated in portal tract and proliferation of the surrounding liver parenchyma and liver Rosette forming cells.Immunohistochemical staining specificity further extend to the surrounding liver parenchyma,and manifest infiltration of CD138 positive plasma cells were detected in portal tract,and infiltration of T lymphocytes CD4 + T cells over CD8 + T cells.Research conclusion:1 Slight liver damage was found in IIM patients,whose elevated ALT excluding from injury liver,the impair muscle may be another source;2 PM-related liver injury is caused by the CD8 + T cell-mediated cellular immunity,CD138+plasma cell-mediated humoral immunity may not play an important role;3 According to serum transaminase levels and autoantibodies can not distinguish AIH liver injury and IIM-related liver injury;There were significantly different between AIH liver injury and IIM-related liver injury in liver histological features, histological examination is the gold standard to identify the two.