Treatment with thymic extract TFX for chronic active hepatitis B

Eighteen patients with biopsy-proven chronic active hepatitis B (CAHB) and a significantly lowered CD4+/CD8+ cell number ratio were treated with thymic factor X (TFX): group I (n = 12) - for 12 months, group II (n = 6) for 6 months. As early as 14 days after starting the treatment a lowering of CD8+ cell numbers with a rise in CD4+/CD8+ cell number ratio were found. These changes continued to progress during the next months and were accompanied by decreased in NK cell numbers with enhancement of NK cell activity. Normalization of the biochemical and immunological parameters occurred after 5-6 months of the treatment. In both groups of patients seroconversion in the HBe system was observed after 9-12 months of the treatment. After two years complete clinical remission persists with normal biochemical and immunological findings and without reversion in the HBe system. The results seem to indicate that TFX has an immunostimulatory action and exerts beneficial effects on the course of CAHB.     

An immunogenetic study of suppressor cell activity in autoimmune chronic active hepatitis

Some patients with autoimmune chronic active hepatitis as well as their disease-free first degree relatives show decreased suppressor cell activity of peripheral blood T lymphocytes. Studies were therefore undertaken in families ascertained by the presence of a single chronic active hepatitis patient to determine if this abnormality of immune regulation represents a genetic phenotype simply controlled by a gene or genes at a putative disease susceptibility locus and, further, if this locus showed linkage to either the HLA or the immunoglobulin constant region loci. In addition to determining circulating autoantibody status and genotyping for HLA and immunoglobulin allotypes, suppressor T cells were evaluated by surface markers and by determining their ability to suppress IgG secretion in vitro. The results suggest that immunoregulatory dysfunction in autoimmune chronic active hepatitis is a familial abnormality, but that this abnormality occurs independent of circulating autoantibody status and of the segregation of genes for HLA or immunoglobulin allotypes.     

Feedback inhibition of thymic secretory activity in mice treated by the thymic extract TP-1 (thymostimulin).

The ultrastructural changes occurring in the medullary epithelium of the thymus of young mice, as a result of repeated injections of thymic extract, TP-1 (thymostimulin) was investigated. After daily injection of TP-1 for 3 weeks, no changes in thymus architecture could be observed by light microscopy. However, by electron microscopy, specific changes were noticed in the epithelial cells. The secretory granules became dilated and engorged; diameter of granules in normal control thymus was approximately 200-250 nm, but reached 1000 nm in treated mice. Degenerative changes appeared in some of these granules, including myelin bodies, distorted configuration and fat droplets. Signs of involution of whole cells and presence of cellular debri within macrophages were observed. Acid phosphatase staining disclosed many lysosomes containing ingested granules. No such findings were observed in control untreated mice, or in mice treated by a heart extract similarly prepared to TP-1. All these findings can be taken as ultrastructural evidence for feedback inhibition of thymic secretory activity, in analogy to the changes occurring other feedback inhibited, peptide hormone secreting glands. The data indicate that (i) the thymus respond to feedback inhibitory stimuli, as other endocrine glands do; (ii)TP-1, the thymic extract under study, contains a physiologically significant thymic hormone, which, when introduced in high doses can exert specific feedback inhibition. This can be taken as an additional, new criterion for the definition of thymic hormones.     

Evidence of impaired antigen non-specific but normal antigen specific suppressor cell function in children with autoimmune chronic active hepatitis.

In autoimmune chronic active hepatitis (aCAH), autoaggression is believed to derive from a defect in immunoregulation. Antigen non-specific Concanavalin A (Con A) induced suppressor cell function has been reported to be impaired. In 11 children with aCAH we have investigated inhibition of production of a specific antibody (anti-tetanus toxoid, anti-TT) by suppressor cells induced either by a non-specific stimulus (Con A) or by the specific antigen (tetanus toxoid, TT). Con A induced suppression of anti-TT was significantly lower in patients (15.7 +/- 2.5%) than in controls (46.7 +/- 4.4%; P less than 0.01). In contrast, high dose tetanus toxoid induced suppression was similar in patients and controls (69.8 +/- 4.2, 72.0 +/- 3.6%, respectively). Both groups had similar serum anti-TT levels and in vitro production of anti-TT in response to optimal dose of TT. Our data indicate that antibody production to a T cell-dependent antigen is under the control of at least two regulatory mechanisms, one antigen specific and one antigen non-specific, only the latter being defective in aCAH.     

Clonal analysis of liver-infiltrating T cells in patients with LKM-1 antibody-positive autoimmune chronic active hepatitis

Autoantibodies against microsomal antigen of liver and kidney (LKM-1) are diagnostic markers for a subgroup of autoimmune chronic active hepatitis (AI-CAH). Cytochrome P450dbl, now classified as cytochrome P450 IID6, is the major antigen of LKM-1 antibodies. Immunohistological studies suggest that hepatic injury in AI-CAH is mediated by liver-infiltrating T cells. In the present study the specificity and function of liver-infiltrating T cells was analysed at the clonal level. Phenotypical characterization of 189 T cell clones isolated from four liver biopsies of LKM-1 antibody-positive patients showed an enrichment of CD4+ CD8- T cell clones proliferated specifically in the presence of recombinant human LKM-1 antigen (rLKM). This reaction was restricted to autologous antigen-presenting cells and to HLA class II molecules. In order to see whether rLKM was also recognized by peripheral blood T lymphocytes (PBL) we tested the proliferative response of PBL from several individuals. PBL from three of the four patients with LKM-1 antibody-positive AI-CAH proliferated to rLKM, whereas no response was seen with PBL from patients with LKM-1 antibody-negative chronic liver diseases and from healthy blood donors. These data demonstrate that the LKM-1 antigen is recognized by liver-infiltrating T cells in LKM-1 antibody-positive AI-CAH. For further functional characterization, liver-derived T cell clones were tested for their cytotoxic activity. In the presence of phytohaemagglutinin 24 out of 26 CD4- CD8+ but also 20 out of 63 CD4+ CD8- T cell clones lysed autologous as well as allogenic EBV-transformed B cell lines or K562 cells. Five CD4- CD8+ T cell clones lysed autologous but not allogenic B cell lines spontaneously in a HLA class I-restricted manner. Although the antigen specificity of these clones is still unknown the data show the presence of autoreactive T cells at the site of inflammation that could contribute in the pathogenesis of AI-CAH.     

In vitro interleukin-1 (IL-1) production in thymic hyperplasia and thymoma from patients with myasthenia gravis

Using a biological test, we measuredin vitro thymic epithelial cell IL-1 activity of 14 thymomas and 8 hyperplasia from myasthenic patients. Our results demonstrate that thymic epithelial cells from hyperplastic thymuses spontaneously produce high amounts of IL-1 compared to controls, while cells from thymomas produce very low amounts of IL-1. After LPS stimulation the difference between patients and controls is no longer significant. Immunohistochemistry studies demonstrate a limited number of IL-1-positive cells on sections from normal thymuses and a variable number of IL-1-positive cells on diseased thymuses, not clearly correlated to thein vitro production. In hyperplastic thymuses an association is found between the level of hyperplasia andin vitro IL-1 production, suggesting a role for IL-1 in the expansion of thymic lymphoid follicles. In thymoma, the low IL-1 production and the loss of corticomedullary organisation raise the possibility that some autoreactive clones could emerge from the lymphocyte pool, present in the abnormal tumoral microenvironment.     

Detection of tissue T-cell in patients with chronic active hepatitis using fragmented sheep red blood cell (SRBC) membrane

Fragmented sheep red blood cell (SRBC) membrane was used for detection of T-cells in liver biopsy specimens from patients with chronic active hepatitis. SRBC was separated with Lymphoprep, sonicated, then filtered through a 3 mu Millipore-membrane as a fragmented SRBC reagent. Tissue T-cells were stained by an indirect immunofluorescent technique using SRBC reagent and fluorescein isothiocyanate (FITC)-labelled rabbit anti-SRBC. Positively staining lymphocytes were present in portal tracts and in areas of piecemeal necrosis. There also seemed to be a positive correlation between the number of positively staining lymphocytes and the activity of chronic hepatitis; numerous lymphocytes being stained in areas of severe piecemeal necrosis. Our findings suggest that the fragmented SRBC technique for detection of T-cells is reliable and reproducible, that it could be used as a clinical routine method, and that it is useful for further elucidating the nature of host immune reactions on tissue levels.     

Acute viral E hepatitis with chronic liver disease (autoimmune hepatitis)

A 36 years old male presented with anorexia, jaundice and ascites. He was suffering from acute viral E hepatitis. In view of ascites, he was investigated for associated asymptomatic chronic liver disease (CLD). The CLD was diagnosed as cirrhosis with autoimmune hepatitis and was treated with steroid with good response. He is maintaining good health with low dose steroid, on follow up for 1 year.     

Transfer factor (TF) treatment of patients with HBs-Ag-positive chronic active hepatitis

Summary It is a clinically and experimentally well supported working hypothesis that infection with hepatitis B virus may result in chronic active hepatitis in patients with suspected immune deficiencies. On this basis, a pilot study was performed in order to evaluate the effect of specific transfer factor (TF) in the treatment of HB_s-Ag-positive chronic active hepatitis. From the leukocytes of 500 ml venous blood each of 40 volunteers that had completely recovered from acute virus hepatitis B within the last 6 months, a unique TF pool (40 units of TF) was prepared according to the method of Lawrence. Preexaminations indicated that this preparation was able to enhance cellular immune reactions in vitro. Thirteen patients with HB_s-antigenemia and chronic active hepatitis (i.e., two liver biopsies within the last 6 or more months with the histological criteria of chronic aggressive hepatitis according to de Groote, elevated serum levels of bilirubin, alkaline phosphatase, transaminase activities, and/or -globulines) were randomized: Seven received s.c. injections of two units of TF each on days 1 and 15, the other six saline. Conversion of skin reactions to some ubiquitous antigens occurred in the TF group, but no significant and constant drop of HB_s-Ag serum titers was observed. Although some of the biochemical parameters seemed to ameliorate in the TF group, the differences versus the control group did not prove to be significant within the limited number of patients under observation. The in vitro reactivity of patients' lymphocytes to HB_s-Ag, tested by means of the³H-thymidine uptake, was never found enhanced after TF application. In the used doses, specific TF was not effective in the treatment of HB_s-Ag-positive chronic active hepatitis; unfavorable side-effects were not observed.     

In vitro effect of alpha-interferon on mononuclear cells of normal and autoimmune patients

The effect of lymphoblastoid interferon (IFN-alpha) on cell-mediated and humoral immunity was studied in patients with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). The base-line natural killer (NK) and antibody dependent cytotoxic cell (ADCC) activity was higher than normal in individuals with RA. The NK and ADCC activities were tested after IFN-alpha pretreatment and the augmentation of NK and ADCC activity was less in these patients than in normals. Lymphoblastoid IFN inhibited antigen induced T-cell proliferation in SLE to a lesser extent than in normal individuals. Finally, the addition of lymphoblastoid IFN was also less effective at suppressing in vitro polyclonal antibody formation by mononuclear cells from patients with SLE than from normals, with enhancement observed in some patients at the lower IFN-alpha concentrations tested.     

The in vitro effect of a calf thymus extract on the peripheral blood lymphocytes of sixty-six melanoma patients.

The in vitro effect of a calf thymus extract (TP-1) on peripheral blood lymphocytes was evaluated in sixty-six melanoma patients. Twenty out of forty-five stage I and seventeen out of twenty-one stage IIb patients presented depressed T lymphocyte values. A statistically significant increase in E-rosette-forming cells (E-RFC) was observed in seventeen (85%) stage I and twelve (70.5%) stage II patients. In thirteen (65%) stage I and eight (47%) stage II patients E-RFC reached normal values. Our results indicate that the fall of T lymphocytes in the peripheral blood of melanoma patients is often related to an increase in TP-1 sensitive cells which suggests the possibility of a thymic hormone deficiency in these patients.     

Steroid economising effects of a calf thymus extract in three patients with juvenile chronic arthritis

Summary Recent therapeutic trials in rheumatology using different immunomodulating agents have given encouraging results. In this study an aqueous calf thymus extract (CTE) was administered to three patients, two with systemic juvenile chronic arthritis (JCA), Still's disease, who could not be weaned from steroids during 2 years of conventional therapy, and one girl with a chronic juvenile monarthritis who had responded unsatisfactorily to nonsteroidal antirheumatics for 19 months. A striking clinical improvement was observed in all three patients. Prednisone (PRED) was discontinued in one case with systemic (JCA) and 0.25 mg/kg body weight/day is presently being given to the other patient. The girl is doing well on 4 mg chloroquin kg body weight/day; indomethacin (IND) was discontinued. Laboratory data including cellular immunoreactivity normalized in all three patients.     

天然多亚型干扰素-αN1治疗慢性乙型肝炎的疗效观察

目的观察干扰素的抗病毒作用。方法用天然多亚型干扰素－αＮ１（惠福仁）治疗５０例慢性乙型肝炎，另５０例为对照组，两组临床与实验室指标均有可比性。治疗组用ＩＦＮ－αＮ１３ＭｕＩＭｑｄ治疗７ｄ，继用３ＭｕＩＭｔｉｗ治疗１１～２３周。结果经３个月治疗，治疗组与对照组的丙氨酸转氨酶复常率分别为８４％（４２５０）和４６％（２３５０），Ｐ＜０．０１；ＨＢｅＡｇ阴转率为５６．７％（１７３０）和１０．８％（４３７）；ＨＢＶＤＮＡ的阴转率为３６．４％（１２３３）和５．９％（１１７），Ｐ＜０．０１。结论ＩＦＮ－αＮ１治疗慢性乙型肝炎是目前疗效比较满意的抗病毒药，且副作用少，适于对其他干扰素疗效不佳的病例     

Effect of CD4+ CD25+ regulatory T-cells on CD8 T-cell function in patients with autoimmune hepatitis

BACKGROUND AND AIMS: CD4 T lymphocytes constitutively expressing the IL-2-receptor alpha-chain (CD25) (T-regs) are central to self-tolerance maintenance, preventing the proliferation and effector function of autoreactive T-cells. In autoimmune hepatitis T-regs are defective in number but maintain the ability to suppress IFNgamma production by CD4+CD25- T-cells. We have studied the ability of CD4+CD25+ (T-regs) to regulate proliferation and cytokine production by CD8 T-cells in patients with autoimmune hepatitis at diagnosis and during remission. METHODS: Twenty-five patients were studied. T-regs were purified from PBMCs by CD4 negative selection followed by CD25 positive selection, using immunomagnetic beads. The ability of T-regs to suppress CD8 T-cell proliferation was assessed by 3H-thymidine incorporation; their ability to regulate cytokine production by intracellular cytokine staining. RESULTS: We found that T-regs are unable to regulate CD8 T-cell proliferation and cytokine production in patients studied at diagnosis, while they suppress CD8 T-cell proliferation and induce an elevation of IL-4 producing CD8 T-cells in patients during drug-induced remission. CONCLUSION: Inability of T-regs to regulate CD8 T-cell function at diagnosis may contribute to the initiation of autoimmune liver damage. The ability of T-regs to regulate CD8 proliferation and IL-4 production during drug-induced remission suggests a role for immunosuppressive treatment at reconstituting T-regs function.     

Neutral effect of prolonged transdermal hormone therapy on liver function of postmenopausal women with chronic active hepatitis

OBJECTIVE: To test whether transdermal hormone therapy can be safely administered to postmenopausal women with chronic viral hepatitis B and/or C. DESIGN: Eighty-one postmenopausal women with chronic viral hepatitis B and/or C and with severe vasomotor symptoms were treated for 5 years with transdermal estradiol (50 microg/day) continuously and with transdermal norethisterone (250 microg/day) for 14 days of every 28-day cycle. Another 95 women with viral chronic hepatitis but without climacteric symptoms were used as controls. Liver enzymes (glutamic-oxalacetic transaminase, glutamic-pyruvic transaminase, gamma-glutamine-transferase, and alkaline phosphatase) were measured every year. RESULTS: At baseline, liver enzymes were similar in the two groups, with the exception of gamma-GT, which was slightly higher in untreated women (P < 0.01). Liver enzymes did not significantly vary with time in hormone-treated and untreated women. No significant difference was observed between the two groups. CONCLUSIONS: Transdermal estradiol and norethisterone can be safely administered for a prolonged period to postmenopausal women with chronic viral B and/or C hepatitis.     

Immunological mechanisms of corticosteroid therapy in chronic active hepatitis: analysis of peripheral blood suppressor T-cell and interleukin 2 activities

To investigate the immunological mechanisms underlying corticosteroid therapy in chronic active hepatitis (CAH), in vitro effects of prednisolone on suppressor T-cell and interleukin 2 (IL-2) activities were examined in six corticosteroid therapy-effective and six therapy-ineffective patients with CAH prior to the therapy. Whereas low suppressor T-cell activity and decreased response to IL-2 in T cells were found in the corticosteroid therapy-effective group, these reductions recovered to the normal range when the activity or response was tested in the presence of prednisolone (1 and 10 micrograms/ml). Corresponding with these recoveries, suppressor T-cell activity arrived at normal values after corticosteroid therapy for 8 weeks. By contrast, in the corticosteroid-ineffective group, no apparent effects of prednisolone on suppressor T-cell activity and the response to IL-2 were observed. The relationship between the clinical effect of corticosteroid therapy and in vitro improvement in suppressor T-cell activity or in the response to IL-2 by prednisolone suggests that, in CAH, the corticosteroid effect is likely to be due to an immunomodulation in T-cell function.     

T-cell subsets in the hyporesponsiveness to hepatitis B surface antigen (HBsAg) and antigen-specific suppressor lymphocytes in chronic hepatitis B virus (HBV) infection

In contrast to convalescent hepatitis B patients, who exhibit the ability to elicit a specific immune response to HBsAg, patients with chronic hepatitis B virus (HBV) infection are markedly hyporesponsive to HBsAg and show a decrease in the normal ratio of OKT4-positive (helper/inducer) to OKT8-positive (suppressor/cytotoxic) lymphocytes. In this study the role of OKT4-positive and OKT8-positive cells on cellular immune response to HBsAg was evaluated in patients with chronic HBV infection and the ability of such patients to develop antigen-specific suppressor lymphocytes after in vitro sensitization to HBsAg. Elimination of OKT8-positive cells markedly improved the in vitro lymphocyte proliferative response to HBsAg without altering the reactivity of cells from the same donor to PPD or Candida. In contrast, the degree of responsiveness to HBsAg was not affected by the depletion of OKT4-positive cells. In vitro co-culture experiments, performed in the seven chronically HBV-infected patients who showed a proliferative response when their PBM were cultured with purified HBsAg or PPD, have demonstrated that lymphocytes from chronic HBV carriers, stimulated with HBsAg, inhibit the response of autologous PBM to HBsAg but not to the unrelated antigen PPD.     

Treatment of combined immunodeficiency with thymic extract (Thymostimulin)

A 14-day-old Chinese male baby was admitted with extensive skin lesions. A wound culture grew Staphylococcus aureus, Acinetobacter anitratus, Enterobacter cloacae, and Candida albicans and a blood culture grew group A beta-Streptococcus hemolyticus. The patient's lymphocyte counts were low and his lymphocytes were unable to produce IgG and IgA in vitro. The immunoglobulin-bearing cell studies also failed to demonstrate IgG and IgA bearing cells. Active Tac+ T cells, total T cells, and T cell subsets were at very low levels. Lymphoproliferative response to mitogens was also poor. Migration inhibitory factor production to Candida antigen was also decreased. The initial lymph node biopsy demonstrated no follicular formation and extensive depletion of lymphocytes in both thymic-dependent and thymic-independent areas. After Thymostimulin (a specific bovine thymic extract, TP-1) treatment, the second lymph node biopsy demonstrated germinal centers containing IgA-bearing cells and IgM-bearing cells and, subsequently, cortical and medullary differentiation. Serum IgG, IgA, and IgM became detectable at low levels and IgG-, IgA-, and IgM-bearing lymphocytes appeared in the peripheral blood. This also correlated with in vitro immunoglobulin synthesis. Active Tac+ T cells, total T cells, T cell subsets and lymphoproliferative response to mitogens increased gradually after thymostimulin therapy. This investigation demonstrated the therapeutic effectiveness of Thymostimulin in combined immunodeficiency both histologically and immunologically and the successful reconstitution of B cell function that did not require continued therapy.