Immune status in Crohn's disease : 3. Peripheral blood B lymphocytes, enumerated by means of F(ab)2-antibody fragments, Null and T lymphocytes

In the peripheral blood of patients with Crohn's disease (CD) the numerical distribution of the three major B lymphocyte subsets was determined by the identification of surface immunoglobulins using F(ab)₂-antibody fragments. T cell counts were also obtained and the number of null cells was calculated. Twenty-eight patients with Crohn's disease including 14 patients with previously untreated and very short-standing disease (group CD 1) and 14 patients with long-standing and/or previous drug treated disease (group CD 2) were compared with 28 sex and age-matched normals as well as with 13 patients with acute inflammatory bowel disease (group D). Patients in group D and inactive patients of group CD 1 showed a significant absolute lymphocytosis due to an increase in both the three B cell subsets and the T cells, without changes in the null cells. While the proportion of T cells was normal, there was a significant relative B lymphocytosis and a relative null cytopenia in these patients. Active CD 1 patients, however, showed significantly lower absolute lymphocyte and T cell numbers. In group CD 2, there was a significant absolute lymphopenia caused by an equal decrease in B and T cells. Highly active CD 2 patients showed higher absolute null cell counts than inactive patients. With increasing disease duration there was a significant decrease of the relative and absolute B cell concentrations. The data obtained suggest that T and B cell populations in the peripheral blood are reduced in certain patients with Crohn's disease and that this occurs secondarily to activity of disease, chronicity of disease, and the effects of therapy.     

T-Lymphocyte Subpopulation in Untreated SLE

An active subpopulation of T lymphocytes characterized by their ability to form early rosettes with sheep erythrocytes (active E-RBL) was studied in the blood of 50 patients with untreated systemic lupus erythematosus (SLE) and in 50 normal controls. The findings were related to the absolute number of circulating lymphocytes and total E-receptor-bearing lymphocytes (total E-RBL). Lupus patients with active disease had markedly decreased absolute lymphocyte counts, but the decrease of both the total and the active E-RBL surpassed what would be expected from the lymphopenia. Patients with inactive disease had moderately decreased absolute lymphocyte counts with a marked and disproportionate decrease in total E-RBL and a moderate decrease in active E-RBL, which seemed to reflect only the absolute lymphopenia. Patients with active disease had significantly lower active E-RBL than those with inactive disease. The changes of these and other lymphocyte subpopulations in relation to disease activity in SLE may reflect the influence of factors leading to T-cell depletion and immaturity. Circulating thymic products may be one of those factors.     

慢性乙型肝炎患者外周血淋巴细胞亚群与病程相关性的研究

目的对慢性乙型肝炎轻中度、重度和肝硬化患者外周血淋巴细胞亚群的百分比和绝对细胞数进行观察,探讨慢性乙型肝炎患者外周血淋巴细胞亚群的变化与病程的关系.方法采集88例慢性乙型肝炎患者柠檬酸钠新鲜抗凝血,经流式细胞仪进行免疫分型检测.结果慢性乙型肝炎重度患者的CD3+CD4+细胞百分比显著低于轻中度患者(P＜0.05),肝硬化患者的CD3+和CD3+CD8+细胞百分比显著低于轻中度患者(P＜0.01).肝硬化患者CD3CD19+细胞百分比显著高于重度和轻中度患者(P＜0.01).CD4/CD8比例在慢性乙型肝炎轻中度、重度和肝硬化患者间无显著差异.肝硬化和重度患者淋巴细胞、CD3+、CD3+CD4+、CD3+CD8+细胞的绝对细胞数均显著低于轻中度患者(P＜0.01),且肝硬化患者CD3-CD16+56+细胞的绝对细胞数显著低于轻中度患者(P＜0.05).肝硬化患者与轻中度患者的DNA载量分布差异有显著性(P＜0.01),其高水平病毒载量的患者比例高于轻中度患者.结论慢性乙型肝炎轻中度发展为重度和肝硬化的过程中,外周血淋巴细胞亚群绝对细胞数随病情的进展显著减少,主要表现为CD3+CD4+和CD3+CD8+的T淋巴细胞亚群的百分比进行性降低.     

Immunological studies in patients with Crohn's disease.

An investigation of immunological parameters was conducted in 38 patients with Crohn's disease. The immunological tests employed included skin tests with dinitrochlorobenzene and a battery of common skin test antigens, lymphocyte transformation with phytohaemagglutinin and pokeweed mitogen, serum immunoglobulins, and absolute lymphocyte counts. Crohn's disease patients were divided into two groups, those treated with immunosuppressive drugs and those not receiving immunosuppressive medications. The latter group was subdivided into patients with active and inactive disease. Immunosuppressed patients with Crohn's disease did not develop sensitivity to dinitrochlorobenzene and had mildly depressed skin test reactivity to common skin test procedures. Non-immunosuppressed patients with active Crohn's disease also reacted less frequently to common skin test antigens, but 16 of 17 such patients developed sensitivity to dinitrochlorobenzene. Lymphocyte transformation with phytohaemagglutinin and pokeweed mitogen was normal in all groups of patients with Crohn's disease. However, when suboptimal incubation periods were used with phytohaemagglutinin stimulation, there was a significant difference between Crohn's disease patients and controls. Serum immunoglobulin levels and absolute lymphocyte counts were normal in all Crohn's disease patients. We conclude that immunity in Crohn's disease is qualitatively normal.     

T lymphocyte subsets in inflammatory bowel disease: peripheral blood

Peripheral blood T lymphocytes and T lymphocyte subsets have been quantified in 28 patients with ulcerative colitis and 26 with Crohn's disease by an indirect immunofluorescence technique using monoclonal antibodies: OKT3, which detects all peripheral blood T lymphocytes; OKT4 (T cells of helper phenotype); and OKT8 (T cells of supressor-cytotoxic phenotype). Eighteen normal subjects and 16 patients with a variety of non-inflammatory gastrointestinal disorders were studied as controls. No significant differences were found between patient and control groups in the proportions of circulating T lymphocytes or their subsets. When compared with normal subjects, absolute numbers of T lymphocytes were reduced in patients with active ulcerative colitis or Crohn's disease (p less than 0.05). OKT4+ T cell numbers were reduced in ulcerative colitis, whether active (p less than 0.02) or inactive (p less than 0.05) and in active Crohn's disease (p less than 0.05) Numbers of OKT8+ T cells were reduced in active Crohn's disease (p less than 0.01). There were no differences in T lymphocyte numbers between the patient groups and the disease control subjects. The OKT4+:OKT8+ ratio in patients with inflammatory bowel disease did not differ from that in controls. No relation was found between any of the parameters studied and disease activity, site, or extent of disease, or treatment with sulphasalazine or corticosteroids. The presence of Ia-like, HLA-DR antigens on T cells was detected using a double marker immunofluorescence technique. In control subjects up to 7% of OKT3+ cells were HLA-DR+. In only three patients was the proportion of HLA-DR+ cells greater than in controls. These results indicate that the pathogenesis of ulcerative colitis or Crohn's disease does not depend upon an alteration in the proportion of circulating T lymphocytes nor upon an imbalance of T lymphocyte subsets as defined by monoclonal antibodies. The reduction in T lymphocyte numbers may result from mucosal infiltration. The findings also suggest that circulating T lymphocytes are not activated.     

Lymphocytotoxicity test against rabbit hepatocytes in chronic liver diseases.

We have studied the cytotoxicity against rabbit liver cells of lymphocytes from the peripheral blood of 71 patients with various liver diseases. The group with chronic active hepatitis and three patients with acute alcoholic hepatitis showed significantly higher mean values of lymphocytotoxicity (P less than 0.001) compared with the other patients with chronic persistent hepatitis, post-necrotic fibrosis and cirrhosis. Wilson's disease, and prolonged viral hepatitis. The mean cytotoxicity of these last groups did not differ significantly from controls. In four out of six patients with chronic active hepatitis a significant decrease of lymphocytotoxicity was found after immunosuppressive therapy with oral prednisolone. A good correlation between the lymphocytotoxicity test and histological signs of activity suggests that a cell-mediated immune aggression is present in this disease.     

Killer cell (K-cell) population in chronic liver disease

Peripheral blood killer cell (K-cell) population of patients with chronic hepatitis was investigated by means of a plaque-assay method using sheep red blood cells. The mean K-cell population of 14 control subjects was 5.1 +/- 2.0% (mean +/- SD), and that of 28 patients with chronic hepatitis was 4.4 +/- 3.1%. These 28 patients were divided into three groups: CPH, CAH 2A and CAH 2B. The mean K-cell population of each group was decreased in order of the severity of the disease. Especially, that of patients with CAH 2B was a statistically significant decrease (p less than 0.01) from control subjects. In the course most patients with CAH, K-cell population did not change for three months after admission. K-cell population was observed to decrease in the patients with active stage of liver cirrhosis, but not in patients with the terminal stage of liver cirrhosis, even in hepatoma patients. It is suggested that the K-cell population may play an important role of pathogenesis of chronic hepatitis.     

Circulating lymphocyte subpopulations in Crohn's disease

Circulating lymphocytes were enumerated in 28 patients with Crohn's disease and in 12 patients with other diseases by rosetting and by immunofluorescent staining using monoclonal antibodies for T-cell surface phenotypic markers [OKT3 (mature), OKT4 (helper), and OKT8 (suppressor/cytotoxic)] or polyvalent antisera for surface immunoglobulins (B cells). Total lymphocyte counts were reduced only in those with non-steroid-treated active Crohn's disease. Circulating monocyte counts, proportions of peripheral T and B cells, and percentages and absolute numbers of mature, helper, and suppressor T-cell subclasses in Crohn's disease were not significantly different than in the controls. Helper to suppressor T-cell ratios were comparable in all subjects, varying directly with numbers of helper T cells (p less than 0.05). Individual ratios of helper to suppressor T cells did not correlate with disease activity or location, the use of steroids, serum albumin, or total lymphocyte or monocyte counts. This study provides no evidence for underlying abnormalities of circulating lymphocyte subpopulations in Crohn's disease when compared to subjects with other illnesses. The characterization of lymphocyte subclasses in affected tissues is an important area of continuing investigation.     

Lymphocyte subpopulations in sarcoidosis: correlation with disease aetivity and duration.

We studied lymphocytes from the peripheral blood of 29 sarcoid patients to relate T- and B-cell populations with disease activity and duration. In patients with acute (less than 1 year) and chronic (greater than or equal to 9 years) active disease, the absolute lymphocyte count was reduced; the absolute number of T cells was reduced; and the proliferative response of lymphocytes to phytohemagglutinin was depressed compared with control subjects. Ten of 21 patients with active disease had 5% to 32% atypical lymphocytes. The proportion of cells bearing surface immunoglobulin (Ig) was increased in patients with active disease, but the absolute number of cells bearing surface Ig did not differ significantly from controls. However, studies using overnight in-vitro culture indicated that a large fraction of these cells bound exogenous Ig. The number of cells indentified by complement receptors was significantly reduced in patients with active disease. In contrast, patients who had complete reasolution of their disease showed no significant differences from controls in either T- or B-cell populations or in the proliferative response of their lymphocytes to phytohemagglutinin.     

T-lymphocyte cytotoxicity to HBsAg-coated target cells in hepatitis B virus infection

The cytotoxic effect of peripheral lymphocytes on chicken red blood cells (ChRBC) coated with purified hepatitis B surface antigen (HBsAg) has been studied as an in vitro parameter of cell-mediated immunity in acute and chronic infection with hepatitis B virus. Using this technique, the mean cytotoxic index of lymphocytes from patients with acute hepatitis B (29·13 ± 20·88) was significantly higher than that obtained with lymphocytes from control subjects (6·53 ± 3·75). Only 33·3% of the patients with HBsAg-positive chronic active hepatitis exhibited lymphocyte cytotoxicity to HBsAg-coated target cells and the mean cytotoxic index (11·66 ± 6·60) was in these cases significantly lower than that found in acute hepatitis B. Healthy chronic carriers of HBsAg failed to show lymphocyte cytotoxicity to target cells. The effector cells detected in acute hepatitis B by this in vitro assay have been demonstrated to be T-lymphocytes, as T-cell depleted subpopulations lacked cytotoxic activity. Target cell lysis could be blocked by addition of HBsAg-coated unlabelled ChRBC as well as of purified HBsAg in the culture tubes. It is suggested that damage to the liver cells in acute hepatitis B is related to the presence of cytotoxic T-lymphocytes reacting with HBsAg on the surface of infected hepatocytes. An inadequate lymphocyte response to the antigen may be responsible for the persistence of the infection in the liver with varied clinical manifestations and associated hepatic lesions.     

Lymphocyte subpopulation in acute viral hepatitis.

Studies of peripheral blood lymphocytes were performed in 41 patients with acute viral hepatitis, in grade III-IV coma; 16 patients were in the third trimester of pregnancy. There were significant reductions in absolute lymphocyte count and T cell number in patients who succumbed to the disease, when compared with those who survived. B cell counts were similar in the two groups and migration inhibition test with BCG antigen was normal. It is postulated that a decrease in the number of cells interacting in cell-mediated immune reactions is related to prognosis in acute viral hepatitis.     

Lack of in vivo activation of the interferon system in HBsAg-positive chronic active hepatitis

The in vivo activation state of the interferon system was biochemically evaluated in patients with HBsAg-positive liver disease by assaying the interferon-induced enzyme, 2'5'-oligoadenylate synthetase, in peripheral blood mononuclear cells. All patients with chronic active hepatitis had normal levels of enzyme activity. Increased values were found in 77% of patients with acute hepatitis, 50% of those with chronic persistent hepatitis and 54% chronic healthy carriers. These results provide evidence for lack of activation of the interferon system in HBsAg-positive chronic active hepatitis and support the hypothesis that an in vivo defective interferon response may aid in development of chronic active hepatitis.     

Azathioprine-Associated acute pancreatitis in the course of chronic active hepatitis

We have described a 22-year-old man with an HBsAg-positive chronic active hepatitis who developed an attack of pancreatitis during the course of treatment with prednisone and azathioprine. Clinical and biochemical abnormalities subsided when azathioprine was stopped and reappeared after rechallenge with the drug. Azathioprine-associated acute pancreatitis is well recognized mostly during treatment of Crohn's disease and after renal transplantation. This adverse effect should, therefore, also be kept in mind during the treatment of chronic active hepatitis.     

Immunosuppressive serum factors in viral hepatitis. III. Prognostic relevance of rosette inhibitory factor and serum inhibition factor in acute and chronic hepatitis

Two immunosuppressive serum factors, serum inhibition factor (SIF) and rosette inhibitory factor (RIF), were studied in sera from patients with acute and chronic viral hepatitis. In a study of 30 patients with acute viral hepatitis, an association was found between RIF, SIF, and biochemical and virological parameters in 27 patients (90%), 25 of whom recovered completely; two had a protracted course. In three patients, the clinical course was not reflected by the immunosuppressive factors. In 26 patients with chronic persistent hepatitis, 3 had RIF and 7 had SIF of low activity. In patients with HBsAg-positive and -negative chronic active hepatitis, 32 of 47 had RIF and 24 had SIF. SIF activity was significantly increased in HBsAg positive as compared to -negative cases. There was no correlation between RIF and SIF activity at any stage of viral hepatitis. Although SIF was demonstrated in patients with various infectious and other inflammatory diseases, RIF was infrequently detected in nonviral liver disorders, and was not present in any of the nonhepatic diseases tested. It was confirmed that RIF is associated with the beta-lipoprotein fraction. RIF was easily separated from SIF by density gradient ultracentrifugation. The evaluation of SIF and RIF may be helpful in determining the outcome of acute viral hepatitis. In chronic hepatitis, RIF was a better indicator of disease activity than was SIF. These clinical data support previous findings that SIF may be related to the immune response whereas RIF is associated with liver cell damage.     

Peripheral blood lymphocyte subsets in patients with chronic hepatitis C – effects of interferon treatment

Abstract: Thirty-three patients with chronic hepatitis non-A, non-B/C were included in a randomized controlled study of recombinant alpha-2b interferon treatment 3 MU three times weekly for 36 weeks. In lysed whole blood, lymphocyte subpopulations were enumerated by flow cytometry detecting fluorescein or phycoerytrin conjugated monoclonal antibodies directed against seven different epitopes. Patients with chronic active hepatitis were significantly older than patients with chronic persistent hepatitis (p>0.05). Before treatment, the proportions of different subsets of lymphocytes were within the normal reference values and the CD4/CD8 ratio was also normal. No increased activation of T-cells was noticed. Patients over 50 years of age, however, had a significantly increased (p>0.01) proportion of HLA-DR + lymphocytes, mainly B-cells. Treatment decreased the absolute number of peripheral blood leukocytes and lymphocytes. There was also a significant decline in the proportion of CD8 + lymphocytes and NK-cells, and a significant increase in the proportion HLA-DR + cells and of the CD4/CD8 ratio. The increased proportion of HLA-DR + cells, however, did not reflect peripheral T-cell activation; instead, it was due to increasing B lymphocyte numbers.     

Immune status in Crohn's disease. VI. Immunoregulation evaluated by multiple, distinct T-suppressor cell assays of lymphocyte proliferation, and by enumeration of immunoregulatory T-lymphocyte subsets

In 28 patients with Crohn's disease, 6 patients with ulcerative colitis, and 34 healthy controls, immunoregulatory function of peripheral blood mononuclear cells was investigated by evaluating the suppression of lymphocyte proliferative responses to mitogens (phytohemagglutinin, concanavalin A, pokeweed mitogen) and to allogeneic lymphocytes (mixed lymphocyte culture) using simultaneously five functional assays as follows: (a) spontaneous T-suppressor-cell activity, (b) concanavalin A-generated T-suppressor-cell activity, both with (3000 rads) and without irradiation of suppressor cells, and (c) allogeneic mixed lymphocyte culture-generated T-suppressor-cell activity, again both with and without irradiation. Concanavalin A- and mixed lymphocyte culture-generated T-suppressor-cell activities were evaluated both in the autologous and the allogeneic system. In addition, using monoclonal antibodies, we determined proportions of T-helper and T-suppressor/cytotoxic lymphocytes. Inactive patients did not differ from normal either in the proportions of immunoregulatory lymphocytes or in the suppression of the various lymphocyte proliferative responses in any of the five T-suppressor-cell assays evaluated. In patients with active disease, however, an impairment of suppression of phytohemagglutinin-, pokeweed mitogen-, and mixed lymphocyte culture-stimulated proliferation of autologous lymphocytes was observed in the concanavalin A-generated, irradiated suppressor assay. In the spontaneous suppressor assay, suppression of phytohemagglutinin- and concanavalin A-stimulated lymphocyte proliferation was significantly lower in active disease than in remission. Thus, in peripheral blood of patients with Crohn's disease who are in remission, there is no indication for an immunoregulatory defect in any of the evaluated assay systems. Single selective, moderate defects in suppression of proliferation of various lymphocyte subpopulations are restricted to active disease.     

T lymphocyte sensitization to HBcAg and T cell-mediated unresponsiveness to HBsAg in hepatitis B virus-related chronic liver disease

Using a newly developed indirect T lymphocyte migration inhibition test, cell-mediated immunity to HBsAg and HBcAg was directly and simultaneously examined in a total of 21 patients with HBsAg-positive chronic liver disease (CLD), and in seven subjects whose sera contained anti-HBs (2 previous acute hepatitis B; 4 hepatitis B vaccine recipients and 1 chronic active hepatitis). T cell sensitization to HBcAg was invariably detected in the HBsAg-positive CLD patients tested (12/12), whereas T cell sensitization to HBsAg was not present in any of the patients (0/21). In contrast, T cell sensitization to HBsAg was present in all anti-HBs-positive subjects. These results support the hypothesis that the cellular immune response to HBcAg, rather than to HBsAg, is implicated in the pathogenesis of HBsAg-positive CLD. Moreover, the observation that the addition of T cells from patients with HBsAg-positive CLD to T cells from anti-HBs positive subjects in a ratio of 1 to 9 reversed their sensitization to HBsAg, suggests that a hyperactivity of HBsAg-specific suppressor T cell population may be responsible for persistent HBs antigenemia.     

Hepatic expression of intercellular adhesion molecule-1 (ICAM-1) in viral hepatitis B

The in situ distribution patterns of intercellular adhesion molecule-1 and human leukocyte antigen-DR antigens were studied in serial sections of 61 liver biopsy specimens from patients with hepatitis B virus infection using immunohistochemical techniques. In addition, the topographical relationship between the display of HBcAg on one hand and the expression of intercellular adhesion molecule-1 by hepatocytes on the other was analyzed with a double-staining immunohistochemical procedure in 14 selected liver biopsy samples showing chronic persistent or chronic active hepatitis and signs of active hepatitis B virus replication as reflected by the presence of variable amounts of HBcAg in a nuclear or cytoplasmic pattern of immunoreactivity. Coexpression of intercellular adhesion molecule-1 and human leukocyte antigen-DR antigens by hepatocytes correlated positively with the site and extent of the inflammatory infiltrate, which was composed of lymphocytes expressing lymphocyte function-associated antigen-1. In healthy HBsAg-positive carriers without inflammatory liver disease, no intercellular adhesion molecule-1 or human leukocyte antigen-DR expression was found on hepatocytes; in acute hepatitis, intercellular adhesion molecule-1 and human leukocyte antigen-DR were strongly expressed throughout the liver parenchyma on liver cell membranes and on sinusoidal lining cells. In chronic persistent and chronic active hepatitis and in active cirrhosis, intercellular adhesion molecule-1 and human leukocyte antigen-DR showed membranous positivity on focal clusters of hepatocytes in areas of periportal or intraacinar inflammation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of HBV replication and delta agent superinfection on T cell subsets and killer (Leu 7+) in chronic hepatitis B virus infection

Peripheral blood T-lymphocyte subsets and cells reacting for Leu 7 antigen, which identifies a subset of killer and natural killer cells, have been examined in 32 patients chronically infected by the hepatitis B or D viruses (HBV, HDV) and in 28 normal subjects. The T8+ lymphocytes were increased and the T4/T8 ratio was decreased in patients with HBV replication (identified by the presence of HBcAg in liver and HBV-DNA in serum) and in patients with HDV infection (HDAg in liver). These patients had more active liver disease than patients without evidence of viral replication, B or D, who showed normal lymphocyte counts. Leu 7+ lymphocytes were also increased in patients with viral replication and active disease and correlated positively with alaninaminotransferase serum levels. These observations suggest the participation of both T8+ and Leu7+ cells in the pathogenesis of liver cell injury in HBsAg-positive chronic liver disease.     

Increase in okm1 + granular lymphocytes in patients with rheumatoid arthritis

Patients with very active rheumatoid arthritis that was being treated only with nonsteroidal antiinflammatory drugs had increased numbers of peripheral blood OKM1 + lymphocytes. In 3 patients, 90° light scatter analysis revealed a double lymphocyte peak. When sorted, the high scatter peak contained a large percentage of granular lymphocytes. Patients with mild-to-moderately active rheumatoid arthritis had normal levels of OKM1 + lymphocytes, but when the drugs were discontinued, the activity of the disease and the numbers of OKM1 + cells increased. Administration of piroxicam was associated with clinical improvement and a decrease in levels of OKM1 + cells. OKM1 + granular lymphocytes are increased in some rheumatoid arthritis patients, and their numbers may correlate with clinical disease activity and/or therapy.