Immunological Profile in Children with Minimal Change Nephrotic Syndrome

ABSTRACT. Peripheral blood from patients with active stage of minimal change nephrotic syndrome (MCNS) was examined for concanavalin A (ConA)-inducible suppressor T cell activity, proliferative response to phytohemagglutinin (PHA) and in the autologous (AMLR) and allogeneic (MLR) mixed lymphocyte reaction, proportions of T cells with receptors for IgM (Tu) or IgG (Tγ) and the levels of serum immunoglobulin M, G and A. Six of 9 patients with MCNS studies showed deficiency of ConA-induced suppressor cell activity. In the AMLR, only one of 9 patients with MCNS demonstrated depressed proliferative response (p<0.05). In the allogeneic MLR, T cells from 5 of 9 patients with MCNS demonstrated poor proliferative response when stimulated with normal control non-T cells. Five of 9 patients with MCNS had depressed proliferative response to PHA. The proportion of total T cells, Tu cells and Ty cells in the patient group were comparable to healthy control group. Serum IgG was significantly decreased in 7 of 11 patients. This study demonstrates multiple immunological abnormalities in patients with MCNS that might play a role in its pathogenesis.     

Enhanced suppressor T cell activity resulting in increased IgM and decreased IgG productions in children with minimal change nephrotic syndrome

In order to clarify the cellular basis accounting for the decreased serum IgG and increased serum IgM in children with minimal change nephrotic syndrome (MCNS), peripheral blood mononulcear cells (MNC) were obtained from 15 children with biopsy-proved MCNS and 15 age- and sex-matched normals. MNC were then separated into helper (OKT4) T cells, suppressor (OKT8) T cells, B cells and monocytes. Different cell populations from patients and normals were then recombined and pokeweed mitogen-stimulated in vitro immunoglobulin biosynthesis was studied. The results strongly suggest the presence of isotype-specific suppressor T cells which may affect the switch of IgM B cells to IgG B cells in patients with MCNS and may be used to explain partly the clinical findings of lowered serum IgG and increased IgM in those individuals.     

Suppressor T-cell activity in newborns and mothers

We studied autologous and allogeneic concanavalin A (Con A)-induced suppression of proliferative responses to phytohemagglutinin (PHA) and Con A in peripheral blood mononuclear cells in 24 normal newborns and compared the results with those obtained from 20 normal older children. Three concentrations of PHA and one of Con A were used to stimulate responder cells. Suppressor activity, elicited in the stimulated cell population after 48 h pre-incubation with Con A, was expressed as percentage inhibition of the proliferative response to PHA. There was an inverse relationship between PHA concentration and suppressor activity, and autologous suppression was greater than allogeneic suppression within each group of patients and at each mitogen concentration. In both the autologous and allogeneic systems, older children showed more suppressor activity than newborns. Feto-maternal pairing showed that newborns efficiently suppress their mother's mitogenic responses, but the mothers do not suppress their own or other newborn's lymphocytes, despite having normal autologous suppressor capability. We suggest that suppressor activity by the fetus and it's inhibition in the mother may play a part in the mechanism for controlling maternal-fetal immune rejection.     

DiGeorge syndrome with hypogammaglobulinaemia: a patient with excess suppressor T cell activity treated with fetal thymus transplantation

A male infant with DiGeorge syndrome had hypogammaglobulinaemia with a normal number of B cells. CD3(+) T cells were reduced and the CD4(+)/CD8(+) ratio was reversed. Proliferative responses of T cells to mitogens and to allogeneic cells were low. The pokeweed mitogen (PWM)-induced B cell differentiation assay revealed a higher than normal suppressor T cell activity. This suggests that some T cells had differentiated into functionally mature cells resulting in an imbalance of regulatory T cell functions and that excess suppressor activity might play a role in hypogammaglobulinaemia. Fetal thymus transplantation improved both cellular and humoral immunity. The patient's susceptibility to viral and bacterial infections, proliferative response of T cells and serum Ig concentration returned to normal. The excess suppressor activity seen before transplantation disappeared. Hypocalcaemia did not improve. These results show that fetal thymus transplantation was effective not only in reconstituting cellular immunity but also in normalizing the imbalance of regulatory T cell functions in this patient with DiGeorge syndrome.Abbreviations MNC mononuclear cells - ConA concanavalin A - PHA phytohaemagglutinin P - PWM pokeweed mitogen - ³H-TdR tritiated thymidine - PBL peripheral blood lymphocytes     

Restoration of suppressor T-cell functions in children with AIDS following intravenous gamma globulin treatment

Suppressor T-cell function was analyzed in seven children with acquired autoimmunodeficiency syndrome (AIDS) or AIDS-related complex (ARC). Four of the patients had markedly increased serum IgG levels. All patients had elevated percentages and absolute numbers of peripheral blood T8 cells. In vitro concanavalin A generation of suppressor cells for T-cell mitogenic responses and suppression of pokeweed mitogen-driven immunoglobulin secretion were diminished in all patients. After intravenous treatment with gamma globulin, four patients regained in vitro suppression of pokeweed mitogen-driven gamma globulin secretion. Treatment with intravenous gamma globulin also modified in vitro suppressor T-cell functions in children with AIDS or ARC.     

Maternal fetal immunological relationship particularly mycobacterial immunity.

Thirty-nine paired maternal and cord blood from normal full term deliveries were tested for lymphocyte function by proliferative response to mitogens-Phytohemagglutinin-P (PHA) and Poke week mitogens (PWM). Monocyte function was assessed by the ability of the monocytes to release hydrogen peroxide (H2O2) in response to standard stimulus (PMA). Mycobacterial immunity was assessed by lymphocyte proliferative response to purified proteins derivative (PPD) and IgM and IgG antibody response to H37Rv and 5 atypical mycobacteria. Lymphocyte functions were significantly lower in cord blood (PHA 20.6, PWM 21.2) as compared with maternal blood (PHA 65.8, PWM 37.8). The capacity of fetal monocytes to release H2O2 was comparable to maternal monocytes. The mean proliferative response of fetal lymphocytes to tubercular protein (PPD) was 0.67 as compared (P less than 0.01) to maternal lymphocytes (3.79). Nearly 86% of the cord blood did not show any response to PPD. None of the cord blood showed IgM antibody response to H37Rv nor to any of the range of 5 atypical mycobacteria though maternal IgM and IgG response was present. There was only passive transfer of IgG antibody from mother to fetus. Hence, though this is a highly endemic area for atypical mycobacteria and M. tuberculosis, there was apparently no transplacental transfer of antigen in normal sensitized mothers.     

Abnormal lymphocyte function in childhood leukemia

Lymphocyte function was evaluated in 26 untreated children with acute lymphoblastic leukemia (ALL) and 5 patients with acute myelogenous leukemia (AML) by stimulation with phytohemagglutinin (PHA) in dose- and time-response studies. The response to PHA correlated positively with the percentage of lymphocytes (r = + 0.786) and negatively with the percentage of lymphoblasts (r = ? 0.728). Sixteen patients with a WBC < 20,000/cu mm (51 ± 5% lymphocytes, 28 ± 7% lymphoblasts, and 39 ± 20% T cells) responded normally (81,156 ± 8,229 cpm) on the fourth or fifth day of culture. Ten patients with a WBC > 20,000/cu mm (10 ± 3% lymphocytes, 88 ± 4% lymphoblasts and 18 ± 28% T cells) had a significantly lower response to PHA on these days (13,609 ± 5,568 cpm). Six of the ten high-WBC patients had a delayed peak response on the sixth or seventh day of culture. This abnormal response to PHA is similar to that which has been described in patients with chronic lymphatic leukemia and is at least partially due to the dilution of a normal lymphocyte population by the proliferation of a population of non-PHA-responsive lymphoblasts. The remaining four high-WBC patients had flat PHA dose- and time-response curves and a poorer clinical course. Two of them had T-cell leukemia. The absence of response to PHA in these children may characterize a group of patients with acute lymphoblastic leukemia who are immunodeficient at the time of diagnosis and may reflect the presence of a unique population of lymphoblasts with suppressor activity.     

Peripheral blood mononuclear cell responses to heat shock proteins in patients undergoing stem cell transplantation

GVHD is a major complication after allogeneic SCT. Etiology of GVHD is multifactorial. Known role of hSPS in antigen presentation could suggest their potential role in the alloreactive process that leads to aGVHD. HSPS represent major immunodominant antigens in a wide spectrum of microbial pathogens. Bacterial and fungal colonization, infection and sepsis are frequent in immunocompromised patients with various malignant and non-malignant diseases. We studied PBMC responses to recombinant human hsp60 (rh-hsp60), rh-hsp70 and Mycobacterium bovis hsp65 (M. bovis hsp65) in relation to aGVHD and infection in 34 pediatric patients with various lympho-hemopoietic malignancies as well as non-malignant disorders subjected to SCT. PBMC of patients before initiation of preparative regimen as well as after engraftment were stimulated with hSPS (1 microg/mL/well, 7-day cultivation). PHA was used as a control of the stimulation ability. Cell responses were measured after the incorporation of 3H-thymidin (pulsing with 1 microCi/well) and were expressed as stimulation indexes (SI). We demonstrated significantly high proliferative response to rh-hsp60 as well as M. bovis hsp65 in a cohort of pretransplant patients with anamnestic and/or actual infection when compared with a cohort of patients without infection and healthy individuals. Strong PBMC cell responses to hSPS were found in patients who were at present colonized with Escherichia coli and Klebsiella pneumoniae or had previously K. pneumoniae infection with subsequent sepsis. Our findings support various studies dealing with immunodominant hSPS in connection with several pathogens and infectious diseases. Although no statistical difference for proliferative response to PHA was observed, PBMC responses against all tested hSPS comparing a cohort of patients with aGVHD and that with no sign of GVHD resulted in significantly lower SI for all tested hSPS in patients with aGVHD. Lower stimulation with hSPS during aGVHD might be explained by the stress-induced upregulation of self-hSPS synthesis that might lead to the inhibition of self-hSPS reactive T-cell response. Vice versa, we hypothesize that increased hsp-specific stimulation may reflect the presence of protecting regulatory T cells preventing the development of Th1-mediated diseases involving aGVHD.     

Juvenile rheumatoid arthritis. Monocyte dysfunction in selected patients

The in vitro parameters of cell-mediated immunity were studied in 20 children with an established diagnosis of Juvenile rheumatoid arthritis (JRA) (age range 4-15 years) and 23 age- and sex-matched healthy children. (No attempt was made to correlate the observed changes with clinical course or treatment). We are not certain, at this time, of clinical relevancy or the generalizability of the findings. The normal level of T-lymphocytes (CD3+) and normal proportions of CD4+ and CD8+ lymphocytes were seen in children with JRA. The in vitro response of lymphocytes to T-cell mitogen phytohemagglutinin (PHA) also was normal. The suppressor activity of JRA monocytes was essentially the same as controls. In contrast, monocytes from patients with JRA showed the following: decreased expression of receptors for Fc part of IgG immunoglobulin (FcR), diminished nitro blue tetrazolium (NBT) reduction activity, and depressed expression of Ia.7 major histocompatibility complex (MHC) class II determinants. This indicates that certain monocyte functions in selected patients with a variety of manifestations of JRA are depressed.     

Immunological study of childhood acute ITP at onset.

We attempted to search for any specific change in the immune system during the onset of childhood acute immune thrombocytopenic purpura (ITP) in order to clarify the pathophysiology of acute ITP by examining the lymphocyte subset, lymphocyte blastogenic response, serum complements, and immunoglobulins in 18 patients with childhood acute ITP and 18 controls (control values after normalization). At the onset of acute ITP, the levels of serum complements and IgG and IgA were found to be within the normal ranges, but serum IgM levels were greater than 200 mg/dL in six cases among 18 patients. Lymphocyte blastogenic response to phytohemagglutinin (PHA) and concanavalin A (ConA) was depressed in patients relative to controls (PHA: p less than 0.05, ConA: p less than 0.01). Lymphocyte blastogenic response to pokewood mitogen (PWM) was lower than that of the control, but no statistical significance was observed. There was no difference in the proportion of CD3, CD4, CD8, SmIg, SmIgG, SmIgM, SmIgA, and SmIgD. The CD4/CD8 ratio was not different from that of controls. The proportion of CD38 was higher than that of control, but no significant difference from the control was observed. Increase in the serum IgM level and proportion of CD38 and depressed lymphocyte blastogenic response may be the influence of preceding infection. It has been reported that the CD4/CD8 ratio is depressed due to an increase in the CD8 level in acute and convalescent phases of viral infection. However, the proportion of CD8 was not necessarily increased in our patient in whom preceding infection was obvious. The immunological status of the patients with acute ITP at onset differs from that after infection.     

T cell subsets in glomerulonephritis

Abnormal lymphocyte function has been postulated to have a pathogenetic role in nephrotic syndrome. In an attempt to investigate the pathogenetic role of lymphocyte subsets in human glomerular disease, we studied 110 children suffering from nephritis during the acute nephrotic phase or nephritis without steroid treatment, 4 weeks later after steroid treatment, in remission and relapse. These patients included minimal change nephrotic syndrome (MCNS) 15 cases, focal segmental glomerular sclerosis (FGS) 6 cases, mesangial cell proliferative nephropathy (MesPGN) 42 cases, membranoproliferative glomerulonephritis (MPGN) 2 cases, hepatitis B surface antigenemia associated with membranous nephropathy (HBVMN) 10 cases, IgA mesangial nephropathy (IgAN) without nephrotic syndrome 7 cases, poststreptococcal glomerulonephritis (PSGN) 24 cases and chronic glomerulonephritis (CGN) 4 cases. There was no significant difference in the total lymphocyte count of each different pathological group of nephritis except that lymphopenia was noted in the CGN patients. When the lymphocyte phenotypic profile was examined, OKT8 cells were significantly increased in the MesPGN patients and both OKT4 and OKT8 cells were significantly increased in HBVMN. Comparison of MCNS and MesPGN during the acute nephrotic phase showed the OKT4/OKT8 ratio decreased significantly in MesPGN. Four weeks after steroid treatment, OKT4 cells decreased both in MCNS and MesPGN being pronounced in MCNS. In the remission stage with steroid treatment the OKT4/OKT8 ratio decreased in MCNS and was mildly elevated in MesPGN. In relapse, the OKT4/OKT8 ratio was the same as it was during the onset of nephrotic phase. MCNS cases were steroid responsive whereas in MesPGN there were frequent relapses or partial steroid response.(ABSTRACT TRUNCATED AT 250 WORDS)     

Varicella arthritis. Report of a case

A rare case of varicella with bilateral acute arthritis of the knee is described. Clinical features and laboratory results of the patient are briefly compared with those of other sixteen cases previously published. The study of immune response shows a decrease of helper T cells (OKT4+) and an increase of suppressor/cytotoxic T cells (OKT8+) with a normal response of lymphocytes to PHA.     

Immunoregulation with levamisole in children with frequently relapsing steroid responsive nephrotic syndrome

The immunological and clinical effects of levamisole were studied in 10 children with frequently relapsing steroid responsive nephrotic syndrome (SRNS). The efficacy of the drug was tested during remission of the disease with all patients on alternate day steroid therapy. The lymphocyte proliferative response to phytohemagglutinin (PHA), concanavalin-A (Con-A) and pokeweed mitogen (PWM) were normal. The Con-A induced suppressor T-lymphocyte activity of 7 patients was low before treatment with levamisole 8 +/- 3.7% and increased to normal values during therapy 34 +/- 6%; p less than 0.001 (control 32 +/- 5%). In these 7 children prednisolone dosage could be decreased significantly or discontinued altogether (44.1 +/- 5.3%). Patients without immunoregulatory abnormalities did not respond to levamisole. In 3 out of 4 children tested the percentage of OKT8+ cells rose during levamisole therapy from 19.7 +/- 2.1 to 37 +/- 2.3 (p less than 0.001), thus correcting the elevated pre-treatment OKT4+/OKT8+ ratio from 3.1 +/- 0.2 to 1.5 +/- 0.2; p less than 0.001 (control 1.47 +/- 0.2). These data support the hypothesis that abnormal immunoregulation may play a role in the pathogenesis of SRNS. Treatment with levamisole can be useful in some patients with the frequently relapsing form of the disease.     

Nephrotic syndrome in mesangial proliferative lupus nephritis

Renal involvement is a major complication of systemic lupus erythematosus (SLE) and occurs in 30-70% of patients with SLE. Lupus nephritis is classified into six classes (I-VI) by the International Society of Nephrology and Renal Pathology Society (ISN/RPS). Although nephrotic syndrome is commonly associated with diffuse (ISN/RPS class IV) or membranous (ISN/RPS class V) lupus nephritis, several reports have described nephrotic syndrome in adult patients with minimal mesangial lupus nephritis (ISN/RPS class I) or mesangial proliferative lupus nephritis (ISN/RPS class II). However, nephrotic syndrome in mesangial proliferative lupus nephritis has rarely been reported in children. Although the pathogenesis of nephrotic syndrome with mesangial lupus nephritis is incompletely understood, three potential mechanisms have been postulated including lupus nephritis itself, non-steroidal anti inflammatory drug (NSAID)-induced minimal change nephrotic syndrome (MCNS) and coincidental occurrence of MCNS. We describe here a child with mesangial proliferative lupus nephritis who developed MCNS.     

Immunological Defects in a Bis(dichloroacetyl) diamine Induced Malformation Complex in Rats Closely Resembling the DiGeorge Syndrome

ABSTRACT: Spleen cells from newborn rats with bisdiamine-induced congenital malformation were examined for subpopulation of lymphocytes and responsiveness of the lymphocytes to phytohemagglutinin (PHA), concanavalin A (ConA) and lipopolysaccharide (LPS). T cells, but not B cells, were significantly diminished in the drug-treated newborn rats. Proliferative response of the lymphocytes to PHA and ConA, but not to LPS, was significantly depressed in the drug-treated newborns. Thus a defect in cell-mediated immunity was suggested to be associated with the congenital anomaly.
It is tempting to consider that this bisdiamine-induced malformation can be another animal model for the DiGeorge syndrome.     

Modulation of Immunological Abnormalities of Growth Hormone-Deficient Children by Growth Hormone Treatment

We studied the immunomodulatory role of growth hormone (GH) treatment in GH-deficient children. CH potentiated natural killer (NK)cell activity and the PHA and Con-A lymphocytoproliferative response. Two-color fluorescence using monoclonal antibodies (CLM, 2H4, CD8 and CD11) showed a moderate reduction of the helper T cells and NK cells, but no changes of suppressor T cells or cytotoxic T-cells during GH therapy. Cancer and slow viral infection in GH-deficient children have been watched for carefully before and after GH therapy.     

Fetal peripheral blood mononuclear cell proliferative responses to mitogenic and allergenic stimuli during gestation

Blood samples were obtained from fetuses and premature babies (n=51) (15-34 weeks gestation) to determine at what stage the fetal immune system was able to produce a positive proliferative response to common allergens. Peripheral blood mononuclear cells (PB MC) were stimulated with the mitogen, phytohaemagglutinin (PHA), and the allergens, house dust mite, cat fur. birch tree pollen, β-lactoglobulin, ovalbumin and bee venom (mellitin). Results were expressed as ratios of stimulated to unstimulated ³H thymidine incorporation, and as percent positive responders. There was an increase in proliferation ratio which correlated with increasing gestational age for PHA (p < 0.0001), cat fur (p=0.042), birch pollen (p=0.022) and β-lactoglobulin (p=0, 006). The point in gestation when cells from some individuals began responding to the allergens with a ratio of 2. 0 was at approximately 22 weeks. PBMC proliferative response ratios were higher from samples from babies > 22 weeks gestation compared to < 22 weeks for the mitogen and all allergens, except mellitin. There was also a greater proportion of positive responders from samples > 22 weeks compared to < 22 weeks for the mitogen and all allergens, except mellitin. Maternal exposure to birch pollen, which has a discrete season, was assessed to determine whether exposure had occurred at 22 weeks gestation or beyond. Results showed a higher prolifera tive response in infant cells stimulated with birch pollen (p=0.005) and higher proportion of positive responders (p=0.01) in the group of babies whose mothers had been exposed to hirch pollen beyond 22 weeks, compared to those whose mothers had not been so exposed. These results suggest that in utero fetal exposure to an allergen from around 22 weeks gestation may result in primary sensitisation to that allergen, leading to positive proliferative responses, at birth.     

Comparison of specificity between IgG,IgE and T cells to three casein components: Implication for the role of circulating allergenspecific T cells in food allergy

In order to investigate the role of food antigen-specific T cells circulating in the blood of patients with food allergy, we compared T cell response to three casein components (αs-, β- and, K-casein) with specificities of IgG and IgE binding to the casein components in four milk-allergic patients (P1-4) with atopic dermatitis. In all patients, the binding activities of IgG antibodies to αs-casein were most dominant, followed by those to β- and to K-casein. The major component of casein bound by IgE antibodies was αs-casein in P1 and P3, K-casein in P2, and αs-casein as well as K-casein in P4; the order of casein components bound by IgE antibodies was different from that by IgG antibodies. Proliferative responses of peripheral blood mononuclear cells (PBMC) to casein components were so low that the dominance of casein recognition could not be clearly demonstrated. However, short-term T cell lines that specifically respond to casein were successfully established from PBMC of the four patients and the proliferative responses of the T cell lines to the three components of casein were in accord with the IgE antibody specificity to casein components but not with that of IgG antibody specificity. When taken together, these results indicate that casein-specific T cells circulating in the blood are involved in or reflect an allergic reaction against casein.     

Immunoregulation with Levamisole in Children with Frequently Relapsing Steroid Responsive Nephrotic Syndrome

ABSTRACT. The immunological and clinical effects of levamisole were studied in 10 children with frequently relapsing steroid responsive nephrotic syndrome (SRNS). The efficacy of the drug was tested during remission of the disease with all patients on alternate day steroid therapy. The lymphocyte proliferative response to phytohemagglutinin (PHA), concanavalin-A (Con-A) and pokeweed mitogen (PWM) were normal. The Con-A induced suppressor T-lymphocyte activity of 7 patients was low before treatment with levamisole 8±3.7% and increased to normal values during therapy 34±6%; p <0.001 (control 32±5%). In these 7 children prednisolone dosage could be decreased significantly or discontinued altogether (44.1±5.3%). Patients without immunoregulatory abnormalities did not respond to levamisole. In 3 out of 4 children tested the percentage of OKT8⁺ cells rose during levamisole therapy from 19.7±2.1 to 37±2.3 ( p <0.001), thus correcting the elevated pre-treatment OKT4⁺/OKT8⁺ ratio from 3.1±0.2 to 1.5±0.2; p <0.001 (control 1.47±0.2). These data support the hypothesis that abnormal immunoregulation may play a role in the pathogenesis of SRNS. Treatment with levamisole can be useful in some patients with the frequently relapsing form of the disease.     

Drugs and depression of lymphocyte blastogenesis in children

Data are presented which demonstrate an inhibitory effect of non anti-blastic drugs with different therapeutical indications: Phenilbutazone (Ph), Thiamphenicol (Th) and Sulfamethopyrazine (Sl) on PHA and mixed stimulation of normal children peripheral blood lymphocytes, when added in vitro. Similarly a depressed response to PHA was observed for the lymphocytes of patients treated with the same drugs. When tested on the same experimental system, Diazepam (Dz) failed to show a significant inhibition.A first trial with a new antiblastic (cis-platinum II diammino dichloride) has also been performed. The in vitro addition of this antiblastic strongly depressed lymphocyte blastogenesis.The viability Trypan-Blue Exclusion Test was performed for all the drugs with inhibiting properties. No citotoxic effect could be seen when the 50% PHA-blastogenesis inhibiting dose was employed.Paper delivered at the 3rd ESPHI Scientific Meeting, Hamburg, September 9/10, 1972.