Critical residues of the Mycobacterium leprae LSR recombinant protein discriminate clinical activity in erythema nodosum leprosum reactions.

We reported earlier (S. Singh, N. P. Shanker Narayan, P. J. Jenner, G. Ramu, M. J. Colston, H. K. Prasad, and I. Nath, Infect. Immun. 62:86-90, 1994) that polyclonal antibodies directed against selective sequences in the Mycobacterium leprae recombinant protein designated LSR were present in lepromatous leprosy patients undergoing erythema nodosum leprosum (ENL) reactions (type 2 reactions). In this study using peptides with single-residue deletions from positions 6 to 24, we define three distinct regions, GVTY, NAA, and RGD, which were important for antibody recognition and for the discrimination of clinically silent and active ENL reactions. Antibodies against NAA were found only in patients undergoing active reactions. This is in contrast to the results for the RGD motif, which was recognized in all ENL patients, irrespective of the clinical status. Though GVTY was recognized in both groups of patients, its recognition was masked by the flanking glutamic acid. These findings point towards a specific molecular recognition pattern that emerges when a lepromatous leprosy patient undergoes immune perturbations leading to ENL reactions. Moreover, the fine specificity of immunological recognition changes during the natural evolution of the host-parasite interaction.     

Natural emergence of antigen-reactive T cells in lepromatous leprosy patients during erythema nodosum leprosum.

Fifteen lepromatous leprosy (LL) patients undergoing erythema nodosum leprosum (ENL) reactions were compared with 13 stable, uncomplicated, anergic individuals of the same leprosy background. ENL patients showed significant antigen-induced leukocyte migration inhibition (migration index = 0.058 +/- 0.01), paralleling the values obtained with a responder tuberculoid leprosy population (migration index = 0.04 +/- 0.004). Both phytohemagglutinin-induced general T-cell proliferation and, more significantly, antigen-induced lymphoproliferation were enhanced during the acute phase of the reaction. Suppressor cell activity, monitored by a costimulant assay, showed enhanced antigen-stimulated suppression of mitogen responses. Interestingly, the improvement in in vitro T-cell responses was not reflected in dermal reactivity, since 48-h delayed-type hypersensitivity responses after intradermal injection of soluble Mycobacterium leprae antigens continued to be poor. After subsidence of reactional lesions, leukocyte migration inhibition, lymphoproliferation, and suppressor cell activity were reduced to the unresponsive state seen in stable LL patients. Significantly, perturbations of T-cell reactivity are detectable in ENL reactions, indicating the natural but transient emergence of antigen-induced T cells in LL.     

Cell-mediated immunity in amyloidosis secondary to lepromatous leprosy.

Cell-mediated immunity in lepromatous leprosy patients with and without amyloidosis has been studied. Amyloidosis occurred mostly in patients with a history of recurrent erythema nodosum leprosum (ENL) reactions. For this reason, two control groups of leprosy patients were included, one having a history of recurrent ENL and the other little or no ENL. The lack of responsiveness to lepromin in vivo and in vitro, characteristic of lepromatous leprosy, was not altered by the presence of amyloidosis or a history of ENL. No significant difference between the patient groups was observed in the response to PPD in vitro, but skin reactivity to PPD was significantly lower in the patients with amyloidosis than in those without amyloidosis. In contrast, the PHA responses of patients with amyloidosis were significantly higher than those of control patients without a history of ENL, but not significantly different from those of control patients with a history of recurrent ENL. Lepromatous leprosy patients who develop amyloidosis thus appear to belong to a group, susceptible to repeated attacks of ENL, whose PHA responses are higher than those of other lepromatous leprosy patients. The lower skin reactivity to PPD observed in the amyloid group may reflect a general impairment in delayed cutaneous hypersensitivity.     

Polymorphonuclear cell function in the various polar types of leprosy and erythema nodosum leprosum.

Polymorphonuclear leukocyte motility, both in vivo and in vitro, and reduction of Nitro Blue Tetrazolium was studied in tuberculoid and lepromatous leprosy patients and a group of lepromatous patients with erythema nodosum leprosum (ENL). A profound defect in random migration, chemotaxis, and chemokinesis was found in lepromatous patients with and without complicating ENL, and marked depletion of skin window migration confirmed these in vitro findings. Tuberculoid patients exhibited a mild defect in polymorphonuclear leukocyte motility. Serum inhibitors of normal polymorphonuclear leukocyte chemotaxis were found in all types of leprosy, but sera from lepromatous and ENL patients were most inhibitory. Resting levels of Nitro Blue Tetrazolium reduction were normal in all three groups. Reconstitution of polymorphonuclear leukocyte cells from normal and ENL patients with ENL serum, however, showed increased Nitro Blue Tetrazolium reduction well above the normal range, whereas reconstitution with normal, lepromatous, and tuberculoid sera failed to increase Nitro Blue Tetrazolium reduction above the normal values.     

Immune complexes and complement hypercatabolism in patients with leprosy.

The occurrence of immune complexes in the serum and the level of the C3 breakdown product C3d in the plasma from patients with leprosy were studied by quantitative methods and the results were compared in various forms of the disease. These studies were performed on sixty-two samples from twenty-six patients. The serum 125I-C1q binding activity was found to be increased by more than 2 s.d., as compared to the normal values, in most of the sera from patients with erythema nodosum leprosum (ENL) (80%) and uncomplicated lepromatous leprosy (82%), but also in the sera from patients with tuberculoid leprosy (58%). In vitro studies suggested that immune complexes involving mycobacterial antigens were present in leprosy sera. An increased C3d level (greater than 2s.d.) was also found in most of the plasma from patients with ENL (70%), but rarely in the plasma from patients with uncomplicated lepromatous leprosy (18%) and never in tuberculoid leprosy patients' plasma. The absence of a significant correlation between the 125I-C1q binding activity and the C3d level in leprosy patients may suggest that extravascular immune complexes are involved in the complement activation occurring in ENL. The quantitation of C3d in plasma may be of some practical interest in the early diagnosis of ENL complications of leprosy.     

Glomerulonephritis in leprosy

A renal biopsy from a patient with lepromatous leprosy but no history of erythrema nodosum leprosum (ENL) showed histologic, immunofluorescent, and ultrastructural features typical of immune-complex glomerulonephritis. A literature review found reports of 187 renal biopsies from leprosy patients, with glomerulonephritis in 31% of the cases, and a variety of other renal lesions. This accumulated evidence demonstrates that, contrary to widely held opinion, there is no association between glomerulonephritis and a history of ENL. Furthermore, it suggests that glomerulonephritis in leprosy patients has a similar incidence in lepromatous and non-lepromatous cases. These observations have an important bearing on concepts regarding the pathogenesis of immune complex glomerulonephritis in leprosy and the humoral immune mechanisms in patients with leprosy.     

Sulphatide-binding properties are shared by serum amyloid P component and a polyreactive germ-line IgM autoantibody,the TH3 idiotype

Serum amyloid P component (SAP) concentration was elevated in sera from leprosy patients, significantly so above endemic controls in lepromatous cases. In the sera of lepromatous leprosy (LL) patients who experienced an erythema nodosum leprosum (ENL) episode the SAP fell at the onset of ENL and remained low throughout, in two of three cases. Changes in SAP concentration parallel anti-sulphatide IgM concentrations. TH3, a monoclonal IgM germ-line antibody derived from a LL patient, and SAP share similar binding patterns. In this study we demonstrate binding to heparin and sulphatide. Moreover, SAP inhibited the binding of TH3 to sulphatide, as well as anti-sulphatide IgM found in a range of sera, and anti-sulphatide IgG in the only sera sample in which it was found. The observation that anti-TH3 idiotype monoclonal and polyclonal anti-SAP antibodies both inhibited the binding of TH3 and IgM in sera (but not IgG) to sulphatide without binding to sulphatide themselves further demonstrated similar binding specificities. The observations of similarity in binding reinforce ideas that SAP may function as a primitive opsonin, but the clear ability to inhibit binding of autoantibodies suggests that SAP may play a role in ameliorating tissue and particularly nerve damage in leprosy patients.     

In situ characterization of T lymphocyte subsets in the reactional states of leprosy.

Using monoclonal antibodies and the immunoperoxidase technique, the numbers and distribution of T lymphocyte subsets in the tissues of reactional states of leprosy (six reversal reaction, nine erythema nodosum leprosum (ENL) and two Lucio's reaction) were determined and compared with those found in stable, non-reactional patients (six tuberculoid, two borderline lepromatous and seven lepromatous). The pattern of segregation of the suppressor/cytotoxic phenotype at the periphery of the granuloma was found in both non-reactional tuberculoid lesions and reversal reactions, but was better developed in the former. In ENL and Lucio's reaction, as well as in non-reactional lepromatous tissue, the helper/inducer and suppressor/cytotoxic phenotypes were both admixed with the aggregated histiocytes. However, the helper/suppressor ratio in ENL (2.1 +/- 0.4) was significantly larger than that in non-reactional lepromatous tissue (0.7 +/- 0.4, P less than 0.001). The immature thymocyte antigen OKT6 was found on scattered large non-lymphoid cells, most commonly in tuberculoid and reversal reaction tissues, less commonly in ENL, but only irregularly in non-reactional lepromatous tissue. The peripheral pattern of the suppressor/cytotoxic phenotype may be an immunohistological reflection of a cell-mediated immune response common to both non-reactional tuberculoid and reversal reaction patients. The reversal of the helper/suppressor ratio in ENL as compared to non-reactional lepromatous disease suggests some role for cell-mediated immunity in the pathogenesis of ENL. The OKT6 positive cell is of unknown origin and function.     

Specificity of IgG subclass antibodies in different clinical manifestations of leprosy.

We analysed specific IgG subclasses levels to Mycobacterium leprae sonicate extract (MSE), lipoarabinomannan B (LAM) and phenolic glycolipid I (PGL-I) in the sera of leprosy patients with different clinical manifestations. IgG2 was found to be the predominant antibody to MSE regardless of clinical manifestations, and IgG1 response was mostly seen in lepromatous patients. IgG3 reacted only rarely but IgG4 reacted relatively more in certain clinical groups such as borderline lepromatous and lepromatous with erythema nodosum leprosum (ENL) reaction. Most of the IgG subclass responses to MSE could be accounted for reactivity with LAM, suggesting that LAM is the major immunogen involved in the pathogenesis of leprosy. In contrast to LAM, PGL-I antigen showed considerably lower reactivities for IgG subclasses. An association between IgG subclass responses and clinical manifestations of leprosy was also seen. Whereas borderline lepromatous patients were found to have significantly higher levels of IgG2 and IgG4 to MSE, lepromatous patients had elevated levels of IgG1 and lower levels of IgG2. An interesting observation, however, was the significantly higher levels of IgG2 to LAM in the pure neuritic leprosy patients.     

Cytokine mRNA expression in leprosy: a possible role for interferon-gamma and interleukin-12 in reactions (RR and ENL)

Leprosy patients during the natural course of the disease may develop reactional episodes, namely reversal reaction (RR) and erythema nodosum leprosum (ENL). Immunological events described as occurring during RR indicate up-regulation of the immune response, whereas in ENL the events are not fully understood. The aim of this study was to analyse the in vivo pattern of cytokine gene expression in the reactional states of leprosy. Peripheral blood mononuclear cells (PBMC, n = 14) and tissue samples (n = 17) obtained from patients with ENL and RR were obtained and assayed by RT-PCR. PBMC obtained from unreactional patients (n = 15) and normal individuals (n = 5) were also assessed. Expression of interferon (IFN)gamma, granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin (IL)-2Rp55, perforin and IL-1beta mRNA in PBMC were detected mostly in ENL/RR patients, but not in unreactional patients. Likewise, cytokines such as IL-6, IL-8, tumour necrosis factor (TNF)alpha and TNFbeta were also present in reactional and tuberculoid patients as opposed to lepromatous leprosy (BL/LL). Interestingly, the majority of ENL/RR patients showed messages for IL-6, IL-10, IL-12 and TNFalpha in the skin. IFNgamma was detected in 84.6% (ENL) and 100% (RR) of the patients, whereas IL-4 was detected only in few individuals (38.5 and 25%, respectively). Although mRNA expression and protein levels may be different, the data reported in this study suggest a cytokine mRNA profile that seems to be indistinguishable for RR and ENL. In addition, it shows up-regulation of immuno-inflammatory cytokines in the blood and tissue of the same patient examined before and during reaction. Furthermore, it is suggested that this pattern of response results from an immunological reactivation that might lead to an acute inflammatory response in both reactional episodes.     

Autoantibodies in lepromatous leprosy

Lepromatous leprosy patients often develop erythema nodusum leprosum (ENL) reactions mainly during treatment of the disease. Hence, this study sought to investigate correlation between the prevalence of certain autoantibodies and ENL reactions in these patients. The patients included in the study were fifty patients with lepromatous leprosy and a similar number of normal controls. Sera were collected from the patients and normal controls and the prevalence of circulating rheumatoid factor antinuclear and antismooth muscle antibodies was determined. The prevalence of autoantibodies was increased in lepromatous leprosy as compared with normal controls. The prevalence of these autoantibodies were affected differently by the ENL reactions. ENL lepromatous leprosy showed a slight decrease in prevalence of antinuclear and antismooth muscle antibodies, whereas there was a slight increase in the prevalence of rheumatoid factor in ENL lepromatous leprosy. The levels of serum immunoglobulins tended to show a decline towards ENL lepromatous patients compared with the uncomplicated lepromatous patients. The significance of these finding is discussed in relation to their possible role in the pathogenesis of ENL reaction. It is concluded that some of these autoantibodies and immunoglobulins may be utilized during ENL reactions in the formation of immune complexes.     

Increased chitotriosidase activity in serum of leprosy patients: Association with bacillary leprosy

Human phagocyte-specific chitotriosidase is associated with several diseases involving macrophage activation. Since macrophage activation plays an important role in the control of Mycobacterium leprae infection, we studied the association of chitotriosidase with leprosy both in serum and in situ in lesional skin biopsies from patients. Serum samples from 78 Indonesian leprosy patients (39 non-reactional and 39 reactional leprosy patients) and 36 healthy controls (HC) from the same endemic region were investigated. The patients were classified as multibacillary (MB, n = 69) or paucibacillary (PB, n = 9) based on the bacterial index in slit-skin smears. Thirty-six of the reactional patients had erythema nodosum leprosum (ENL), while only 3 had reversal reaction (RR). Follow-up serum samples after corticosteroid treatment were also obtained from 17 patients with ENL and one with RR. Multibacillary (MB) patients showed increased chitotriosidase activity in serum as compared to paucibacillary (PB) patients and healthy controls. Although no significant difference was observed between reactional and the corresponding non-reactional groups, ENL showed significantly higher chitotriosidase activity as compared to HC. Furthermore, corticosteroid treatment resulted in significant decline of enzyme activity in ENL sera. Chitotriosidase activity correlated with levels of neopterin, another macrophage activation marker, but not with IL-6, IFN-γ, TNF-α and IL-10. Immunohistochemical staining of 6 MB (LL = 5, BL = 1) lesional skin sections from stored material showed positive staining for chitotriosidase within lipid-laden macrophages suggesting that macrophages are the source of the enzyme detected in serum. Thus, serum chitotriosidase activity is potentially useful in distinguishing MB from PB leprosy and in monitoring response to therapy in ENL.     

Tumor necrosis factor production in patients with leprosy.

The spectrum of host responses to Mycobacterium leprae provides a model for investigating the role of cytokines in the pathogenesis of mycobacterial disease. Of particular interest is tumor necrosis factor (TNF), a cytokine which may have both antimycobacterial and immunopathologic effects. To evaluate the potential role of TNF in leprosy, we measured TNF production in response to M. leprae and its defined constituents by peripheral blood mononuclear cells from patients across the spectrum of disease. The levels of TNF induced through the stimulation of cells with M. leprae or its dominant "lipopolysaccharide," lipoarabinomannan, were higher in patients with the tuberculoid form of the disease than in those with the lepromatous form. In patients with erythema nodosum leprosum (ENL), a reactional state of lepromatous leprosy, the levels of TNF release by peripheral blood mononuclear cells were higher than in any other form of the disease. Treatment of ENL patients with thalidomide reduced TNF secretion by more than 90%. The mycolylarabinogalactan-peptidoglycan complex of Mycobacterium species, the protein-peptidoglycan complex, and muramyl dipeptide all elicited significant TNF release. Therefore, TNF release appears to be triggered by at least two major cell wall structural constituents of M. leprae, lipoarabinomannan and segments of the cell wall skeleton. The prominent TNF release in patients with the paucibacillary tuberculoid form of the disease compared with that in patients with the multibacillary lepromatous form suggests that this cytokine contributes to a resistant immune response to mycobacterial infection. However, the marked TNF release in patients with ENL indicates that TNF may also mediate immunopathologic effects, such as fever and tissue damage.     

Differences in predominant T cell phenotypes and distribution pattern in reactional lesions of tuberculoid and lepromatous leprosy.

The nature and histological pattern of the cutaneous infiltrates of 17 leprosy patients in reversal reactions (Type I) and erythema nodosum leprosum (Type II, ENL) were compared with tissues from 18 non-reactional borderline leprosy (BT, BL) and lepromatous leprosy (LL) patients using monoclonal antibodies and immunofluorescence. Reactional BT lesions showed a mild increase in OKT11+ pan T cells as compared to non-reactional tissues and a significant influx of OKT8+ (suppressor/cytotoxic) cells which were peripherally localized in the lymphocyte mantle surrounding the epithelioid cells. The Leu 3a+ (helper/inducer) cells were scattered amongst the lymphocytes and macrophages. The mean ratio (+/- s.d.) of Leu 3a+/OKT8+ cells was 1.88 +/- 0.64 in Type I BT reactions as compared to 2.95 +/- 0.95 in BT lesions. In contrast, lesions of BL reversal reactions and ENL showed a more marked increase in pan T cells with a preponderance of the helper/inducer subset, Leu 3a+/OKT8+ ratio being 2.26 +/- 0.61 and 0.93 +/- 0.57 in BL reactional and non-reactional lesions, respectively. Interestingly, this increase in the numbers of the T cells reached levels observed in BT lesions. The distribution pattern of OKT8+ cells was similar to Leu 3a+, both being diffusely scattered amongst the bacilli laden macrophages. Ia like antigens were present in all granulomas and were abundant on lymphocytes and macrophages and less conspicuous on epithelioid cells. T6+ Langerhans cells were uniformly increased in all reactional lesions. It would appear that the changes observed in both Type I and Type II reactions are similar in the lepromatous group of patients. They differ significantly from the BT reversal reaction in terms of the dominant T cell subset and the microanatomical distribution of the OKT8+ cells in the lesions.     

Serum lymphocytotoxic activity in leprosy.

Sera from 167 patients across the spectrum of leprosy and 46 endemic controls were screened for lymphocytotoxic activity (LCA). The Terasaki microdroplet lymphocytotoxicity assay was performed at 37 degrees C and 15 degrees C to test sera for LCA against a panel of lymphocytes from 50 donors which represented most known HLA-ABC antigens. Raised complement-dependent LCA at 15 degrees C was seen in leprosy patients with histories of erythema nodosum leprosum (ENL) or reversal/Type I (I) reactions. Eighty-six per cent of lepromatous (LL) patients with a history of ENL (n = 21, P less than 0.001), 83% of borderline lepromatous (BL) and 88% of borderline tuberculoid patients (BT) with a history of Type I reactions (n = 12, P less than 0.01 and n = 24, P less than 0.001 respectively) had LCA compared to 39% of endemic controls (n = 46). LCA was attributed to IgM on the basis of reduced activity when serum was treated with both dithiothreitol or absorbed with antiserum for IgM. Removal of immune complexes and rheumatoid factor did not influence LCA. LCA-positive sera reacted similarly with allogeneic lymphocytes from either healthy donors or leprosy patients. Moreover LCA-positive sera reacted with autologous lymphocytes. Specificities for HLA-ABC antigens were not identified. The potential role of these autoantibodies, manifested in leprosy patients with hypersensitivity reactions remains speculative.     

High antibody levels to the mycobacterial fibronectin-binding antigen of 30-31 kD in tuberculosis and lepromatous leprosy.

Immunoblot assays showed that mycobacterial fibronectin-binding antigens are important targets of the humoral immune response in tuberculosis and leprosy. Using culture filtrate antigens of Mycobacterium tuberculosis, strong reactivity with the fibronectin-binding of 30-31 kD (Fn 30-31) was demonstrated in 55.9% of tuberculosis sera and in 56.5% of lepromatous leprosy sera. Sera from patients with tuberculoid leprosy and control sera gave very weak binding. Reactivity of tuberculosis and lepromatous leprosy sera with the fibronectin-binding antigen of 58-60 kD (Fn 58-60) was less conspicuous. The ability to react with fibronectin of the antigens of 58-60 and 30-31 kD was demonstrated by parallel labelling with a fibronectin-biotin conjugate. Fn 30-31 was purified to homogeneity by a two-step procedure and used for ELISA. Positive titres were found in 63% out of 65 tuberculosis sera and in 60.5% out of 43 lepromatous leprosy sera. Antibody titres in lepromatous leprosy sera were higher than in tuberculosis sera. Our observations indicate indirectly that M. leprae possess a highly immunogenic molecule homologous to M. tuberculosis Fn 30-31, which elicits a high antibody response in lepromatous leprosy but not in tuberculoid leprosy. In this investigation, direct evidence for the presence of this antigen in M. leprae was obtained by immunochemistry of lepromatous leprosy lesions with a monospecific antibody raised against M. tuberculosis Fn 30-31.     

Association of C4B deficiency (C4B*Q0) with erythema nodosum in leprosy

A considerable number of studies have postulated significant associations between susceptibility to the different clinical manifestations of leprosy and the MHC, In this investigation, the association between the MHC class III complement proieins C2, BF, C4A and C4B and leprosy in a patient population of Southern Brazil was studied. A total of 109 non-related leprosy patients was investigated; 73 presented wilh lepromatous leprosy (LL), 46 of Ihem had the immunopathological reaction of erythema nodosum (ENL), the remaining 36 were tuberculoid, borderline and indeterminate leprosy (TIBL) patients. The control group included 172 healthy individuals matched with the patients according lo their ethnic and geographical origin, C2, BF, C4A and C4B allotypes were determined by slandard technologies including Western blots for C2 and C4 variant alleles wilh monoclonal and polyclonal antibodies. Non-expressed (‘silent’) C4 alleles in hemizygously deficient individuals were estimated semiquantitatively on the basis of the C4A and C4B isolype ratio and by the M ASC (‘minimal chi-square’) method. The results showed a significantly elevated presence of the non-expressed C4B allele (C4B*Q0) in the LL and ENL patient groups in comparison with the controls. The most signifieant difference was observed in the ENL group when compared with the controls. In addition, all patients who were homozygously C4B-deficient had ENL, and most of them had the BF*F1 allele. The comparison between LL patients with and without ENL also showed a statistically significant difference in the presence of C4B*Q0, indieating thai C4B deficiency itself is associated with EN L. The relative risk of LL patients with the C4B*Q0 allele suffering from ENL was 53 compared with LL palients without C4B*Q0, Since immune complexes (IC) are considered to be the palhogenic cause of ENL, our findings indicate thai C4B deficieney may play an important role in the abnormal immune response against Myeobaeterium leprae and in the lack of IC clearance, leading to ENL reactions. Individuals wilh this allele seem to be at a higher risk of developing pathologieal immune reactivity in lepromatous leprosy.     

An isoelectric focusing method for the study of the humoral response against the antigen 85 complex of Mycobacterium bovis BCG in the different forms of leprosy.

Isoelectric focusing was used to separate the three components of the antigen 85 complex of Mycobacterium bovis BCG. Antibody responses of leprosy patients against each Mycobacterium bovis BCG. Antibody responses of leprosy patients against each component were quantitated by densitometric analysis of immunoblot assays. The 85A component was recognized by 40% (8/20) of the lepromin positive and negative healthy subjects, by 76% (19/25) of the tuberculoid and by 96% (24/25) of the lepromatous leprosy sera. In contrast, the 85B component was not stained by the control sera, nor by the tuberculoid leprosy sera but by 64% (16/25) of the lepromatous leprosy sera. The results suggest that antigen 85B contains one or several epitopes that are specifically recognized by sera of lepromatous leprosy patients only.     

Suppressed natural killer cell activity during episodes of erythema nodosum leprosum in lepromatous leprosy.

Natural killer (NK) cell activity was investigated in peripheral blood mononuclear cells of patients with leprosy. The NK activity of patients with borderline or lepromatous leprosy did not differ significantly from that of normal subjects. However, in a group of patients with lepromatous leprosy undergoing an episode of erythema nodosum leprosum (ENL), NK activity was significantly depressed. In four patients with ENL, NK activity was virtually abolished. Depressed NK activity could not be attributed to the effects of corticosteroid therapy, nor did a serum factor appear to be responsible. Evidence was obtained that depressed NK activity in patients with ENL was not due to dysfunction of the NK cells themselves; they functioned normally when separated from the individual's total mononuclear cell population. Additional cell depletion studies suggested that the patients' monocytes were responsible for the observed depression of NK activity.