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To investigate the pathogenesis of low CD4 T-cell count in subjects who are immunological non responders (InR) to HAART.Thirty-five HIV-positive subjects on HAART for at least 1 year, all with undetectable HIV-1 RNA, were studied. Patients were defined as InR according to a CD4 cell increase20% from CD4 cell baseline or CD4 cell count200/microl; subjects with a CD4 T-cell increase20% from baseline and a CD4 cell count200/microl were defined as immunological responders (IR). We performed a comprehensive study to characterize the immune response of InR.The immunological phenotype of peripheral blood mononuclear cells, thymic naive T cells, T-cell receptor Vbeta repertoire, serum concentration of interleukin (IL)-7, the expression of IL-7Ralpha on naive and memory CD4 and CD8 T cells, and regulatory T cells (Treg) were studied.In InR a significant reduction (P0.0001) of naive and thymic naive CD4 T cells was associated with a reduced expression of IL-7Ralpha in both cell subsets, with an increased serum concentration of IL-7 was observed. Furthermore, an increased immune activation with a reduced Treg frequency and increased number of expansions of Vbeta families was observed.The reduced expression of IL-7Ralpha associated with the persistent immune activation and the alteration of Treg frequencies in part explains the low level of CD4 T cells observed in InR.

Primary effusion lymphomas (PELs) are invariably infected by human herpesvirus type 8 (HHV8) and often co-infected by Epstein-Barr virus (EBV). We found that expression of major histocompatibility complex class I (MHC-I) surface molecules was significantly decreased in PEL cells when compared with HHV8 negative lymphomas, irrespective of EBV infection. MHC-I downregulation rendered PEL cells sensitive to recognition and killing by natural killer (NK) cells. Intriguingly, analysis of MHC-I non-restricted cytotoxicity in two PEL patients indicated a reduced NK cell activity when compared with healthy individuals. These data suggest that PEL outgrowth may require an impaired NK cell function.

Background: Idiopathic CD4+ lymphocytopenia is defined by a stable decrease of CD4+ T cells in the absence of any known cause of immune deficiency. The mechanisms responsible for the immunological impairment are still unknown, but a regenerative failure of hematopoietic stem/progenitor cells has been hypothesized. Methods: We evaluated in the bone marrow (BM) of 5 patients with idiopathic CD4+ lymphocytopenia the phenotype of BM progenitor cells, their differentiation capacity with colony-forming cells and long-term culture-initiating cell assays, in parallel with the spontaneous IL-7 production in the patient sera. Results: Compared with controls, a regenerative failure of hematopoietic stem cells has been observed, both in ‘committed’ and in ‘uncommitted’ progenitor cells, despite high IL-7 serum levels. The percentage of phenotypically primitive CD34+CD38–DR+ cells (this includes the lymphoid precursor cells) was decreased, suggesting an involvement of the more primitive BM compartment in the de novo T cell generation. Conclusions: Despite the low number of patients, due to the low incidence of the disease, the decrease of primitive precursors sustains the possibility that diminished stem cell precursors might contribute to the development of CD4+ T cell depletion.

Kaposi's sarcoma (KS) is an angioproliferative disease of multifactorial origin arising in different clinic-epidemiologic forms, which show the same histopathological features. It generally starts as a hyperplastic reactive-inflammatory and angiogenic process, which may evolve into monomorphic nodules of KS cells that can be clonal (late-stage lesions) and resemble a true sarcoma. Infection with the human herpesvirus 8, cytokine- and angiogenic factor-induced growth together with an immuno-dysregulated state represent fundamental conditions for the development of this tumor. Several local therapies are used to eradicate early and confined skin lesions, whereas widely disseminated, progressive or symptomatic disease requires a more aggressive treatment. Although different chemotherapeutic agents have been used to treat aggressive KS, the growing understanding of the pathogenetic factors participating in KS development has provided a strong rationale for using less- or non-cytotoxic agents that block the mechanisms involved in KS pathogenesis. The angiogenic nature of KS makes it particularly suitable for using therapies based on anti-angiogenic agents. Of note on this goal, recent studies indicate that the highly active anti-retroviral therapy, including at least one human immunodeficiency virus (HIV) protease inhibitor (PI), is associated with a dramatic decrease in the incidence of AIDS-KS and with a regression of KS in treated individuals. Consistent with this, results from preclinical studies indicate that PIs have potent and direct anti-angiogenic and anti-KS activities, suggesting that they should be further investigated, alone or combined with other therapies, as a novel treatment for KS in both HIV seropositive or seronegative individuals.

Background Common variable immunodeficiency (CVI) is a primary defect of the immune system. Infections, persistent diarrhoea and malabsorption may result in malnutrition, which may in turn contribute to increased morbidity. In this paper, the prevalence of malnutrition in CVI was evaluated. Patients and methods Forty CVI patients (20 male, 20 female, aged 17–75 years) underwent anthropometric measurements from which body mass index, arm fat and muscle area were calculated. Body mass index values

PurposeTo facilitate increased use of keratolimbal allograft transplantation (KLAL) for severe ocular surface disease (OSD) by informing clinicians and eye banks of differences in tissue requirements and preparation for the procedure and to describe the surgical technique of KLAL transplantation.Met

A survey for antibodies to a recombinant small viral capsid antigen (sVCA) of human herpesvirus type 8 (HHV‐8) was conducted in Sardinia, one of the world's highest incidence areas for classic Kaposi's sarcoma (KS). Prevalence of antibodies to HHV‐8 sVCA was greatest in patients with KS (95%), followed by family members (39%) and a Sardinian control population age‐ and sex‐matched to the relatives (11%). Within families, prevalence of antibodies was about equal among spouses, children, and siblings of KS patients, a finding that raises the possibilities of intrafamilial person‐to‐person or vertical transmission. Antibodies were detected 2–3 times more frequently in males than in females. The data show that prevalence of antibodies to HHV‐8 sVCA correlates with the distribution of classic KS in a high‐ incidence area. Clustering of seroprevalence within some families suggests the presence of familial risk factors for active HHV‐8 infection.

Kaposi's sarcoma (KS) was a rare disease in Europe and North America until a decade ago, when it became the most common neoplasm complicating the acquired immunodeficiency syndrome (AIDS), where it acquires an aggressive course. Clinical and experimental data suggest that, at least in early stage, KS may not be a true sarcoma, but an hyperplastic-proliferative lesion that may regress. At least three components characterize KS lesions: (1) neoangiogenesis and proliferation of spindle-shaped cells of endothelial and macrophage cell origin, some of which may originate from a circulating precursor; (2) a cellular infiltrate represented by macrophages, lymphoid cells, mast cells, and neutrophils; and (3) the infection of spindle cells and mononuclear cells with a new virus of the Herpesvirinae family defined KS-associated herpesvirus or human herpesvirus-8 (HHV-8). KS lesions are highly responsive, in terms of growth, to inflammatory cytokines (IC) and many lesional cell components are able to secrete cytokines and chemokines, which induce paracrine-autocrine mechanisms of growth, angiogenesis, and promote further cellular recruitment. The association between HHV-8 and KS is close; however, the role of the virus in KS development is yet unknown. Nevertheless, the virus has the potential to encode for homologs of cellular cytokines and some chemokines and its reactivation is sensitive to stimuli provided by IC. This review focuses on these aspects of KS pathogenesis, trying to reconcile many of the clinical and experimental observations. Finally, the role of the HIV-1 Tat protein as a factor of progression in AIDS-KS as well as the role of cellular and HHV-8 encoded proto-oncogenes as factors and markers of progression of KS to a true malignancy is reviewed.

The aim of this study was to assess the effect of natural killer stimulatory factor (NKSF)/IL-12) on the cytolytic potential of natural killer (NK) cells from normal donors or HIV-1 infected individuals, at different stages of disease. The results obtained confirm the powerful ability of IL-12 to enhance PBLs' cytotoxic activity to tumour cells in normals and in those HIV-1 infected patients, with maintained cytotoxic activity. IL-12 was however poorly effective in enhancing the cytolytic potential of any donors. Similar results were obtained adding IL-2, even at doses as high as 100 IU/ml in patients with CD4 number below 200/mm3. These results should be taken into account when designing therapeutic trials of HIV-1 infection based on cytokines.

The "natural killer" (NK) cells preferentially kill targets lacking surface major histocompatibility complex class I (MHC-I) molecule expression. NK cells recognize these targets through membrane receptors, which can trigger activating or inhibitory signals for killing. Several tumors or virus-infected cells downregulate MHC-I expression as a mechanism to evade recognition and killing by cytotoxic T lymphocytes (CTL). They, however, become targets for NK cells cytotoxic activity. NK cell activity is reduced during disease progression in human immunodeficiency virus (HIV) infection, and in individuals with AIDS-associated tumors linked with infection by the oncogenic human herpes virus type-8 (HHV8), including Kaposi's sarcoma (KS) and primary effusion lymphomas (PEL). We have demonstrated that AIDS-related KS (AIDS-KS) is characterized by an increased expression of inhibitory receptors by T lymphocytes, and that HIV-non-infected patients with KS (classic KS, C-KS) have a substantial number of NK cells bearing these same receptors. NK cells from patients with C-KS are normally equipped with cytolytic molecules including granzyme A and perforin. However, the cytotoxic activity of NK cells is reduced in patients with C-KS, AIDS-KS, or PEL patients, who are all infected by the HHV8, and this correlates with disease severity. Moreover, we have found that HHV8-infected cell lines established from PELs have a reduced surface expression of MHC-I molecules and are sensitive to the lysis mediated by NK cells. Since PEL cells express the same HHV8 latency program as KS cells, these data point to MHC-I downregulation by HHV8 as a primary immune evasion mechanism against CTL responses, further reinforced by upregulation of inhibitory receptors on T and NK cells in the setting of HIV and/or HHV8 infection. Thus, studies on killing receptor regulation and signaling in T and NK cells may shed light on the pathogenesis of HHV8-associated tumors both in HIV-infected or -noninfected patients.

The case of a 20-year-old patient, affected by X-linked agammaglobulinaemia (XLA), who developed severe pneumonia from Pneumocystis jiroveci (formerly Pneumocystis carinii) (PCP), is reported. This infection usually affects patients with AIDS, children affected by severe combined immunodeficiency or hypogammaglobulinaemia with hyperimmunoglobulin M, or patients undergoing severe immunosuppression. The XLA patient developed PCP during therapy with steroids and cyclosporine A, carried out for several months, due to an extended skin vasculitis, accompanied by general symptoms. The pneumonia had a severe clinical course, requiring a long hospitalization. At the diagnosis of PCP, immunosuppressive therapy was suspended and the patient recovered after a long-term trimethoprim/sulfamethoxazole therapy. Immunological studies revealed an unexpected normal number of CD4+ and CD8+ T cells. The two subsets had an exclusive naïve phenotype (95% CD4+CD45RA+CD62L+ and 89% CD8+CD45RA+CD62L+ cells), with an absence of primed cells. Lymphoproliferative responses to P. carinii and recall antigens as well as to mitogens were extremely deficient. During the follow-up, memory cells appeared with recovery of the lymphoproliferative response to mitogens and maintained defective responses to antigens. This is one of the few reported XLA cases experiencing severe PCP. In this patient, the infection became clinically evident during immunosuppressive therapy. We believe that the absence of functional activities, despite a normal level of T lymphocyte counts, sustained this long-lasting infection. Thus, the CD4+ and CD8+ T cell count evaluation, without functional studies, may not be per se sufficient for predicting the risk of a severe clinical course of PCP in patients undergoing immunosuppression.

Kaposi's sarcoma (KS) develops upon reactivation of human herpesvirus 8 (HHV8) infection and virus dissemination to blood and tissue cells, including endothelial and KS spindle cells where the virus is mostly present in a latent form. However, this may likely require the presence of compromised host immune responses and/or the evasion of infected cells from the host immune response. In this regard, mechanisms of evasion of productively infected cells from both CTL and NK cell responses, and resistance of latently infected cells from specific CTL, have already been shown. Here we show that cells which are latently infected by HHV8 are indeed efficiently lysed by NK cells from individuals with a normal immune response. Notably, NK cell-mediated immunity was found to be significantly reduced in AIDS patients with progressing KS as compared to both HIV-negative patients with indolent classic KS or normal blood donors. However, it was restored after treatment with the highly active antiretroviral therapy (HAART) in AIDS-KS patients, that showed regression and clearance of HHV8 from PBMC. By contrast, AIDS-KS patients with a more aggressive disease and no clinical response had persistent HHV8 viremia associated with reduced NK cell cytotoxicity. These results suggest a key role for NK cells in the control of HHV8 latent infection, KS development, and in disease remission upon HAART.

Receptors interacting with Major Histocompatibility Complex class I molecules have been initially found on the surface of human natural killer (NK) cells, where they deliver inhibitory signals to the lysis, being thus defined killer inhibitory receptors (KIR). Subsequently, they were detected also on the surface of T-CD8+ lymphocytes and are particularly expanded during human immunodeficiency virus (HIV) infection, where they downregulate HIV-specific cytolysis. The expression of KIR recognizing human leukocyte antigen-C alleles was assessed in HIV-infected patients, undergoing highly active antiretroviral therapy (HAART). To this end, the combined expression of CD16/CD56, of CD3 and CD8 as well as of KIR (CD158a and CD158b) surface molecules was analyzed on peripheral blood mononuclear cells by monoclonal antibodies, and flow cytometry. An increase of CD3+CD8+CD158b+ cells was found after 6 months of HAART. This finding may have implications for the regulation of T-cell mediated cytolysis during HAART.

Kaposi's sarcoma (KS) is an angioproliferative disease occurring in several different clinical-epidemiological forms that, however, share the same histological traits and are all associated with infection by the human herpesvirus 8 (HHV8). KS initiates in a context of immune dysregulation characterised by CD8+ T cell activation and the production of Th1-type cytokines that induce a generalised activation of endothelial cells leading to adhesion and tissue extravasation of lympho-monocytes, spindle cell formation and angiogenesis. These phenomena are triggered or enhanced by infection with HHV8 that, in turn, is reactivated by the same cytokines. Productively-infected circulating cells are recruited into 'activated' tissue sites where HHV8 finds an optimal environment for establishing a persistent, latent infection of KS spindle cells (KSC). HHV8 dissemination is favoured by virus escape mechanisms and immune dysregulation, and leads to immune responses that are not effective against the virus but, paradoxically, exacerbates the reactive process. Although early KS is a reactive process of polyclonal nature that can regress, in time it can progress in to a true sarcoma. The progression of KS appears to be due to the deregulated expression of oncogenes and oncosuppressor genes, to the long-lasting expression of the HHV8 latency genes and, for AIDS-KS, is promoted by the proliferative and angiogenic effects of the HIV-1 Tat protein.

A human immunodeficiency virus-negative woman with severe classic Kaposi's sarcoma, idiopathic leukopenia, and massive spread of human herpesvirus 8 (HHV-8) in circulating cells showed stable disease remission in response to systemic interferon-alpha treatment that was accompanied by increased CD3(+) and CD4(+) T cell numbers and complete clearance of HHV-8 from the circulation. These results suggest a direct relationship between HHV-8 clearance from blood and regression of Kaposi's sarcoma and are consistent with the in vitro inhibitory effects of interferon-alpha on HHV-8 infection.

The purpose of this study was to evaluate, in Kaposi's sarcoma patients, the correlation between antibody titers to the lytic antigens of human herpesvirus 8, as assessed by immunofluorescence assay, and values obtained by an enzyme immunoassay. The methods showed a stringent correlation, r = 0.625 ( P < 0.001).

Vol 349 • January 25, 1997 255 cultures with the endothelial cell marker VE-cadherin and the tissue-macrophage marker PAM-1 (40–60% in KS patients and 0·5–6% in controls) (table). 4 weeks later, culture were examined for HHV-8 DNA sequences by nested PCR. None of the control cultures was positive, whereas HHV-8 was detected in all T-KS and AIDS-KS cases and in 11/14 cultures derived from patients with C-KS (table). HHV-8 was also detected in the patient with C-KS after bleomycin therapy. Our results indicate that an expansion of circulating KS-like cells or their progenitors is present in all forms of KS; KS-derived cultures are infected by HHV-8; and the presence of HHV-8 in these cells correlates with KS. Since these cells are capable of chemotaxis and can induce KS-like lesion formation in nude mice, our data suggest that they may localise into tissues, transmit HHV-8 infection to neighbour cells, and participate in the formation of KS lesions.

The aim of this study was to assess the cytolytic potential of natural killer (NK) cells from human immunodeficiency virus type 1 (HIV-1)-infected patients, at different stages of the disease. Twenty HIV-1 seronegative donors as well as sixty HIV-1 seropositive patients were studied. Phytohemagglutinin and/or the anti-CD16 monoclonal antibody Kd1 were used to redirect the peripheral blood lymphocyte lysis of these patients to the 51Cr-labeled Fc gamma receptor-positive P815 murine mastocytoma target cell line. In parallel, NK cytotoxicity to tumor targets was investigated. Seronegative as well as HIV-1 Center for Disease Control (CDC) stage II patients showed maintained cytolytic activity. The cytolytic potential declined with disease progression, starting with CDC IVC2 patients, and was strongly diminished in acquired immunodeficiency syndrome stage patients. This defect was accompanied by decreased cytolytic activity to tumor targets and was not corrected by the in vitro addition of interleukin-2. The number of cells bearing a mature NK phenotype was normal in all the study groups. Our data suggest that the impaired NK cytotoxicity to tumor targets described during the progression of HIV-1 disease may be related to the progressive loss of function of surface receptors involved in NK cell triggering.

The effect of somatostatin-14 (SST) and the SST analogues SMS and RC160 on human natural killer (NK) activity mediated by large granular lymphocytes (LGL), as well as on IL-2-and/or anti-CD16 monoclonal antibody (mAb)-induced activation of these cells, was investigated. The NK activity of LGL was studied by the release of 51Cr by the erythroleukemia-derived cell line K562, whereas 51Cr release by the P815 murine mastocytoma-derived cell line, for which lysis was redirected by the use of an anti-CD16 mAb, was used to study the cytolytic potential of these cells. IL-2 was used at the final concentration of 100 IU and was incubated overnight with LGL. SST and the analogues, added to these systems at final doses ranging from 10−12 to 10−5 M , were inhibitory of the NK cell activity to K562, with a dose-response curve starting from 10−8 M and reaching a significant level at 10−6 M . On the contrary, no effect was observed on the redirected killing assay to P815 and on the IL-2-induced activation of NK cells. These results provide additional evidence for the immunomodulatory action of somatostatin.

Background. We evaluated haematological and immunological characteristics of four thalassemia patients after T-cell-depleted HLA-haploidentical stem cell transplantation Methods. We evaluated the clonogenic capability by the colony forming cell assay (CFC) and the long term culture-initiating cell (LTC-IC) assay at baseline and 20 days after transplant. Stromal cells were obtained from long term culture of bone marrow mononuclear cells (BMMCs) and analysed by immunohystochemistry. Lymphocyte subsets were studied by flow cytometry; and stromal IL-7 production by BMMCs was analysed by ELISA. Results. At baseline, no significant differences were observed in haematological and in immunological parameters in thalassemia patients when compared with a group of normal subjects Day + 20 after transplant, a reduced clonogenic capability was observed (4 ± 2 vs. 41 ± 40 CFU-E, 17 ± 9 vs. 109 ± 22 BFU-E, 3 ± 1 vs. 9 ± 6 CFU-GEMM and 16 ± 10 vs. 66 ± 23 CFU-GM). The number of primitive bone marrow (BM) progenitor cells was also decreased (1.8 ± 1.4 vs. 15.4 ± 3.6 LTC-CFC/106 BMMCs). In addition, stromal cells secreted lower IL-7 levels (0.3 + 0.1 pg/mL vs. 0.8 + 0.1 pg/mL, in controls) and displayed by immunohistochemistry an altered phenotype. Upon light microscopy examination, the majority (75%) of these cells appeared as moderately large cells, frequently rounded, with abundant cytoplasm, whereas in control subjects about 90% of the stromal cells exhibited a different morphology characterized by irregular or spindle shape and branching cytoplasmic processes (fibroblast-like). Compared with normal subjects, thalassemia patients showed: reduction of naïve CD4+ T-cells (2 ± 0.5% vs 50 ± 10%), reduction of thymic naïve CD4+ T-cells (1 ± 0.2% vs 40 ± 12%,) and a significant increase of CD4+ cells activation markers (CD95, HLA-DR and CCR5). IL-7 receptor (CD127) expression was also significantly decreased on CD4+ T-cells and on naïve CD4+ T-cells (CD4+/CD45RA+CD62L+/CD127+). NK cells were among the first lymphocytes to repopulate the peripheral blood, and up to 70% of these cells were CD56 brigh whereas CD16+ NK cells were decreased. Conclusions. Twenty days post transplant, an impaired growth and differentiation capacity of stem/progenitor cells were observed in thalassemia patients, in parallel with an altered homeostasis of T-cells and a reduction of T-cell naïve compartment. We hypothesize that the damage of T cell compartment may be at least partially due to an altered production of new T cells starting from the haematopoietic stem/progenitor cells. CD56+ NK cells develop more rapidly than other lymphocytes, but CD16+ NK cells (with cytotoxic potential) require more prolonged exposure to maturation factors (IL-2) in the bone marrow. An IL7/IL7R pathway dysregulation has been also observed, possibly involving bone marrow stromal cells. In vitro studies are ongoing about the use of cytokines (IL-2, IL-7, IL-2 plus IL-7) supporting T cell development.

The investigation of the effects of inflammatory cytokines (IC) on the growth and differentiation of neural cells has provided new insights on the role of such soluble mediators in nervous system development and/or plastic remodeling as well as in the pathogenesis of inflammatory neurodegenerative disorders, which are characterized by chronic IC dysregulation in the central nervous system (CNS). Thus, the study of the interaction between CNS and immune-derived soluble signals in physiological or pathological conditions is of increasing interest. This review first discusses experimental evidence supporting the instructive/permissive role of immune-derived cytokines on CNS development and plasticity. Next, we focus on human neurological disease states such as multiple sclerosis and the neurodegeneration associated to the acquired immune deficiency syndrome in which different inflammatory cytokines have been proposed as potential neuropathogenic mediators.

Deficiency in natural killer (NK) cell activity is a common feature of acquired immune deficiency syndrome (AIDS) 1,2 . This is part of a general immune dysfunction in AIDS and may lead to progression of the disease, since NK cells are known to be involved in protection against tumors and against viral infections 3 . The lack of immunological surveillance by NK cells of the growth of pathogens that activate the HIV-1 tat infectivity gene may also favor progression to AIDS 4 . The pathogenesis of NK cell deficiency in AIDS is not known. Previous studies have shown that NK cells from AIDS patients are able to bind but not to lyse the target cell line K562 (Ref. 5). This results from an inability to rearrange the cytoskeleton microtubular (MT) system 6 and to release the natural killer cytotoxic factor (NKCF) 7 . This report by Maria Caterina Sirianni and colleagues evaluates the possible mechanisms leading to this NK cell deficiency.

Thirteen patients affected with unexplained lymphoadenopathy, fever, weight loss, and diarrhea (lymphoadenopathy syndrome; LAS) were evaluated for the possible appearance of acquired immunodeficiency syndrome (AIDS) and for immunological and virological characterization. The patients belonged to categories of individuals at risk for AIDS and were homosexual and/or drug abusers or hemophiliacs. Lymph node biopsy showed the histological picture of a follicular hyperplasia. The study of cell-mediated immunity (CMI), humoral immune response, and natural killer (NK) activity demonstrated a significant decrease in T cells with the helper/inducer phenotype (OKT4+ cells) and a relatively increased number of lymphocytes with the suppressor/cytotoxic phenotype (OKT8+ cells). NK activity was significantly lower than in normal controls. Thein vitro response to policlonal activators (phytohemagglutinin; PHA) and the cutaneous responsiveness to recall skin tests were impaired, whereas immunoglobulin production was increased, mainly in the IgG fraction. Virological studies showed high serum antibody titers to cytomegalovirus (CMV) but a lack of specific CMI as assayed by the leukocyte migration inhibition test (LMIT). CMV was also isolated from the urine specimen of one patient. The antibody pattern to Epstein-Barr virus (EBV) showed the uncommon contemporary presence of both Epstein-Barr nuclear antigen (EBNA) and early antigen (EA) antibodies. Antibodies to human T-lymphotropic retroviruses (HTLV III) were positive in 10 patients and the virus was isolated in 3 of them. In some patients the presence of serum antibodies to HTLV III was not associated with an impairment of the immune function. A group of individuals at risk for AIDS without LAS was also evaluated for the presence of HTLV III antibodies; the percentage of positive sera was 11.4. Nevertheless, individuals without specific antibodies had immunological abnormalities resembling those of LAS HTLV III-positive patients. The possible implications of these findings are discussed.

Summary Eighteen healthy adult male subjects, living in a work environment with hepatitis B surface antigen (HBsAg) carriers, were vaccinated against hepatitis B with French vaccine. Immunological monitoring, performed on days 0, 15, 30, 45, 60, 90 and 120, concerned (1) specific humoral and cell-mediated immunity to HBsAg (cell-mediated immunity was explored in only 13 subjects by the leukocyte migration inhibitory test, or LMIT), (2) autologous mixed lymphocyte reaction (AMLR), (3) OKT3-, T4- and T8-positive lymphocytes, (4) the study of phagocytic respiratory activity, and (5) autoantibodies and circulating immune complexes (CIC). Absolute clinical, hepatic and immunological safety, as documented by the absence of clinical side-effects, no change in serum transaminase levels and the lack of appearance of CIC and/or autoantibodies, was observed. Specific seroconversion, absent on the 15th day, reached 89% in subjects by the 90th day, whereas specific cell-mediated conversion seemed to occur earlier (many subjects already showed LMIT positivity by day 15). For monoclonal antibodies and AMLR, an increase in OKT8+ lymphocytes on day 60, seen only among normal responders, was observed with a subsequent reduction in the OKT4/OKT8 ratio and a reduction of the proliferative response in AMLR on the 45th day (except in the only non-responding subject tested). No significant variations in granulocyte respiratory activity were evident. These results are discussed, focusing on the role of cellular immunity during anti-HBsAg vaccination.

Rosette formation of human lymphoid cells with mouse erythrocytes has recently been proposed as a marker for a subpopulation of B lymphocytes. In this work we studied the percentage of mouse rosette forming cells (MRFC) in normal and pathological conditions and compared them to the percentage of sheep rosette forming cells (SRFC) a marker for T lymphocytes. Peripheral blood lymphocytes (PBL) from normal donors contained 6.2 ± 1.1% (mean ±1 S.D.) MRFC. High percentages of MRFC were found in CLL patients, and a slight increase was observed in patients with systemic lupus erythematosus. MRFC were absent in Bruton's type agammaglobulinaemia, but were normally present in patients with T cell defects. Cryopreservation of lymphocytes in 10% DMSO did not significantly affect the mean percentages of SRFC and MRFC, though a slight increase of the former and a small reduction of the latter was observed. Double binding experiments on peripheral blood lymphocytes showed a predominant association of MRFC with cells staining for surface IgM and/or IgD. In all samples tested, we also observed a small population of MRFC negative for sIgM or sIgD and a few sIgM or sIgD positive cells that did not rosette with mouse erythrocytes.

Peripheral blood mononuclear cells from three patients with severe combined immunodeficiency (SCID), three with SCID whether B cell positive or with pure T-cell defect, and two with DiGeorge syndrome were analyzed with a panel of monoclonal antibodies against immature and mature T-cell subsets. Natural killer (NK) cells were enumerated by the use of the HNK-1 monoclonal antibody. NK activity against the K562 and MOLT 4 cell lines was also investigated. According to the monoclonal antibodies profile and the NK activity, patients could be divided into three groups. Patients with classical SCID had no detectable circulating T cells as well as NK cells and activity, probably due to an early block in stem cell differentiation. Patients affected by SCID with B cells or pure T cell defect showed a decrease in lymphocytes with mature phenotypes but prothymocytes or immature thymocytes circulated in peripheral blood. Children with DiGeorge syndrome had a decrease in mature thymocytes and, in this study, NK cells were normal. These data help to clarify both the preeminent immunologic features of SCID and related syndromes and the character of NK cells.

Twenty-one immunodeficient patients with recurrent herpes simplex labialis (HSL) were randomly allocated to either the saline or the bovine thymus extract Thymostimulin (TS) group and treated for a period of 6 months. A total of 17 recurrences with a mean severity index of 266.2 +/- 192 in the TS-treated group versus a total of 62 recurrences in the placebo group (score 458.8 +/- 299.6) were observed after 12 months of follow-up. A significant increase of total WBC (P less than 0.05 versus control group), lymphocyte count, and T-cell number (P less than 0.001) were detected in the TS group after 6 months as well as after 12 months. In vitro lymphoproliferative responses to herpes simplex virus antigen and natural killer cell activity were also significantly higher (P less than 0.05) in the TS group, whereas no significant difference in antibody titers to herpes simplex could be detected between the two groups. TS may be potentially useful in preventing viral reactivation in immunocompromised hosts by potentiating cell-mediated immune responses. Further studies evaluating TS for prophylaxis of infection in patients at risk should be considered.

The ability of two preparations of dialyzable leukocyte extracts (DLE, formerly termed “dialyzable transfer factor”) to induce specific responsiveness by cord blood leukocytes (CBL) in the direct leukocyte migration inhibition (LMI) assay in vitro was studied. One donor of DLE (DLE-A) was immune to cytomegalovirus (CMV), as judged by positive LMI test, and negative to candida as judged by skin test. The other donor (DLE-B) was immune to both CMV and candida. Several concentrations of DLE and of antigen were tested. High concentrations of DLE nonspecifically inhibited leukocyte migration, while low concentrations had no effect on antigen-induced LMI. Antigen-specific conversion was produced by intermediate concentrations of DLE. Both preparations transferred reactivity to CMV, but only DLE-B transferred reactivity to candida in vitro.

Blood lymphocytes (BL) of eleven patients with lymphoadenopathy syndrome (LAS) were studied for natural killer (NK) activity against the K562 cell line (using both the standard 51Cr release assay and the single-cell cytotoxicity assay on poly-L-lysine-coated coverslips) and for surface phenotype (employing OKT4, OKT8 and Leu7 monoclonal antibodies). A significant reduction in NK activity and in NK active cells was detected, while the percentage of target binding cells was not affected. Furthermore, the OKT4/OKT8 ratio was found to be inverted, and the Leu7+ subpopulation expanded. The patients had high titers of anti-HTLV-III antibodies. This study indicates that defective NK activity in LAS is secondary to an abnormality in the lytic event itself and not in target binding.

The range of immunological abnormalities which underlie and sustain active infections by herpesviruses is wide (1) and makes the use of immuno- stimulating agents a promising therapeutic approach to them. Herpetic keratitis represent a quite severe manifestation of herpetic infection, since patients, if not treated, can develop blindness. Some patients with herpetic keratitis have been found to have impaired cell-mediated immunity (CMI) (1,2) and an immunostimulating therapy has also been proposed in the treatment of these patients.

Natural killer (NK) activity against K-562 and MOLT-4 cell lines and the distribution of T lymphocytic subpopulations were analyzed in 24 patients with primary immunodeficiencies. Theophylline sensitive sheep rosette-forming cells (T-sens) and Fc-IgG receptor-positive (FcIgG+) lymphocytes were normal in the majority of patients with immunoglobulins and B-cell abnormalities. NK activity was decreased in two patients with pure T-cell deficiency. One patient with severe combined immunodeficiency (SCID) lacked T-sens, FcIgG+ cells, and NK activity. In this case, TP-5, a synthetic thymopoietin-derived pentapeptide, partially corrected these abnormalities. These data suggest that the maturation of NK function can be induced by thymic hormones. The significant positive correlation between T-sens, FcIgG+ cells, and NK activity suggests a partial identity of the lymphocytic subpopulations detected by these surface markers and that such populations could mediate NK function in humans.

Dialyzable leukocyte extracts (DLE) have been widely used in attempts to restore cell-mediated immunity (CMI) in patients affected by diseases associated with impairment of cellular (delayed) immune reactivity. After the initial report made by Lawrence1 27 years ago about the ability of DLE to transfer CMI from positive to negative donors, several groups have published conflicting results on both in vitro and in vivo effects of DLE. We shall discuss some of the problems involved in evaluating these results and comparing different DLE, with particular emphasis on our own studies.

Alpha- and gamma-interferon (IFN) production by peripheral blood mononuclear cells (PBMC) from 18 patients affected by primary immunodeficiency syndromes was examined and compared with that of 20 normal donors. Patients included 8 with common variable immunodeficiency (CVI), 2 with congenital agammaglobulinemia, 4 with ataxia-telangiectasia, 2 with hyper-IgE syndrome, 1 with chronic EBV infection, 1 with combined immunodeficiency, and 1 with immunodeficiency with hyper-IgM. No spontaneous IFN production was observed in either patients and controls. Newcastle disease virus-induced alpha-IFN production was found to be normal in all patients. Gamma-IFN was induced by both galactose oxidase and staphylococcal enterotoxin (B). Gamma-interferon production was low or undetectable in patients with ataxia-telangiectasia, in immunodeficiency with hyper-IgM, and in hyper-IgE syndrome. No major defect of gamma-IFN was found in other types of immunodeficiency, despite the presence of occasional low producers (1 of 8 CVI patients and 1 case of congenital agammaglobulinemia). No correlation was found between IFN production and natural killer activity in individual patients. The analysis of lymphocyte subsets by monoclonal antibodies revealed gross imbalances of helper/inducer and suppressor/cytotoxic subpopulations, but no overall correlation could be established with gamma-IFN production. The observation of major defects in gamma-IFN yield only in diseases with depression of T cell-mediated immunity might contribute to a better understanding of the pathogenetical mechanisms in these diseases. Moreover, future studies should monitor these in vitro functions and their modifications by in vitro or in vivo manipulations.

The complement system was examined in a group of eight patients (six with lymphoadenopathy syndrome (LAS); two with acquired immunodeficiency syndrome (AIDS)-related complex (ARC], who were found to be human immunodeficiency virus (HIV)-positive, for the presence of specific HIV-anti-HIV complexes. A significant impairment of the classical and/or alternative pathway was found associated with the presence of cleavage fragments of C3 and/or B and a significant reduction in the complement factors studied. Ultracentrifugation fractions of serum samples obtained from one of the patients were assessed for the detection of specific HIV-anti-HIV (GP41-anti-GP41) complexes and were incubated with normal human serum to determine their complement activation capacity. A clear complement activation was found with the fraction in which a clear peak of HIV-anti-HIV (GP41-anti-GP41) immune complexes was present. The results demonstrate that specific immune complexes and complement activation are sometimes concomitantly present in patients with AIDS-related disease and that specific immune complexes may be one of the causal factors of the pathogenesis of complement activation in these patients. Possible consequences for the severe immune regulation with relevance to the dramatic failure in treating the virus effectively are discussed.

Twelve hundred children with convulsions when feverish were studied during a period of five years. Among them 52 subjects (4.33%) developed nonfebrile seizures after a period of eight months to five years from the first febrile convulsion (group A). Twenty-three children had neither afebrile seizures nor EEG abnormalities during the period of observation (group B). The two groups were comparable for age of the first febrile convulsion onset, sex, and socioeconomic status. None had risk factors for subsequent epilepsy or clinical signs of congenital cytomegalovirus infection. The isolation rate of CMV from urine was 53.84% in patients of group A, 26.09% in children of group B, and 26.83% in healthy control children. Twelve CMV-positive children from group A were followed for one to more than three years. In five of seven children with persisting EEG abnormalities, cytomegaloviruria was still present 13 to 41 months after the first isolation, whereas none of five patients with normal electroencephalograms had viruria after a comparable period. We found that CMV-positive children generally lacked cell-mediated immunity to the virus, whereas CMV-negative patients had positive reactions. Our data suggest a correlation between persistence of neurologic abnormalities and CMV excretion in children with nonfebrile seizures and CMV infection.

During recent years the identification of functionally active T cell subpopulations has become possible in humans by surface markers, 1,2 reactivity with heteroantisera3,4 and by a number of monoclonal antibodies (MoAb) that selectively bind to regulatory T cells.5,6

This study was undertaken with the aim of elucidating the mechanisms underlying the cell-mediated immunodeficiency seen in the acquired immunodeficiency syndrome (AIDS). An intrinsic functional defect in the in vitro surviving T lymphocytes from patients with AIDS has been described. This defect is reflected by profound reductions in both the cloning efficiency of these cells and in the number of precursor cells for response to lectins. Since many patients affected by AIDS present active cytomegalovirus (CMV) infections and impairment in CMV-specific cellular immunity, we examined the number of CMV-specific precursor cells in patients affected by the AIDS-related complex (ARC), who had serum antibodies to CMV and to the human-T-lymphotropic retrovirus-type III (HTLV-III), recently termed human immunodeficiency virus (HIV). Their responses were compared to those of patients with AIDS and to those of healthy-CMV-seropositive and HTLV-III seronegative controls. We detected a significant reduction of precursors for cell-mediated immune response to CMV in AIDS, in comparison to normal controls and a reduction in ARC, even if not significant. In parallel, we assayed the response to phytohemagglutinin, which was maintained in ARC and depressed in AIDS. Our results show a defect of specific cell-mediated immunity to CMV in ARC and AIDS patients.