Frank M. Collins

Adoptive protection of the Mycobacterium tuberculosis-infected lung: Dissociation between cells that passively transfer protective immunity and those that transfer delayed-type hypersensitivity to tuberculin

Adoptive transfer of protective immunity to an aerogenic infection with the facultative intracellular bacterium Mycobacterium tuberculosis was mediated by a population of T cells acquired in the spleen of donor mice at the height of the primary cell-mediated immune response to an immunizing infection with M. bovis bacillus Calmette-Guerin. Successful adoptive immunotherapy was ablated by prior exposure of immune donor cells to ionizing radiation or by treatment of these cells with antibody raised against the Ly-2 marker. In contrast, however, the capacity of immune donor cells to passively transfer delayed-type hypersensitivity (DTH) responses to tuberculin was unaffected by prior treatment with antibody to Ly-2, but was completely ablated by treatment by antibody to Ly-1. These results indicate, that DTH and protective anti-tuberculous immunity are dissociable phenomena, mediated by separate populations of T lymphocytes.

Immunization of mice with mycobacterial culture filtrate proteins.

Culture filtrate proteins were obtained from Mycobacterium tuberculosis cultures after 7 days growth in Proskauer and Beck medium. The protein yield increased substantially to peak about the time the number of viable organisms reached its maximum level (day 8). Examination of the protein concentrate by SDS PAGE revealed the presence of at least 12 separate protein bands varying from 10 to 90 kD. Mice were injected subcutaneously with 20μg of M. tuberculosis culture filtrate (MTCF) protein suspended in saline or Freunds complete or incomplete adjuvant. The vaccinated mice were subjected to an aerogenic challenge with 103 colony‐forming unit (CPU) M. tuberculosis Erdman and a significant reduction in the number of viable organisms was observed in the spleens and lungs determined over a 21‐day period compared with age‐matched normal controls. Mice immunized with the same culture filtrate proteins bound to nitrocellulose particles also showed some resistance to the virulent challenge, suggesting that individual antigens present in the culture filtrate were able to induce a protective T cell‐mediated immune response in appropriately immunized mice.

The relationship of delayed hypersensitivity to acquired antituberculous immunity: I. Tuberculin sensitivity and resistance to reinfection in BCG-vaccinated mice☆

Abstract Specific pathogen-free mice were infected intravenously with different strains of BCG or with M. tuberculosis H 37 R v . Despite marked differences in the resulting levels of tuberculin sensitivity, mice of all groups were comparable in their resistance to challenge with an isoniazid-resistant strain of BCG. A highly virulent strain of M. tuberculosis (Erdman) was no more resistant to inactivation in the tissues of immunized mice than was the drug-resistant strain of BCG. Mice which had been infected with 10 5 M. tuberculosis (Erdman) showed extensive growth of the organism in lung, liver, and spleen; tuberculin sensitivity appeared early, increased rapidly, and then declined. This phenomenon of desensitization was not observed in animals infected with comparable numbers of BCG, but did occur with larger doses of BCG (10 8 ). Spleen cells from these animals were as capable of conferring tuberculin sensitivity upon normal recipients as were cells from tuberculin-sensitive donors. Thus, an absence of peripheral sensitivity does not imply that immunologically committed cells are lacking from the tissues. When animals were challenged with virulent tubercle bacilli 3, 12, and 28 weeks after vaccination, they responded promptly with a major rise in tuberculin sensitivity, thereby demonstrating a persisting memory for mycobacterial antigens. This would explain why specific resistance showed little tendency to wane despite loss of tuberculin sensitivity.

CELLULAR ANTIMICROBIAL IMMUNITY

Acquired resistance to infectious disease may be expressed by a predominantly humoral or a cellular mechanism or, more frequently, by a combination of the two. The cellular interactions which are responsible for the induction of the immune response in the skin, lung, intestinal mucosa, genitourinary tract, conjunctiva, and peritoneal cavity are discussed and the role of living or dead vaccines in the induction of acquired resistance is outlined. The host response involves three different cell types: the phagocytic cell (polymorphs or macrophages), the thymus-dependent (T) lymphocyte, and the thymus-independent (B) lymphocyte-plasma cell line. The normal unstimulated phagocytic cell is capable of killing most nonpathogenic bacteria that gain entry to the tissues. However, the presence of opsonic antibodies and activated macrophages is required to eliminate the pathogenic intracellular parasites. Such immunological activation involves the presence of sensitized T-lymphocytes in the lesion. The cellular response is also characterized by the simultaneous development of a state of delayed-type hypersensitivity (DTH), along with the antimicrobial CMI response. A rising humoral response normally develops subsequently. Killed bacterial cells (except when incorporated into Freunds complete adjuvant) induce the humoral response without the CMI reaction so that such vaccines are not able to fully protect the host against the naturally acquired disease. With the development of cell fractionation methods as well as the identification of distinctive cell surface markers, suspensions of B- and T-cells and macrophages can now be prepared for use in increasingly sophisticated transfer and reconstitution studies. The role of the different cell types in the expression of humoral and cellular immunity has been determined, and the effect of various immunopotentiating and immunosuppressive regimens on the immune system as a whole has been evaluated quantitatively. These studies have led to an appreciation of the role played by suppressor B- and T-cells in the interplay of both humoral and cellular components of the host defense system during the development of immune tolerance, desensitization, anergy, autoimmunity, and the expression of an anamnestic immune response following reinfection.

The In Vitro Induction of Human Immunodeficiency Virus (HIV) Replication in Purified Protein Derivative-Positive HIV-Infected Persons by Recall Antigen Response to Mycobacterium tuberculosis Is the Result of a Balance of the Effects of Endogenous Interleukin-2 and Proinflammatory and Antiinflammatory Cytokines

Coinfection with Mycobacterium tuberculosis and human immunodeficiency virus (HIV) is a serious problem, particularly in developing countries. Recently, M. tuberculosis and purified protein derivative (PPD) were demonstrated to induce HIV replication in CD8 T cell-depleted peripheral blood mononuclear cells from HIV-positive, PPD-positive persons but not in cells from PPD-negative persons. The role of endogenous and exogenous cytokines in modulating M. tuberculosis-induced HIV replication was evaluated. M. tuberculosis-induced HIV replication decreased following simultaneous inhibition of endogenous interleukin (IL)-2, IL-1beta, and tumor necrosis factor-alpha by the addition of soluble receptors and receptor antagonists or following exogenous IL-10 and transforming growth factor (TGF)-beta. In contrast, neutralization of endogenous IL-10 and TGF-beta augmented M. tuberculosis-induced HIV replication by increasing cellular activation. Thus, the balance between IL-2 and proinflammatory and antiinflammatory cytokines plays a major role in M. tuberculosis-induced replication of HIV.

Tuberculosis: The Return of an Old Enemy

Tuberculosis is an ancient human scourge that continues to be an important public health problem worldwide. The increasing number of multidrug-resistant (MDR) M. tuberculosis isolates from both AIDS and non-AIDS patients is an ominous trend that threatens tuberculosis eradication programs both in the U.S. and overseas. New antituberculosis vaccines with therapeutic properties are urgently needed for human immunodeficiency virus-infected individuals, as well as health care professionals likely to be exposed to MDR tubercle bacilli. Recombinant DNA vaccines bearing protective genes from virulent M. tuberculosis are being developed using shuttle phasmids to transfer genetic material from one mycobacterial species to another. Improved assay procedures are needed to measure the protection afforded by these new vaccines under experimental and field test conditions. Tuberculosis vaccine development should be given a high priority in current medical research goals.

Effects of in vivo T lymphocyte subset depletion on mycobacterial infections in mice

The relative importance of CD4+ and CD8+ T cell subsets in the expression of acquired resistance to systemic infection by Mycobacterium kansasii was determined. T cell subsets were depleted in thymectomized C57BL/6 mice by the intravenous administration of monoclonal antibodies directed against the relevant T cell determinants. Depletion of the CD4+ subset exacerbated the severity of the infection in intravenously challenged mice. This effect was apparent in the first 2 weeks of the infection and persisted throughout the 12 weeks of the study. On the other hand, depletion of the CD8+ cells had no apparent effect on the growth curves. Infections by Mycobacterium tuberculosis Erdman or bacille Calmette‐Guérin (BCG) Pasteur were also substantially enhanced by CD4 depletion, but not by the depletion of CD8+ cells. The effect of subset depletion on infections by M. tuberculosis and BCG was examined in both innately susceptible C57BL/6 mice and innately resistant B6D2 mice.

Protection to mice afforded by bcg vaccines against an aerogenic challenge by three mycobacteria of decreasing virulence

Specific pathogen-free mice were vaccinated subcutaneously with 10(7) CFU of BCG Pasteur or BCG Glaxo and 30 or 90 days later, the mice were challenged aerogenically with Mycobacterium tuberculosis (Erdman or South Indian strains) or with M. avium. Both vaccines induced substantial levels of resistance to tuberculosis and tuberculin hypersensitivity. There was no detectable difference in the host response to the three aerogenic challenges which could be related in any way to the immunogenicity of the BCG strain or to the mouse virulence of the challenge organism. These results do not support the hypothesis that the protective activity of BCG vaccines varies, depending upon the virulence of the infecting organism.

The restorative effect of peritoneal macrophages on delayed hypersensitivity following ionizing radiation

Abstract Sublethal whole-body irradiation in the guinea pig had little demonstrable effect on the development of delayed hypersensitivity but caused a profound, though transient depression of dermal reactivity in previously sensitized animals. Macrophage-rich peritoneal cell suspensions from non-sensitive donors when injected intradermally with eliciting antigen, resulted in the restoration of a significant degree of reactivity following irradiation. Inocula of lymphocytes, on the other hand, failed to yield similar results. These findings, when taken with the persistence of low levels of reactivity following irradiation and the ability to transfer reactivity with peritoneal exudate cells from animals so treated, warrant the conclusion that the presence of macrophages is necessary for the expression of cutaneous delayed hypersensitivity. The spontaneously renewed activity which follows the depressed phase is thus more of a reflection of the recovery of macrophage precursors from radiation injury, rather than the emergence of a new population of sensitized cells. The results, in addition, substantiate the belief that the expression of delayed hypersensitivity requires at least two cell populations, only one of which carries the property of specificity at the outset. The presence of the sensitized population cannot always be excluded by a weak or absent skin test.

Aerogenic vaccination of mice with Mycobacterium bovis BCG

The course of infection with Mycobacterium bovis BCG Pasteur was followed against time in groups of mice vaccinated by either the aerogenic or subcutaneous route. The generation of acquired protective immunity and immunological memory was determined in each group by adoptive immunisation procedures. In addition, subcutaneously vaccinated mice were tested for their ability to resist an aerogenic challenge with a lethal dose of M. tuberculosis. No overall qualitative differences in the magnitude or longevity of antituberculosis immunity in mice vaccinated by the two procedures were observed. It is concluded that aerogenic vaccination offers no immunological advantage over vaccination by the subcutaneous route.