Serologically defined and lymphocyte-defined components of the major histocompatibility complex in the mouse.

The mixed leukocyte culture (MLC) test is an in vitro model of the recognition phase of the homograft response. For the most part, activation in MLC is dependent on differences of the major histocompatibility complex (MHC). Our present studies in the mouse suggest that activation is primarily associated with differences of genetic regions of the MHC other than those which control the serologically defined (H-2) antigens. These differences do not lead to cytotoxic or agglutinating antibody formation despite extensive immunization; we have called these differences lymphocyte-defined (LD) differences. The strongest stimulation in MLC is associated with differences of the Ir region. It is possible that the Ir product is the T cell receptor and that it is this same molecule which can act as the stimulatory agent in MLC. Other possibilities are discussed.

Cell-mediated lympholysis. Importance of serologically defined H-2 regions.

The cell-mediated lympholytic capability of mouse spleen cells stimulated in mixed lymphocyte culture is related to the major histocompatibility complex genotype on target lymphocytes. The strain combinations AQR-B10. T(6R) and B10.A(4R)-B10.A(2R) that result in significant mixed lymphocyte culture activation do not mediate cell-mediated lympholysis on sensitizing target lymphocytes; serologically defined regions (H-2K and H-2D) are identical within each combination. H-2K or H-2D region disparity alone does not cause cell-mediated lympholysis. However after mixed lymphocyte culture activation as seen with B10.A-B10.T(6R), a target cell bearing only an H-2K region difference from the effector cell is sensitive to cell-mediated lympholysis. Likewise an H-2D region difference is an adequate target after mixed lymphocyte culture activation of the effector cell in the combination B10.A(2R)-B10.D2.

Genetic aspects of canine mixed leukocyte cultures.

Recent data stress the importance of matching donor and recipient of an organ graft for both the serologically defined (SD) and lymphocyte-defined (LD) determinants. To allow experimental evaluation of the effect of these SD and LD structures in a noninbred experimental animal, mixed leukocyte culture tests were performed between SD identical and nonidentical dogs to clarify the LD system in these animals. The results of these experiments can be summarized as follows: (a) In the dog there is a LD locus distinct from the known SD loci, which in all probability is localized outside the first (SD-1) series locus on the chromosome. (b) The crossing-over frequency between the SD and LD loci on the chromosome is low. (c) Studies in SD identical unrelated dogs and random unrelated dogs show an apparent high linkage disequilibrium between SD and LD loci. (d) The LD system in dogs is polymorphic.

Three HLA-D region antigens defined by primed LD typing.

We have recently described a new method, primed LD typing or PLT, for specific identification of HLA-D antigens. Highly discriminatory PLT cells have been developed which clearly differentiate between cells of individuals that restimulate strongly and those that restimulate weakly. Seven such discriminatory PLT cells have been used to define three antigens called PL1, PL2, and PL3; two more PLT cells may define antigen(s) PL4.

Secondary cell-mediated lympholysis: importance of H-2 LD and SD factors.

Lymphocytes stimulated in mixed leukocyte cultures and left for 13-17 days, i.e. beyond their peak proliferative and cytotoxic reactivities, can be restimulated to give a secondary-type rapid and strong proliferative and cytotoxic response when confronted with cells of the original sensitizing cell donor. We have concerned ourselves primarily with the requirements of restimulation for the presence of LD and/or SD stimuli on the restimulating cells. (a) The low level cell-mediated lympholysis (CML) associated with LD differences in a primary CML can be restimulated to give a secondary-type response by those same LD antigens. (b) If the original sensitizing cells differ from the responding cells by both LD and SD antigens, restimulation with only the LD antigens, or third-party cells presumably carrying cross-reactive LD antigens, can restimulate the secondary CML responses directed against the SD antigens on the original sensitizing cells. (c) The presence of SD antigens on the restimulating cells that are cross-reactive with the primary sensitizing SD antigens (as determined in a primary CML) leads to the preferential activation of cytotoxic T lymphocytes reactive to those antigens although maximum cytotoxicity is still directed at cells carrying the original sensitizing SD antigens. A model to explain these results is presented.

Leukemia-associated antigens in the mixed leukocyte culture test.

Patients with acute leukemia (blast cells in the peripheral blood) manifest antigens in the one-way mixed leukocyte culture test. Leukocytes from normal patients and leukocytes of patients in remission do not clearly show these antigens. These antigens could be leukemia-associated antigens in man.

Chloral hydrate: a solvent for biological membranes.

A buffer system conitainin chloral hydrate, taurine, and bromopyridinium lactate was used to dissolve several biological membranes and separate their protein components by polyacrylamide gel electrophoresis. This solvent system was capable of separating molecules of similar size on the hasis of their charge and allows easy recovery of the proteins Thus. aqueous chloral hydrate is an effective solvent for biological membranes.

HL-A LD (lymphocyte defined) typing: a rapid assay with primed lymphocytes.

When human lymphocytes are cultured for 9 to 14 days with stimulating cells of a family member differing by a single HL-A haplotype they become "primed" to recognize specific HL-A LD (mixed lymphocyte culture) antigens. These primed lymphocytes respond specifically and rapidly when "restimulated" with cells of a person that contain the same LD antigens as those of the priming haplotype. Specific HL-A LD antigens can be detected within 24 hours by this primed LD typing.

Genetic and immunological complexity of major histocompatibility regions.

There are genetic differences within the major histocompatibility complex of the mouse which lead to skin graft rejection but which cannot be detected serologically. When confronted with these differences on allogeneic cells, lymphocytes proliferate in vitro. In other cases, in vitro lymphocyte proliferation but no skin graft rejection is associated with loci that are linked to but genetically separable from the loci controlling the serologically defined antigens.

Beta 2-microglobulin: association with lymphocyte receptors.

beta(2)-Microglobulin (beta(2)m) is a low-molecular-weight protein constituent of lymphocyte membranes. Amino acid sequence analysis has revealed a high degree of homology between the beta(2)m and certain regions of immunoglobulin molecules, suggesting a possible recognition function for the beta(2)m, in analogy with the immunoglobulins. The data presented demonstrate that highly specific antiserum against beta(2)m blocks lymphocyte reactivity against allogeneic cells in mixed leukcocyte cultures and against phytohemagglutinin, both of which processes presumably function via a cell surface receptor on thymus-derived (T) lymphocytes. There is very little inhibition of T lymphocyte rosette formation with sheep red blood cells. The findings suggest a possible relation between the beta(2)m and recognition units on the T lymphocyte surface.

Cell mediated immunity: separation of cells involved in recognitive and destructive phases.

The mixed leukocyte culture (MLC) and the cell mediated lympholysis (CML) assays are used as in vitro models of the afferent, or recognitive, and efferent, or destructive, phases of the homograft reaction. Activity in both of these tests has been related to differences at the major histocompatibility complex, HL-A in man and H-2 in mouse. Recent evidence suggests that the presumed cell surface differences which lead to cell proliferation in MLC are different from those which act as a target for CML. Data are presented providing further support for this hypothesis; in addition separate cell populations may respond to the differences which activate cells in MLC and to the differences which serve as targets for CML. There thus appears to be a dichotomy both for genetic control of, and cell populations involved in, the recognitive and destructive phases of cell mediated immunity.

Clinical evaluation of a new test of hemostasis: the Filter Bleeding Time.

The clinical value of a new in vitro test of hemostasis, which we have called Filter Bleeding Time (FBT), was determined in 59 patients referred because of a suspected bleeding disorder. FBT is based on the progressive slowing of the drop rate of citrated blood through a filter of woven Dacron under constant pressure as platelet aggregates occlude the filter. The value for FBT is defined as the time when the blood drop interval has reached 1 minute. The Mayo modification of the Ivy bleeding time (IBT) was performed in all patients; platelet response to ADP, collagen, epinephrine and arachidonate was performed in 24 patients. In 30 normal volunteers FBT measured 1-3 hr after venipuncture was 2.8 +/- 1.5 (means +/- 1SD) min. The FBT was prolonged in 3 of 3 patients with Glanzmann's thrombasthenia, 2 with disseminated intravascular coagulation, 1 with chronic lymphocytic leukemia, 1 with myelofibrosis, and 1 who had taken aspirin. In 6 patients FBT was prolonged while IBT was normal: 4 after taking aspirin, 2 with polycythemia vera. All 6 had reduced platelet aggregation (PA) to ADP (5 microM), collagen (2 mg/ml), epinephrine (5 microM) and/or arachidonate (1.7 mM). In 3 patients FBT was normal while IBT was abnormal: 1 with disseminated intravascular coagulation, 2 undiagnosed; 1 of these 3 had abnormal PA. Of 6 patients with von Willebrand's disease, FBT was prolonged in 5 and borderline in 1; IBT was prolonged in 3, normal in 1, and not done in 2 infants.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of a heparin assay method using a fluorogenic synthetic peptide substrate for thrombin.

A heparin assay method based on thrombin cleavage of a fluorogenic synthetic peptide has been evaluated in normal subjects, in patients with chronic renal disease on hemodialysis, and in patients with other disorders. The method has satisfactory accuracy, sensitivity, and reproducibility. In contrast to methods commonly used to monitor heparin therapy, it measures heparin directly and is uninfluenced by patient variables such as low antithrombin III. The method may provide valuable information in selected clinical settings such as cardiopulmonary bypass and in patients with chronic renal disease who receive heparin during hemodialysis or prior to surgery.

Appropriate and necessary healthcare: new language for a new era.

Conceptual and language changes are necessary to accompany the paradigm shift from fee-for-service medicine to managed care. Medical necessity is an inadequate and ambiguous term defined differently by providers, payers, patients, and legislators. The attempt by legislators in Minnesota to develop a universal standard benefits set for healthcare services strikingly underscores the need to define relevant terminology to accompany the transition to managed care. We suggest the term appropriate and necessary healthcare as a state-of-the-art term for the new era of managed care.

Past, present, and future aspects of histocompatibility.

We have stressed problems attendant on studies of the MHC, ignoring non-HLA factors and their role in allograft immunity. Many other topics could have been chosen for discussion is any such overview; our selection reflects our own interests. We felt assured, however, that excellent coverage by our many colleagues of the diverse and varied aspects of histocompatibility not approached in this summary has allowed us this freedom.

Ethics and Medicaid: a new look at an old problem.

Recent proposals to reform Medicaid, driven primarily by the need for cost containment, rarely pay explicit attention to values. This paper presents the Medicaid Values Framework, the authors' interpretation of a set of societal ideals embodied in Title XIX of the Social Security Amendments of 1965. The Framework comprises seven interlocking values that are stratified into three interdependent tiers--access, quality, and equity. We use the access and equity tiers to analyze treatment of Aid to Families with Dependent Children (AFDC) and Supplemental Security Income (SSI) recipients under Medicaid. We document striking inequities in eligibility standards and in funding for the two groups--inequities that unexpectedly fail to translate into marked disparities in access to Medicaid. In conclusion, we comment on why the present inequities exist and why they are ethically unacceptable.