Enhanced T-cell activation and differentiation in lymphocytes from transgenic mice expressing ubiquitination-resistant 2KR LAT molecules.

Linker for activation of T cells (LAT) is critical for the propagation of T-cell signals upon T-cell receptor (TCR) activation. Previous studies demonstrated that substitution of LAT lysines with arginines (2KR LAT) resulted in decreased LAT ubiquitination and elevated T-cell signaling, indicating that LAT ubiquitination is a molecular checkpoint for attenuation of T-cell signaling. To investigate the role of LAT ubiquitination in vivo, we have generated transgenic mice expressing WT and ubiquitin-defective 2KR LAT. On TCR stimulation of T cells from these mice, proximal signaling and cytokine production was elevated in 2KR versus wild-type (WT) LAT mice. Enhanced cytolytic activity as well as T-helper responses were observed on LAT expression, which were further elevated by 2KR LAT expression. Despite greater T-effector function, WT or 2KR LAT expression did not have any effect on clearance of certain pathogens or tumors. Our data support the model that lack of tumor clearance is due to increased differentiation and acquisition of effector phenotype that is associated with suboptimal immunity in an immunotherapy model. Thus, our data further reinforce the role of LAT ubiquitination in TCR signaling and uncovers a novel role for LAT in driving T-cell differentiation.

Inhibition of TGF-ß signaling in genetically engineered tumor antigen-reactive T cells significantly enhances tumor treatment efficacy.

Transforming growth factor ß (TGF-ß) is a cytokine with complex biological functions that may involve tumor promotion or tumor suppression. It has been reported that multiple types of tumors secrete TGF-ß, which can inhibit tumor-specific cellular immunity and may represent a major obstacle to the success of tumor immunotherapy. In this study, we sought to enhance tumor immunotherapy using genetically modified antigen-specific T cells by interfering with TGF-ß signaling. We constructed three γ-retroviral vectors, one that expressed TGF-ß-dominant-negative receptor II (DNRII) or two that secreted soluble TGF-ß receptors: soluble TGF-ß receptor II (sRII) and the sRII fused with mouse IgG Fc domain (sRIIFc). We demonstrated that T cells genetically modified with these viral vectors were resistant to exogenous TGF-ß-induced smad-2 phosphorylation in vitro. The functionality of antigen-specific T cells engineered to resist TGF-ß signaling was further evaluated in vivo using the B16 melanoma tumor model. Antigen-specific CD8+ T cells (pmel-1) or CD4+ T cells (tyrosinase-related protein-1) expressing DNRII dramatically improved tumor treatment efficacy. There was no enhancement in the B16 tumor treatment using cells secreting soluble receptors. Our data support the potential application of the blockade of TGF-ß signaling in tumor-specific T cells for cancer immunotherapy.

Effect of cholecystokinin on human cholangiocarcinoma xenografted into nude mice.

Gastrointestinal polypeptide hormones regulate growth of various normal gastrointestinal tissues as well as certain visceral cancers. Since cholecystokinin (CCK) promotes growth of normal biliary tract, we sought to determine whether CCK affects the growth and metabolism of human cholangiocarcinoma line SLU 132. Twenty-six nude mice with s.c. xenografts of this cancer received either CCK octapeptide (50 micrograms/kg/dose) or 0.9% NaCl solution (saline) twice a day i.p. for 14 days. Tumor volume was calculated from Vernier caliper measurements. At sacrifice on Day 15, tumors were excised, weighed, and examined histologically. DNA, RNA, and protein were measured in the xenografted carcinomas. Because this cholangiocarcinoma produces carcinoembryonic antigen (CEA), we obtained serum at sacrifice for CEA radioimmunoassay and also tumor tissue for CEA immunolabeling with murine anti-CEA monoclonal antibody. Serum CEA levels were 90% higher in the CCK-treated group. Tumor tissue in the CCK-treated group also contained more CEA than did the controls. Mean tumor volume increased significantly in the saline group during the 14-day treatment period, whereas mean tumor volume did not increase significantly in the CCK group. Exogenous high-dose CCK thus appears to increase production and release of CEA from SLU-132; it also appears to retard growth of this tumor line in the nude mouse.

High incidence of myocarditis by endomyocardial biopsy in patients with idiopathic congestive cardiomyopathy.

Thirty-five patients with unexplained congestive heart failure were evaluated with endomyocardial biopsy. Utilizing microscopic, ultrastructural and immunofluorescent studies, samples were classified as exhibiting either no inflammation (cardiomyopathy) or active lymphocytic myocarditis, grade I to IV. Twenty-two (63%) of the patients had inflammatory changes. Of these 22 patients, 18 had low grade I or II inflammation, 7 were treated with immunosuppressive agents with improvement in 5 and stabilization in 1. One patient died of progressive congestive heart failure. Three of four patients with high grade III or IV myocarditis died after a fulminant course. The results suggest that inflammatory myocarditis may be more common than previously suspected and add evidence that there may be ongoing inflammation in many cases of congestive cardiomyopathy.

Heart graft arteriosclerosis. An ominous finding on endomyocardial biopsy.

Heart graft arteriosclerosis remains a severe and irreversible complication of allogeneic heart transplantation despite prophylactic therapy. Immunologically mediated endothelial damage has been proposed as a stimulus for the development of graft arteriosclerosis. The vascular lesions may accumulate large amounts of lipid resembling atheromas, or may be purely proliferative, as illustrated in the case of a 42-year-old heart transplant patient who developed slowly progressive graft dysfunction at eight months posttransplantation. Endomyocardial biopsy ten months posttransplantation revealed proliferative arteriolar occlusion, while changes on the coronary angiogram were minimal. Repeat biopsy at eleven months showed ischemic myocardial necrosis. The patient expired shortly thereafter. On postmortem examination, proliferative graft arteriosclerosis affecting both intramural and epicardial vessels was present, along with massive biventricular infarction. Tissue immunofluorescence studies demonstrated extensive vascular deposition of immunoglobulin and complement. We propose that (1) the presence of proliferative arteriolar occlusion on endomyocardial biopsy is predictive of poor heart graft survival; (2) proliferative graft arteriosclerosis may appear as advanced small vessel disease before extensive large vessel involvement is detected by coronary angiogram; and (3) immunofluorescence results support an immune-mediated mechanism of vascular injury in proliferative heart graft arteriosclerosis.

Clinical temporary ventricular assist. Pathologic findings and their implications in a multi-institutional study of 41 patients.

Forty-one patients, distributed among four centers, had left (33 patients), right (five), or bilateral (three) temporary ventricular assistance with textured (24) or smooth (17) surfaced diaphragm pumps, during an evaluation supported by the National Institutes of Health. Cardiac failure had occurred in 39 postoperative patients (after aorta-coronary bypass [23], valve replacement [four], both [nine], or other [three]), with total cardiopulmonary bypass time mean 306 minutes (range 69 to 600). Two patients had cardiomyopathy. Death of 35 nonsurvivors was due to myocardial necrosis (14), hemorrhage (nine), cerebrovascular accidents (three), infection (three), and other (six). Mean duration of support in all patients was 62 hours. In 16 patients (40%) whose condition improved, cardiac assist duration was mean 127 hours (range 48 to 264), compared with mean 19 hours (range 1 to 120) in 25 who did not. Of 17 patients in whom duration of support exceeded 72 hours, 15 (88%) improved, 11 were weaned, and six survived long term. Tissue examination (in 33 patients) by biopsy at pump implantation or autopsy revealed coagulation or contraction band myocyte necrosis, with or without hemorrhage, in 26 patients; of these, 10 improved and six were long-term survivors. Pump-related complications (two) included pulmonary embolism, most likely related to a cannulation site thrombus, and an aortic cannulation site infection in one patient each. This study suggests that mechanical cardiac assist may be accomplished with a low complication rate; should not necessarily be denied to patients with existing necrosis, because myocardial necrosis does not preclude improvement or survival; and frequently leads to functional myocardial recovery if patients survive early noncardiac complications, often the result of long duration of cardiopulmonary bypass.

The prognostic importance of steroid receptors in endometrial carcinoma.

Three hundred forty-nine cases of primary endometrial carcinoma (endometrioid, adenosquamous, and clear-cell) were studied to investigate the relative prognostic importance of age, menopausal status, stage, histology, myometrial invasion, and estrogen and progesterone receptor content. Excluding menopausal status, all of these variables had a significant relationship to overall survival in a univariate analysis. Using a Cox multivariate regression analysis, stage, age, and an estrogen receptor value of more than 70 fmol/mg protein, combined with a progesterone receptor value of more than 30 fmol/mg protein, were independently associated with survival. The results demonstrate that for maximum prognostic information, both estrogen and progesterone content of tumors should be measured. Maximum prognostic information is obtained by using cutoff levels that are much higher than those traditionally accepted. This has particular relevance for patient stratification in clinical trials investigating receptor information and response to adjuvant or therapeutic treatment.

Response to exogenous cholecystokinin of six human gastrointestinal cancers xenografted in nude mice.

Gastrointestinal hormones regulate growth of cancers as well as normal tissues. We investigated whether long-term cholecystokinin (CCK) administration might affect growth or metabolism of human tumors xenografted in nude mice. In each experiment, approximately 20 nude mice bearing subcutaneous xenografts of the particular cancer line being studied were used. Half received CCK and half received saline solution intraperitoneally twice daily for 14 days. Tumor volume and body weight were measured every 3 days. If the tumors produced marker substances, these were measured in nude mouse serum and also in the xenografts. Tumor growth was significantly retarded by CCK in two of the six cancers studied. In each case, DNA, RNA, and protein reflected tumor volumes. In one of these tumors (SLU 077), serum carcinoembryonic antigen (CEA) levels paralleled the tumor volumes. In another tumor (SLU 132), serum CEA levels and tumor immunolabeling for CEA and pancreatic oncofetal antigen increased in response to CCK administration, whereas tumor volumes did not. These findings suggest that exogenous highdose CCK altered the growth and metabolism in two of six human cancers studied.

Essentialism and selectionism in cognitive science and behavior analysis.

Contingencies of selection, be they phylogenetic or ontogenetic, merely set boundaries on units; they do not provide blueprints. Thus, variability is fundamental to all products of selection. Skinner, by characterizing the units of analysis in behavior as generic in nature, established his science squarely within the selectionist paradigm, thereby avoiding the tendency, common throughout psychology, to slip into essentialist analyses. The distinction between essentialism and selectionism is refined in this article, and prominent examples of essentialism in linguistics, theories of memory, theories of representation, associationism, and even in behavior analysis are identified. Recent trends in cognitive science--specifically, research on adaptive networks--is amenable to a selectionist interpretation, suggesting the possibility of future fruitful interactions with behavior analysis.

A selectionist approach to reinforcement.

We describe a principle of reinforcement that draws upon experimental analyses of both behavior and the neurosciences. Some of the implications of this principle for the interpretation of behavior are explored using computer simulations of adaptive neural networks. The simulations indicate that a single reinforcement principle, implemented in a biologically plausible neural network, is competent to produce as its cumulative product networks that can mediate a substantial number of the phenomena generated by respondent and operant contingencies. These include acquisition, extinction, reacquisition, conditioned reinforcement, and stimulus-control phenomena such as blocking and stimulus discrimination. The characteristics of the environment-behavior relations selected by the action of reinforcement on the connectivity of the network are consistent with behavior-analytic formulations: Operants are not elicited but, instead, the network permits them to be guided by the environment. Moreover, the guidance of behavior is context dependent, with the pathways activated by a stimulus determined in part by what other stimuli are acting on the network at that moment. In keeping with a selectionist approach to complexity, the cumulative effects of relatively simple reinforcement processes give promise of simulating the complex behavior of living organisms when acting upon adaptive neural networks.

The S-R issue: its status in behavior analysis and in Donahoe and Palmer's learning and complex behavior.

The central focus of this essay is whether the effect of reinforcement is best viewed as the strengthenng of responding or the strengthening of the environmental control of responding. We make the argument that adherence to Skinner's goal of achieving a moment-to-moment analysis of behavior compels acceptance of the latter view. Moreover, a thoroughgoing commitment to a moment-to-moment analysis undermines the fundamental distinction between the conditioning process instantiated by operant and respondent contingencies while buttressing the crucially important differences in their cumulative outcomes. Computer simulations informed by experimental analyses of behavior and neuroscience are used to illustrate these points.

Discriminated interresponse times: role of autoshaped responses.

When discriminated interresponse-time (IRT) procedures have been used to assess preference relations among temporally extended operants, deviations from matching have been obtained. Using a yoked-control procedure, the present study found that key pecking in a discriminated IRT procedure has two sources of strength--that arising from the response-reinforcer contingency that is explicitly arranged, and that arising from a stimulus-reinforcer contingency that is a by-product of the explicitly arranged contingency. The key pecking of all lead birds, and that of 3 of the 4 birds exposed to a yoked autoshaping procedure, was controlled by the keylight that signaled the lead birds' criterion IRTs. Because stimulus control of key pecking by the keylight, whether autoshaped or discriminative, fosters deviations from matching, the discriminated IRT procedure does not provide an appropriate basis for conclusions about preference relations among IRTs.

The speaker as listener: The interpretation of structural regularities in verbal behavior.

Regularities in word order not specifically addressed by Skinner require behavioral interpretation if our field is to become more influential among students of language. Three such phenomena are briefly described in traditional structural terms and are offered as test cases: subtle differences in dative verbs, transformational traces, and the formation of compound nouns. It is argued that the variables that control such regularities derive from the speaker's repertoire as listener. Intraverbal frames are established as verbal responses in the listener through reinforcement by parity. Transitions from element to element in such frames are controlled, moment to moment in time, partly by the speaker's responses as a listener to his or her own verbal behavior. Although this account offers only a tentative interpretation of grammar and syntax in a limited domain, it suggests that the conceptual tools of behavior analysis are adequate to the task of explaining even the most subtle of grammatical rules.

Jaundice: a 10 year review of 41,000 live born infants.

A review is presented of jaundiced newborn infants during the 10-year period to 1980. Included are those whose serum bilirubin level was 154 mumol/l or more. Of 41,057 live births, 4,406 (10.7%) infants had hyperbilirubinaemia. The most common (19.9;%) aetiological factor was prematurity, followed by ABO erythroblastosis 7.1%; sepsis 3.4%; Rhesus erythroblastosis 2.7%; bruising 2.2%; multifactorial 1.0% and glucose-6-phosphate dehydrogenase deficiency 0.5%. Treatment was not undertaken in 2,855 (64.7%) infants, but 1,419 (32.2%) received phototherapy alone, 122 (2.7%) infants received both exchange transfusion and phototherapy and 10 (0.2%) infants received exchange transfusion alone. Of the infants requiring exchange transfusion 50.0% had Rhesus erythroblastosis, 28.0% ABO erythroblastosis, 10.6% jaundice of prematurity and the remainder were due to a variety of causes. Sixty-three (1.4%) infants died, with two deaths being related to the hyperbilirubinaemia, as their death was due to necrotizing enterocolitis following exchange transfusion. Phototherapy proved safe with no deaths directly attributable to its use.