Narrative interpretations for clinical laboratory evaluations: an overview.

As the clinical laboratory test menu has significantly expanded in volume and complexity, there is a rapidly growing need by clinicians for narrative interpretations of complex studies that resemble those provided in anatomic pathology and radiology. In this report, the impact of advice on laboratory test selection and interpretation is presented with regard to providing adequate quality of care, reducing medical error, and reducing the cost for health care. In addition, past and current attempts to address the physician's need for advice on laboratory test selection and interpretation are also described. These include curbside consultations, intelligent laboratory information systems, and medical information from the Internet. Each is presented with examples from the literature and with its advantages and disadvantages for practicing clinicians confronting large, expensive test menus and the results of esoteric assays.

The generation of narrative interpretations in laboratory medicine: a description of service-specific sign-out rounds.

The logistical details for organizing effective interpretive rounds in a laboratory medicine subspecialty must be carefully established so that expert opinions are provided in a timely fashion in a patient-specific report, rather than as a collection of fixed comments associated with a particular laboratory result generated by a computer This report describes the test batteries for interpretations, the billing for interpretations, clinical examples of interpretations, and interpretations for which billing is not typically performed in several clinical or laboratory areas in our institution. These include coagulation disorders, hemoglobin and anemia evaluations, autoimmune disorders, serum protein analysis, toxicology, molecular diagnostics, and transfusion medicine. The information in this report should provide sufficient detail to allow development of interpretive services with successful billing for the areas in laboratory medicine described.

Selenomonas lipolytica sp. nov., an obligately anaerobic bacterium possessing lipolytic activity.

A novel, oligately anaerobic bacterium capable of hydrolysing lipids was isolated from a tropical anaerobic lagoon receiving waste water from an edible oil mill. The isolate had many characteristics similar to those of members of the genus Selenomonas. The isolate showed lipolytic activity on tributyrin, triolein and groundnut oil in qualitative plate clearance assays, which has not been reported for the type strain of the genus Selenomonas. It did not require n-valerate supplementation for growth on glucose. Acetate and propionate were the only volatile fatty acids produced from glucose fermentation with propionate as the major end product. The isolate could grow optimally at pH 6.8 and at a temperature of 40 degrees C. It could tolerate NaCl concentrations of up to 40 g l-1. The G&C content of the DNA was 40 mol% as determined by thermal denaturation analysis. Comparison of partial 165 rRNA gene sequences revealed that the isolate was most closely related to genus Selenomonas with 91% sequence similarity (250 bp compared) to Selenomonas ruminantium strain GA 192. On the basis of the results obtained in the present investigation, it is suggested that a new species of Selenomonas should be created for this novel isolate and the name Selenomonas lipolytica is proposed for this new species. The type strain is strain CF1BT (= MCMB 505T).

Demonstration of an interferon gamma-dependent tumor surveillance system in immunocompetent mice.

This study demonstrates that endogenously produced interferon gamma (IFN-gamma) forms the basis of a tumor surveillance system that controls development of both chemically induced and spontaneously arising tumors in mice. Compared with wild-type mice, mice lacking sensitivity to either IFN-gamma (i.e., IFN-gamma receptor-deficient mice) or all IFN family members (i.e., Stat1-deficient mice) developed tumors more rapidly and with greater frequency when challenged with different doses of the chemical carcinogen methylcholanthrene. In addition, IFN-gamma-insensitive mice developed tumors more rapidly than wild-type mice when bred onto a background deficient in the p53 tumor-suppressor gene. IFN-gamma-insensitive p53(-/-) mice also developed a broader spectrum of tumors compared with mice lacking p53 alone. Using tumor cells derived from methylcholanthrene-treated IFN-gamma-insensitive mice, we found IFN-gamma's actions to be mediated at least partly through its direct effects on the tumor cell leading to enhanced tumor cell immunogenicity. The importance and generality of this system is evidenced by the finding that certain types of human tumors become selectively unresponsive to IFN-gamma. Thus, IFN-gamma forms the basis of an extrinsic tumor-suppressor mechanism in immunocompetent hosts.

Regulation of the interleukin (IL)-12R beta 2 subunit expression in developing T helper 1 (Th1) and Th2 cells.

The developmental commitment to a T helper 1 (Th1)- or Th2-type response can significantly influence host immunity to pathogens. Extinction of the IL-12 signaling pathway during early Th2 development provides a mechanism that allows stable phenotype commitment. In this report we demonstrate that extinction of IL-12 signaling in early Th2 cells results from a selective loss of IL-12 receptor (IL-12R) beta 2 subunit expression. To determine the basis for this selective loss, we examined IL-12R beta 2 subunit expression during Th cell development in response to T cell treatment with different cytokines. IL-12R beta 2 is not expressed by naive resting CD4+ T cells, but is induced upon antigen activation through the T cell receptor. Importantly, IL-4 and IFN-gamma were found to significantly modify IL-12 receptor beta 2 expression after T cell activation. IL-4 inhibited IL-12R beta 2 expression leading to the loss of IL-12 signaling, providing an important point of regulation to promote commitment to the Th2 pathway. IFN-gamma treatment of early developing Th2 cells maintained IL-12R beta 2 expression and restored the ability of these cells to functionally respond to IL-12, but did not directly inhibit IL-4 or induce IFN-gamma production. Thus, IFN-gamma may prevent early Th cells from premature commitment to the Th2 pathway. Controlling the expression of the IL-12R beta 2 subunit could be an important therapeutic target for the redirection of ongoing Th cell responses.

Targeted disruption of the Stat1 gene in mice reveals unexpected physiologic specificity in the JAK-STAT signaling pathway.

The JAK-STAT signaling pathway has been implicated in mediating biological responses induced by many cytokines. However, cytokines that promote distinct cellular responses often activate identical STAT proteins, thereby raising the question of how specificity is manifest within this signaling pathway. Here we report the generation and characterization of mice deficient in STAT1. STAT1-deficient mice show no overt developmental abnormalities, but display a complete lack of responsiveness to either IFN alpha or IFN gamma and are highly sensitive to infection by microbial pathogens and viruses. In contrast, these mice respond normally to several other cytokines that activate STAT1 in vitro. These observations document that STAT1 plays an obligate and dedicated role in mediating IFN-dependent biologic responses and reveal an unexpected level of physiologic specificity for STAT1 action.

Tissue-specific targeting of cytokine unresponsiveness in transgenic mice.

The ubiquitous cellular distribution of certain cytokine receptors has hampered attempts to define the physiologically important cell-specific functions of cytokines in vivo. Herein, we report the generation of transgenic mice that express a dominant-negative IFN gamma receptor alpha chain mutant under the control of either the human lysozyme promoter or the murine lck proximal promoter, which display tissue-specific unresponsiveness in the macrophage or T cell compartments, respectively, to the pleiotropic cytokine, IFN gamma. We utilize these mice to identify previously undefined cellular targets of IFN gamma action in the development of a murine antimicrobial response and the mixed lymphocyte reaction. Moreover, we identify the macrophage as a critical responsive cell in manifesting the effects of IFN gamma in regulating CD4+ T helper subset development. These studies thus represent a novel approach to studying the cell-specific actions of an endogenously produced pleiotropic cytokine in vivo.

Ligand-induced autoregulation of IFN-gamma receptor beta chain expression in T helper cell subsets.

Interferon gamma (IFN-gamma) responsiveness in certain cells depends on the state of cellular differentiation or activation. Here an in vitro developmental system was used to show that IFN-gamma produced during generation of the CD4+ T helper cell type 1 (TH1) subset extinguishes expression of the IFN-gamma receptor beta subunit, resulting in TH1 cells that are unresponsive to IFN-gamma. This beta chain loss also occurred in IFN-gamma-treated TH2 cells and thus represents a specific response of CD4+ T cells to IFN-gamma rather than a TH1-specific differentiation event. These results define a mechanism of cellular desensitization where a cytokine down-regulates expression of a receptor subunit required primarily for signaling and not ligand binding.

Developmental commitment to the Th2 lineage by extinction of IL-12 signaling.

Developmental-commitment to Th1 or Th2 responses critically influences host susceptibility to particular pathogens. We describe a novel mechanism governing stable commitment to Th2 differentiation. Naive T cells develop strongly polarized Th1 and Th2 profiles by 7 days after activation. However, commitment of these developing cells differs substantially. Although IL-4 reverses early Th1 differentiation, IL-12 cannot reverse early Th2 differentiation. Th1 reversibility results from maintenance of IL-4 signal transduction, whereas Th2 commitment results from rapid loss of IL-12 signaling. The IL-12 signaling defect in Th2 cells results in failure to phosphorylate Jak2, Stat3, and Stat4. Since Th2 cells express the mRNA for the cloned murine IL-12 receptor beta subunit, the signaling defect may involve expression or function of unidentified receptor components. The rapid extinction of IL-12 signaling in Th2 cells provides a demonstration of a mechanism for the stable commitment to a T helper phenotype.

Enhanced in vivo growth and resistance to rejection of tumor cells expressing dominant negative IFN gamma receptors.

Using a neutralizing monoclonal antibody specific for murine IFN gamma we show that endogenously produced IFN gamma plays an obligate role in mediating LPS-induced rejection of the Meth A fibrosarcoma tumor in syngeneic BALB/c mice. To examine the cellular targets of IFN gamma action, we generated IFN gamma-insensitive tumor cells by stably overexpressing in Meth A a truncated dominant negative form of the murine IFN gamma receptor alpha chain. When implanted in BALB/c mice, IFN gamma-insensitive Meth A cells displayed enhanced tumorigenicity compared with control Meth A cells and were not rejected when tumor-bearing mice were treated with concentrations of LPS that eliminated control tumors. In Meth A immune mice, IFN gamma-insensitive Meth A did not establish tumors while IFN gamma-insensitive tumors grew in a progressive manner. In addition, the IFN gamma-insensitive tumor cells were unable to elicit strong protective immunity to subsequent wild-type tumor challenge. These results show that IFN gamma has direct effects on tumor cell immunogenicity and thus plays an important role in promoting tumor cell recognition and elimination.

Inhibition of cellular responsiveness to interferon-gamma (IFN gamma) induced by overexpression of inactive forms of the IFN gamma receptor.

Herein, we report that overexpression of either human or murine interferon-gamma (IFN-gamma) receptors lacking their entire intracellular domains in cells bearing functionally active IFN gamma receptors ablates responsiveness to homologous ligand in a dominant negative manner. Unresponsiveness could also be induced in murine cells overexpressing murine IFN gamma receptor mutants that either lack 39 COOH-terminal amino acids or contain an alanine substitution for a functionally critical tyrosine. Overexpression of the full-length receptor did not alter cellular responsiveness to IFN gamma. The inhibitory activities of the receptor mutants were dose-dependent, generalizable to a variety of cellular responses, and specific. Cells expressing 100:1 ratios of mutant to wild-type receptor were unresponsive to IFN gamma even at doses 30,000 times greater than that required to induce a maximal response in wild-type cells. These results provide an example of a dominant negative mutation that effects the complete inactivation of a transmembrane receptor lacking a kinase domain and suggest a more general utility for dominant negative mutations in the study of cytokine receptor function.