Characterization of KIR intermediate promoters reveals four promoter types associated with distinct expression patterns of KIR subtypes.

The human killer cell immunoglobulin-like receptor (KIR) genes contain multiple promoters that control the process of gene activation and variegated expression of KIR on natural killer (NK) and T cells. Specific subfamilies of KIR genes have differences in the timing and tissue specificity of expression: however, previous studies of the proximal KIR promoters have not shown significant differences in activity between differentially expressed KIR gene subsets. The recent identification of an intermediate KIR promoter (ProI) associated with KIR2DL1 expression suggested a central role for this element in KIR expression. The current study identifies ProI elements in all of the KIR genes, revealing four classes of ProI that correspond with four distinct expression phenotypes of KIR subgroups: KIR2DL2/S2/L3 that are expressed early in reconstituting NK after transplant; KIR2DL4 that is expressed by CD56-bright NK in a non-variegated manner; KIR3DL3 that is not expressed by circulating NK cells; and the remaining KIR that are expressed by subsets of CD56-dim NK. The four classes of ProI are structurally diverse and display distinct functional properties. Altogether, these results indicate that KIR ProI elements contribute to the tissue/cell-type specificity of KIR transcription and cooperate with the probabilistic proximal promoter to control KIR expression.

The impact of human immune deficiency virus and hepatitis C coinfection on white matter microstructural integrity.

The purpose of the present study is to examine the integrity of white matter microstructure among individuals coinfected with HIV and HCV using diffusion tensor imaging (DTI). Twenty-five HIV+ patients, 21 HIV+/HCV+ patients, and 25 HIV- controls were included in this study. All HIV+ individuals were stable on combination antiretroviral therapy (cART; ≥3 months). All participants completed MRI and neuropsychological measures. Clinical variables including liver function, HIV-viral load, and CD4 count were collected from the patient groups. DTI metrics including mean diffusivity (MD), axial diffusivity (AD), radial diffusivity (RD), and fractional anisotropy (FA) from five subregions of the corpus callosum were compared across groups. The HIV+/HCV+ group and HIV+ group were similar in terms of HIV clinical variables. None of the participants met criteria for cirrhosis or fibrosis. Within the anterior corpus callosum, significant differences were observed between both HIV+ groups compared to HIV- controls on DTI measures. HIV+ and HIV+/HCV+ groups had significantly lower FA values and higher MD and RD values compared to HIV- controls; however, no differences were present between the HIV+ and HIV+/HCV+ groups. Duration of HIV infection was significantly related to DTI metrics in total corpus callosum FA only, but not other markers of HIV disease burden or neurocognitive function. Both HIV+ and HIV+/HCV+ individuals had significant alterations in white matter integrity within the corpus callosum; however, there was no evidence for an additive effect of HCV coinfection. The association between DTI metrics and duration of HIV infection suggests that HIV may continue to negatively impact white matter integrity even in well-controlled disease.

Characterization of a weakly expressed KIR2DL1 variant reveals a novel upstream promoter that controls KIR expression.

Members of the human KIR (killer cell immunoglobulin-like receptor) class I major histocompatibility complex receptor gene family contain multiple promoters that determine the variegated expression of KIR on natural killer cells. In order to identify novel genetic alterations associated with decreased KIR expression, a group of donors was characterized for KIR gene content, transcripts and protein expression. An individual with a single copy of the KIR2DL1 gene but a very low level of gene expression was identified. The low expression phenotype was associated with a single-nucleotide polymorphism (SNP) that created a binding site for the inhibitory ZEB1 (Zinc finger E-box-binding homeobox 1) transcription factor adjacent to a c-Myc binding site previously implicated in distal promoter activity. Individuals possessing this SNP had a substantial decrease in distal KIR2DL1 transcripts initiating from a novel intermediate promoter located 230 bp upstream of the proximal promoter start site. Surprisingly, there was no decrease in transcription from the KIR2DL1 proximal promoter. Reduced intermediate promoter activity revealed the existence of alternatively spliced KIR2DL1 transcripts containing premature termination codons that initiated from the proximal KIR2DL1 promoter. Altogether, these results indicate that distal transcripts are necessary for KIR2DL1 protein expression and are required for proper processing of sense transcripts from the bidirectional proximal promoter.

Identification of a KIR antisense lncRNA expressed by progenitor cells.

Human NK cells express cell surface class I MHC receptors (killer cell immunoglobulin-like receptor, KIR) in a probabilistic manner. Previous studies have shown that a distal promoter acts in conjunction with a proximal bidirectional promoter to control the selective activation of KIR genes. We report here the presence of an intron 2 promoter in several KIR genes that produce a spliced antisense transcript. This long noncoding RNA (lncRNA) transcript contains antisense sequence complementary to KIR-coding exons 1 and 2 as well as the proximal promoter region of the KIR genes. The antisense promoter contains myeloid zinc finger 1 (MZF-1)-binding sites, a transcription factor found in hematopoietic progenitors and myeloid precursors. The KIR antisense lncRNA was detected only in progenitor cells or pluripotent cell lines, suggesting a function that is specific for stem cells. Overexpression of MZF-1 in developing NK cells led to decreased KIR expression, consistent with a role for the KIR antisense lncRNA in silencing KIR gene expression early in development.

Glial cells of the oligodendrocyte lineage express both kainate- and AMPA-preferring subtypes of glutamate receptor.

mRNAs for AMPA- and kainate-preferring glutamate receptor subunits are expressed abundantly in the CNS, yet functional studies of neurons and glia from brain suggest selective expression of AMPA receptors. We now show that glial cells of the O-2A lineage express rapidly desensitizing responses to kainate, mRNAs for GluR6, GluR7, KA-1, and KA-2, rapidly desensitizing responses to AMPA, and mRNAs for GluR-B, -C, and -D. Analysis of glutamate receptor currents in single cells reveals two receptor populations with high and low affinity for kainate and different sensitivity for potentiation by concanavalin A and for block of desensitization by cyclothiazide. Our experiments describe the characterization of native kainate-preferring receptors in glia and reveal coexpression in single cells of functional AMPA- and kainate-preferring receptors.

Immunity to tumor antigens: potential implications in human neuroblastoma.

Immune reactions to tumor-specific and tumor-associated antigens have been demonstrated in animals with neoplasms with in vitro and in vovo techniques. Some of the antigens detected in vitro induce transplantation resistance in vivo, while others do not. Human neuroblastoma cells cultivated in vitro have been shown to possess common antigens to which lymphocytes from neuroblastoma patients react. Whether it is possible to augment the immune reactivity of patients with neuroblastoma to these common antigens and, if so, whether this heightened immune reactivity would have clinically beneficial effects are as yet unknown. These reactions are complex, involving both cellular and humoral mechanisms. The fact that one type of immune response can be detected to one type of antigen present in a tumor in vitro does not necessarily mean that the immune response is effective in vivo. Responses to other tumor antigens may be deficient, or the immune response may be depressed. This may be due to active suppression of and/or selective deficiencies in critical cell populations required for an augmented immune response; this possibility may be evaluated with techniques allowing for in vitro sensitization to tumor antigens.

Synergy in proliferative and cytotoxic responses of WF rats to syngeneic Gross virus-induced lymphoma cells in vitro.

A synergistic interaction between rat lymphoid cell subsets was found for the in vitro immune response to the syngeneic Gross virus-induced lymphoma (C58NT)D. Mixtures of thymocytes and lymph node cells from inbred WF rats primed in vivo to the lymphoma demonstrated significantly greater proliferative and cytotoxic reactivities in vitro than would be expected from the sum of the reactivities of the two cell types tested separately. A soluble extract of nonimmune syngeneic thymocytes, shown in previous studies to amplify the in vitro responses to alloantigen, was also demonstrated to increase significantly proliferative and cytotoxic responses in vitro to syngeneic lymphoma cells. Preliminary evidence indicated that this soluble thymus amplifier factor differed from previously described thymus factors.

Inhibition of Fas (CD95) expression and Fas-mediated apoptosis by oncogenic Ras.

The ras oncogene plays an important role in the multistep progression to cancer by activation of signal transduction pathways that contribute to aberrant growth regulation. Although many of these effects are cell autonomous, the ras oncogene also regulates the expression of genes that alter host/tumor interactions. We now extend the mechanisms through which ras promotes tumor survival by demonstrating that oncogenic Ras inhibits expression of the fas gene and renders Ras-transformed cells resistant to Fas-induced apoptosis. A panel of Ras-transformed clones exhibited a marked inhibition in fas mRNA and Fas cell surface expression as compared with untransformed parental cell lines. Fas expression was induced by culture in the presence of IFN-gamma + tumor necrosis factor alpha; however, the maximal level attained in Ras transformants was approximately 10-fold below the level of untransformed cells. Whereas untransformed cells were sensitive to apoptotic death induced by cross-linking surface Fas (especially after cytokine treatment), Ras-transformed cells were very resistant to Fas-induced death even under the most stringent assay conditions. To demonstrate that this resistance was mediated by oncogenic Ras and not secondary genetic events, pools of Ras-transformed cells were generated using a highly efficient retroviral transduction technique. Transformed pools were assayed 6 days after infection and demonstrated a marked decrease in fas gene expression and Fas-mediated apoptosis. Oncogenic Ras did not promote general resistance to apoptosis, because ectopic expression of a fas cDNA in Ras-transformed cells restored sensitivity to Fas-induced apoptosis. These data indicate that oncogenic Ras inhibits basal levels of expression of the fas gene, and although cytokine signal transduction pathways are functional in these cells, the level of surface Fas expression remains below the threshold required for induction of apoptosis. These data identify a mechanism by which Ras-transformed cells may escape from host-mediated immune destruction.

Long-term survival in small-cell carcinoma of the lung: a population experience.

Small-cell lung carcinoma (SCLC) is a rapidly progressive and fatal disease. Historically, surgical resection or radiotherapy of the primary tumor has done little to prolong survival, although the use of combination chemotherapy is more effective. Reported here is the survival experience of 1,538 incident cases of SCLC identified through the Surveillance, Epidemiology and End Results Program in western Washington State from 1974 to 1982. The survival experience of this population series is similar to that reported from specialized referral centers. For 71 of 78 persons surviving at least 24 months, the original diagnostic slides were independently reviewed, 47 cases being confirmed as SCLC. No differences were found in actuarial survival estimates between those confirmed and those not confirmed as SCLC. Multivariate survival analysis was conducted to estimate the effects on survival of stage, therapy, age, sex, primary site, and histologic type. All factors except primary site and histologic type significantly influence initial survival rates. However, the only factor related to post--two-year (ie, long-term) survival, once stage is accounted for, is whether surgery was received as a first course of therapy. Those not receiving surgery were at four times the risk of death as those who did. These results indicate that long-term survival can be achieved in patients with SCLC treated in the community, and that the chance of surviving an additional two years for such patients is approximately 40%.

Persistent pollutants urban rivers sediment survey: implications for pollution control.

The impacts of diffuse urban sources of pollution on watercourses are quantified. A survey of nine urban streams in Scotland for persistent pollutants in stream sediments is described, together with sediments from SUDS ponds. Determinands reported are: PAHs, total hydrocarbons, and toxic metals (As, Zn, Ni, Pb, Cu, Cr, Cd). Results highlight hydrocarbons as a major urban pollutant, and show significant sediment contamination by toxic metals. The metals that occurred in the highest concentrations varied across the nine streams, but Pb, Cr, Ni, Zn and Cu most frequently present exceeded sediment quality standards. The pattern of contamination by PAHs suggested that pyrolytic sources were more ubiquitous and present in greater quantities than oil spill sources in these urban catchments. Exceptions were the sites below industrial estates. The findings indicate that four levels of activity will be needed to control urban diffuse sources of pollution: reductions in quantities of toxic pollutants used by manufacturers in the motor and construction industries; housekeeping measures to minimise storage and handling risks for oil and chemicals; public engagement to minimise polluting activities such as dumping oil and chemicals, and private car use; use of SUDS technology, including retro-fits in the worst affected urban areas.

A descriptive study of 42 cases of Branhamella catarrhalis pneumonia.

PURPOSE: We studied the clinical and laboratory findings of patients with pneumonia due to Moraxella (Branhamella) catarrhalis to better characterize the types of patients who develop this pneumonia, the clinical features of the illness, and the type of and response to drug therapy, as well as the immediate and long-term survival of these patients. PATIENTS AND METHODS: Patients with sputum samples that met cellular criteria as quality samples and that grew B. catarrhalis as the sole pathogen were identified retrospectively from microbiology records at a regional referral hospital for cardiac and pulmonary diseases. Records of these patients were reviewed to identify patients with radiographic findings of pneumonia. Clinical and laboratory characteristics of these patients were then studied in detail. RESULTS: Forty-two patients who met the criteria for B. catarrhalis pneumonia were identified. Most patients were elderly (over 65 years; 55 percent), malnourished (69 percent), and had severe chronic obstructive pulmonary disease or another serious underlying disease (98 percent). The seasonal incidence of this pneumonia was October through April (88 percent), with the annual number of cases having increased since 1982. The clinical presentation was typically mild. Interstitial or mixed interstitial-alveolar infiltrates superimposed on pre-existing chronic lung disease was the most common radiographic finding. Approximately 90 percent of sputa were acceptable for Gram stain and contained 10 to more than 50 gram-negative cocci/1,000 x field. All cultures produced a heavy growth of B. catarrhalis, with 67 percent of strains positive for beta-lactamase. No patient had identified bacteremia (zero of 25 tested). Therapy with numerous agents including cefotaxime, amoxicillin/clavulanic acid, and trimethoprim/sulfamethoxazole resulted in a good clinical and bacteriologic response. However, 45 percent of patients died of their underlying diseases on this admission or within three months. CONCLUSION: These findings provide a good profile of B. catarrhalis pneumonia. Despite the mild character of the illness, the pneumonia occurs in patients with end-stage pulmonary or malignant disease and almost 50 percent of patients die of their underlying diseases within three months.

Imaging of melanoma with L-131-labeled monoclonal antibodies.

Mouse monoclonal antibodies and Fab fragments specific for p97, a melanoma-associated antigen, were used to image metastatic human melanoma. Preclinical studies in athymic mice showed antigen-specific uptake in melanoma xenografts, and toxicity tests in rabbits gave no evidence for tissue damage after injection of up to 100 times the amount of antibody used in humans. Six patients received 1 mg labeled antibody, and one patient received 1 mg of labeled Fab. No. toxic side effects were observed. All of the six patients had positive scans, visualizing 22 of 25 (88%) of lesions larger than 1.5 cm. In tumors from two patients, greater uptake of p97-specific, versus control IgG and Fab, respectively, was documented by biopsy. Antibodies to mouse immunoglobulin appeared in three patients receiving 1 mg or more of radiolabeled mouse antibody.

Obstruction of the left ventricular outflow tract by the mitral valve due to a muscle band.

This paper details a rare cause of subaortic obstruction--a muscle band tethering the anterior leaflet of the mitral valve to the ventricular septum. Excision of this band released the leaflet and cured the obstruction. The patient also had a discrete subaortic membranous obstruction, the membrane being excised.

Staged bilateral thoracotomies for multiple pulmonary arteriovenous malformations complicating hereditary hemorrhagic telangiectasia.

A patient with hereditary hemorrhagic telangiectasia (Osler-Weber-Rendu syndrome) with multiple bilateral pulmonary arteriovenous malformations was hypoxic, cyanotic, polycythemic, and severely limited by dyspnea on exertion. Following staged bilateral thoracotomies, with removal of 23 major fistulas (12 from the right lung and 11 from the left), marked improvement in symptoms, blood gases, and objective measurements of exercise tolerance occurred.

A case of recurrent typhoid fever in the United States: importance of the grandmother connection and the use of large restriction fragment pattern analysis of genomic DNA for strain comparison.

An 8-year old girl was infected for a second time with Salmonella typhi by contact with her grandmother, a known typhoid carrier. The S. typhi from both patient and grandmother had closely related genomic pulsed field gel electrophoresis patterns that differed from epidemiologically unrelated strains. The girl responded well to a 14-day course of oral trimethoprimsulfamethoxazole. The grandmother was treated successfully with a 28-day regimen of oral ciprofloxacin. Typhoid fever remains an endemic disease in the United States, largely because of recognized chronic stool carriers. Most of these carriers had typhoid in the preantibiotic era and remain potential sources of disease when they provide meals for others, not uncommonly grandchildren. The importance of this "grandmother" connection to endemic typhoid fever is reviewed, as is the potential use of pulsed field gel electrophoresis pattern analysis for comparison of strains of S. typhi.

Initial clinical assessment of the comatose patient: cerebral malaria vs. meningitis.

One hundred twenty-one Liberian children were admitted in coma to the ELWA Hospital, Monrovia, Liberia. Admitting diagnoses, before lumbar puncture, were compared with discharge diagnoses. Ninety-four children were discharged with a final diagnosis of cerebral malaria and 27 with a diagnosis of meningitis. The admitting diagnosis was correct in 76.6% (72 of 94) of patients with cerebral malaria and 59.3% (16 of 27) of patients with meningitis. The cerebrospinal fluid leukocyte count was the single most significant factor in determining the correct diagnosis. Without the cerebrospinal fluid analysis, the discriminant accuracy (77%), i.e. definitive separation of the two illnesses, was comparable to the physician's admission diagnosis (73%). Other data contributing to the differential diagnosis of cerebral malaria and meningitis included the number of days of fever before admission, the presence or absence of nuchal rigidity, fontanelle fullness and peripheral blood malaria smear. Mortality rates for cerebral malaria and meningitis were 14.9 and 29.6%, respectively. These data suggest that physicians cannot reliably discriminate between cerebral malaria and meningitis without cerebrospinal fluid analysis.

Photothrombotic lesions of the frontal cortex impair the performance of the delayed non-matching to position task by rats.

The effects of photochemically induced lesions of the frontal cortex on the short-term memory capacity of the rat have been investigated using the delayed non-matching to position task. Pretrained animals received lesions and were tested 4 days after surgery and twice per week for 3 weeks. The lesions produced a profound impairment of performance of this task which was still evident 3 weeks after surgery. Spontaneous locomotor activity was recorded 7 days after surgery and no difference was found between the control and lesion group. These effects indicated a generalized disruption of performance of this task in the absence of motor dysfunction. These results suggest that photothrombotic lesions of the frontal cortex can produce reliable, long-term behavioural deficits.

The effects of adenosine triphosphate and related purines on arterial resistance vessels in vitro and in vivo.

The distribution of P2-purinoceptors in pre-capillary resistance vessels was studied in vitro, using Krebs perfused rabbit ears and in vivo, in autoperfused hindquarters, intestinal and renal vasculatures of pentobarbitone anaesthetised cats. ATP (10(-10)-10(-6) mol i.a.) caused dose-dependent vasodilatation which, in the rabbit ear, was antagonised by reactive blue 2 (10(-5)-10(-4) M). At the highest concentration of reactive blue 2, ATP responses were reversed and a dose-dependent vasoconstriction was seen. Reactive blue 2, also reduced the vasodilator responses to carbachol and to a lesser extent papaverine which suggests that the antagonist has limited selectivity. The rank order of potency of ATP analogues as vasodilators, 2-methylthio ATP greater than ADP greater than ATP greater than alpha,beta-methylene and beta,gamma-methylene ATP, suggests P2y purinoceptors are involved. The selective P2x-purinoceptor agonist, alpha,beta-methylene ATP, caused pronounced vasoconstriction in the rabbit ear and cat intestinal vasculature which was not antagonised by phenoxybenzamine. In contrast, alpha,beta-methylene ATP had little effect in the autoperfused hindquarters and renal vasculatures suggesting a very heterogeneous distribution of P2x-purinoceptors in the cat. The results are consistent with the proposal that two distinct types of P2-purinoceptors are present on blood vessels.

Selection of skin test antigens to evaluate PPD anergy.

BACKGROUND: A purified protein derivative (PPD) tuberculin skin test may be nonreactive because of cutaneous anergy, technical problems with the test, or absence of tuberculosis infection. This study investigated the sensitivity and specificity of five test agents in measuring cutaneous anergy when the PPD test is nonreactive. Agents evaluated include antigens for Candida, mumps, histoplasmin, tetanus, and Trichophyton. METHODS: Delayed-type hypersensitivity skin test records were analyzed in 1113 patients admitted to the University of Texas Health Center at Tyler from December 1988 through June 1993. These patients were admitted with initial diagnoses of diseases other than active tuberculosis or human immunodeficiency virus infection. RESULTS: Patients with a negative PPD test reacted most often to the control skin test Candida (63.5%), followed by mumps (52.2%), histoplasmosis (37.2%), tetanus (35.7%), and Trichophyton (6.1%). Analysis of these data indicates that the use of more than three of the four most commonly reactive control tests (Candida, mumps, and histoplasmin or tetanus) yielded minimal additional precision in the determination of skin test anergy compared with using all five control skin tests. This finding remained constant whether the PPD was considered negative at < 5 mm, < 10 mm, or < 15 mm of induration. CONCLUSIONS: In controlling for false-negative PPD tests, the use of three skin test antigens, Candida, mumps, and tetanus, should provide reliable control for delayed-type hypersensitivity anergy.