Dissecting natural killer cell activation pathways through analysis of genetic mutations in human and mouse

Summary:  Natural killer (NK) cell cytotoxicity is mediated by multiple germ line-encoded activating receptors that recognize specific ligands expressed by tumor cells and virally infected cells. These activating receptors are opposed by NK inhibitory receptors, which recognize major histocompatibility complex class I molecules on potential targets, raising the threshold for NK cell activation. Once an abnormal cell has been detected, NK cells are the sentinel source of cytolytic mediators, such as granzymes and perforins, as well as interferon-γ, which can polarize the immune response to a T-helper 1 cell type. Activation signals are transmitted by adhesion-dependent pathways, immunoreceptor tyrosine-based activation motif (ITAM)-dependent pathways, DAP10 ITAM-independent pathways, and by signaling through immunoreceptor tyrosine-based switch motifs. These pathways activate downstream signaling partners to trigger NK cell cytotoxicity. Some of these downstream molecules are unique to the various pathways, and some of these molecules are shared. Because of the complexity of signals involved in NK cell–target cell interaction, the generation of mice with targeted mutations in signaling molecules involved in adhesion, activation, or inhibition is essential for a precise dissection of the mechanisms regulating NK cell effector functions. Here we review recent advances in the genetic analysis of the signaling pathways that mediate NK cell killing.     

Humoral immune response to thymidylate synthase in colon cancer patients after 5-FU chemotherapy

Thymidylate synthase (TYMS), the critical enzyme for DNA synthesis and a target for chemotherapy, was recently characterized as an oncogene and a potential target for specific immunotherapy. Here we report TYMS-specific antibody response in a fraction of colon cancer patients. Humoral immune response to TYMS is induced by chemotherapy using TYMS inhibitors, such as 5-fluorouracil (5-FU), and may be associated with tumor burden. Therefore, TYMS may serve as a useful serological biomarker for monitoring the course of disease and treatment in cancer patients.     

Bioactive glasses for in situ tissue regeneration

Historically the function of biomaterials has been to replace diseased or damaged tissues. Recent findings show that controlled release of the ionic dissolution products of bioactive glasses results in regeneration of tissues. The mechanism for in situ tissue regeneration involves upregulation of seven families of genes that control the osteoblast cell cycle, mitosis and differentiation. In the presence of critical concentrations of Si and Ca ions, within 48 h osteoblasts that are capable of differentiating into a mature osteocyte phenotype begin to proliferate and regenerate new bone. Osteoblasts that are not in the correct phase of the cell cycle and unable to proceed towards differentiation are switched into apoptosis by the ionic dissolution products. A controlled release of soluble Ca and Si from bioactive glass--resorbable polymer composites leads to vascularised soft tissue regeneration. Gene activation by controlled ion release provides the conceptual basis for molecular design of a third generation of biomaterials optimised for in situ tissue regeneration.     

Evaluation of R-Mix shell vials for the diagnosis of viral respiratory tract infections.

Respiratory viruses cause significant morbidity and mortality. The management of these infections can be improved by a rapid diagnosis and administration of available virus-specific therapy. The goal of this study was to compare R-Mix, an engineered tissue monolayer for rapid shell vial (SV) diagnosis of viral respiratory infections, with conventional tissue culture (TC) and conventional respiratory SV (primary rhesus monkey kidney (RhMK) and Hep2 monolayers). The primary outcome measure was sensitivity for detection of influenza A and B, respiratory syncytial virus, parainfluenza 1-3, and adenovirus. The study was performed in two phases: (1) the three methods were compared using 250 nasal washes from children with lower respiratory tract infections; (2) a modified R-Mix SV harvesting schedule (SV were harvested at 24 and 120 h) was compared with TC and conventional RhMK/Hep2 SV using 311 respiratory specimens. A total of 110 viruses were identified in the first and 55 in the second phase. Diagnostic accuracies of R-Mix harvested at 24, 48, and 120 h were 98%, whereas for TC varied between 99 and 100%, and for RhMK/Hep2 SV between 98 and 99%. Sensitivities of R-Mix harvested at 24, 48, and 120 h were 26, 75, and 47%, respectively, whereas for TC varied between 60 and 94%, and for RhMK/Hep2 SV between 62 and 85%. R-Mix harvested at 48 h represent a valuable substitute for RhMK/Hep2 SV because they have comparable sensitivities and diagnostic accuracies, but R-Mix offers several technical advantages. In contrast, R-Mix harvested at 24h did not seem a very useful diagnostic tool. The utility of R-Mix harvested at 120 h, which accelerated the diagnosis of 16% of positive specimens in study phase 2, needs further investigation.     

Clonal chromosome abnormalities in human breast carcinomas I. Twenty-eight cases with primary disease

Cytogenetic analysis was performed on a selected series of short-term cultures of primary breast carcinomas from 28 patients. All patients had histopathologically confirmed malignancies, with the majority (25/28 cases) demonstrating infiltrating ductal carcinoma. All 28 cases evidenced clonal chromosome abnormalities, with 10/28 displaying only numeric aberrations, whereas 18/28 displayed clonal structural alterations. In near-diploid tumors the most common numeric changes were — 17 and — 19. However, trisomy 7 was the only numeric change in two near-diploid tumors. Structural chromosome alterations were primarily isochromosomes, apparent terminal deletions, and unbalanced non-reciprocal translocations. Chromosomes 1 (10/18–56%) and 6 (8/18–44%) were most frequently altered in this series. Breakpoints of clonal structural abnormalities were shown to cluster to several chromosome segments, including 1p22-q11, 3p11, 6p11–13, 7p11-q11, 8p11-q11, and 19q13. Analysis of the gain or loss of specific chromosome segments revealed that the most consistent tendency was over-representation of 1q, 3q, and 6p. © 1993 Wiley-Liss, Inc.     

Prevention of polyglutamine oligomerization and neurodegeneration by the peptide inhibitor QBP1 in Drosophila

Polyglutamine (polyQ) diseases are a growing class of inherited neurodegenerative diseases including Huntington's disease, which are caused by abnormal expansions of the polyQ stretch in each unrelated disease protein. The expanded polyQ stretch is thought to confer toxic properties on the disease proteins through alteration of their conformation leading to pathogenic protein-protein interactions including oligomerization and/or aggregation. Hypothesizing that molecules with selective binding affinity to the expanded polyQ stretch may interfere with the pathogenic properties, we previously identified Polyglutamine Binding Peptide 1 (QBP1) from combinatorial peptide phage display libraries. We show here that a tandem repeat of the inhibitor peptide QBP1, (QBP1)(2), significantly suppresses polyQ aggregation and polyQ-induced neurodegeneration in the compound eye of Drosophila polyQ disease models, which express the expanded polyQ protein under the eye specific promoter. Most importantly, (QBP1)(2) expression dramatically rescues premature death of flies expressing the expanded polyQ protein in the nervous system, resulting in the dramatic increase of the median life span from 5.5 to 52 days. These results suggest that QBP1 can prevent polyQ-induced neurodegeneration in vivo. We propose that QBP1 prevents polyQ oligomerization and/or aggregation either by altering the toxic conformation of the expanded polyQ stretch, or by simply competing with the expanded polyQ stretches for binding to other expanded polyQ proteins. The peptide inhibitor QBP1 is a promising candidate with great potential as a therapeutic molecule against the currently untreatable polyQ diseases.     

A locus for posterior polymorphous corneal dystrophy (PPCD3) maps to chromosome 10

Posterior polymorphous corneal dystrophy (PPCD) is an autosomal dominant disorder characterized by corneal endothelial abnormalities, which can lead to blindness due to loss of corneal transparency and sometimes glaucoma. We mapped a new locus responsible for PPCD in a family in which we excluded the previously reported PPCD locus on 20q11, and the region containing COL8A2 on chromosome 1. Results of a 317-marker genome scan provided significant evidence of linkage of PPCD to markers on chromosome 10, with single-point LOD scores of 2.63, 1.63, and 3.19 for markers D10S208 (at (circumflex)theta = 0.03), D10S1780 (at (circumflex)theta = 0.00), and D10S578 (at (circumflex)theta = 0.06). A maximum multi-point LOD score of 4.35 was found at marker D10S1780. Affected family members shared a haplotype in an 8.55 cM critical interval that was bounded by markers D10S213 and D10S578. Our finding of another PPCD locus, PPCD3, on chromosome 10 indicates that PPCD is genetically heterogeneous. Guttae, a common corneal finding sometimes observed along with PPCD, were found among both affected and unaffected members of the proband's sib ship, but were absent in the younger generations of the family. Evaluation of phenotypic differences between family members sharing the same affected haplotype raises questions about whether differences in disease severity, including differences in response to surgical interventions, could be due to genetic background or other factors independent of the PPCD3 locus.     

Identifying undiagnosed diabetes: cross-sectional survey of 3.6 million patients' electronic records.

BACKGROUND: Around 1% of the UK population has diabetes that is either undiagnosed or unrecorded on practice disease registers. AIM: To estimate the number of people in UK primary care databases with biochemical evidence of undiagnosed diabetes. To develop simple practice-based search techniques to support early recognition of diabetes. DESIGN OF STUDY: Cross-sectional survey of 3 630 296 electronic records. SETTING: Four hundred and eighty UK practices contributing to the QRESEARCH database. METHOD: Electronic searches to identify people with no diabetes diagnosis in one of two categories (A and B), using the most recently recorded blood glucose measurement: random blood glucose level >or=11.1 mmol/l or fasting blood glucose level >or=7.0 mmol/l (A); either a random or a fasting blood glucose level >or=7.0 mmol/l (B). An additional outcome measure was the proportion of the population with at least one blood glucose measurement in the record. RESULTS: The number (percentage) identified in category A was 3758 (0.10% of the total population); the number in category B was 32 785 (0.90%). Projected to a practice of 7000 patients, around eight patients have biochemical evidence of undiagnosed diabetes, and 68 have results suggesting the need for further follow-up. One-third of people aged over 40 years without diabetes have a blood glucose measurement in the past 2 years in their record. CONCLUSION: People with possible undiagnosed diabetes are readily identifiable in UK primary care databases through electronic searches using blood glucose data. People with borderline levels, who may benefit from interventions to reduce their risk of progression to diabetes, can also be identified using practice-based software.