ABSTRACT
BACKGROUND: Bronchiectasis can result from infectious, genetic, immunological and allergic causes. 60-80% of cases are idiopathic, but a well-recognised genetic cause is the motile ciliopathy, primary ciliary dyskinesia (PCD). Diagnosis of PCD has management implications including addressing comorbidities, implementing genetic and fertility counselling and future access to PCD-specific treatments. Diagnostic testing can be complex; however, PCD genetic testing is moving rapidly from research into clinical diagnostics and would confirm the cause of bronchiectasis. METHODS: This observational study used genetic data from severe bronchiectasis patients recruited to the UK 100,000 Genomes Project and patients referred for gene panel testing within a tertiary respiratory hospital. Patients referred for genetic testing due to clinical suspicion of PCD were excluded from both analyses. Data were accessed from the British Thoracic Society audit, to investigate whether motile ciliopathies are underdiagnosed in people with bronchiectasis in the UK. RESULTS: Pathogenic or likely pathogenic variants were identified in motile ciliopathy genes in 17 (12%) out of 142 individuals by whole-genome sequencing. Similarly, in a single centre with access to pathological diagnostic facilities, 5-10% of patients received a PCD diagnosis by gene panel, often linked to normal/inconclusive nasal nitric oxide and cilia functional test results. In 4898 audited patients with bronchiectasis, <2% were tested for PCD and <1% received genetic testing. CONCLUSIONS: PCD is underdiagnosed as a cause of bronchiectasis. Increased uptake of genetic testing may help to identify bronchiectasis due to motile ciliopathies and ensure appropriate management.
Subject(s)
Bronchiectasis , Ciliary Motility Disorders , Ciliopathies , Kartagener Syndrome , Humans , Mutation , Bronchiectasis/diagnosis , Bronchiectasis/genetics , Cilia , Ciliary Motility Disorders/diagnosis , Ciliary Motility Disorders/genetics , Ciliopathies/complications , Kartagener Syndrome/diagnosis , Kartagener Syndrome/geneticsABSTRACT
Glioma is a rare brain tumor with a poor prognosis. Familial glioma is a subset of glioma with a strong genetic predisposition that accounts for approximately 5% of glioma cases. We performed whole-genome sequencing on an exploratory cohort of 203 individuals from 189 families with a history of familial glioma and an additional validation cohort of 122 individuals from 115 families. We found significant enrichment of rare deleterious variants of seven genes in both cohorts, and the most significantly enriched gene was HERC2 (P = 0.0006). Furthermore, we identified rare noncoding variants in both cohorts that were predicted to affect transcription factor binding sites or cause cryptic splicing. Last, we selected a subset of discovered genes for validation by CRISPR knockdown screening and found that DMBT1, HP1BP3, and ZCH7B3 have profound impacts on proliferation. This study performs comprehensive surveillance of the genomic landscape of familial glioma.
Subject(s)
Brain Neoplasms , Glioma , Humans , Glioma/genetics , Glioma/pathology , Brain Neoplasms/genetics , Brain Neoplasms/pathology , Genomics , Genetic Predisposition to Disease , Whole Genome Sequencing , Calcium-Binding Proteins/genetics , DNA-Binding Proteins/genetics , Tumor Suppressor Proteins/geneticsABSTRACT
The primary antigen-specific antibody response of various strains of mice to TEPC-15/PnC immune complexes has been examined. We found that both BALB/c and C3H mice were good responders to the PnC antigen; however, C3H mice were low responders, whereas BALB/c mice were high responders to the TEPC-15/PnC complexes. Using congenic strains on the C3H and BALB/c background, we have shown that the response to the complexes is not restricted by gene products of the H-2 complex or by the Igh (allotype) locus. However, responsiveness may be controlled by genes linked to the Igh locus, since we have shown that strains that are Ighj, Ighd, and Ighf are low responders, whereas strains that are Igha, Ighb, and Ighe are high responders to the immune complex. Moreover, responsiveness correlates with the expression of the T15 Id as measured using the anti-T15 monoclonal antibody, AB1-2. Thus, strains such as BALB/c, BALB.B, BALB.K, and CB-20, which express high levels of T15 (AB1-2) Id in their PFC response to PnC are relatively high responders to TEPC-15/PnC complexes, whereas C3H, C3H.SW, and C3H-OH, which express low levels of the T15 (AB1-2) Id, are low responders to the complexes. Finally, we found that BALB/c mice are high responders to complexes formed with T15+ antibodies, whereas they are low responders to complexes formed using T15- antibodies. The results suggest that the antigen-specific response to these immune complexes is Id-restricted.
Subject(s)
Antigen-Antibody Complex/immunology , Immunoglobulin Idiotypes/biosynthesis , Animals , Chromosome Mapping , Female , Genetics , H-2 Antigens/genetics , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, Inbred Strains , T-Lymphocytes/immunologyABSTRACT
The effects of immune complexes on the antibody response of BALB/c mice to Streptococcus pneumoniae R36a (Pn) were investigated. The cell wall polysaccharide (PnC) extracted from Pn was used to form complexes with TEPC-15, a myeloma protein that binds to phosphorylcholine determinants on the PnC. Complexes formed at equivalence were cultured with splenic T cells from BALB/c mice for 2 d, and then the cells were added to fresh BALB/c spleen cell cultures to test their effect on the antibody response to Pn, a response dominated by the T15 idiotype family. The results indicate that TEPC-15/PnC complexes induced potent suppressor T cells (Ts) whereas cells cultured with free antigen or free antibody alone had no effect on the plaque-forming cell response to Pn. The suppression was specific since the response to control antigens such as sheep erythrocytes was unaffected. The suppression appears to be idiotype-specific since the Ts had a relatively weak (and in some cases no) effect on the anti-Pn response of BALB/c mice that had been suppressed for T15 idiotopes by neonatal injection of a monoclonal anti-T15 antibody, MaId 5-4. Furthermore, cells cultured with TEPC-15/PnC complexes were shown to express specific receptors for TEPC-15 idiotopes. The results indicate that antigen/antibody complexes may have important immunoregulatory effects because they are potent inducers of idiotype-specific Ts.
Subject(s)
Antigen-Antibody Complex/immunology , Immunoglobulin Idiotypes/immunology , T-Lymphocytes, Regulatory/immunology , Animals , Cells, Cultured , Fluorescent Antibody Technique , Immunoglobulin A/immunology , Mice , Mice, Inbred BALB C , Myeloma Proteins/immunology , Polysaccharides, Bacterial/immunology , Streptococcus pneumoniaeABSTRACT
In the past year, substantial progress has been made in both mapping and fine mapping the genes involved in blood pressure regulation. Genome scans have been carried out in humans and mice and these reveal many new potential chromosomal locations for blood pressure susceptibility loci. The chromosomal regions containing blood pressure genes for many of the inbred hypertensive rat models have been refined using new congenic strains. Further genetic studies support a role for antiotensinogen, aldosterone synthase and a region close to the epithelial sodium channel in blood pressure regulation. Finally, comprehensive single-nucleotide polymorphism analysis of cardiovascular genes has been undertaken using chip technology.
Subject(s)
Hypertension/genetics , Angiotensinogen/genetics , Animals , Cytochrome P-450 CYP11B2/genetics , Epithelial Sodium Channels , Genome, Human , Humans , Mice , Rats , Rats, Inbred SHR , Sodium Channels/geneticsABSTRACT
The Salmonella-microsome assay for mutagenesis was used to determine the effect of ozone on the mutagenesis of selected carcinogens and mutagens in water. Short periods of ozonation were shown to completely inactivate the mutagenicity of several polyaromatic amine mutagens including acriflavine, proflavine, and beta-naphthylamine. Selected polyaromatic hydrocarbons were also sensitive to ozonation. Kinetic studies revealed that the mutagenicity of benzo(a)pyrene, 3-methylcholanthrene, and 7,12-dimethylbenz(a)anthracene was destroyed after short periods of ozonation. To correlate loss of mutagenicity with loss of carcinogenicity, two polyaromatic hydrocarbons were treated with ozone, extracted from water with hexane, and tested for carcinogenicity in mice. When 7,12-dimethyl-benz(a)anthracene and 3-methyl-cholanthrene were treated with ozone, there was a substantial reduction in carcinogenicity compared to control groups treated with oxygen alone. However, a small number of tumors developed in the group of animals receiving a hexane extract of ozonated 7,12-dimethylbenz(a)anthracene. This activity may be due to breakdown products of 7,12-dimethylbenz(a)anthracene that are not mutagenic.
Subject(s)
Amines , Carcinogens, Environmental , Ozone , Polycyclic Compounds , Water Pollutants, Chemical , Water Pollutants , Water Supply , 2-Naphthylamine , 9,10-Dimethyl-1,2-benzanthracene , Acridines , Acriflavine , Animals , Croton Oil , Drug Evaluation, Preclinical , Female , Male , Methylcholanthrene , Mice , Mutagens , Neoplasms, Experimental/chemically induced , ProflavineABSTRACT
The effect of ozonation on the mutagenicity of selected chemicals in water was determined. The use of the Salmonella-microsome assay for mutagensis allowed kinetic studies to be performed on the ozonation of all chemicals tested. The results indicate that the mutagenicity of certain pesticides, including captan and Dexon, was inactivated by short periods of ozonation. The mutagenicity of certain alkylating agents including bis(2-chloroethyl)amine and sodium azide was rapidly inactivated by ozonation while other alkylating agents such as beta-propiolactone, propanesultone, and N-methyl-N'-nitro-N-nitrosoguanidine were unaffected by treatment with ozone. The mutagenicity of aflatoxin B1 was rapidly inactivated by treatment with ozone. Three chemicals were shown to be converted to direct mutagens by ozone treatment. Under certain conditions, dimethylhydrazine could be converted to a mutagen that was stable for 3 weeks. A similar chemical, 2-hydroxyethylhydrazine, was converted to an unstable mutagen that was inactive after 24 hr at room temperature. When benzidine was treated with ozone, there was a transient increase in mutagenicity which was lost after longer treatment with ozone.
Subject(s)
Carcinogens, Environmental , Mutagens , Ozone , Water Pollutants, Chemical , Water Pollutants , Water Supply , Aflatoxins , Alkylating Agents , Benzidines , Drug Evaluation, Preclinical , PesticidesABSTRACT
Autologous immunoglobulin was detected on the cell surface of tumor cells freshly isolated from cancerous kidneys of patients with renal cell carcinoma by flow cytometry after staining with murine anti-human IgG monoclonal antibodies. Cells isolated in parallel from macroscopically normal regions of the tumorous kidneys were not specifically stained with the anti-human IgG reagents. In further studies, tumor cells were stained with an antibody to decay-accelerating factor (DAF), a known inhibitor of complement. Flow cytometry of these cells revealed that nearly all tumor cells expressed DAF, and that the intensity of staining with the anti-DAF monoclonal antibody correlated with the staining of cells with anti-IgG. The results suggest that tumor cells coated with autologous antibody may be resistant to complement-mediated cytotoxicity in vivo through the expression of high levels of DAF.
Subject(s)
Autoantibodies/analysis , Complement Inactivator Proteins/analysis , Kidney Neoplasms/immunology , Membrane Proteins/analysis , Receptors, Antigen, B-Cell/analysis , Antibodies, Monoclonal , CD55 Antigens , Flow Cytometry , Humans , Kidney Neoplasms/pathology , Lymphocytes/immunology , Lymphocytes/pathologyABSTRACT
Two human renal carcinoma cell lines have been established from the same patient. One cell line (CCF-RC1) was obtained from the primary tumor and the second (CCF-RC2) was established from cells of the renal vein effluent of the perfused tumorous kidney. Although they were established from the same patient, the cell lines differed in certain biological properties. They have been passaged up to 50 times in vitro for about two years. Each has an epithelial morphology and exhibits mutilayering. Cell cycle time of CCF-RC1 and CCF-RC2 was 34 and 36 h, respectively. They exhibited anchorage independent growth, and the plating efficiency of CCF-RC2 in soft agar was higher than that of CCF-RC1. Both lines induced tumors in nude mice at the site of s.c. injection closely resembling the original tumor in histological examination. Electron microscopic features of both tumors in nude mice were consistent with epithelial origin. Doubling time of CCF-RC1 and CCF-RC2 in nude mice was 11 and 12 days, respectively. CCF-RC1 and CCF-RC2 have hypotetraploid karyotype and modal numbers of 83 and 73, presenting two and three marker chromosomes, respectively. Immunocytology with commercial monoclonal antibodies against renal carcinoma (URO-3) and cytokeratin (Mac 6) showed positive reactions with both lines, suggesting that these cell lines derived from renal epithelium. A murine monoclonal antibody (2E11) was generated against CCF-RC2 by the hybridoma technique; 2E11 reacted with CCF-RC2, but not with CCF-RC1. These cell lines may provide a useful model for the study of tumor heterogeneity and its relationship to metastasis.
Subject(s)
Carcinoma, Renal Cell/genetics , Kidney Neoplasms/genetics , Tumor Cells, Cultured/pathology , Animals , Antibodies, Monoclonal/immunology , Carcinoma, Renal Cell/pathology , Carcinoma, Renal Cell/ultrastructure , Cell Cycle , Cryopreservation , DNA, Neoplasm/genetics , Flow Cytometry , Humans , Immunohistochemistry , Karyotyping , Kidney Neoplasms/pathology , Kidney Neoplasms/ultrastructure , Male , Mice , Mice, Nude , Middle Aged , PloidiesABSTRACT
Newborn germ-free (GF) and conventional (CV) BALB/c mice were infected with murine leukemia virus-Moloney (MuLV-M) and subsequently monitored for virus expression and leukemia development. GF mice expressed more than 10-fold less virus in peripheral blood compared with CV mice, despite equivalent numbers of infected cells in the spleens, lymph nodes, thymi, and bone marrow of both groups. In addition to lower levels of virus expression, the latency period before the onset of fatal leukemias was greatly extended in GF mice; the first and last fatalities were recorded at 25 and 43 weeks postinfection, respectively, with a mean survival time of approximately 36 weeks. In CV mice, the first and last fatalities occurred at 8 and 17 weeks, respectively, with a mean survival time of approximately 13.5 weeks. Finally, the gross pathology of involved lymphoid organs varied in the two groups. GF mice experienced severe splenomegaly with or without lymphadenopathy but without thymoma; CV mice, in contrast, developed splenomegaly, lymphadenopathy, and severe thymoma. Collectively, these results indicate a marked resistance of GF animals to MuLV-M and suggest that the level of immune system activation may influence the pathogenicity of nontransforming retroviruses.
Subject(s)
Germ-Free Life , Leukemia, Experimental/immunology , Moloney murine leukemia virus/pathogenicity , Tumor Virus Infections , Animals , Leukemia, Experimental/microbiology , Mice , Mice, Inbred BALB C , T-Lymphocytes/immunology , Virus ReplicationABSTRACT
The combination of investigation of rare Mendelian forms of hypertension, candidate gene studies, comparative mapping and genome-wide screening in both animal models and man has led to significant progress in determining new mechanisms of blood pressure control. In this review, the newly discovered blood pressure/cardiovascular genes, WNK kinases and angiotensin converting enzyme 2 and the development of a new anti-hypertensive agent PST2238 are discussed. Major genes causing essential hypertension have yet to be discovered, however, there are now over 20 published genome-wide screens for blood pressure controlling genes. Several regions demonstrate suggestive linkage to the trait and there is some overlap of regions between the different studies. It is hoped that new blood pressure genes will ultimately be discovered using this method. Pharmacogenetic studies in hypertension have only been initiated recently, some are described in this paper. Small studies upon single candidate genes, suggest that the contribution of genetics to the inter-individual variation in blood pressure response to anti-hypertensive therapy, is small, approximately 3-5%. Recently micro-arrays with multiple polymorphisms in multiple genes have been used. After accounting for the additive affects of multiple blood pressure loci, an individual's genetic profile appeared to explain up to 50% of the variation in blood pressure response to therapy. Knowledge of the genetic variants that cause hypertension and influence response to anti-hypertensive therapy will ultimately provide a greater understanding of the molecular mechanisms underlying blood pressure control.
Subject(s)
Antihypertensive Agents/pharmacology , Blood Pressure/genetics , Hypertension/genetics , Androstanols/pharmacology , Androstanols/therapeutic use , Angiotensin-Converting Enzyme 2 , Animals , Antihypertensive Agents/therapeutic use , Calmodulin-Binding Proteins/pharmacology , Calmodulin-Binding Proteins/therapeutic use , Carboxypeptidases/genetics , Genomics/trends , Humans , Hypertension/drug therapy , Intracellular Signaling Peptides and Proteins , Minor Histocompatibility Antigens , Oligonucleotide Array Sequence Analysis , Peptidyl-Dipeptidase A , Pharmacogenetics/trends , Protein Serine-Threonine Kinases/genetics , Renin-Angiotensin System/genetics , WNK Lysine-Deficient Protein Kinase 1ABSTRACT
A new computer program is described which calculates titers and antibody concentrations from ELISA data. Optical densities are measured in 96-well microtiter plates using an automated colorimeter and simultaneously fed into a microcomputer. The data can then be arranged and printed in an 8 X 12 format corresponding to the format of a 96-well microtiter plate. The computer program can also compute the titers of samples if the samples are arranged and titrated in one of the suggested formats. In addition, the titers of unknown samples can be automatically compared with the titer of a standard to obtain concentrations. An ELISA designed to measure the concentration of murine antibodies to the cell wall polysaccharide (PnC) extracted from Streptococcus pneumoniae was performed to document the use of the program.
Subject(s)
Computers , Enzyme-Linked Immunosorbent Assay/instrumentation , Immunoenzyme Techniques/instrumentation , Polysaccharides, Bacterial/analysis , Software , Immunoglobulin M , Microchemistry , Streptococcus pneumoniae/immunologyABSTRACT
In this study, we describe a simple and efficient method for both the monitoring of antigen-specific CD4 and CD8 T cell responses as well as the identification of novel CD4 and CD8 T cell epitopes using a modified ELISpot assay and pools of 20mer peptides. We have demonstrated that pools containing as many as 64 20mer peptides may be used to screen for CD4 and CD8 T cell responses to HPV16 L1, E1, and E7 in mice. Using arrays of pools of overlapping 20mer peptides, we have identified novel CD4 and CD8 epitopes in both HPV16L1 and HPV16E1 which are presented in Balb/c mice. We have further shown that the use of 20mer peptides is equivalent to using minimal 9mer epitopes for the stimulation of CD8 T cell responses in our assay. While our experiments are conducted in mice, the use of peptide pool arrays allows for the identification of epitope-specific responses using far fewer cells than is required for testing a panel of overlapping peptides individually, making this strategy particularly useful in clinical settings where immune cells may be limiting.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Capsid Proteins , Immunologic Techniques , Oncogene Proteins, Viral/immunology , Oncogene Proteins/immunology , Animals , Enzyme-Linked Immunosorbent Assay/methods , Epitope Mapping , Epitopes, T-Lymphocyte/genetics , Epitopes, T-Lymphocyte/immunology , Humans , Interferon-gamma/biosynthesis , Mice , Mice, Inbred BALB C , Oncogene Proteins/genetics , Oncogene Proteins, Viral/genetics , Papillomavirus E7 Proteins , Peptides/immunologyABSTRACT
Essential hypertension is a major risk factor for cardiovascular disease in humans, and originates from both genetic and environmental factors. Data from animal and more recently human studies have indicated the presence of a gene influencing blood pressure on human chromosome 17. This study tested for linkage of markers located on chromosome 17q to essential hypertension in African Caribbean hypertensive families. No support of linkage was found between the markers studied and hypertension, however only genes of a lamda sib value of less than 1.8 could be excluded Journal of Human Hypertension (2000) 14, 385-387
Subject(s)
Black People/genetics , Chromosome Mapping , Chromosomes, Human, Pair 17/genetics , Hypertension/genetics , Africa/ethnology , Caribbean Region , Cohort Studies , Genetic Linkage , Humans , Hypertension/ethnologyABSTRACT
This study evaluated whether hypertensive siblings had excess sharing of RsaI and SstI alleles of the insulin receptor gene compared with a random population. Thirty families consisting of 60 affected individuals with established hypertension were genotyped for the RsaI and SstI restriction fragment length polymorphisms and the resulting genotype data was analysed using the affected pedigree member method of linkage analysis. The hypertensive siblings were found to have increased sharing of INSR alleles; however, this linkage could not be confirmed using a maximum LOD score method. Thus, the results from this study do not support a role for the INSR gene in the genesis of essential hypertension in the population studied.
Subject(s)
Hypertension/genetics , Receptor, Insulin/genetics , Aged , Female , Genetic Markers , Humans , Hypertension/etiology , Male , Middle Aged , Pedigree , Polymorphism, Genetic/genetics , Receptor, Insulin/analysis , Sensitivity and SpecificityABSTRACT
Atrial natriuretic peptide (ANP) which alters sodium balance, blood volume and vascular tone represents an important candidate for investigating the genetic basis of essential hypertension (EH). Accordingly, we have studied Bgl1 and Xho1 restriction fragment length polymorphisms (RFLPs) of the ANP gene in 147 hypertensive, 141 normotensive and 67 population-based control subjects from a homogenous population of West African origin from St Vincent and the Grenadines. We found no association of either Bgl1 and Xho1 RFLPs with EH. This study suggests that the ANP locus may not exert a major gene effect on EH amongst the black people of St Vincent and the Grenadines.
Subject(s)
Atrial Natriuretic Factor/genetics , Black People , Hypertension/genetics , Adult , Aged , Female , Humans , Hypertension/ethnology , Male , Middle Aged , Polymorphism, Genetic , Saint Vincent and the Grenadines/ethnologyABSTRACT
A monoclonal antibody (H-99) was prepared that reacts specifically with the leukemia cells from a patient with stage IV chronic lymphocytic leukemia of B-cell origin. Escalating doses of the antibody were administered to the patient in two courses of therapy. The first course did not result in any significant change in total lymphocyte count; however, the platelet count (which was initially abnormal) rose steadily during treatment to a level greater than 250,000 per mm3. A second course of treatment was begun approximately three months after the first course, and the protocol was modified to consist of five doses of antibody given over a 10-day period, with the highest dose being 500 mg per day. The second course of treatment did not result in any change in total lymphocyte or platelet levels, and there was no significant toxicity associated with the treatment. The failure to induce an anti-tumor response in the patient was probably related to the large concentration (approximately 400 micrograms/mL) of free serum idiotype in addition to the severely compromised immune system of the patient at the time of treatment.
Subject(s)
Antibodies, Monoclonal/therapeutic use , Immunoglobulin Idiotypes , Leukemia, Lymphocytic, Chronic, B-Cell/therapy , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/immunology , Male , Middle AgedABSTRACT
INTRODUCTION: This article attempts to summarise the genetic research that has taken place during the past decade to determine the identity of genes causing high blood pressure. METHODS: Candidate gene studies and genome-wide scanning have been the methods primarily employed, and studies have been performed in both experimental models (rats and mice) and human volunteers (sibling-pairs and case-control). Key studies from the past 10 years are discussed, in addition to the congenic strains. RESULTS: Genome-wide scans and candidate gene studies in both rat and man have generated many chromosomal regions and loci involved in blood pressure regulation. However, much work is still required to fine map the large chromosomal regions found in the genome-wide scans and to isolate variants in candidate genes and prove that they are disease-causing. CONCLUSIONS: It is anticipated that within the next 5 to 10 years at least one blood pressure susceptibility gene will be identified in rat and possibly some in man. It is hoped that the identification of genes controlling blood pressure will enable investigators to determine physiological/biochemical pathways defective in hypertensive patients. This information may then be utilised to identify specific hypertensive phenotypes, to tailor therapy appropriately for patients and hopefully to develop novel therapeutic agents for hypertension.