Screen for alterations of iron related parameters in N-ethyl-N-nitrosourea-treated mice identified mutant lines with increased plasma ferritin levels.

Iron is essential for numerous cellular processes. For diagnostic purposes iron-related parameters in patients are assessed by clinical chemical blood analysis including the analysis of ferritin, transferrin and iron levels. Here, we retrospectively evaluated the use of these parameters in the phenotype-driven Munich N-ethyl-N-nitrosourea mouse mutagenesis project for the generation of novel animal models for human diseases. The clinical chemical blood analysis was carried out on more than 10,700 G1 and G3 offspring of chemically mutagenized inbred C3H mice to detect dominant and recessive mutations leading to deviations in the plasma levels of iron-related plasma parameters. We identified animals consistently exhibiting altered plasma ferritin or transferrin values. Transmission of the phenotypic deviations to the subsequent generations led to the successful establishment of three mutant lines with increased plasma ferritin levels. For two of these lines the causative mutations were identified in the Fth1gene and the Ireb2 gene, respectively. Thus, novel mouse models for the functional analysis of iron homeostasis were established by a phenotype-driven screen for mutant mice.

Basic Exploratory Study of Bisphenol A (BPA) Dietary Administration to Istrian Pramenka Rams and Male Toxicity Investigation.

Bisphenol A (BPA), an endocrine-disrupting chemical and environmental pollutant, has been reported by many researchers to induce male reproductive toxicity in different experimental models. In this study, we investigated whether long-term exposure for two months to 25 µg/kg body weight (low dose) of BPA affects spermatogenesis or sperm quality in young Istrian Pramenka rams exposed via diet. We evaluated body and testicular weights, histopathology of testes and epididymides, and sperm analyses, and compared these parameters between the group of treated rams and the control group of rams. Although there were some differences between the two groups, these differences were not large or statistically significant. The only statistically significant difference was the lower epithelial height of seminiferous tubules in treated rams, compared to control rams. In addition to assessing toxicity, BPA concentrations in the blood plasma of treated rams were determined after the first administration, and the toxicokinetic parameters of total BPA were calculated. In this study, no major signs of altered reproduction in rams were detected.

Generation of N-ethyl-N-nitrosourea-induced mouse mutants with deviations in hematological parameters.

Research on hematological disorders relies on suitable animal models. We retrospectively evaluated the use of the hematological parameters hematocrit (HCT), hemoglobin (HGB), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), mean corpuscular volume (MCV), red blood cell count (RBC), white blood cell count (WBC), and platelet count (PLT) in the phenotype-driven Munich N-ethyl-N-nitrosourea (ENU) mouse mutagenesis project as parameters for the generation of novel animal models for human diseases. The analysis was carried out on more than 16,000 G1 and G3 offspring of chemically mutagenized inbred C3H mice to detect dominant and recessive mutations leading to deviations in the levels of the chosen parameters. Identification of animals exhibiting altered values and transmission of the phenotypic deviations to the subsequent generations led to the successful establishment of mutant lines for the parameters MCV, RBC, and PLT. Analysis of the causative mutation was started in selected lines, thereby revealing a novel mutation in the transferrin receptor gene (Tfrc) in one line. Thus, novel phenotype-driven mouse models were established to analyze the genetic components of hematological disorders.

Association of vitamin D binding protein variants with chronic mucus hypersecretion in Iceland.

BACKGROUND: Previous studies of vitamin D binding protein (VDBP, also known as group-specific component, Gc, encoded by the GC gene) have implicated two gene variants, GC*2 and GC*1F, as possible contributors with chronic obstructive pulmonary disease (COPD) protection and susceptibility, respectively. The objective of this study was to examine the association of VDBP to different subtypes of COPD. STUDY DESIGN: The association of the various GC genotypes to the COPD phenotype was examined in Icelandic COPD patients who were followed by pulmonary physicians at the University Hospital of Iceland. METHODS: All patients were genotyped for the known alleles of the GC gene. The single nucleotide polymorphisms (SNPs) were identified by a restriction fragment length polymorphism procedure. Study power was estimated based on allele frequencies of the variants, and risk ratios were calculated from the prevalence of genotypes in the affected group divided by its prevalence in the control population. Statistical analyses were performed using the 2-tailed Fisher's Exact Test and chi(2) test, where appropriate. PATIENT GROUP: One hundred and two COPD patients and 183 controls, together with 46 asthma patients and 48 patients with chronic mucous hypersecretion (CMH) were examined. MAIN OUTCOME MEASURE AND RESULTS: The results demonstrate similar allele and genotype frequencies of GC in COPD patients overall and healthy controls. However, there was a higher prevalence of genotypes carrying a GC*1F allele and lower prevalence of genotypes with a GC*2 allele in the CMH patients than in controls. This difference was most notable in the homozygous form: 8.3% vs 1.1% for the GC*1F/*1F, and 0.0% vs 7.6% for the GC*2/*2 genotypes, respectively. When controlled for smoking, only the non-smoking CMH patients demonstrated a significantly altered frequency of the GC*1F/*1F genotype (p = 0.0001). The prevalence of the GC*2/*2 genotype was also significantly lower in patients with bronchial hypersecretion with airflow obstruction compared with the control group (2.9% vs 7.6%). Taken together, these results demonstrate that the GC*1F and GC*2 alleles are associated with sputum hypersecretion in individuals who are at increased risk of developing COPD.

Autoimmunity and inflammation due to a gain-of-function mutation in phospholipase C gamma 2 that specifically increases external Ca2+ entry.

The identification of specific genetic loci that contribute to inflammatory and autoimmune diseases has proved difficult due to the contribution of multiple interacting genes, the inherent genetic heterogeneity present in human populations, and a lack of new mouse mutants. By using N-ethyl-N-nitrosourea (ENU) mutagenesis to discover new immune regulators, we identified a point mutation in the murine phospholipase Cg2 (Plcg2) gene that leads to severe spontaneous inflammation and autoimmunity. The disease is composed of an autoimmune component mediated by autoantibody immune complexes and B and T cell independent inflammation. The underlying mechanism is a gain-of-function mutation in Plcg2, which leads to hyperreactive external calcium entry in B cells and expansion of innate inflammatory cells. This mutant identifies Plcg2 as a key regulator in an autoimmune and inflammatory disease mediated by B cells and non-B, non-T haematopoietic cells and emphasizes that by distinct genetic modulation, a single point mutation can lead to a complex immunological phenotype.

Vestibular defects in head-tilt mice result from mutations in Nox3, encoding an NADPH oxidase.

The vestibular system of the inner ear is responsible for the perception of motion and gravity. Key elements of this organ are otoconia, tiny biomineral particles in the utricle and the saccule. In response to gravity or linear acceleration, otoconia deflect the stereocilia of the hair cells, thus transducing kinetic movements into sensorineural action potentials. Here, we present an allelic series of mutations at the otoconia-deficient head tilt (het) locus, affecting the gene for NADPH oxidase 3 (Nox3). This series of mutations identifies for the first time a protein with a clear enzymatic function as indispensable for otoconia morphogenesis.