Molecular monitoring of bleomycin-induced pulmonary fibrosis by cDNA microarray-based gene expression profiling.

Pulmonary fibrosis is a progressive disorder whose molecular pathology is poorly understood. Here we developed an in-house cDNA microarray ("lung chip") originating from a lung-normalized cDNA library. By using this lung chip, we analyzed global gene expression in a murine model of bleomycin-induced fibrosis and selected 82 genes that differed by more than twofold intensity in at least one pairwise comparison with controls. Cluster analysis of these selected genes showed that the expression of genes associated with inflammation reached maximum levels at 5 days after bleomycin administration, while genes involved in the development of fibrosis increased gradually up to 14 days after bleomycin treatment. These changes in gene expression signature were well correlated with observed histopathological changes. The results show that microarray analysis of animal disease models is a powerful approach to understanding the gene expression programs that underlie these disorders.

[A model for clinical study based on genome science--trials in disorders of the immune system and allergies].

Disorders of the immune system, such as allergies, have multi-factorial etiologies that include both genetic and environmental components. The recent advances in genome science have facilitated two strategies for studying the genetic basis of disease: (1) systematic analysis of gene expression profiles and (2) comprehensive analysis of gene variations, such as polymorphisms. Here, we describe a unique research institute, Genox Research Inc., that can relate the clinical profile of a patient to genotyping and molecular profiling. Systematic gene expression analyses using differential display have been performed to explore genes related to allergy, and revealed 93 differentially expressed candidate genes in T-cells. Also, a single nucleotide polymorphism(SNP) analysis project has been designed to mine disease-related and/or drug-response-related genes involved in allergic disorders using biochip technologies. As exemplified above, clinical studies based on these applications of genome science would be of considerable value in clarifying our understanding of multi-gene disorders.

Functional genomic research of alpha 1-adrenoceptors.

The Human Genome Project is now almost completed, and we are about to move into the post-genome sequence era of functional genomics. The advent of genome science has markedly changed the way life science research including pharmacological study is conducted; thus, systematic and integrated 'genome-wide' survey is feasible. The stream of 'Genome-->Transcriptome--> Proteomics' is logical and, in each aspect, approaches for functional genomics are now pursued at a high pace. We have recently developed a standardized technical platform (in various levels, such as transcription, cell and whole animal levels, etc.), and applied these techniques to the study of functional genomics of G-protein-coupled receptors, particularly alpha1-adrenoceptors as a model. Combining the genome information and technology, future pharmacological studies would become the genome-based search and research.

Genomic analysis of a mouse model of immunoglobulin A nephropathy reveals an enhanced PDGF-EDG5 cascade.

The molecular mechanism of immunoglobulin A nephropathy (IgAN), the most common primary renal glomerular disease worldwide, is unknown. HIGA (high serum IgA) mouse is a valid model of IgAN showing almost all of the pathological features, including mesangial cell proliferation. Here we elucidate a pattern of gene expression associated with IgAN by analyzing the diseased kidneys on cDNA microarrays. In particular, we showed an enhanced expression of several genes regulating the cell cycle and proliferation, including growth factors and their receptors, as well as endothelial differentiation gene-5 (EDG5), a receptor for sphingosine 1-phosphate (SPP). One of the growth factors, platelet-derived growth factor (PDGF) induces a marked upregulation of EDG5 in proliferative mesangial cells, and promotes cell proliferation synergistically with SPP. The genomic approach allows us to identify families of genes involved in a process, and can indicate that enhanced PDGF-EDG5 signaling plays an important role in the progression of IgAN.

Lead and zinc concentrations in plasma, erythrocytes, and urine in relation to ALA-D activity after intravenous infusion of Ca-EDTA.

Lead and zinc concentrations in plasma, erythrocytes, and urine, urinary ALA concentration, and ALA-D activity in blood were studied for four hours in two male lead workers during and after a one hour infusion of Ca-EDTA 2Na. Urinary and plasma lead concentrations increased as a result of administering Ca-EDTA 2Na, and the ratios of lead concentrations in plasma to those in urine were greatly increased. The increase of plasma lead concentration was not due to the haemolytic effect of Ca-EDTA 2Na but was mobilised lead, rapidly excreted in the urine. ALA-D activity in blood increased at the end of the experiment with a transient decrease during the infusion of Ca-EDTA 2Na. As zinc concentrations in erythrocytes and plasma did not decrease during the infusion despite an increase in the urinary excretion of zinc, the transient decrease of ALA-D activity was not due to a loss of zinc caused by Ca-EDTA 2Na. From the results of additional experiments in vitro, this transient decrease could be related neither to Ca-EDTA 2Na nor to lead in the blood.

Hippuric acid and o-cresol in the urine of workers exposed to toluene.

Factory workers, 74 males and 56 females exposed predominantly to toluene up to 129 ppm, were examined for the urinary excretion of hippuric acid and o-cresol. The time-weighted averages (TWA) of toluene exposure were measured by personal sampling with carbon felt dosimeters. A preliminary study revealed that the concentrations of hippuric acid and o-cresol in urine increased during work and both reach their peaks at the end of the shift. Correlation coefficients between the TWA of toluene concentration in air and hippuric acid concentration in urine collected at the end of the shift were 0.803 for the 74 males, and 0.830 for the 56 females, while the counterpart correlation coefficients between toluene and o-cresol were 0.607 for the 74 males, and 0.627 for the 56 females, suggesting that hippuric acid is more reliable than o-cresol as an index of toluene exposure. In the urine samples (4 to 8 samples per subject) collected during 8-h worktime from 11 males and 13 females, the urinary levels of o-cresol increased as a function of exposure time in parallel with those of hippuric acid, and the correlation coefficients between o-cresol and hippuric acid were significant (r = 0.834 approximately 0.987; P less than 0.05) when the urine samples from the same subjects were examined. The comparison of the slopes of 24 regression lines between o-cresol and hippuric acid in urine revealed that the maximal slope was almost 8 times as large as the minimal one. From 8 female workers, five urine samples each were collected during 8-h worktime on two consecutive Mondays and analyzed for the two metabolites. The slopes of the regression lines between o-cresol and hippuric acid in the samples from the same subject were identical, regardless of variation in exposure intensity. The findings indicate that an individual difference exists in the pattern of toluene metabolism, and that the ratio between aliphatic and aromatic oxidation is presumably set congenitally. Possible toxicological significance is discussed.

Acute renal failure and maneb (manganous ethylenebis[dithiocarbamate]) exposure.

A 62-year-old man spread maneb on about 200 sq m of garden and subsequently was taken to the emergency clinic with complaints of oliguria, diarrhea, and hoarseness. Based on the clinicobiochemical data, he was found to have acute renal failure; the serum levels of BUN, creatinine, and potassium were 144.3 mg/dL, 14 mg/dL, and 5.8 mEq/L, respectively. The ST segment depression in V4-6, reciprocal ST segment elevation in V1-3, and inverted T waves in V5 and V6 were recorded on ECGs. Both the renal failure and the ECG abnormalities disappeared after hemodialysis. The possibility exists that the maneb caused the acute renal failure.