Shigehiko Taniguchi

Expression of hepatocyte growth factor gene in endothelial and Kupffer cells of damaged rat livers, as revealed by in situ hybridization

Hepatocyte growth factor (HGF) has been demonstrated to be synthesized and secreted by non-parenchymal liver cells for liver regeneration after hepatic injury. We performed in situ hybridization to identify HGF-producing cell types in rat liver hepatitis induced by administrating carbon tetrachloride as a hepatotoxin. We found that transcripts of the HGF gene are localized in the Kupffer and endothelial cells in normal livers and increased remarkably in the Kupffer cells of the damaged livers. Thus, HGF is concluded to be synthesized in the Kupffer and endothelial cells to repair the liver tissue in paracrine fashion. No significant increase in the transcripts of the HGF gene was observed in livers after partial hepatectomy, indicating that a mechanism on liver regeneration after the hepatectomy differs from that on liver repairs. Since the HGF gene expression was also found in lung and kidney, HGF may be a ubiquitous factor for tissue repairs.

Expression pattern of acidic and basic fibroblast growth factor genes in adult rat eyes

Although the retinal angiogenic and mitogenic factors have been identified to be acidic and basic fibroblast growth factors (aFGF and bFGF), little information has so far been available about the cells producing them and their function in retinal tissues. We found, by in situ hybridization, that the expression pattern of the aFGF gene differed remarkably from that of the bFGF gene in adult rat eyes. Our results demonstrated that the aFGF gene was produced by photoreceptor visual cells, neuronal cells in the inner nuclear layer and ganglion cells of the retina, in addition to pigment epithelial cells of the choroid, iris and ciliary body, and epithelial cells of the cornea, conjunctiva and lens, while bFGF was synthesized solely by the photoreceptor visual cells.

Expression of retinoic acid receptor genes in neural crest-derived cells during mouse facial development

Retinoic acid (RA) is known as a teratogen that induces abnormalities in facial structures which are made up mainly of neural crest‐derived mesenchyme. We investigated expression patterns of RA receptor (RAR) genes (subtypes α, β, γ) during mouse facial development. The expression of the rarβ gene is specific for the mesenchyme around developing eyes and nose, whereas the RARγ gene is expressed in the mesenchyme differentiating to facial cartilages and bones. In contrast, the RARα gene is expressed weakly and uniformly over the facial region. These results suggest that crucial roles of endogenous RA in facial development depend on differential functions of the RAR subtypes.

The narK gene product participates in nitrate transport induced in Escherichia coli nitrate-respiring cells

The nucleotide sequence of the Escherichia coli narK gene, which is located in the upstream region of the narCHJI operon, was determined. The narK gene encodes a very hydrophobic protein with 463 amino acid residues (M r 49 693). A narK deletion mutant, under conditions for the induction of nitrate respiration, was unable to perform nitrate transport. Loss of transport activity was recovered by transforming the mutant with a narK + plasmid. Thus, we conclude that the narK gene encodes a transmembrane protein participating in nitrate transport. In the narK promoter region, we defined a unique sequence that we designate as a ‘nitrate box’, functioning as a putative NarL‐binding site, in addition to the consensus sequence of the ‘anaero‐box’.

Bacterial endotoxin-induced expression of metallothionein genes in rat brain, as revealed by in situ hybridization

In order to clarify acute-phase response in brain, we investigated induction of metallothionein (MT) genes by administrating an endotoxin (lipopolysaccharide) in rat intraperitoneum. We performed in situ hybridization on the serial brain sections to identify the cells expressing the MT genes in acute-phase. After endotoxin administration, transcripts of MT genes were detected in the arachnoideal, ependymal cells and glial cells around the Purkinje cells of the cerebellum, while no significant induction of the MT genes by zinc ion was observed in brain. These results suggest that the acute-phase response occurs specifically in at least these 3 non-neuronal cells.

Elevation in metallothionein messenger RNA in rat tissues after exposure to X-irradiation

Metallothionein (MT) mRNA levels in tissues were measured in rats following whole-body X-irradiation (2 and 20 Gy). When compared with control rats, the elevation in MT mRNA levels of liver, kidney, and thymus was observed in irradiated rats at 9 or 72 hr after irradiation. However, the elevation in MT mRNA levels was not observed in brain, spleen, lung, heart, and testis. When compared with other tissues, testicular MT mRNA levels in control rats were extremely high, and treatment with X-irradiation produced a slight decrease of testicular MT mRNA levels. Time-course experiments indicated that hepatic and renal MT mRNA reached a maximum at 6 hr after irradiation. In low-dose (2 Gy) irradiated rat, these values were returned to control values by 4 days after irradiation. However, in high-dose (20 Gy) irradiated rat, the values were not decreased to control values. These data indicate that treatment with X-irradiation produces an elevation in MT mRNA in rat tissues.

Expression pattern of the activin receptor type IIA gene during differentiation of chick neural tissues, muscle and skin

To elucidate target cells of activins during embryogenesis we isolated cDNAs of chick activin receptor type II (cActR‐II) and studied expression patterns of thecActR‐II gene by in situ hybridization. Transcripts ofcActR‐II were observed in neuroectoderm developing to spinal cord, brain and eyes, in surface ectoderm differentiating to epidermis, and in myotomes differentiating to muscles. The expression patterns ofcActR‐II suggest that activin and its receptor are involved in differentiation of chick neural tissues, muscle and skin after inducing the dorsal mesoderm.

Quantitative Assay of Epidermal Growth Factor Receptor in Human Squamous Cell Carcinomas of the Oral Region by an Avidin‐Biotin Method

A quantitative assay method for epidermal growth factor receptors (EGFRs) of human tumor tissues was established, based on enzyme‐labeled avidin‐biotin (LAB) interaction with anti‐human EGFR monoclonal antibody 52SIgG. A standard calibration curve for EGFR estimation in human tumor tissues was obtained with A431#8 cells cloned from A431 human epidermoid carcinoma cell line. The coefficient of variance for the standard curve was below 35% in the application to tumor tissues from nude mice implanted with human tumor cell lines. The minimum tissue amount required for the quantitative assay was around 0.1 g (wet weight). Using the LAB method, the correlation between the level of EGFR number and tumor malignancy was examined for 14 human squamous cell carcinomas (SCCs) from the oral region. Seven of the SCCs showed a more than two‐fold higher EGFR number compared to normal gingival tissues. Three highly aggressive carcinomas with poor prognosis possessed five to ten times higher levels of EGFR number than normal tissues. The elevated EGFR level in the SCCs seems to correlate to increasing tumor size and the stage of SCCs as clinically classified according to the 1987 UICC TNM system.

Functional modes of retinoic acid in mouse osteoblastic clone MC3T3-E1, proved as a target cell for retinoic acid

Mouse osteoblastic clone MC3T3‐E1 was proved as a target cell for retinoic acid (RA) in bone tissues through the demonstration of RA‐receptor gene expression by the northern blot analysis. The effect of RA on the cell growth of MC3T3‐E1 was repressive for both subconfluent and confluent growth, whereas RA enhancement of alkaline phosphatase expression was observed at the confluent stage. This implies that RA is a regulatory factor leading osteogenesis of the cells after the confluent stage. RA exhibited simultaneously the stage‐dependent effects on EGF‐dependent mitogenesis: promotive at the subconfluent, but repressive at the confluent stage.

Spatial and temporal expression pattern of retinoic acid receptor genes during mouse bone development

Retinoic acid receptor gene; Hybridization, in situ; Bone formation; (Mouse, Forelimb)