Transforming growth factor-alpha expression in rat experimental hepatocarcinogenesis.

Growth factors in general and transforming growth factor-alpha in particular have been related to cell proliferation and cell differentiation. This study was designed to clarify the distribution pattern of TGF-alpha in chemically-induced hepatocarcinogenesis. Sprague-Dawley rats were subjected to different non-intensive or intensive carcinogenic treatments using diethylnitrosamine (DEN) as carcinogen and ethinyl estradiol (EE) as promoter. The livers were fixed in 2% paraformaldehyde, dehydrated in a series of ethanol solutions, embedded in paraffin and sectioned. In the preneoplastic lesions no TGF-alpha immunoreactive cells were identified, but in some hepatic tumours cell immunostained with TGF-alpha antibody were observed. These results suggest that the cells capable of expressing TGF-alpha constitutively may be involved in neoplastic development in vivo.     

Stage-specific expression of a developmentally regulated gene in Dirofilaria immitis.

A previously reported cDNA clone encoding 34 kDa antigenic polypeptide of Dirofilaria immitis (lambda cD34) was studied to elucidate the mechanism of stage-specific gene expression. The 34 kDa polypeptide was a larva-specific antigen and the mRNA was detectable in microfilariae but not in adult worms and eggs. The lambda cD34 gene was not sex linked and was contained in the genome of D. immitis at each stage. The stage-specific expression of the developmentally regulated gene in D. immitis may be controlled primarily at the mRNA level.     

Transforming growth factor-alpha expression in benign and malignant human prostatic disease.

Investigation of biological variables in prostatic disease may not only prevent patients with a good prognosis being overtreated, but allow better selection of appropriate therapy, and may identify potential targets for novel therapies. This study investigates the growth factor transforming growth factor-alpha (TGF alpha) expression in benign and malignant prostatic biopsies using both radioimmunoassay and immunohistochemistry, considering its role in malignant epithelial transformation and as a prognostic indicator. Biochemical methods were less satisfactory than the more selective immunohistochemical methods, due to the heterogeneity of prostatic tissue. Seventy-one percent of benign biopsies (range 0-18.62ng/mg DNA) and 69% of malignant biopsies (range 0-11.1ng/mg DNA) had detectable levels of TGF alpha using radioimmunoassay. Immunohistochemical staining for TGF alpha identified expression in 15% of benign (4 out of 27) and 53% malignant biopsies (18 out of 34). Positive staining was also identified in premalignant lesions and within stromal elements, thus implying the factor's role in autocrine/paracrine growth and/or malignant transformation. Immunostaining for TGF alpha may enhance detection of premalignant lesions and small foci of malignant glands which are otherwise difficult to identify using standard histopathological techniques.     

Transforming growth factor-alpha and epidermal growth factor receptor gene expression and action during pubertal development of the seminiferous tubule.

The potential role of transforming growth factor-alpha (TGF-alpha) as a mediator of cell-cell interactions in the growth and development of the testis was examined. Developing rat testes were collected, and preparations of mesenchymal-derived peritubular cells and epithelial-like Sertoli cells were isolated from prepubertal, midpubertal, and late pubertal rat testes. The developmental expression of TGF-alpha and its receptor, the epidermal growth factor receptor (EGFR), in whole testis and isolated cell types was determined using a nuclease protection assay. TGF-alpha and EGFR gene expression were predominant early in testis development and decreased during pubertal development. TGF-alpha expression was greatest in prepubertal peritubular cells. Sertoli cell TGF-alpha expression remained relatively constant during development, with a slight decline at the later pubertal stages. EGFR gene expression was predominant in peritublar cells throughout development. A low level of EGFR expression was detected in Sertoli cells. Scatchard analysis confirmed the presence of high affinity receptors on peritubular cells; however, no functional receptors were detected on Sertoli cells from any stage of development examined. Interestingly, low-level EGFR gene expression was also detected in pachytene spermatocytes and round spermatids. TGF-alpha was found to stimulate [3H] thymidine incorporation into DNA and increase cellular proliferation of peritubular cells from each developmental stage, while having no effect on Sertoli cells. The in vivo physiological significance of TGF-alpha was evaluated in a line of transgenic mice which overexpress TGF-alpha in the mature testis. These transgenic animals had no abnormal testicular morphology or alterations in spermatogenesis. Observations demonstrate that gene expression of TGF-alpha and its receptor is high during early pubertal stages when somatic cell growth is predominant and low at late pubertal stages when somatic cell proliferation is reduced. TGF-alpha can act as an autocrine/paracrine mitogen for the mesenchymal-derived peritubular cell, while actions on the Sertoli cell population are not evident. The observation that spermatogenic cells express the EGFR gene, although the protein remains to be identified, implies that TGF-alpha may potentially mediate Sertoli-germinal cell interactions.     

Pleiotrophin gene expression is highly restricted and is regulated by platelet-derived growth factor.

Pleiotrophin (PTN) is a growth and neurite extension promoting polypeptide which is highly expressed in brain and in tissues derived from mesenchyme. The PTN gene is developmentally regulated and is closely related to the MK and RI-HB genes, both of which are developmentally regulated and induced by retinoic acid. We now have screened 17 cell lines and report that expression of the PTN gene in these cells is restricted to embryo fibroblasts and intestinal smooth muscle cells. However, NIH 3T3 cells stimulated by the platelet-derived growth factor (PDGF) express a marked increase in levels of PTN mRNA whereas retinoic acid failed to increase levels of PTN mRNA in NIH 3T3 cells or in F9 embryonal carcinoma cells within 72 hours of exposure. The results suggest that expression of the PTN gene is highly restricted and that the PTN gene is a new member of the PDGF-induced cytokine family.     

Tissue-specific and developmentally regulated expression of a chimeric actin-globin gene in transgenic mice.

A chimeric plasmid containing about 2/3 of the rat skeletal muscle actin gene plus 730 base pairs of its 5' flanking sequences fused to the 3' end of a human embryonic globin gene (D. Melloul, B. Aloni, J. Calvo, D. Yaffe, and U. Nudel, EMBO J. 3:983-990, 1984) was inserted into mice by microinjection into fertilized eggs. Eleven transgenic mice carrying the chimeric gene with or without plasmid pBR322 DNA sequences were identified. The majority of these mice transmitted the injected DNA to about 50% of their progeny. However, in transgenic mouse CV1, transmission to progeny was associated with amplification or deletion of the injected DNA sequences, while in transgenic mouse CV4 transmission was distorted, probably as a result of insertional mutagenesis. Tissue-specific expression was dependent on the removal of the vector DNA sequences from the chimeric gene sequences prior to microinjection. None of the transgenic mice carrying the chimeric gene together with plasmid pBR322 sequences expressed the introduced gene in striated muscles. In contrast, the six transgenic mice carrying the chimeric gene sequences alone expressed the inserted gene specifically in skeletal and cardiac muscles. Moreover, expression of the chimeric gene was not only tissue specific, but also developmentally regulated. Similar to the endogenous skeletal muscle actin gene, the chimeric gene was expressed at a relatively high level in cardiac muscle of neonatal mice and at a significantly lower level in adult cardiac muscle. These results indicate that the injected DNA included sufficient cis-acting control elements for its tissue-specific and developmentally regulated expression in transgenic mice.     

Transforming growth factor-alpha binds to the epidermal growth factor receptor in gastric mucosa.

The ability of transforming growth factor-alpha (TGF-alpha) to interact with the gastric mucosal epidermal growth factor (EGF) receptor was investigated using a mucosal membrane preparation. TGF-alpha inhibited specific binding of [125I]EGF to its receptor, but the IC50 for TGF-alpha was at least 100 fold greater than that observed for unlabeled EGF. Cross-linking studies revealed no attachment of [125I]TGF-alpha to EGF-receptor size components, and the unlabeled TGF-alpha was only weakly effective in inhibiting cross-linking of [125I]EGF to the 170 kDa receptor. However, when the cytosolic fraction was reconstituted with the membrane preparation, an enhancement in binding of [125I]TGF-alpha to the EGF receptor occurred in a manner dependent on the concentration of cytosolic protein. Hence the binding characteristics of TGF-alpha to the EGF receptor in gastric mucosa are different from those for EGF.     

Interplay of developmentally regulated gene expression and heterochromatic silencing in trans in Drosophila

The brown(Dominant) (bw(D)) allele of Drosophila contains a heterochromatic block that causes the locus to interact with centric heterochromatin. This association silences bw(+) in heterozygotes (trans-inactivation) and is dependent on nuclear organizational changes later in development, suggesting that trans-inactivation may not be possible until later in development. To study this, a P element containing an upstream activating sequence (UAS)-GFP reporter was inserted 5 kb from the bw(D) insertion site. Seven different GAL4 driver lines were used and GFP fluorescence was compared in the presence or the absence of bw(D). We measured silencing in different tissues and stages of development and found variable silencing of GFP expression driven by the same driver. When UAS-GFP was not expressed until differentiation in the eye imaginal disc it was more easily trans-inactivated than when it was expressed earlier in undifferentiated cells. In contrast to some studies by other workers on silencing in cis, we did not find consistent correlation of silencing with level of expression or evidence of relaxation of silencing with terminal differentiation. We suggest that such contrasting results may be attributed to a potentially different role played by nuclear organization in cis and trans position-effect variegation.