Molecular misreading in non-neuronal cells.

+1 Frame-shifted proteins such as amyloid precursor protein(+1) and ubiquitin-B(+1) have been identified in the neuropathological hallmarks of Alzheimer's disease. These frameshifts are caused by dinucleotide deletions in GAGAG motifs of messenger RNA encoded by genes that have maintained the unchanged wild-type DNA sequence. This process is termed 'molecular misreading'. A key question is whether this process is confined to neurons or whether it could also occur in non-neuronal cells. A transgenic mouse line (MV-B) carrying multiple copies of a rat vasopressin minigene as a reporter driven by the MMTV-LTR promotor was used to screen non-neuronal tissues for molecular misreading by means of detection of the rat vasopressin(+1) protein and mutated mRNA. Molecular misreading was demonstrated to occur in several organs (e.g., epididymis and the parotid gland) where transgenic vasopressin expression is abundant, but its penetrance is variable both between and within tissues. This implies that non-neural tissues too, could be affected by cellular derangements caused by molecular misreading.

Ectopic vasopressin expression in MMTV-v-Ha-ras transgenic mice delays the onset of mammary tumorigenesis.

Neuropeptides are often ectopically expressed by non-endocrine tumours. We used transgenic mice to assess the effect of ectopic expression of the neuropeptide, vasopressin, in mammary tumours induced by the transgenetic expression of an activated ras oncogene. Mice bearing a mouse mammary tumour virus-vasopressin (MMTV-VP) fusion transgene synthesise authentic VP in mammary ducts and alveoli. Bitransgenic mice bearing both MMTV-VP and MMTV-v-Ha-ras transgenes developed tumours that were histologically indistinguishable from those of single MMTV-v-Ha-ras animals. However, tumour onset was significantly delayed in the bitransgenic animals. These data provide evidence that an ectopic neuropeptide can slow the development of ras tumours in vivo.

Ectopic vasopressin expression in MMTV-Wnt-1 transgenic mice modifies mammary tumor differentiation and pathology.

A transgenic mouse model has been developed to test the involvement of ectopic neuropeptide production as a secondary factor in cancer. Mice bearing a mouse mammary tumor virus-vasopressin (MMTV-VP) fusion transgene synthesized authentic vasopressin in mammary ducts and alveoli, but this had no effect on mammary gland development and growth. Mice bearing the MMTV-VP transgene were then mated with mice bearing the MMTV-Wnt-1 transgene to produce bitransgenic animals. Two types of mammary tumor develop in MMTV-Wnt-1 mice; type A mammary adenocarcinomas are uniform with fine acinar structure composed of small epithelial cells arranged to form round cavities and elongated tubules, while adenocarcinoma type B tumors have acinar areas, cystic spaces filled with blood or fluid, intracystic papillary projections, and cords as well as sheets of cells. Compared to the MMTV-Wnt-1 mice, the bitransgenic animals developed proportionally less type B tumors. Further, type B mammary adenocarcinomas from bitransgenic mice exhibited increased proliferation and growth, as judged by mitotic index and argyrophilic nucleolar organizer region counts, compared to type B tumors from MMTV-Wnt-1 mice. These data provide evidence that ectopic neuropeptide production can modulate the development of tumors in vivo.

Postmortem examination of transgenic mice.

Postmortem examination of transgenic animals should ideally be performed by or with the help of a trained pathologist. This is because although the steps in the examination can be manually performed by anyone, the interpretation of the findings can only be done with the knowledge of veterinary medicine. This chapter describes the steps in a postmortem examination (necropsy), and does not attempt to describe pathological lesions and their significance. This method is a systematic approach to detecting gross abnormalities. These abnormalities can then be further examined at the microscopic level or be submitted to other laboratory tests (see Note. 1). Prior knowledge of the anatomy of the animal species examined is necessary (1). Always perform necropsies on freshly killed animals or as soon as they are discovered dead. Necropsies performed on animals that have been dead for more than 24 h generally do not yield much useful information or material for further study.

Histological examination of transgenic mice.

Histological examination is important to detect and differentiate among different types of tissue changes at the microscopic level. An enlarged organ can be hyperplastic, hypertrophied, or neoplastic. It can also be merely inflamed or infected. Only a histological examination can differentiate among the various growth disturbances and pathological changes at the microscopic level. Histopathology is indispensable when neoplasia induced by transgenes is the subject of study.

Decrease in hypothalamic vasopressin mRNA poly(A) tail length following physiological stimulation.

1. The vasopressin mRNA in the adult male rat hypothalamus is modulated in two distinct ways by a dehydration stimulus. In addition to the well-established increase in transcript abundance, it has recently been demonstrated that the vasopressin mRNA poly(A) tail increases in length. 2. We have studied the ontogeny of poly(A) tail length modulation in neonates in response to milk deprivation and found that poly(A) tail length changes are age dependent. In neonates older than 12 days of age, the vasopressin mRNA poly(A) tail length increased with milk deprivation and this effect became more marked in older animals. However, in rats 5 to 9 days old, milk deprivation resulted in a detectable though not significant decrease in vasopressin mRNA poly(A) tail length. 3. As milk deprivation is a combination of dehydration and starvation, we investigated the effect of the latter stimulus in more mature animals. We found that starvation modifies the length of the vasopressin mRNA poly(A) tail in a manner opposite that due to dehydration. 4. Our data indicate a novel mode of regulation of the vasopressin mRNA, namely, poly(A) tail shortening. This system provides a model for future studies concerning the adaptive role of poly(A) tail length modulation in response to physiological stimuli.