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CRE-mediated expression of the expected protein His-HA-N17-Rap1b ; was verified after treatment of mice with tamoxifen for five days. Thyroid extracts were prepared, and His-HA-N17-Rap1b was affinity purified on NTA-agarose and analyzed by Western blot with anti-HA antibodies. Only the Rap1bV12LoxP-N17 CRE-ER-Tx mice expressed the expected tamoxifen-dependent HA-Rap1b immunoreactivity Fig.4A, lower panel ; . These results demonstrate the success of the strategy used to generate the Gprotein "activity-state switch" mouse model. Rap1-GTP is required to maintain TSH- and cAMP-mediated hyperplasia. Finally, we tested whether the enhanced cAMP-mediated tumorigenesis seen in thyroids of the Rap1bV12-LoxPN17 mice was causally linked to G12V-Rap1b expression. Two-month-old Rap1bV12-LoxP-N17 CREER-Tx mice were treated with goitrogen, and a five days tamoxifen oil regime then tested either at the beginning establishment ; or during maintenance ; the goitrogen protocol. After four and six months, animals were sacrificed and their thyroids analyzed. The thyroids of tamoxifen-treated mice were significantly smaller than controls Fig.4B ; . Thyroid cross-section areas were ~50% decreased in mice subjected to the tamoxifen regime, both at establishment and during maintenance of the hyperplastic state Fig.4B ; , indicating that Rap1-GTP is required to maintain this state. Moreover, the Ki67 Fig. 4C ; and BrdU not shown ; labeling indices, as well as the total fraction of cycling thyrocytes measured by PCNA staining not shown ; were markedly reduced upon induction of the tamoxifen switch. The partial tamoxifen-dependent inhibition correlated well with the partial penetrance of the recombination event. None of these changes were seen in Rap1bV12-LoxP-N17 mice not shown ; , indicating that the effects of tamoxifen were all CRE-dependent. In addition, TUNEL and DNA ladder analysis failed to reveal any differences in the apoptotic indices of tamoxifen treated animals whether these were the Rap1bV12.
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Julie Kaye, Parkinson's Disease Nurse Specialist, Guildford Research Group ; , Royal Surrey County Hospital, Guildford. Julie Kaye reported on a pilot study done with Patrick Trend and other members of the Guildford Research Group. She said that the aims of their pilot study into the impact of constipation on the quality of life of people with Parkinson's were to investigate the prevalence and burden of constipation in Parkinson's and to explore the need for further research. Questionnaires on bowel habits were given to 156 people with Parkinson's in their dedicated Parkinson's clinic and 148 people attending a local GP surgery who were over 65 and without known neurological problems. Questions were asked about frequency of the four symptoms of constipation using the Rome criteria use of laxatives; how often constipation caused concern to them. The results showed that constipation meeting the Rome criteria was nearly three times more prevalent amongst people with Parkinson's n 92 ; than in the non-Parkinson's group n 31 ; . The most frequently reported symptoms were straining on defecation and lumpy and hard stools. People with Parkinson's reported taking more laxatives than those in the non-Parkinson's group. The study confirmed that constipation is a significant problem for people with Parkinson's and it does severely impact on their quality of life. The Guildford Parkinson's Disease Research Group is now undertaking more detailed research into constipation in Parkinson's. They are investigating the quality of life implications and exploring the socio-demographic, economic and clinical aspects of constipation subjects will complete a `stool diary'. Recruitment is going well and they expect to have completed the work by the end of 2005 and to be able to report on the results during 2006 and docusate.
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