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Biomedical subjects

T Crook

Publications and source records attributed to T Crook.

At least 37 records · Page 2Linked to original sources
PubMed1995

Aging and memory: corrections for age, sex and education for three widely used memory tests.

The associate learning subtest from the Wechsler Memory Scale; Benton's Visual Retention test and a Controlled Word Association Task (FAS) were administered to a random sample of normal, healthy individuals whose age ranged from 20 to 79 years, recruited within the Italian peninsula. The neuropsychological examination took place on a mobile unit and the tests were given by the same team of neuropsychologists to reduce variability among examiners. The Research Project was known as Progetto Memoria. Corrections to the scores of these tests were calculated for age, sex, and education. These corrected values will allow clinicians to screen for memory impairment with greater precision among normally aging individuals, thus improving differential diagnosis between physiologic and pathologic deterioration of cognitive functions.

Adult↗
PubMed1994

Transcriptional activation by p53 correlates with suppression of growth but not transformation.

The tumor suppressor protein p53 shows growth and transformation suppression functions that are frequently lost by mutant proteins detected in cancers. Using a large series of p53 mutants, we have demonstrated an excellent correlation between transcriptional activation and growth suppression in p53-null human cells. Not all transcriptionally active mutants retain the ability to suppress transformation in primary rodent cells, however, and two tumor-derived point mutants displayed some evidence of both transforming and transactivating activity. Transformation by these mutants was not mediated by transdominant repression of endogenous p53 transactivating function, and cell lines expressing these p53 proteins showed elevated p53 transcriptional activity. Our results suggest that activation of transcriptional regulation by p53 will not necessarily result in tumor suppression.

Animals↗
PubMed1994

HPV16 E7 oncoprotein deregulates B-myb expression: correlation with targeting of p107/E2F complexes.

HPV16 is a human tumour virus encoding two principal oncoproteins, E6 and E7. Expression of E7 can induce DNA synthesis in quiescent cells and this property coincides with its ability to bind to the cell proteins pRb and p107. As these cell proteins are regulators of the transcription factor E2F, we have investigated whether the interaction with E7 could result in induction of cell cycle regulated genes. We show that B-myb, whose induction at the G1/S boundary is regulated by release from E2F mediated transcriptional repression, is a target for transcriptional activation by E7 and is the first E7 responsive cell gene to be identified. E7 transactivation leads to both inappropriate transcription of B-myb during G1 and constitutive over-expression in cycling cells. B-Myb plays an essential role in cell cycle progression, and activation by E7 is likely to contribute to the mitogenic activity of the viral oncoprotein. Regulation of the B-myb promoter in NIH3T3 cells correlates with binding of distinct p107-containing complexes at the E2F binding site, and analysis of E7 mutants confirms that B-myb transcription in these cells is regulated through interactions with p107 rather than pRb. These results provide the first example of a potentially specific role for p107 in the regulation of the cell cycle.

3T3 Cells↗
PubMed1994

Posttransplant skin cancer: a possible role for p53 gene mutation but not for oncogenic human papillomaviruses.

BACKGROUND: Loss of p53 tumor suppressor function is a critical step in the development of diverse malignancies, including skin cancers in nonimmunosuppressed patients where UV-specific p53 gene mutations have been identified. In tumors associated with human papillomavirus (HPV), such as cervical carcinoma, p53 may be inactivated instead by binding to a viral oncoprotein. OBJECTIVE: Our purpose was to examine the hypothesis that HPV may play an analogous role in the development of posttransplant skin cancer. METHODS: p53 Immunoreactivity, suggestive of p53 gene mutation, was examined by immunocytochemistry. Oncogenic HPV DNA was detected by polymerase chain reaction. RESULTS: Comparable p53 immunoreactivity was seen in skin tumors from both transplant and nontransplant patients. HPV DNA was not demonstrated in any tumor specimen. CONCLUSION: Our data do not implicate oncogenic HPV in posttransplant skin cancer. p53 Gene mutation, rather than HPV-induced p53 degradation, may be more significant in the development of these tumors.

Basal Cell Carcinoma↗
PubMed1994

Detergent extraction and enzymatic analysis for fecal long-chain fatty acids, triglycerides, and cholesterol.

We report a procedure for determining fecal long-chain fatty acids (LCFA), triglycerides, and cholesterol after detergent extraction with 10 mL/L Triton X-100, 6 mL/L Brij 30, and 0.1 mol/L HCl in isotonic saline by enzymatic analysis of the extraction supernates. Mean recoveries of calibrators ranged from 105% to 117%. Assays of fecal extracts were linear with concentration from 0 to 1000 mumol/L for LCFA, from 10 to 90 mumol/L for triglycerides, and from 0 to 300 mumol/L for cholesterol. Within-run CVs were < 3% for medium and high concentrations, and 3%, 14%, and 28% for low concentrations of LCFA, cholesterol, and triglycerides, respectively. Correlation with gravimetric determination resulted in coefficients (r2) of 0.72 for total fat, 0.63 for neutral fat, and 0.66 for nonesterified fat. Assays of extracts and residues prepared for gravimetric determination indicated that approximately 90% of triglycerides, cholesterol, and LCFA were extracted by organic solvents and that the extracts contain a significant amount of other fats. The proposed method appears accurate, precise, specific, and suitable for routine analysis.

Cholesterol↗
PubMed1994

Interaction of p53 with MDM2 is independent of E6 and does not mediate wild type transformation suppressor function.

Using a new series of p53 mutants targeting the conserved regions we have analysed the relationship of various activities of the protein. Mdm-2 and human papillomavirus (HPV) E6, two proteins which interact with and abrogate p53 function, were shown to bind independently. Deletion of the conserved regions of the protein in which most of the naturally occurring mutations are found (boxes II-V) abrogated transcriptional activity and the ability to interact with E6, supporting the importance of this DNA binding domain to these activities. Nevertheless, these mutants retained the ability to interact with mdm2. One mutant, deleted of all the C-terminal sequences, showed loss of mdm2 binding, E6 binding and transcriptional activity. More subtle mutations within the C-terminus of the protein, including alterations of the cdc2 and CKII phosphorylation sites, had no effect on the transcriptional trans-activation, mdm-2 or E6 binding functions, indicating that phosphorylation of these sites is not essential for these activities. Deletion of conserved box I sequences abolished the interaction with mdm-2 without loss of transcriptional activation or transformation suppressor activity, suggesting that mdm-2 is not a downstream effector of p53 function.

CDC2 Protein Kinase↗
PubMed1994

Modulation of transcriptional regulatory properties of p53 by HPV E6.

The E6 protein encoded by human papillomavirus type 16 (HPV16), a genital virus with oncogenic potential, can target cellular p53 for rapid degradation following the formation of a complex including the two proteins. Some studies suggest that the E6 proteins encoded by HPV6 and 11, viral types which are normally limited to benign lesions, may also interact with p53, although the association is weaker than that seen with HPV16 E6. The present study demonstrates that E6 proteins from HPV16 and HPV6 can modulate the transcriptional regulatory functions of p53 in several cell types. A series of E6 mutants was used to show that association between E6 and p53 is necessary for this activity and that E6 proteins which retain the ability to associate with p53 but show no detectable degradation activity in vitro can, to some extent, abrogate p53 mediated transcriptional trans-regulation. This activity is augmented, however, by the ability of the E6 protein to target bound p53 for rapid degradation. These results suggest that some degree of modulation of p53 function is necessary in the normal viral life cycle but also demonstrate a correlation between the efficiency of this activity and oncogenic potential of the virus.

3T3 Cells↗
PubMed1993

Biological activities of p53 mutants in Burkitt's lymphoma cells.

Wild-type human p53 and a series of p53 point mutants isolated from Burkitt's lymphoma (BL) cell lines were tested for their ability to inhibit DNA synthesis in a p53-negative BL cell line and to bind and be degraded by the human papillomavirus type 16 E6 protein. All the mutants lost the wild-type ability to inhibit DNA synthesis, demonstrating that they are all functionally altered. Binding to E6 and consequent degradation of the p53 mutants frequently correlated with changed suppressor properties in BL cells.

Animals↗
PubMed1993

Human papillomavirus type 16 E7 associates with a histone H1 kinase and with p107 through sequences necessary for transformation.

The transforming function of human papillomavirus type 16 (HPV16) E7 has been shown to depend on activities additional to the ability to bind RB. In this paper we describe two further properties of E7 which may also contribute to transformation, an association with a histone H1 kinase at the G2/M phase of the cell cycle and an ability to bind the RB-related protein p107. The region of E7 identified previously as important for RB binding was found to be involved in the association with the kinase and complex formation with p107, although analysis of E7 point mutants within this region revealed a difference in the precise sequence requirement for RB and p107 binding. Association with the kinase activity correlated with the ability to bind RB, but the restriction of the kinase association to the G2/M phase of the cell cycle implies that this activity might not be directly mediated by RB binding. Since kinase-binding-deficient E7 mutants are also transformation defective, this may represent an independent function of E7 which plays a role in the G2/M phase of the cell cycle.

Cell Line↗
PubMed1993

Human papillomavirus type 16 E7 regulates E2F and contributes to mitogenic signalling.

We have produced human papillomavirus type 16 E7 protein in a bacterial expression system and examined the mitogenic activity of this protein in Swiss 3T3 cells after scrape loading. The ability of E7 to induce cellular DNA synthesis in quiescent mouse fibroblasts is strongly enhanced by the presence of a single growth factor such as insulin. Although only weakly mitogenic, introduction of E7 alone resulted in the rapid induction of the transcriptionally active form of E2F, which was not enhanced further by the addition of insulin. Mutant E7 proteins defective for RB binding failed to induce the active form of E2F or act synergistically with insulin to stimulate DNA synthesis. The ability of E7 to regulate E2F may therefore be necessary, but is not sufficient, for full induction of DNA synthesis.

3T3 Cells↗
PubMed1992

Clonal p53 mutation in primary cervical cancer: association with human-papillomavirus-negative tumours.

Analyses of cancer cell lines and of anal cancers suggest an inverse correlation between infection with human papillomavirus (HPV) and somatic mutation of the p53 tumour-suppressor gene. We have investigated this association in primary cervical tumours. Tumour-tissue samples from 28 women with primary cancer of the cervix were analysed for presence of HPV sequences and for somatic mutations of the p53 gene. Southern blot analysis and the polymerase chain reaction (PCR) showed that 25 of the tumours contained HPV sequences; 20 were HPV16 positive and 5 HPV18 positive. 17 tumours subjected to restriction fragment length polymorphism analysis for the short arm of chromosome 17 showed no evidence of allelic deletion. Sequencing of the entire coding region of the p53 gene by asymmetric PCR detected heterozygous point mutations in only 3 HPV-negative tumours. By contrast, in 21 HPV-positive cancers the p53 sequence was wild-type throughout. Our data indicate that loss of wild-type p53 function is important in the pathology of cervical cancer and that in the absence of an HPV-encoded gene product that mediates loss of p53 function, somatic mutation of the gene is required. This pattern of p53 mutation may partly explain the apparently worse prognosis of HPV-negative cervical cancers.

Adenocarcinoma↗
PubMed1992

Human papillomavirus E6 proteins bind p53 in vivo and abrogate p53-mediated repression of transcription.

The transforming proteins of DNA tumor viruses SV40, adenovirus and human papillomaviruses (HPV) bind the retinoblastoma and p53 cell cycle regulatory proteins. While the binding of SV40 large T antigen and the adenovirus E1B 55 kDa protein results in the stabilization of the p53 protein, the binding of HPV16 and 18 E6 results in enhanced degradation in vitro. To explore the effect of viral proteins on p53 stability in vivo, we have examined cell lines immortalized in tissue culture by HPV18 E6 and E7 or SV40 large T antigen, as well as cell lines derived from cervical neoplasias. The half-life of the p53 protein in non-transformed human foreskin keratinocytes in culture was found to be approximately 3 h while in cell lines immortalized by E6 and E7, p53 protein half-lives ranged from 2.8 h to less than 1 h. Since equivalent levels of E6 were found in these cells, the range in p53 levels observed was not a result of variability in amounts of E6. In keratinocyte lines immortalized by E7 alone, the p53 half-life was found to be similar to that in non-transformed cells; however, it decreased to approximately 1 h following supertransfection of an E6 gene. These observations are consistent with an interaction of E6 and p53 in vivo resulting in reductions in the stability of p53 ranging between 2- and 4-fold. We also observed that the expression of various TATA containing promoters was repressed in transient assays by co-transfection with plasmids expressing the wild-type p53 gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Properties of p53 mutations detected in primary and secondary cervical cancers suggest mechanisms of metastasis and involvement of environmental carcinogens.

Primary human papillomavirus (HPV) positive anogenital cancers normally develop without somatic mutation within the p53 gene. In this study, however, we have identified p53 point mutations in metastases arising from HPV positive cervical carcinomas, suggesting that acquisition of p53 mutation may play a role in the progression of some HPV associated primary cancers. p53 mutants identified in anogenital cancers exhibit a dominant transforming phenotype and increased resistance to HPV16 E6 directed degradation. The association of p53 mutation with metastases may explain the poor prognosis reported for HPV negative primary cancers, many of which already contain mutant p53. A high proportion of p53 mutations detected in both primary and metastatic cancers are GC-->TA transversions, strongly suggesting a role for external carcinogens in the development of these cancers.

Animals↗
PubMed1992

Lack of immortalizing activity of a human papillomavirus type 16 variant DNA with a mutation in the E2 gene isolated from normal human cervical keratinocytes.

The oncogenic potential of a human papillomavirus type 16 (HPV16) variant cloned from normal human cervical keratinocytes has been tested in vitro using primary rodent epithelial cells and human cervical keratinocytes. The HPV16 variant was able to extend the lifespan of, but failed to immortalize, human keratinocytes. It could however cooperate with an activated ras oncogene to transform primary rodent cells. Radioimmunoprecipitation assays of the rodent cells showed that they expressed the E7 protein. DNA sequence analysis of the URR/E6/E7 and E5 regions of the HPV16 showed them to be fully functional, but a deletion in the viral E2 open reading frame was detected. This truncated E2 only weakly stimulated transcription of the viral regulatory region. Complementation assays using the HPV16 variant and a full-length E2 enabled the cloned variant to immortalize human cervical keratinocytes with wild-type efficiency. These results suggest that other viral gene products in addition to E6/E7 may play an important role in the in vitro immortalization of cervical keratinocytes in HPV16 and the development of cervical cancer.

Amino Acid Sequence↗
PubMed1992

Effects of phosphatidylserine in Alzheimer's disease.

We studied 51 patients meeting clinical criteria for probable Alzheimer's disease (AD). Patients were treated for 12 weeks with a formulation of bovine cortex phosphatidylserine (BC-PS; 100 mg t.i.d.) or placebo, and those treated with the drug improved on several cognitive measures relative to those administered placebo. Differences between treatment groups were most apparent among patients with less severe cognitive impairment. Results suggest that phosphatidylserine may be a promising candidate for study in the early stages of AD.

Aged↗
PubMed1992

Noradrenergic intervention in Alzheimer's disease.

We report results of a double-blind, 13-week trial of the alpha2 noradrenergic agonist guanfacine in 29 patients meeting clinical criteria for probable Alzheimer's disease (AD). Patients studied were relatively young (mean = 71 years), well educated (mean = 13.5 years), and in the early stages of cognitive deterioration (mean Mini-Mental State score = 22). We found no evidence of improvement with a single .5-mg daily dose of guanfacine and some evidence of a modest negative effect of the drug on neuropsychological tests. We concluded that noradrenergic intervention alone is unlikely to be effective in Alzheimer's disease.

Aged↗