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PubMed2005

Improvement of a FRET-based indicator for cAMP by linker design and stabilization of donor-acceptor interaction.

Förster resonance energy transfer (FRET) technology has been used to develop genetically encoded fluorescent indicators for a variety of intracellular molecular events. Often, however, the poor dynamic range of such reporters prevents detection of subtle but physiologically relevant signals. Here we present a strategy for improving FRET efficiency between donor and acceptor fluorophores in a green fluorescent protein (GFP)-based protein indicator for cAMP. Such indicator is based on protein kinase A (PKA) and was generated by fusion of CFP and YFP to the regulatory and catalytic subunits of PKA, respectively. Our approach to improve FRET efficiency was to perform molecular dynamic simulations and modelling studies of the linker peptide (L11) joining the CFP moiety and the regulatory subunit in order to define its structure and use this information to design an improved linker. We found that L11 contains the X-Y-P-Y-D motif, which adopts a turn-like conformation that is stiffly conserved along the simulation time. Based on this finding, we designed a new linker, L22 in which the YPY motif was doubled in order to generate a stiffer peptide and reduce the mobility of the chromophore within the protein complex, thus favouring CFP/YFP dipole-dipole interaction and improving FRET efficiency. Molecular dynamic simulations of L22 showed, unexpectedly, that the conformational behaviour of L22 was very loose. Based on the analysis of the three principal conformational states visited by L22 during the simulation time, we modified its sequence in order to increase its rigidity. The resulting linker L20 displayed lower flexibility and higher helical content than L22. When inserted in the cAMP indicator, L20 yielded a probe showing almost doubled FRET efficiency and a substantially improved dynamic range.

Amino Acid Sequence↗
PubMed1999

Face-to-face diagnosis vs telediagnosis of pigmented skin tumors: a teledermoscopic study.

BACKGROUND: Teledermoscopy uses telecommunication technologies to transfer images of pigmented skin lesions, including clinical and anamnestic data, via e-mail to specialized centers for teleconsultation. DESIGN: Sixty-six pigmented skin lesions examined on a face-to-face basis in a skin lesion clinic in L'Aquila, Italy, were sent via e-mail on a standard-resolution color monitor for consultation at a university dermatology department in Graz, Austria. INTERVENTION: Digital photographs of the clinical and dermoscopic images of all pigmented tumors were taken with a stereomicroscope connected to a high-resolution video camera in Truevision advanced graphic array (Targa) format file and converted successively into a Joint Photographic Expert Group (PEG) format file. All lesions were excised surgically and diagnosed histopathologically. MAIN OUTCOME MEASURE: Diagnostic concordance between face-to-face diagnosis and telediagnosis. RESULTS: The diagnostic concordance was 60 (91%) of 66 cases. The number of correct telediagnoses was lower, but the difference was not statistically significant (Wilcoxon test, P = .10). The accuracy of the telediagnoses was not related to the quality of the images, but highly depended on the level of diagnostic difficulty of a given pigmented skin tumor (Spearman correlation, P= .01). CONCLUSION: Teleconsultation of clinical and dermoscopic images of skin tumors via e-mail provides a similar degree of diagnostic accuracy as face-to-face diagnosis.

Adolescent↗
PubMedDate not supplied

Human tissue research in the genomic era of medicine: balancing individual and societal interests.

Advances in DNA sequencing technology and in our understanding of the human genome are ushering in a new era of genomic medicine, one with dramatic potential to not only benefit society through research involving human tissue, but also to cause economic or psychosocial harms to tissue donors and their families. This delicate situation requires that the needs of tissue donors be carefully considered and balanced with those of the medical research community, especially on issues concerning confidentiality, consent, and compensation. We analyzed the tensions between tissue donors and researchers over the research use of human tissue. We also reviewed several approaches, including the establishment of tissue-trustee infrastructures at academic medical centers, aimed at achieving a more equitable balance between individual donor protection and societal benefits derived from tissue-based research. Arch Intern Med. 2000;160:3377-3384.

Academic Medical Centers↗
PubMed2000

Policies on faculty conflicts of interest at US universities.

CONTEXT: Despite federal regulations on faculty conflicts of interest in federally funded research, academic-industry ties are common, and evidence exists that financial considerations bias the research record. Public scrutiny of these ties is increasing, especially in cases where researchers have financial interests in the corporate sponsors of their clinical research. OBJECTIVE: To review policies on conflict of interest at major biomedical research institutions in the United States. DESIGN: Cross-sectional survey and content analysis study conducted from August 1998 to February 2000. SETTING AND PARTICIPANTS: The 100 US institutions with the most funding from the National Institutes of Health in 1998 were initially sampled; policies from 89 institutions were available and included in the analysis. MAIN OUTCOME MEASURES: Process for disclosure, review, and management of conflicts of interest and specified management strategies or limitations, according to the institutions' faculty/staff conflict of interest policies. RESULTS: Content of the conflict of interest policies varied widely across institutions. Fifty-five percent of policies (n = 49) required disclosures from all faculty while 45% (n = 40) required them only from principal investigators or those conducting research. Nineteen percent of policies (n = 17) specified limits on faculty financial interests in corporate sponsors of research, 12% (n = 11) specified limits on permissible delays in publication, and 4% (n = 4) prohibited student involvement in work sponsored by a company in which the faculty mentor had a financial interest. CONCLUSIONS: Most policies on conflict of interest in our sample of major research institutions in the United States lack specificity about the kinds of relationships with industry that are permitted or prohibited. Wide variation in management of conflicts of interest among institutions may cause unnecessary confusion among potential industrial partners or competition among universities for corporate sponsorship that could erode academic standards. It is in the long-term interest of institutions to develop widely agreed-on, clear, specific, and credible policies on conflicts of interest. JAMA. 2000;284:2203-2208.

Biomedical Research↗
PubMed2005

University-based science and biotechnology products: defining the boundaries of intellectual property.

The pharmaceutical and biotechnology industries have long relied on patenting as the primary means of allocating ownership and control over new discoveries. Yet, patent protection is a double-edged sword that has major implications for the future of innovation in biomedical science in the United States. Excessive "upstream" patenting of genes and molecular targets could hinder further research by creating a need for expensive and inefficient cross-licensing. However, limiting such basic science patenting could allow private entities to use the results of years of costly publicly funded research to produce and market lucrative products without compensating university- or public sector-based innovators. Academic and other nonprofit research centers would, therefore, be deprived of revenue for pursuing novel therapeutics or other seminal research work that may not be patentable. Recent court cases illustrate the inherent conflicts in allocating ownership and control of basic biomedical discoveries. Several options exist to avoid the complex problems of overlapping basic science patents while still rewarding pivotal discoveries and encouraging further innovation. These include establishing basic science patent pools and mandating arbitration arrangements that would assign credit and royalties for biotechnology innovations that depend on prior research that was performed, financed, or both in the public sector.

Academic Medical Centers↗
PubMed1996

Encouraging small businesses to adopt effective technologies to prevent exposure to health hazards.

Small businesses are heterogeneous and the prospects are low for direct OSHA inspection and enforcement. Opportunities are explored to encourage voluntary adoption of new technology to reduce workplace exposures. The case of radiator repair shops is used in this paper to illustrate an approach to the dissemination of control technology to small businesses that will encourage these companies to adopt controls. Several behavioral theories are applied to the case.

Automobiles↗